Human APOE variants in Alzheimer’s Disease and type III hyperlipoproteinemia: insights from Long-Evans knock-in rat models with humanized App and APOE

Human APOE variants in Alzheimer’s Disease and type III hyperlipoproteinemia: insights from Long-Evans knock-in rat models with humanized App and APOE | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Human APOE variants in Alzheimer’s Disease and type III hyperlipoproteinemia: insights from Long-Evans knock-in rat models with humanized App and APOE Metin Yesiltepe, Tao Yin, Marc Tambini, Hanmei Bao, Meixia Pan, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4789248/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 27 Sep, 2024 Read the published version in Cell Communication and Signaling → Version 1 posted 10 You are reading this latest preprint version Abstract APOE is a major genetic factor in late-onset Alzheimer's disease (LOAD), with APOE4 significantly increasing risk, APOE3 acting as a neutral isoform, and APOE2 offering protective effects. The primary hypothesis links APOE isoforms to LOAD through their impact on Aβ production and deposition, which is thought to be related to their effects on lipid metabolism. Specifically, APOE4 enhances Aβ production and accumulation in amyloid plaques more than APOE3. In contrast, APOE3-Aβ complexes, which promote Aβ clearance and reduce Aβ aggregation, are approximately 20 times more prevalent than APOE4-Aβ complexes, highlighting differences in their functional interactions. APOE is also important in lipid metabolism, affecting both peripheral and central systems. It is involved in the metabolism of lipoproteins and plays a key role in triglyceride and cholesterol regulation. APOE2 is notably associated with Hyperlipoproteinemia type III (HLP), which is characterized by mixed hypercholesterolemia and hypertriglyceridemia due to impaired binding to Low Density Lipoproteins receptors. To explore the impact of human APOE isoforms on lipid metabolism and LOAD, we developed Long-Evans rats with the rat Apoe gene replaced by human APOE2 , APOE3 , or APOE4 . These rats were crossed with those carrying a humanized App allele, enabling the expression of human Aβ, which is more aggregation-prone than rodent Aβ. This model offers a significant advancement for studying APOE-Aβ interactions. We found that APOE2 hAβ rats had the highest levels of APOE in serum and brain, with no significant transcriptional differences among isoforms, suggesting variations in protein translation or stability. Elevated Aβ43 levels in male APOE4 hAβ rats compared to APOE2 hAβ rats highlight the model’s utility for amyloid pathology studies. Additionally, a lipidomic analysis of 222 lipid molecular species in serum samples showed that APOE2 hAβ rats displayed elevated triglycerides and cholesterol, making them a valuable model for studying HLP. These rats also exhibited elevated levels of phosphatidylglycerol, phosphatidylserine, phosphatidylethanolamine, sphingomyelin, and lysophosphatidylcholine. Minimal differences in lipid profiles between APOE3 hAβ and APOE4 hAβ rats reflect findings from mouse models. Future studies will include comprehensive lipidomic analyses in various CNS regions to further validate these models and explore the effects of APOE isoforms on lipid metabolism in relation to AD pathology. Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 INTRODUCTION The Apolipoprotein E ( APOE ) gene plays an important role in both lipid metabolism and neurological functions. In humans, there are three forms of APOE : APOE2 , APOE3 , and APOE4 . These forms are distinguished by the presence of either arginine or cysteine residues at positions 112 and 158. APOE4 , the ancestral form of APOE , is has arginine residues at positions 112 and 158 [ 1 ] and is present in about 12% of the population. APOE3 is the most common variant in contemporary human populations with a prevalence of 69–82% depending on geolocation [ 2 ]. APOE3 emerged later through an arginine-to-cysteine substitution at position 112 of APOE4 , dating back approximately 200,000 years based on time-depth analysis and natural selection assumptions [ 3 ]. The fact that APOE3 is more recent compared to APOE4 , yet significantly more prevalent, implies a positive selection pressure favoring APOE3 . APOE2 , the least common variant, originated around 80,000 years ago from an arginine-to-cysteine substitution at position 158 of the APOE3 gene. APOE is predominantly expressed in hepatocytes, macrophages, and astrocytes, a type of glial cell in the brain [ 4 , 5 ]. APOE is genetically linked to late-onset sporadic Alzheimer’s disease (LOAD), with APOE4 being a significant risk factor, APOE3 considered the "neutral" allele, and APOE2 exerting a protective role against LOAD [ 6 , 7 ]. The prevailing hypothesis linking APOE to LOAD suggests that Aβ production and deposition vary with APOE isoforms. Aβ, the primary component of amyloid plaques that characterize AD pathology, originates from sequential cleavage of the Amyloid-β Precursor Protein (APP) by β- and γ-secretases (known as the amyloidogenic processing pathway). APOE isoforms likely influence Aβ production by modulating lipid metabolism and the lipid composition of cellular membranes, with APOE4 promoting amyloidogenic APP cleavage and Aβ production more than APOE3 [ 8 – 10 ]. Moreover, APOE facilitates Aβ clearance and inhibits its aggregation through the formation of APOE-Aβ complexes, with APOE3-Aβ complexes being approximately 20 times more prevalent than APOE4-Aβ complexes [ 11 , 12 ]. Additionally, APOE isoforms may impact neurite growth differently [ 13 ] and affect neuronal cell survival in an isoform-dependent manner [ 14 ]. To investigate the impact of human APOE isoforms on LOAD, we developed Long-Evans rats with the rat Apoe gene replaced by human APOE2 , APOE3 , or APOE4 variants. Given that the pathogenic mechanisms linked to APOE4 may significantly influence LOAD pathogenesis through its effects on Aβ production and metabolism via direct interaction with Aβ, we crossed these human APOE replacement rats with rats carrying a humanized App allele ( App h allele) [ 13 ]. Humanization of App targeted the Aβ region via humanization of the three amino acid differences between rodent and human Aβ. Therefore, these rat models, designated as APOE2 hAβ , APOE3 hAβ and APOE4 hAβ , are expected to physiologically express both human APOE isoforms and human Aβ. These amino acids differences in Aβ may be crucial as human Aβ is more prone to aggregation compared to its rodent counterpart. In addition, rodent Aβ may not interact as effectively with human APOE as human Aβ does, which could reduce the ability to accurately study pathogenic and physiological mechanisms based on APOE-Aβ interactions. This setup enables the investigation of mechanisms underlying the interaction between human APOE and human Aβ. These new rat models represent an advancement compared to earlier rodent models carrying human APOE variants, as they now allow for a comprehensive exploration of the interplay between human APOE and human Aβ that was not possible in these earlier models [ 14 – 16 ]. APOE also plays a crucial role in lipid metabolism, both peripherally and in the central nervous system. Plasma APOE circulates in the bloodstream and is associated with chylomicron, very low-density lipoprotein (VLDL), and high-density lipoprotein (HDL) particles, playing a crucial role in their metabolism. Chylomicrons, which are derived from the intestine, and VLDL particles, which come from the liver, are lipolyzed in the bloodstream by an enzyme called lipoprotein lipase (LPL). APOE on the remnant lipoprotein particles binds to low-density lipoprotein (LDL) receptors, LDL receptor-related proteins (LRP), and heparan sulfate proteoglycans (HSPG) on the surface of liver cells [ 17 ]. These remnant particles are then endocytosed by the liver cells and removed from the bloodstream. Some VLDL remnants are cleared quickly, while others undergo further lipolysis and are gradually converted into intermediate-density lipoprotein (IDL) and eventually into LDL [ 18 ]. LDL particles do not contain APOE, and their removal from the bloodstream is facilitated by the binding of another protein, APOB, to the LDL receptor (LDLR) [ 19 ]. APOE is also crucial for the production of VLDL particles. Its expression within liver cells promotes the assembly and secretion of VLDL particles. Optimal expression of APOE is essential for the normal metabolism of triglyceride (TG)-rich lipoproteins. However, overexpression or accumulation of APOE stimulates the production of VLDL triglycerides [ 20 ], leading to hypertriglyceridemia. Additionally, an excess of APOE on VLDL particles can hinder their lipolysis [ 21 ], resulting in elevated plasma triglyceride levels. The critical role of APOE in lipid metabolism is underscored by evidence showing that APOE2 homozygosity can lead to Hyperlipoproteinemia type III (HLP), characterized by mixed hypercholesterolemia and hypertriglyceridemia [ 22 – 26 ]. This is attributed to the fact that the cysteine change at position 158 in APOE2, near the LDLR binding region, hinders APOE2 binding to the LDLR [ 17 , 27 ]. In addition, APOE4 is linked to hypercholesterolemia and increased risk of cardiovascular disorders (CVD) [ 7 , 28 ], although the specific underlying mechanism remains unclear [ 26 , 29 ]. In this study, we provide an initial characterization of APOE2 hAβ , APOE3 hAβ and APOE4 hAβ rats, focusing on early effects of these APOE isoforms on APP processing and plasma lipidomics as an indicator of their influence on peripheral lipid metabolism. While the lipidomics studies do not directly focus on the LOAD- APOE link, our findings may shed light on how APOE isoforms modulate LOAD risk, potentially through effects on lipid metabolism pathways. MATERIALS AND METHODS Animals. All experiments were done according to policies on the care and use of laboratory animals of the Ethical Guidelines for Treatment of Laboratory Animals of the NIH. Relevant protocols were approved by the Rutgers Institutional Animal Care and Use Committee (Protocol #201702513). All efforts were made to minimize animal suffering and reduce the number of rats used. Generation of Long-Evans Rat Models Expressing Human APOE Variants with Human APP Gene . gRNAs targeting vectors and the donor vector, which is flanked by homologous arms, were constructed and confirmed by sequencing (designed as shown below). The vectors’ sequences are available at the indicated links. gRNA designed: gRNA1 (matches forward strand of gene): AATCACAACTGGGAAGATGAAGG gRNA2 (Cas9_D10A) (matches reverse strand of gene): TTCATCTTCCCAGTTGTGATTGG gRNA3 (Cas9_D10A) (matches forward strand of gene): AATCACAACTGGGAAGATGAAGG Links the the gRNA and donor vectors on VectorBuilder: gRNA1: https://www.vectorbuilder.com/vector/VB171212-1056smp.html gRNA2 (Cas9_D10A): https://www.vectorbuilder.com/vector/VB171212-1329ath.html gRNA3 (Cas9_D10A): https://www.vectorbuilder.com/vector/VB171212-1058ubb.html Donor vector for APOE2 plus SV40 late polyadenylation site flanked by homologous arms: https://www.vectorbuilder.com/vector/VB171225-1186ayg.html Donor vector for APOE3 plus SV40 late polyadenylation site flanked by homologous arms: https://www.vectorbuilder.com/vector/VB171225-1189pmt.html Donor vector for APOE4 plus SV40 late polyadenylation site flanked by homologous arms: https://www.vectorbuilder.com/vector/VB171225-1191dnt.html Cas9 mRNA, gRNA generated by in vitro transcription, and oligo donor were co-injected into fertilized eggs to generate knock-in (KI) rats. The PCR primer pairs used to determine correct KI insertion and the integrity of the APOE sequences' insertions were as follows (5' to 3'): SPF1: CACCCGTGGCAGAGGAATCAAC x SPR1: TTCTAGCGGGTCGGGTCGTCT SPF2: CCAACCCCCTTCATCTGGATTTC x SPR2: AAAGGTCAGAATTAGGGTGGGAGG’ KI-1-F:TGCTCTATTGTGGAGATGTTTGTGATG x KI-1-R: GTGTGGGGGTGATGGAGAATAAAGATC KI-2-F: CCACACCCGACTAACTTTTTTGTATTTTC x KI-2-R: TCAACTCCTTCATGGTCTCGTCCATC KI-3-F: GCCTCCTAGCTCCTTCTTCGTCTCTG x KI-3-R: CAGGCGTATCTGCTGGGCCTG KI-4-F: TAAGCGGCTCCTCCGCGATG x KI-4-R: AGCAGAATCGCTTGAACCCAAGAG KI-5-F: CCTCAGTTTCTCTTTCTGCCCACATA x KI-5-R: TATTATGGATAGGGAAAGACAAGGCC The primers used for the Southern blot analysis were: 5’ arm Probe: F: CCAAGATTATACATCCGGCAACCG x R: GGCTGGAGGCTTAAATGGAAATAGG 3’ arm Probe: F: TGTTGGTCCCATTGCTGACAGGTA x R: AAGCAACAGTGCGTCTGGAAGTCAG To generate APOE2 hAβ , APOE3 hAβ and APOE4 hAβ rats, we doubly crossed humanized APOE rats described above with App h/h rats that carries App genes with the humanized Aβ sequence. The App h allele enables the physiological production of human Aβ instead of rodent Aβ from the endogenous rat App gene [ 30 – 36 ]. Protein preparation. These procedures were performed as previously described [ 37 ]. Briefly, the rats were first put under anesthesia using isoflurane, followed by perfusion through intracardiac catheterization using ice-cold PBS. Brains were extracted and homogenized with a glass-teflon homogenizer in 250 mM Sucrose, 20 mM Tris-base pH 7.4, 1 mM EDTA, 1 mM EGTA plus protease and phosphatase inhibitors (Thermo Scientific). All steps were carried out on the ice. Homogenates were solubilized with 1% NP-40 for 30 min rotating and spun at 20,000 g for 10 min. Supernatants were collected and protein content was quantified using the Bradford method. Quantitative RT-PCR . Total brain RNA was extracted using the RNeasy RNA Isolation kit (Qiagen) and converted to cDNA using the High-Capacity cDNA Reverse Transcription Kit (Thermo Fisher) with oligo dT priming. For each reaction, 50 ng of cDNA, TaqMan™ Fast Advanced Master Mix (Thermo Fisher 4444556), and the appropriate TaqMan probes (Thermo Fisher) were used. Real-time PCR was conducted on a QuantStudio 6 Flex Real-Time PCR System (Thermo Fisher). Relative RNA quantification was performed using LinRegPCR software (hartfaalcentrum.nl). The rat Apoe transcript was detected using probe Rn00593680_m1 targeting exon junction 3–4, while human APOE was detected using probe Hs00171168_m1 for the same exon junction. Amplification data were normalized to rat Gapdh expression, assessed using probe Rn01775763_g1. ELISA. For analysis of human APOE, Aβ38, Aβ40, Aβ42, sAPPα and sAPPβSw, the following Meso Scale Discovery kits were used: levels of APOE were measured with R-PLEX Human ApoE Assay (K151AMLR) (serum samples were diluted 1:20,000), Aβ38, Aβ40, and Aβ42 were measured with V-PLEX Plus Aβ Peptide Panel 1 6E10 (K15200G); sAPPα and sAPPβ were measured with sAPPα/sAPPβ kit (K15120E). Measurements were performed according to the manufacturer’s recommendations. Plates were read on a MESO QuickPlex SQ 120. For analysis of Aβ43, IBL Human Amyloidβ (1–43) (FL) Assay Kit (27710) was used according to the manufacturer’s recommendations. Western blots (WB). WB were performed as follows: proteins were diluted with PBS and LDS Sample Buffer (Invitrogen NP0007) containing 10% β-mercaptoethanol, and 4.5M urea to a concentration of 1 µg/µl. Samples were loaded onto a 4–12% Bis-Tris polyacrylamide gel (Biorad 3450125) and transferred onto nitrocellulose membranes at 25 V for 7 minutes using the Trans-Blot Turbo system (Biorad). Blotting efficiency was confirmed by red Ponceau staining of the membranes. Membranes were blocked for 45 minutes in 5% milk (Biorad 1706404), followed by extensive washing in PBS/Tween20-0.05%. Primary antibodies (anti-APOE Rabbit mAb, Cell Signaling Technology, 10197SF; anti-APP (Y188) Rabbit mAb, Abcam, Ab32136; anti-GAPDH Rabbit mAb, Sigma, g9545; anti-PSD95, Cell Signaling Technology, 3450) were applied overnight at 4°C. After washing three times for 10 minutes each with PBS/Tween20-0.05%, membranes were incubated with a mixture of HRP-conjugated anti-rabbit secondary antibodies (Southern Biotech, OB405005 and Cell Signaling Technology, 7074) diluted 1:1,000 in 5% milk for 45 minutes at room temperature with shaking. Blots were developed using Clarity Western ECL reagent (Bio-rad 1705061) and visualized on a ChemiDoc MP Imaging System (Bio-Rad). For WB quantifications, signal intensities were analyzed with Image Lab software (Bio-Rad). Blood glucose and lipid profile measurements. Blood glucose levels were measured using the ACCU-CHEK Guide Me system (Roche, mg/dL). For the blood lipid profile, including total cholesterol (TC, range 100–450 mg/dL), HDL (range 25–95 mg/dL), LDL (calculated), and TG (range 45–450 mg/dL), a CURO L7 Lipid Analyzer (CUROfit, CA) cholesterol home test kit was utilized. Briefly, blood was collected via cardiac puncture from 80-day-old rats (with a sample size of 4 per sex per genotype). 35 µL of fresh blood was applied to a lipid profile test strip and measured after 3 minutes. Measurements falling below the specified range were rounded up to the closest integer within the lower end, while those above the range were rounded down to the closest integer within the upper end. LDL levels were calculated using the Friedewald equation: LDL (mg/dL) = TC - HDL - (TG/5). Lipidomic analysis. Serum for lipidomic analysis was collected from the same rats used for blood glucose and lipid profile measurements. Blood was drawn into serum separator tubes (BD Becton Dickinson vacutainers, SSTTM) and left to incubate at room temperature for 30 minutes. The tubes were then centrifuged at 2000×g for 10 minutes to separate the serum, which was subsequently stored at -80°C. For lipidomic analysis, the protein content of the thawed serum samples was quantified using a BCA protein assay kit (Pierce, Rockford, IL, USA). A mixture of approximately 20 internal standards was added to serum samples based on the protein content for quantification of individual lipid molecular species as previously described [ 38 ]. The extraction of lipids was carried out using a modified Bligh and Dyer extraction method, as previously described [ 39 ]. Multi-dimensional mass spectrometry (MDMS)-based shotgun lipidomics (MDMS-SL) was performed using electrospray ionization mass spectrometry (ESI/MS) to measure individual lipid molecular species [ 40 – 42 ]. Instrumentation utilized was a triple-quadruple mass spectrometer (Thermo Scientific TSQ Altis, San Jose, CA, USA) equipped with a NanoMate device (Advion Bioscience Ltd., Ithaca, NY, USA). Xcalibur system software was utilized for this process. Data processing included several steps such as ion peak selection, baseline correction, data transfer, peak intensity comparison, 13C deisotoping, and quantitation. These steps were conducted using a custom-programmed Microsoft Excel macro, as previously described [ 43 ]. The concentrations of the total lipid class populations were calculated by summing the individually detected analytes that belonged to the class. Statistical analysis. Data were analyzed using GraphPad Prism software and expressed as mean ± SEM. Statistical tests used to evaluate significance and statistical data are shown in Figure legends and Results section. Significant differences were accepted at P < 0.05. RESULTS Generation of Long-Evans rat models expressing either human APOE2 , APOE3 or APOE4 variant genes and the humanized App rat gene. The rat Apoe gene, with GenBank accession number NM_138828.3 and Ensembl ID ENSRNOG00000018454, is on rat chromosome 1. The gene comprises 4 exons, with the ATG start codon located in exon 2 and the TGA stop codon located in exon 4. Long-Evans rat models expressing human APOE2 , APOE3 , and APOE4 variants were separately generated using CRISPR/Cas-mediated genome engineering. To achieve this, the ATG start codon in exon 2 of the rat Apoe gene was replaced with coding sequences for human APOE2 , APOE3 , or APOE4 (Fig. 1 a). These human APOE coding sequences were linked to the SV40 late polyadenylation site. By employing the gene manipulation strategy as described above, the expression of human APOE2 , APOE3 , and APOE4 variants, which replaces the expression of the rat Apoe gene, is controlled using the regulatory elements of the rat Apoe gene. This allows for the accurate and specific regulation of the human APOE variants' expression in the Long-Evans rat models. To confirm CRISPR-induced mutations, the resulting pups underwent genotyping by PCR, followed by sequence analysis. PCR was initially performed using primers SPF1 x SPR1 and SPF2 x SPR2 to identify founder rats (F0). Based on this screening, rat 10 was selected as an F0 rat for APOE2 , rat 26 as an F0 for APOE3 , and rats 6 and 7 as F0s for APOE4 (Fig. 1 b). Subsequently, the same PCR method was used to identify and designate certain offspring of these F0 rats as APOE2 F1s (rats 1, 5, 6, 7, and 9), APOE3 F1s (rats 58, 62, and 63), and APOE4 F1s (rats 59, 67, 76, and 87) based on their respective parentage (Fig. 1 b). The selection of F0 and F1 rats was further validated by PCR using 5 primer pairs (KI-1-F x KI-1-R, KI-2-F x KI-2-R, KI-3-F x KI-3-R, KI-4-F x KI-4-R, and KI-5-F x KI-5-R) followed by sequencing to confirm the genotyping results. The accurate gene targeting in F1 animals was verified through Southern blot analysis of the tail DNA samples. The Southern blot analysis strategy is depicted in Fig. 1 c. The results demonstrated that all F1 rats analyzed expressed both the rat Apoe allele and the human APOE knock-in (KI) allele in a 1:1 ratio (Fig. 1 d). F1 rats were crossed to Long Evans for 5 generations. The probability that F5 rats carry unidentified off-target insertions/mutations (except those that may be on Chr. 1) is ~ 1.5625%. APOE isoforms have been associated with the risk of LOAD, with APOE4 increasing the risk and APOE2 reducing it. Considering the important role of APP in LOAD pathogenesis and the interplay between APP, its metabolic product Aβ, and APOE isoforms, we crossed APOE2 , APOE3 , and APOE4 rats with animals carrying a rat App allele humanized specifically in the Aβ region ( App h/h rats). The resulting progeny, double heterozygous for APOE and App h alleles, were further bred to generate rats homozygous for humanized APOE and App h , thereby exclusively producing human Aβ species in a physiological manner. These models were designated as APOE2 hAβ , APOE3 hAβ and APOE4 hAβ . APP is expressed in virtually all types of cells, suggesting broader functions beyond the central nervous system (CNS). Studying these double humanized rats could offer significant advantages, particularly in exploring how human APOE's systemic functions related to lipid metabolism are influenced by its interactions with human Aβ. Utilizing Apoe hAβ , APOE2 hAβ , APOE3 hAβ and APOE4 hAβ rats for this lipid metabolism study may yield valuable insights into their potential interactions without apparent drawbacks. Serum and brain APOE levels are highest in APOE2 hAβ rats. To characterize these models, we first measured human APOE levels in blood serum and brain. Since Apoe hAβ rats have rat Apoe , we excluded them from this analysis and used them as negative control. Serum APOE levels were highest in APOE2 hAβ rats in both males and females compared to APOE3 hAβ and APOE4 hAβ rats (Fig. 2 a) (2-way ANOVA summary; sex factor, F (1, 18) = 0.2930, P = 0.5949; genotype factor, F (2, 18) = 95.68, P < 0.0001; sex × genotype interaction, F (2, 18) = 0.5054, P = 0.6116. For males APOE2 hAβ vs. APOE3 hAβ p < 0.0001****; APOE2 hAβ vs. APOE4 hAβ p < 0.0001****; APOE3 hAβ vs. APOE4 hAβ p = 0.6589. For females APOE2 hAβ vs. APOE3 hAβ p < 0.0001****; APOE2 hAβ vs. APOE4 hAβ p < 0.0001****; APOE3 hAβ vs. APOE4 hAβ p = 0.9979). Remarkably, this pattern of APOE isoform levels mirrors that found in human serum [ 44 ]. Brain APOE levels measured by ELISA were also the highest in APOE2 hAβ rats compared to APOE3 hAβ and APOE4 hAβ rats in both sexes (Fig. 2 b) (2-way ANOVA summary; sex factor, F (1, 25) = 15.13, P = 0.0007; genotype factor, F (2, 25) = 68.7, P < 0.0001; sex × genotype interaction, F (2, 25) = 5.765, P = 0.0087. For males APOE2 hAβ vs. APOE3 hAβ p = 0.0022**; APOE2 hAβ vs. APOE4 hAβ p < 0.0001****; APOE3 hAβ vs. APOE4 hAβ p = 0.8481. For females APOE2 hAβ vs. APOE3 hAβ p < 0.0001****; APOE2 hAβ vs. APOE4 hAβ p < 0.0001****; APOE3 hAβ vs. APOE4 hAβ p = 0.4170.). Additionally, in the brains of APOE2 hAβ females, APOE level was higher than male APOE2 hAβ rats (female vs male p < 0.0001****). The ELISA results were confirmed by WB analysis of brain APOE levels (Fig. 2 c) (2-way ANOVA summary; sex factor, F (1, 25) = 0.6498, P = 0.4278; genotype factor, F (2, 25) = 75.87, P < 0.0001; sex × genotype interaction, F (2, 25) = 2.743, P = 0.0837. For males APOE2 hAβ vs. APOE3 hAβ p = 0.0013**; APOE2 hAβ vs. APOE4 hAβ p < 0.0001****; APOE3 hAβ vs. APOE4 hAβ p = 0.2056. For females APOE2 hAβ vs. APOE3 hAβ p < 0.0001****; APOE2 hAβ vs. APOE4 hAβ p < 0.0001****; APOE3 hAβ vs. APOE4 hAβ p = 0.90380). To investigate whether the increase in APOE2 protein levels is due to enhanced transcription, we measured mRNA levels of human APOE2 , APOE3 , and APOE4 in APOE2 hAβ , APOE3 hAβ and APOE4 hAβ rats. Overall, these mRNA levels were comparable, with the notable exception of a decrease in APOE2 mRNA levels in male APOE2 hAβ compared to male APOE3 hAβ rats (Fig. 2 d), despite APOE2 protein levels being significantly higher than APOE3 levels in male rats (Fig. 2 a and 2 b). Therefore, differences in protein expression are likely attributable to translational and/or protein stability variances. Sex- and APOE isoform-dependent variations in brain APP metabolites . APP is a substrate of several proteases. α-Secretase cleaves APP to produce a soluble ectodomain (sAPPα) and a membrane-bound C-terminal fragment (αCTF). Alternatively, β-secretase cleaves APP to generate the soluble sAPPβ ectodomain and the membrane-tethered βCTF stub. Subsequent cleavage of βCTF by γ-secretase leads to the production of Aβ peptides and the short intracellular domain of APP [ 45 ]. APOE modulates APP metabolism in an isoform-dependent manner through distinct mechanisms: 1) influencing APP processing via effects on lipid metabolism [ 8 – 10 ], and 2) modulating Aβ clearance and aggregation via the formation of APOE-Aβ complexes [ 11 , 12 ]. Consequently, we investigated APP metabolites in the CNS of Apoe hAβ , APOE2 hAβ , APOE3 hAβ and APOE4 hAβ rats. ELISA experiments showed no significant differences in Aβ38 levels among humanized APOE variants (Fig. 3 a; 2 -way ANOVA summary; sex factor, F (1, 34) = 1.717, P = 0.1989; genotype factor, F (3, 34) = 1.978, P = 0.1358; sex x genotype interaction, F (3, 34) = 1.978, P = 0.1358). In female rats, Aβ40 levels were higher in Apoe hAβ compared to APOE4 hAβ rats (Fig. 3 b) (2-way ANOVA summary; sex factor, F (1, 34) = 10.53, P = 0.0026; genotype factor, F (3, 34) = 5.083, P = 0.0051; sex x genotype interaction, F (3, 34) = 1.677, P = 0.1902. For females Apoe hAβ vs. APOE4 hAβ p = 0.0238*). There was no significant difference in Aβ42 levels across the humanized APOE variants (Fig. 3 c; 2 -way ANOVA summary; sex factor, F (1, 34) = 0.449, P = 0.5073; genotype factor, F (3, 34) = 2.009, P = 0.1311; sex x genotype interaction, F (3, 34) = 1.901, P = 0.1480). However, the levels of Aβ43, a determining factor in the onset of pathological amyloid deposition [ 37 ], were higher in APOE4 hAβ males compared to APOE2 hAβ males (Fig. 3 d) (2-way ANOVA summary; sex factor, F (1, 34) = 1.840, P = 0.1838; genotype factor, F (3, 34) = 2.781, P = 0.0558; sex x genotype interaction, F (3, 34) = 2.735, P = 0.0587. For males APOE2 hAβ vs. APOE4 hAβ p = 0.0146*). The levels of sAPPα showed gender-specific variations (Fig. 3 e; 2 -way ANOVA summary; sex factor, F (1, 34) = 39.75, P < 0.0001; genotype factor, F (3, 34) = 1.152, P = 0.3424; sex x genotype interaction, F (3, 34) = 3.222, P = 0.0347). Apoe hAβ and APOE2 hAβ female rats exhibited higher sAPPα levels compared to males of the same genotype (Fig. 3 e) (for Apoe hAβ male vs female p = 0.0195*; For APOE2 hAβ male vs female p < 0.0001****). Finally, sAPPβ levels were higher in APOE2 hAβ females compared to all other female groups and APOE2 hAβ male rats (Fig. 3 f) (2-way ANOVA summary; : sex factor, F (1, 34) = 25.04, P < 0.0001; genotype factor, F (3, 34) = 6.863, P = 0.0010; sex x genotype interaction, F (3, 34) = 7.099, P = 0.0008. For females Apoe hAβ vs. APOE2 hAβ p = 0.0059**; APOE2 hAβ vs. APOE3 hAβ p = 0.0023**; APOE2 hAβ vs. APOE4 hAβ p < 0.0001****. For APOE2 hAβ male vs. female p < 0.0001****). Levels of APP, αCTF and βCTF were quantified by WB, as shown in Fig. 3 g. APP levels were similar except for APOE2 hAβ males which showed significant lower levels compared to APOE2 hAβ females (Fig. 3 h)(2-way ANOVA summary; sex factor, F (1, 25) = 12.08, P = 0.0019; genotype factor, F (2, 25) = 0.0983, P = 0.9067; sex x genotype interaction, F (2, 25) = 0.2287, P = 0.7972. Males APOE2 hAβ vs. females APOE2 hAβ p = 0.0106*). In contrast, αCTF levels were significantly higher in APOE4 hAβ males compared to APOE4 hAβ females (Fig. 3 h)(2-way ANOVA summary; sex factor, F (1, 25) = 8.588, P = 0.0071; genotype factor, F (2, 25) = 0.7281, P = 0.4928; sex x genotype interaction, F (2, 25) = 2.612, P = 0.0933. Males APOE4 hAβ vs. females APOE4 hAβ p = 0.0029**). Finally, βCTF showed a more complex pattern (Figure h) (2-way ANOVA summary; sex factor, F (1, 25) = 0.6420, P = 0.4306; genotype factor, F (2, 25) = 2.157, P = 0.1368; sex x genotype interaction, F (2, 25) = 8.787, P = 0.0013). APOE2 hAβ males showed significantly lower levels compared to APOE4 hAβ males (males, APOE2 hAβ vs. APOE4 hAβ p = 0.0006***) and APOE2 hAβ females ( APOE2 hAβ male vs. female p = 0.0092**), and APOE4 hAβ males showed significantly higher levels compared to APOE4 hAβ females ( APOE4 hAβ male vs. female p = 0.0065**), Overall, these findings demonstrated distinct regulation of APP metabolites in relation to APOE variants, with notable sex differences. Increased triglyceride and cholesterol levels in APOE2 hAβ rats. Next, we measured blood glucose, triglyceride, cholesterol, LDL and HDL levels using home test kits while collecting blood via cardiac puncture, to assess the metabolic health of our rats after a 16-hour fasting period. No significant variance was observed in the levels of blood glucose (Fig. 4 a; 2 -way ANOVA summary; sex factor, F (1, 24) = 10.40, P = 0.0036; genotype factor, F (3, 24) = 0.6423, P = 0.5953; sex × genotype interaction, F (3, 24) = 1.389, P = 0.2701). In contrast, both blood triglyceride (Fig. 4 b; 2 -way ANOVA summary; sex factor, F (1, 24) = 1.697, P = 0.2051; genotype factor, F (3, 24) = 35.47, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.1583, P = 0.9233. For males Apoe hAβ vs. APOE2 hAβ p 0.9999. For females Apoe hAβ vs. APOE2 hAβ p < 0.0001****; APOE2 hAβ vs. APOE3 hAβ p = 0.0019**; APOE2 hAβ vs. APOE4 hAβ p = 0.0002***; APOE3 hAβ vs. APOE4 hAβ p = 0.9848) and cholesterol levels (Fig. 4 c; 2 -way ANOVA summary; sex factor, F (1, 24) = 1.798, P = 0.1925; genotype factor, F (3, 24) = 35.47, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.5164, P = 0.6749. For males Apoe hAβ vs. APOE2 hAβ p = 0.0006***; APOE2 hAβ vs. APOE3 hAβ p = 0.0037**; APOE2 hAβ vs. APOE4 hAβ p = 0.0102*; APOE3 hAβ vs. APOE4 hAβ p = 0.9998. For females Apoe hAβ vs. APOE2 hAβ p 0.9999) were significantly higher in APOE2 hAβ compared to Apoe hAβ , APOE3 hAβ and APOE4 hAβ rats. In addition, the serum of APOE2 hAβ rats exhibited a turbid appearance (Fig. 4 g) reminiscent of human cases of type III HLP [ 22 ]. This observation, coupled with evident mixed hypercholesterolemia and hypertriglyceridemia, supports the characterization of APOE2 hAβ rats as a model for type III HLP, a condition prevalent in a significant fraction of individuals homozygous for APOE2 [ 17 , 23 – 26 ]. HDL levels were similar in all humanized APOE groups, but lower compared to Apoe hAβ group (Fig. 4 d) (2-way ANOVA summary; sex factor, F (1, 24) = 0.8298, P = 0.3714; genotype factor, F (3, 24) = 24.69, P < 0.0001; sex × genotype interaction, F (3, 24) = 2.178, P = 0.1168. For males Apoe hAβ vs. APOE2 hAβ p < 0.0001****; Apoe hAβ vs. APOE3 hAβ p = 0.0090**; Apoe hAβ vs. APOE4 hAβ p = 0.0020**. For females Apoe hAβ vs. APOE2 hAβ p = 0.0076**; Apoe hAβ vs. APOE3 hAβ p = 0.0065**; Apoe hAβ vs. APOE4 hAβ p = 0.0010***). Moreover, the level of LDL is similar in all humanized APOE groups. But LDL levels were higher in APOE2 hAβ rats compared to Apoe hAβ (Fig. 4 e) (2-way ANOVA summary; sex factor, F (1, 24) = 0.4476, P = 0.5098; genotype factor, F (3, 24) = 11.07, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.4133, P = 0.7450. For males Apoe hAβ vs. APOE2 hAβ p = 0.0163*. For females Apoe hAβ vs. APOE2 hAβ p = 0.0058**). As a result, the LDL/HDL ratio was much higher in male APOE2 hAβ compared to male Apoe hAβ and APOE3 hAβ rats, but there was no difference between groups in females (Fig. 4 f) (2-way ANOVA summary; sex factor, F (1, 24) = 0.0244, P = 0.8771; genotype factor, F (3, 24) = 9.385, P = 0.0003; sex × genotype interaction, F (3, 24) = 1.372, P = 0.2752. For males Apoe hAβ vs. APOE2 hAβ p = 0.0040**; APOE2 hAβ vs. APOE3 hAβ p = 0.0288*; APOE2 hAβ vs. APOE4 hAβ p = 0.0796). The LDL/HDL ratio is used to predict Coronary Heart Disease (CHD), with a higher ratio indicating a higher risk of CHD [ 46 ]. Sex- and APOE isoform-dependent variations in serum lipidomics. A comprehensive lipidomic analysis was conducted to measure 222 lipid molecular species in serum samples of Apoe hAβ , APOE2 hAβ , APOE3 hAβ and APOE4 hAβ rats, after scanning thousands of lipid molecular species. The lipid species included 3 phosphatidylglycerol (PG) species, 3 phosphatidylserine (PS) species, 21 phosphatidylethanolamine (PE) species, 13 plasmalogen PE (pPE) species, 12 lyso PE (LPE) species, 11 acylcarnitine (CAR) species, 6 phosphatidylinositol (PI) species, 20 sphingomyelin (SM) species, 31 phosphatidylcholine (PC) species, 4 plasmalogen PC (pPC) species, 12 lyso PC (LPC) species, 59 types of triacylglycerol (TAG) without deconvolution of individual molecular species, 11 types of fatty acyl chains in TAG (FA), total cholesterol, free cholesterol, and 14 types of cholesterol esters. The results revealed significant differences in levels of various lipid molecular species between the different APOE genotypes. Notably, APOE2 hAβ rats displayed markedly elevated levels of serum PG, PS, PE, pPE, SM, LPC, TAG, FA, total cholesterol, free cholesterol, and cholesterol esters compared to Apoe hAβ , APOE3 hAβ , and APOE4 hAβ rats in both males and females(PG is shown in Fig. 5 a; sex factor, F (1, 24) = 2.492, P = 0.1275; genotype factor, F (3, 24) = 125.3, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.1789, P = 0.9096. PS is shown in Fig. 5 b; sex factor, F (1, 24) = 0.3257, P = 0.5735; genotype factor, F (3, 24) = 260.1, P < 0.0001; sex × genotype interaction, F (3, 24) = 2.486, P = 0.0849. PE is shown in Fig. 5 c; sex factor, F (1, 24) = 1.382, P = 0.2512; genotype factor, F (3, 24) = 63.46, P < 0.0001; sex × genotype interaction, F (3, 24) = 2.978, P = 0.0515. pPE is shown in Fig. 5 d; sex factor, F (1, 24) = 2.117, P = 0.1586; genotype factor, F (3, 24) = 69.44, P < 0.0001; sex × genotype interaction, F (3, 24) = 2.014, P = 0.1388 SM is shown in Fig. 5 h; sex factor, F (1, 24) = 0.0049, P = 0.9444; genotype factor, F (3, 24) = 82.17, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.08844, P = 0.9657. LPC is shown in Fig. 5 k; sex factor, F (1, 24) = 1.024, P = 0.3217; genotype factor, F (3, 24) = 28.38, P < 0.0001; sex × genotype interaction, F (3, 24) = 1.421, P = 0.261. TAG is shown in Fig. 5 l; sex factor, F (1, 24) = 2.519, P = 0.1256; genotype factor, F (3, 24) = 70.46, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.2196, P = 0.8818. FA is shown in Fig. 5 m; sex factor, F (1, 24) = 2.318, P = 0.141; genotype factor, F (3, 24) = 71.29, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.2084, P = 0.8896. Cholesterol-Total is shown in Fig. 5 n; sex factor, F (1, 24) = 1.158, P = 0.2927; genotype factor, F (3, 24) = 74.78, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.8491, P = 0.4807. Cholesterol-Free is shown in Fig. 5 o; sex factor, F (1, 24) = 0.0239, P = 0.3460; genotype factor, F (3, 24) = 127.6, P < 0.0001; sex × genotype interaction, F (3, 24) = 0.2969, P = 0.8273. Cholesterol-Ester is shown in Fig. 5 p; sex factor, F (1, 24) = 1.103, P = 0.3041; genotype factor, F (3, 24) = 45.94, P < 0.0001; sex × genotype interaction, F (3, 24) = 1.105, P = 0.3666.). This indicates a distinct lipid profile associated with the APOE2 hAβ genotype (Fig. 5 ) (see Table 1 for multiple comparisons). Table 1 Post hoc Tukey’s Analysis: p-Values for Multiple Comparisons of Various Lipids. PG PS PE pPE LPE CAR PI SM PC pPC LPC TAG FA ♂: Apoe hAβ vs.♂: APOE2 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0031 0.5878 0.4875 < 0.0001 < 0.0001 < 0.0001 0.0235 < 0.0001 0.9999 0.9901 > 0.9999 0.6910 0.9265 0.5522 0.9434 > 0.9999 0.7490 0.7492 0.8712 0.8751 ♂: Apoe hAβ vs. ♂: APOE4 hAβ > 0.9999 > 0.9999 0.9388 0.9998 0.9696 0.1423 0.0392 0.7406 0.9667 0.3953 0.0893 0.9918 0.9928 ♂: Apoe hAβ vs. ♀: Apoe hAβ 0.8432 0.9904 0.4333 0.5589 0.0424 0.0050 0.9999 0.0033 0.9526 0.9630 > 0.9999 > 0.9999 ♂: Apoe hAβ vs. ♀: APOE2 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0007 < 0.0001 < 0.0001 < 0.0001 0.0004 0.6346 < 0.0001 0.9999 0.7642 0.9941 0.2140 0.3535 0.1697 0.1800 ♂: Apoe hAβ vs. ♀: APOE4 hAβ > 0.9999 0.9988 0.9991 0.9352 0.9734 0.9999 0.9656 0.7112 0.6869 0.3542 0.0285 0.8855 0.9061 ♂: APOE2 hAβ vs. ♂: APOE3 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.1396 0.0861 0.0120 < 0.0001 < 0.0001 < 0.0001 0.0005 < 0.0001 < 0.0001 ♂: APOE2 hAβ vs. ♂: APOE4 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0003 0.0021 0.0003 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 ♂: APOE2 hAβ vs. ♀: Apoe hAβ < 0.0001 < 0.0001 0.0002 0.0006 0.9473 0.2633 0.0002 < 0.0001 0.1521 0.0002 0.2001 < 0.0001 0.9999 0.8368 0.8801 0.5854 0.9721 0.9817 ♂: APOE2 hAβ vs. ♀: APOE3 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0339 0.8409 0.6325 < 0.0001 < 0.0001 < 0.0001 0.0001 < 0.0001 < 0.0001 ♂: APOE2 hAβ vs. ♀: APOE4 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0003 0.3461 0.0867 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.9999 > 0.9999 > 0.9999 0.1720 0.7425 0.7973 0.9997 0.9968 0.9987 0.8292 0.9991 0.9991 ♂: APOE3 hAβ vs. ♀: Apoe hAβ 0.9244 0.9998 0.8919 0.3764 0.6909 0.0003 < 0.0001 0.8510 0.0014 0.1753 0.1923 0.9644 0.9669 ♂: APOE3 hAβ vs. ♀: APOE2 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0024 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0431 < 0.0001 0.9999 > 0.9999 0.9968 0.7248 0.4121 0.9998 0.9999 0.9733 0.9972 0.8679 0.8772 ♂: APOE3 hAβ vs. ♀: APOE4 hAβ 0.2661 > 0.9999 0.8567 0.9877 0.1797 0.9928 0.9840 0.9993 0.8652 0.9972 0.5188 > 0.9999 > 0.9999 ♂: APOE4 hAβ vs. ♀: Apoe hAβ 0.6409 0.9998 0.9748 0.3066 0.0041 < 0.0001 < 0.0001 0.5802 0.0003 0.0542 0.0088 0.9997 0.9997 ♂: APOE4 hAβ vs. ♀: APOE2 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0014 < 0.0001 0.9999 > 0.9999 0.4931 0.0580 0.0232 > 0.9999 > 0.9999 > 0.9999 0.9930 0.5586 0.5678 ♂: APOE4 hAβ vs. ♀: APOE4 hAβ 0.9993 > 0.9999 0.6820 0.9957 > 0.9999 0.2929 0.2865 > 0.9999 0.9968 > 0.9999 0.9993 0.9994 0.9997 ♀: Apoe hAβ vs. ♀: APOE2 hAβ < 0.0001 < 0.0001 < 0.0001 0.9999 < 0.0001 0.0067 0.0058 0.9944 < 0.0001 0.9999 0.9629 0.2485 0.2972 0.0143 0.9999 0.1781 0.0833 0.0044 0.0018 < 0.0001 0.5488 < 0.0001 0.0460 0.0024 0.9704 0.9791 ♀: APOE2 hAβ vs. ♀: APOE3 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0005 0.0021 < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0094 < 0.0001 < 0.0001 ♀: APOE2 hAβ vs. ♀: APOE4 hAβ < 0.0001 < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0003 < 0.0001 < 0.0001 < 0.0001 < 0.0001 0.0004 < 0.0001 0.9999 0.7253 0.9988 0.5078 0.9872 0.9045 > 0.9999 0.9774 > 0.9999 0.8879 0.8524 0.8416 Table 2 Post hoc Tukey’s Analysis: p-Values for Multiple Comparisons of Cholesterol Species Cholesterol-Total Cholesterol-Free Cholesterol-Esters ♂: Apoe hAβ vs. ♂: APOE2 hAβ < 0.0001 0.9999 0.9996 ♂: Apoe hAβ vs. ♂: APOE4 hAβ 0.9929 0.9994 0.9889 ♂: Apoe hAβ vs. ♀: Apoe hAβ 0.9966 0.9870 0.9991 ♂: Apoe hAβ vs. ♀: APOE2 hAβ < 0.0001 < 0.0001 0.9999 0.9950 ♂: Apoe hAβ vs. ♀: APOE4 hAβ 0.9803 0.9996 0.9592 ♂: APOE2 hAβ vs. ♂: APOE3 hAβ < 0.0001 < 0.0001 0.0001 ♂: APOE2 hAβ vs. ♂: APOE4 hAβ < 0.0001 < 0.0001 < 0.0001 ♂: APOE2 hAβ vs. ♀: Apoe hAβ < 0.0001 < 0.0001 0.0019 ♂: APOE2 hAβ vs. ♀: APOE2 hAβ 0.6360 0.9638 0.5012 ♂: APOE2 hAβ vs. ♀: APOE3 hAβ < 0.0001 < 0.0001 < 0.0001 ♂: APOE2 hAβ vs. ♀: APOE4 hAβ < 0.0001 < 0.0001 0.9999 > 0.9999 > 0.9999 ♂: APOE3 hAβ vs. ♀: Apoe hAβ 0.9484 0.9602 0.9606 ♂: APOE3 hAβ vs. ♀: APOE2 hAβ < 0.0001 < 0.0001 0.9999 > 0.9999 > 0.9999 ♂: APOE3 hAβ vs. ♀: APOE4 hAβ 0.9995 > 0.9999 0.9991 ♂: APOE4 hAβ vs. ♀: Apoe hAβ 0.8189 0.8556 0.8525 ♂: APOE4 hAβ vs. ♀: APOE2 hAβ < 0.0001 < 0.0001 0.9999 > 0.9999 > 0.9999 ♂: APOE4 hAβ vs. ♀: APOE4 hAβ > 0.9999 > 0.9999 > 0.9999 ♀: Apoe hAβ vs. ♀: APOE2 hAβ < 0.0001 < 0.0001 < 0.0001 ♀: Apoe hAβ vs. ♀: APOE3 hAβ 0.8983 0.9594 0.8956 ♀: Apoe hAβ vs. ♀: APOE4 hAβ 0.7383 0.8685 0.7378 ♀: APOE2 hAβ vs. ♀: APOE3 hAβ < 0.0001 < 0.0001 < 0.0001 ♀: APOE2 hAβ vs. ♀: APOE4 hAβ < 0.0001 < 0.0001 0.9999 > 0.9999 > 0.9999 The levels of LPE, CAR, PI, and PC showed gender-dependent variations in Apoe hAβ rats (LPE is shown in Fig. 5 e; sex factor, F (1, 24) = 5.143, P = 0.0326; genotype factor, F (3, 24) = 27.96, P < 0.0001; sex × genotype interaction, F (3, 24) = 3.351, P = 0.0357. CAR is shown in Fig. 5 f; sex factor, F (1, 24) = 33.52, P < 0.0001; genotype factor, F (3, 24) = 21.55, P < 0.0001; sex × genotype interaction, F (3, 24) = 1.325, P = 0.2894. PI is shown in Fig. 5 g; sex factor, F (1, 24) = 82.76, P < 0.0001; genotype factor, F (3, 24) = 49.39, P < 0.0001; sex × genotype interaction, F (3, 24) = 7.462, P = 0.0011. PC is shown in Fig. 5 i; sex factor, F (1, 24) = 5.798, P = 0.0241; genotype factor, F (3, 24) = 71.09, P < 0.0001; sex × genotype interaction, F (3, 24) = 5.659, P = 0.0044. pPC is shown in Fig. 5 j; sex factor, F (1, 24) = 0.4159, P = 0.5251; genotype factor, F (3, 24) = 59.09, P < 0.0001; sex × genotype interaction, F (3, 24) = 1.165, P = 0.3436.). In male rats, the levels of these lipid species were lower compared to female rats. Moreover, within the APOE2 hAβ rats, males exhibited lower levels of PI compared to females (Fig. 5 ) (see Table 1 for multiple comparisons). Furthermore, a comparison among the humanized APOE genotypes revealed additional insights. Female APOE3 hAβ and APOE4 hAβ rats exhibited lower levels of LPE and CAR compared to female APOE2 hAβ rats. Additionally, male APOE2 hAβ rats displayed higher levels of LPE compared to male Apoe hAβ and APOE4 hAβ rats, and higher levels of CAR compared to male APOE4 hAβ rats (Fig. 5 ). In APOE3 hAβ and APOE4 hAβ rats, females showed reduced levels of PI, PC, and pPC compared to Apoe hAβ and APOE2 hAβ rats of the same sex. Additionally, male APOE3 hAβ and APOE4 hAβ rats had lower levels of PI compared to male APOE2 hAβ rats, with male APOE4 hAβ rats also displaying lower levels of PI compared to male Apoe hAβ . Moreover, male APOE2 hAβ rats exhibited higher levels of PC, pPC, and LPC compared to male Apoe hAβ , APOE3 hAβ , and APOE4 hAβ rats (Fig. 5 ). We also analyzed the ratios of TAG to PC and FA 18:1 to FA 18:2 which are indicators of the size of lipoproteins and FA profiles in TAG pools, respectively, since FA 18:1 largely represents the de novo synthesized pool and FA 18:2 represents the portion from dietary uptake. APOE2 hAβ rats have much higher TAG/PC ratio compared to all other variants in male animals, indicating a relatively larger size of lipoprotein particles in APOE2 hAβ male rats compared to the other counterparts (TAG/PC is shown in Fig. 5 q; sex factor, F (1, 24) = 2.472, P = 0.1290; genotype factor, F (3, 24) = 20.62, P < 0.0001; sex × genotype interaction, F (3, 24) = 1.071, P = 0.3798.). But, in females TAG/PC ratio was higher in both APOE2 hAβ and APOE3 hAβ compared to Apoe hAβ rats (Fig. 5 q). Moreover, FA18:1/FA 18:2 ratio was lower in male Apoe hAβ rats compared to females (FA18:1/ FA18:2 is shown in Fig. 5 r; sex factor, F (1, 24) = 23.33, P < 0.0001; genotype factor, F (3, 24) = 20.65, P < 0.0001; sex × genotype interaction, F (3, 24) = 2.608, P = 0.0749 ) . Also, FA18:1/FA 18:2 ratio was lower in APOE4 hAβ compared to Apoe hAβ in both sexes. A lower FA18:1/FA 18:2 ratio was manifest in APOE2 hAβ and APOE3 hAβ females compared to Apoe hAβ animals (Fig. 5 r). These different FA ratios clearly indicate the different lipid metabolism at the FA levels. Given the markedly elevated lipid levels in APOE2 hAβ relative to other variants, it becomes impractical to compare lipid changes among Apoe hAβ , APOE3 hAβ , and APOE4 hAβ . Consequently, in Fig. 6 , we have selectively excluded the APOE2 hAβ rats, thereby replicating the analytical approach employed in Fig. 5 without incorporating the APOE2 hAβ dataset. First, we observed that both sexes of APOE4 hAβ rats had lower levels of PG compared to APOE3 hAβ rats (Fig. 6 a) (PG is shown in Fig. 6 a; sex factor, F (1, 18) = 2.437, P = 0.1359; genotype factor, F (2, 18) = 11.39, P = 0.0006; sex × genotype interaction, F (2, 18) = 0.3179, P = 0.7317.) (For male APOE3 hAβ vs. APOE4 hAβ p = 0.0415*. For females APOE3 hAβ vs. APOE4 hAβ p = 0.0429*). In male Apoe hAβ animals, the levels of PE, pPE, LPE, CAR, PI, PC, total cholesterol, and free cholesterol were found to be lower compared to their female counterparts. In APOE4 hAβ animals, females exhibited lower levels of PE, pPE, LPE, CAR, PI, SM, PC, pPC, LPC, total cholesterol, free cholesterol, cholesterol esters, and the ratio of FA18:1/FA18:2 when compared to female Apoe hAβ animals (PE is shown in Fig. 6 c; sex factor, F (1, 18) = 0.3314, P = 0.5719; genotype factor, F (2, 18) = 1.552, P = 0.2388; sex × genotype interaction, F (2, 18) = 8.314, P = 0.0028. pPE is shown in Fig. 6 d; sex factor, F (1, 18) = 1.776, P = 0.1993; genotype factor, F (2, 18) = 21.43, P < 0.0001; sex × genotype interaction, F (2, 18) = 12.23, P = 0.0004. LPE is shown in Fig. 6 e; sex factor, F (1, 18) = 2.997, P = 0.1005; genotype factor, F (2, 18) = 10.30, P = 0.0010; sex × genotype interaction, F (2, 18) = 5.70, P = 0.0121. CAR is shown in Fig. 6 f; sex factor, F (1, 18) = 20.87, P = 0.0002; genotype factor, F (2, 18) = 13.27, P = 0.0003; sex × genotype interaction, F (2, 18) = 1.718, P = 0.2076. PI is shown in Fig. 6 g; sex factor, F (1, 18) = 64.51, P < 0.0001; genotype factor, F (2, 18) = 52.32, P < 0.0001; sex × genotype interaction, F (2, 18) = 12.77, P = 0.0004. SM is shown in Fig. 6 h; sex factor, F (1, 18) = 0.2447, P = 0.6268; genotype factor, F (2, 18) = 52.96, P < 0.0001; sex × genotype interaction, F (2, 18) = 1.818, P = 0.1910. PC is shown in Fig. 6 i; sex factor, F (1, 18) = 7.169, P = 0.0154; genotype factor, F (2, 18) = 26.70, P < 0.0001; sex × genotype interaction, F (2, 18) = 15.82, P = 0.0001. pPC is shown in Fig. 6 j; sex factor, F (1, 18) = 0.0014, P = 0.9701; genotype factor, F (2, 18) = 32.94, P < 0.0001; sex × genotype interaction, F (2, 18) = 3.648, P = 0.0468. LPC is shown in Fig. 6 k; sex factor, F (1, 18) = 0.01269, P = 0.9116; genotype factor, F (2, 18) = 22.77, P < 0.0001; sex × genotype interaction, F (2, 18) = 1.368, P = 0.2799. Cholesterol-Total is shown in Fig. 6 n; sex factor, F (1, 18) = 1.041, P = 0.3212; genotype factor, F (2, 18) = 34.85, P < 0.0001; sex × genotype interaction, F (2, 18) = 5.507, P = 0.0136. Cholesterol-Free is shown in Fig. 6 o; sex factor, F (1, 18) = 6.462, P = 0.0204; genotype factor, F (2, 18) = 22.53, P < 0.0001; sex × genotype interaction, F (2, 18) = 5.915, P = 0.0106. Cholesterol-Ester is shown in Fig. 6 p; sex factor, F (1, 18) = 0.0808, P = 0.7794; genotype factor, F (2, 18) = 39.83, P < 0.0001; sex × genotype interaction, F (2, 18) = 5.055, P = 0.0181. FA18:1/FA18:2 is shown in Fig. 6 r; sex factor, F (1, 18) = 19.17, P = 0.0004; genotype factor, F (2, 18) = 25.58, P < 0.0001; sex × genotype interaction, F (2, 18) = 2.596, P = 0.1022). Similarly, male APOE4 hAβ rats had lower levels of PI, SM, pPC, LPC, total cholesterol, and cholesterol esters compared to male Apoe hAβ rats. Additionally, female APOE3 hAβ rats showed lower levels of pPE, CAR, PI, SM, PC, pPC, LPC, total cholesterol, free cholesterol, cholesterol esters and ratio of FA18:1/FA18:2 compared to female Apoe hAβ rats. Conversely, female APOE3 hAβ rats showed a higher ratio of TAG to PC (TAG/PC is shown in Fig. 6 q; sex factor, F (1, 18) = 2.756, P = 0.1140; genotype factor, F (2, 18) = 8.524, P = 0.0025; sex × genotype interaction, F (2, 18) = 1.102, P = 0.3537). Male APOE3 hAβ rats, on the other hand, only had lower levels of SM compared to male Apoe hAβ animals. PS, TAG and FA levels were comparable in Apoe hAβ , APOE3 hAβ and APOE4 hAβ (PS is shown in Fig. 6 b; sex factor, F (1, 18) = 3.876, P = 0.0646; genotype factor, F (2, 18) = 0.1970, P = 0.8230; sex × genotype interaction, F (2, 18) = 0.4870, P = 0.6223. TAG is shown in Fig. 6 l; sex factor, F (1, 18) = 2.321, P = 0.1450; genotype factor, F (2, 18) = 5.081, P = 0178; sex × genotype interaction, F (2, 18) = 0.4433, P = 0.6488. FA is shown in Fig. 6 m; sex factor, F (1, 18) = 2.233, P = 0.1524; genotype factor, F (2, 18) = 5.030, P = 0184; sex × genotype interaction, F (2, 18) = 0.4383, P = 0.6519). This comprehensive lipidomic analysis across nine lipid classes (CAR, FA, PC, PE, PI, PG, PS, TAG, and SM) revealed distinct patterns of lipid abundance in different rat samples, identified through the generation of heatmaps. Each heatmap provided a color-coded representation of the relative abundance of each lipid type within each sample. Our major finding was that APOE2 hAβ male rats consistently displayed unique lipidomic profiles across all lipid classes. This pattern underscores the substantial influence of genotype on lipid metabolism and suggests that specific genotypes and sex, such as APOE2 hAβ males, might uniquely influence lipid metabolism. This insight has potential implications for understanding disease susceptibilities or responses to treatments in these genetic models. We observed substantial heterogeneity in lipid composition across samples and lipid species, which underlined the diversity of the lipidomic landscape in the studied rat models. Certain lipid species like PS P-18:0/22:6 and PS O-16:0/22:6 in the PS lipid data, and TAG 48:0 and TAG 50:1 in the TAG lipid data, consistently exhibited lower relative abundances across all samples. These patterns suggest that these lipids might have less dominant role in the overall lipid metabolism, or their functions might be conserved across different rat models. Conversely, certain lipids were found to be consistently present at higher levels across all samples, indicating their prominent role in lipid metabolism across different rat genotypes and sexes. For instance, in the PC lipid data, lipid species such as PC 32:0 and PC 34:1 were observed to have higher relative abundances in most samples (Figs. 7 and 8 ). Correlation heatmaps (Fig. 9 ) depicted relationships among lipid species in Apoe hAβ , APOE2 hAβ , APOE3 hAβ , and APOE4 hAβ rats. We used the Pearson correlation coefficient to calculate all correlations and p-values between 222 lipid species in each group. We identified the top 20 correlations based on absolute values of correlation coefficients with p < 0.05. The heatmaps reveal strong positive correlations between different types of lipid species across all groups. The strongest positive correlations were between TAG C52:2/C53:9 and PE D18:0–20:4/D16:0–22:4 in Apoe hAβ rats ( \(\:r\) = 0.97), between PE P18:1–20:4/P16:0–22:5 and PE A20:0–20:4/P18:0–22:3 in APOE2 hAβ rats ( \(\:r\) = 0.98), between CAR 16:2 and CAR 18:2 in APOE3 hAβ rats ( \(\:r\) = 0.97), between CAR 18:0 and CAR 18:1 in APOE4 hAβ rats ( \(\:r\) = 0.97). There are strong positive correlations between LPE 20:4 and pPE P18:0–22:6/P18:1–22:5 with correlation coefficients of 0.97, 0.45 and 0.33 in Apoe hAβ , APOE2 hAβ and APOE3 hAβ rats, respectively. Conversely, there are strong negative correlations between CAR 7:0 and PE P16:0–20:3/P18:1–18:2 in APOE3 hAβ ( \(\:r\) = -0.91) and between SM N18:0 and PE D16:0–20:4/D18:2–18:2 in APOE4 hAβ animals ( \(\:r\) = -0.94). The heatmaps also show that there are several opposite correlations between the different lipid types in the different groups of animals. A strong positive correlation exists between SM N20:0 and PC D18:0–22:5 with correlation coefficients of 0.76 and 0.32 in Apoe hAβ and APOE2 hAβ , respectively but in APOE4 hAβ animals there is negative correlation ( \(\:r\) = -0.55). Similarly, there is a strong negative correlation between PC D18:2–18:2/D16:0–20:4 and SM N24:2 in APOE4 hAβ rats with correlation coefficients of -0.68, while there is positive correlation in Apoe hAβ ( \(\:r\) = 0.32) and APOE3 hAβ rats ( \(\:r\) = 0.36). The correlation between CAR 7:0 and CAR 18:1 is more robust in APOE3 hAβ rats ( \(\:r\) = 0.91) than in APOE4 hAβ animals ( \(\:r\) = 0.29), suggesting a more pronounced relationship between these two lipid species in APOE3 hAβ rats. Another intriguing observation is that the correlation between PE D16:0–22:6 and SM N18:0, while there is strong negative correlation in APOE4 hAβ animals ( \(\:r\) = -0.56), there is slightly positive correlation in APOE2 hAβ rats ( \(\:r\) = 0.018). All other correlation coefficients are reported in Supplemental Table 1. In conclusion, our findings offer a complex picture of lipid homeostasis across APOE variants of rat models. The substantial variations in lipid composition could have significant implications for understanding lipid metabolism in these models, potentially serving as a basis for the identification of lipidomic biomarkers for different physiological or pathological states. DISCUSSION In this study, we have characterized Long-Evans rat models that express human APOE2 , APOE3 , and APOE4 variant genes, as well as humanized App h alleles. We observed that APOE2 hAβ rats exhibited the highest levels of APOE in both serum and brain compared to APOE4 hAβ and APOE3 hAβ rats, which showed comparable levels of APOE. These changes were unlikely due to transcriptional differences, as mRNA levels of human APOE2 , APOE3 , and APOE4 were comparable. Therefore, differences in protein expression are likely attributed to translational and/or protein stability variances. While further investigation into these mechanisms is warranted, these findings closely mirror observations from human studies, underscoring the validity and translatability of these rat models [ 44 , 47 , 48 ]. The humanization of both APP/Aβ and APOE in model organisms that express these human proteins in a physiological manner enables a comprehensive study of the multifaceted APOE-Aβ interaction. Our findings revealed a significant elevation in Aβ43 levels in male APOE4 hAβ rats compared to their APOE2 hAβ counterparts. This observation is particularly noteworthy as Aβ43 has high propensity for oligomerization and is a primary determinant of amyloid pathology [ 37 ], while APOE4 is the most prominent genetic risk factor for LOAD, increasing the risk up to 12-fold, whereas APOE2 is a protective factor [ 6 , 49 ]. This finding also aligns with evidence that Aβ oligomer levels in APOE4 AD patients' brains are 2.7 times higher than those in APOE3 AD patients [ 50 ] and is consistent with findings from mouse models expressing APOE4 and APOE2 [ 51 ]. APOE2 homozygosity can lead to type III HLP in humans. Our findings suggest that APOE2 hAβ rats serve as a suitable model for studying this condition, showing elevated triglyceride and cholesterol levels. This led us to investigate how different human APOE isoforms affect lipid metabolism in models expressing humanized APOE variants and humanized APP/Aβ in a more comprehensive manner. Our findings confirm the elevated lipid levels previously observed in humanized APOE2 mice compared to those carrying APOE3 and APOE4 [ 52 ]. Moreover, we identified significant differences with the mouse models, including notably higher serum levels of total PG, pPE, LPE, CAR, PI, PC, pPC, LPC, FA in TAG, free-cholesterol, and cholesterol-esters. These observations are consistent with data showing that brain samples from APOE2 LOAD patients exhibit elevated levels of CAR, SM, PC, and LPE compared to APOE3 and APOE4 LOAD patients [ 53 ]. Thus, rats, known for their more human-like metabolism compared to mice [ 54 ], and engineered to carry human APOE2 along with the humanized App gene, may offer enhanced modeling capabilities for type III HLP compared to their mouse counterparts. A follow-up analysis aimed at discerning differences between APOE3 hAβ and APOE4 hAβ rats revealed that only PG levels were lower in both male and female APOE3 hAβ rats compared to APOE4 hAβ animals. All other lipid categories showed no significant variation between these two groups. This serum lipidomic similarity between APOE4 hAβ and APOE3 hAβ rats mirrors the lipid homogeneity observed in synaptosomes isolated from 2-month-old mice expressing human APOE3 and APOE4 [ 54 ]. Conclusions In this study, we utilized Long-Evans knock-in rats with humanized App and APOE2 , APOE3 , and APOE4 isoforms to investigate their roles in LOAD type III HLP. Our findings reveal distinct profiles of APOE expression and lipid metabolism associated with each APOE isoform. Specifically, APOE2 hAβ rats exhibited higher APOE levels in both serum and brain compared to APOE4 hAβ and APOE3 hAβ rats, suggesting isoform-specific differences in protein expression likely due to translational or stability factors rather than transcriptional differences. Notably, elevated Aβ43 levels were observed in male APOE4 hAβ rats, which could be instrumental in studying amyloid pathology due to Aβ43's high aggregation propensity. Furthermore, APOE2 hAβ rats showed elevated triglyceride and cholesterol levels, supporting their utility as a model for type III HLP. Minimal lipid profile differences between APOE4 hAβ and APOE3 hAβ rats reflect previously observed patterns in mouse models Overall, these Long-Evans rat models provide a valuable platform for exploring the interactions between human APOE isoforms and Aβ in the context of LOAD and lipid metabolism. Future studies will expand lipidomic analyses to various CNS regions, offering deeper insights into the effects of human APOE isoforms on lipid metabolism and Alzheimer’s disease pathology. Comparing the brain lipidomic of APOE2 hAβ , APOE3 hAβ and APOE4 hAβ rats with lipidomic profiles from postmortem AD brains will further validate the translatability of the models described here [ 53 ]. Declarations Funding: MY: Alzheimer's Association , 24AARFD-1243865 XH: National Institute on Aging, R01AG061729; National Institutes of Health, P30 AG013319 LD: National Institute on Aging , R01AG073182; National Institute on Aging , R01AG063407; National Institute on Aging , RF1AG064821 Author Contribution MY and LD wrote the first draft of the manuscript. MY, TY, MT, HB, MP, and LD contributed to the material preparation, data collection, and figure drawing. MY, TY, MT, XH, and LD contributed to the critical review of the manuscript. All authors reviewed and approved the final version. References Yamazaki, Y., et al., Apolipoprotein E and Alzheimer disease: pathobiology and targeting strategies. Nature Reviews Neurology, 2019. 15 (9): p. 501-518. Huebbe, P. and G. Rimbach, Evolution of human apolipoprotein E (APOE) isoforms: Gene structure, protein function and interaction with dietary factors. Ageing Research Reviews, 2017. 37 : p. 146-161. Fullerton, S.M., et al., Apolipoprotein E variation at the sequence haplotype level: implications for the origin and maintenance of a major human polymorphism. The American journal of human genetics, 2000. 67 (4): p. 881-900. Kayden, H.J., F. Maschio, and M.G. Traber, The secretion of apolipoprotein E by human monocytederived macrophages. Archives of biochemistry and biophysics, 1985. 239 (2): p. 388-395. Gee, J.R. and J.N. Keller, Astrocytes: regulation of brain homeostasis via apolipoprotein E. The International Journal of Biochemistry & Cell Biology, 2005. 37 (6): p. 1145-1150. West, H.L., G.W. Rebeck, and B.T. Hyman, Frequency of the apolipoprotein E ε2 allele is diminished in sporadic Alzheimer disease. Neuroscience letters, 1994. 175 (1-2): p. 46-48. Farrer, L.A., et al., Effects of Age, Sex, and Ethnicity on the Association Between Apolipoprotein E Genotype and Alzheimer Disease: A Meta-analysis. JAMA, 1997. 278 (16): p. 1349-1356. Kojro, E., et al., Low cholesterol stimulates the nonamyloidogenic pathway by its effect on the α-secretase ADAM 10. Proceedings of the National Academy of Sciences, 2001. 98 (10): p. 5815-5820. Grösgen, S., et al., Role of amyloid beta in lipid homeostasis. Biochimica et Biophysica Acta (BBA)-Molecular and Cell Biology of Lipids, 2010. 1801 (8): p. 966-974. Cordy, J.M., et al., Exclusively targeting β-secretase to lipid rafts by GPI-anchor addition up-regulates β-site processing of the amyloid precursor protein. Proceedings of the National Academy of Sciences, 2003. 100 (20): p. 11735-11740. LaDu, M.J., et al., Isoform-specific binding of apolipoprotein E to beta-amyloid. J Biol Chem, 1994. 269 (38): p. 23403-6. Sharman, M.J., et al., APOE genotype results in differential effects on the peripheral clearance of amyloid-β 42 in APOE knock-in and knock-out mice. Journal of Alzheimer's Disease, 2010. 21 (2): p. 403-409. Tambini, M.D., W. Yao, and L. D'Adamio, Facilitation of glutamate, but not GABA, release in Familial Alzheimer's APP mutant Knock-in rats with increased beta-cleavage of APP. Aging Cell, 2019. 18 (6): p. e13033. Piedrahita, J.A., et al., Generation of mice carrying a mutant apolipoprotein E gene inactivated by gene targeting in embryonic stem cells. Proceedings of the National Academy of Sciences, 1992. 89 (10): p. 4471-4475. Sullivan, P.M., et al., Targeted replacement of the mouse apolipoprotein E gene with the common human APOE3 allele enhances diet-induced hypercholesterolemia and atherosclerosis. J Biol Chem, 1997. 272 (29): p. 17972-80. Wu, Y., et al., Features of Lipid Metabolism in Humanized ApoE Knockin Rat Models. International Journal of Molecular Sciences, 2021. 22 (15): p. 8262. Mahley, R.W., Y. Huang, and S.C. Rall, Pathogenesis of type III hyperlipoproteinemia (dysbetalipoproteinemia): questions, quandaries, and paradoxes. Journal of lipid research, 1999. 40 (11): p. 1933-1949. Mahley, R.W. and Z.-S. Ji, Remnant lipoprotein metabolism: key pathways involving cell-surface heparan sulfate proteoglycans and apolipoprotein E. Journal of Lipid Research, 1999. 40 (1): p. 1-16. Lund-Katz, S., et al., Apolipoprotein B-100 Conformation and Particle Surface Charge in Human LDL Subspecies: Implication for LDL Receptor Interaction. Biochemistry, 1998. 37 (37): p. 12867-12874. Huang, Y., et al., Overexpression and Accumulation of Apolipoprotein E as a Cause of Hypertriglyceridemia *. Journal of Biological Chemistry, 1998. 273 (41): p. 26388-26393. Rensen, P.C.N. and T.J.C. van Berkel, Apolipoprotein E Effectively Inhibits Lipoprotein Lipase-mediated Lipolysis of Chylomicron-like Triglyceride-rich Lipid Emulsions in Vitro and in Vivo*. Journal of Biological Chemistry, 1996. 271 (25): p. 14791-14799. Fredrickson, D.S., R.I. Levy, and R.S. Lees, Fat Transport in Lipoproteins — An Integrated Approach to Mechanisms and Disorders. New England Journal of Medicine, 1967. 276 (3): p. 148-156. Utermann, G., M. Jaeschke, and J. Menzel, Familial hyperlipoproteinemia type III: deficiency of a specific apolipoprotein (apo E-III) in the very-low-density lipoproteins. FEBS lett, 1975. 56 (2): p. 352-355. Zannis, V.I. and J.L. Breslow, Human very low density lipoprotein apolipoprotein E isoprotein polymorphism is explained by genetic variation and posttranslational modification. Biochemistry, 1981. 20 (4): p. 1033-1041. Mahley, R.W., Apolipoprotein E: Cholesterol Transport Protein with Expanding Role in Cell Biology. Science, 1988. 240 (4852): p. 622-630. Davignon, J., R.E. Gregg, and C.F. Sing, Apolipoprotein E polymorphism and atherosclerosis. Arteriosclerosis: An Official Journal of the American Heart Association, Inc., 1988. 8 (1): p. 1-21. Phillips, M.C., Apolipoprotein E isoforms and lipoprotein metabolism. IUBMB Life, 2014. 66 (9): p. 616-623. Gustavsson, J., et al., Interaction of apolipoprotein E genotype with smoking and physical inactivity on coronary heart disease risk in men and women. Atherosclerosis, 2012. 220 (2): p. 486-492. Weisgraber, K.H., T.L. Innerarity, and R.W. Mahley, Abnormal lipoprotein receptor-binding activity of the human E apoprotein due to cysteine-arginine interchange at a single site. Journal of Biological Chemistry, 1982. 257 (5): p. 2518-2521. Pham, H., T. Yin, and L. D'Adamio, Initial assessment of the spatial learning, reversal, and sequencing task capabilities of knock-in rats with humanizing mutations in the Aβ-coding region of App. PLoS One, 2022. 17 (5): p. e0263546. Tambini, M.D., W. Yao, and L. D'Adamio, Facilitation of glutamate, but not GABA, release in Familial Alzheimer's APP mutant Knock-in rats with increased β-cleavage of APP. Aging Cell, 2019. 18 (6): p. e13033. Yesiltepe, M., et al., Late-long-term potentiation magnitude, but not Abeta levels and amyloid pathology, is associated with behavioral performance in a rat knock-in model of Alzheimer disease. Front Aging Neurosci, 2022. 14 : p. 1040576. Tambini, M.D., et al., Abeta43 levels determine the onset of pathological amyloid deposition. J Biol Chem, 2023. 299 (7): p. 104868. Yin, T., et al., A familial Danish dementia rat shows impaired presynaptic and postsynaptic glutamatergic transmission. J Biol Chem, 2021. 297 (3): p. 101089. Ren, S., et al., TNF-alpha-mediated reduction in inhibitory neurotransmission precedes sporadic Alzheimer's disease pathology in young Trem2(R47H) rats. J Biol Chem, 2021. 296 : p. 100089. Ren, S., et al., Microglia TREM2(R47H) Alzheimer-linked variant enhances excitatory transmission and reduces LTP via increased TNF-alpha levels. Elife, 2020. 9 . Tambini, M.D., et al., Aβ43 levels determine the onset of pathological amyloid deposition. Journal of Biological Chemistry, 2023. 299 (7). Han, X., Lipidomics: Comprehensive mass spectrometry of lipids . 2016: John Wiley & Sons. Palavicini, J.P., et al., Novel molecular insights into the critical role of sulfatide in myelin maintenance/function. Journal of neurochemistry, 2016. 139 (1): p. 40-54. Han, X., K. Yang, and R.W. Gross, Microfluidics-based electrospray ionization enhances the intrasource separation of lipid classes and extends identification of individual molecular species through multi-dimensional mass spectrometry: development of an automated high-throughput platform for shotgun lipidomics. Rapid Commun Mass Spectrom, 2008. 22 (13): p. 2115-24. Yang, K., et al., Automated lipid identification and quantification by multidimensional mass spectrometry-based shotgun lipidomics. Anal Chem, 2009. 81 (11): p. 4356-68. Wang, M. and X. Han, Multidimensional mass spectrometry-based shotgun lipidomics. Methods Mol Biol, 2014. 1198 : p. 203-20. Yang, K., et al., Automated lipid identification and quantification by multidimensional mass spectrometry-based shotgun lipidomics. Analytical chemistry, 2009. 81 (11): p. 4356-4368. Cai, C., Z. Wen, and L. Li, The relationship between ApoE gene polymorphism and the efficacy of statins controlling hyperlipidemia. Am J Transl Res, 2021. 13 (6): p. 6772-6777. Passer, B., et al., Generation of an apoptotic intracellular peptide by gamma-secretase cleavage of Alzheimer's amyloid beta protein precursor. J Alzheimers Dis, 2000. 2 (3-4): p. 289-301. Kinosian, B., H. Glick, and G. Garland, Cholesterol and coronary heart disease: predicting risks by levels and ratios. Annals of internal medicine, 1994. 121 (9): p. 641-647. Martínez-Morillo, E., et al., Total apolipoprotein E levels and specific isoform composition in cerebrospinal fluid and plasma from Alzheimer’s disease patients and controls. Acta Neuropathologica, 2014. 127 (5): p. 633-643. Giannisis, A., et al., Plasma apolipoprotein E levels, isoform composition, and dimer profile in relation to plasma lipids in racially diverse patients with Alzheimer’s disease and mild cognitive impairment. Alzheimer's Research & Therapy, 2023. 15 (1): p. 119. Michaelson, D.M., APOE ε4: The most prevalent yet understudied risk factor for Alzheimer's disease. Alzheimer's & Dementia, 2014. 10 (6): p. 861-868. Hashimoto, T., et al., Apolipoprotein E, Especially Apolipoprotein E4, Increases the Oligomerization of Amyloid β Peptide. The Journal of Neuroscience, 2012. 32 (43): p. 15181-15192. Hudry, E., et al., Gene Transfer of Human Apoe Isoforms Results in Differential Modulation of Amyloid Deposition and Neurotoxicity in Mouse Brain. Science Translational Medicine, 2013. 5 (212): p. 212ra161-212ra161. Sharman, M.J., et al., Profiling brain and plasma lipids in human apoe ε2, ε3, and ε4 knock-in mice using electrospray ionization mass spectrometry. Journal of Alzheimer's Disease, 2010. 20 (1): p. 105-111. Lefterov, I., et al., APOE2 orchestrated differences in transcriptomic and lipidomic profiles of postmortem AD brain. Alzheimer's Research & Therapy, 2019. 11 (1): p. 113. Igbavboa, U., et al., Murine synaptosomal lipid raft protein and lipid composition are altered by expression of human apoE 3 and 4 and by increasing age. Journal of the Neurological Sciences, 2005. 229 : p. 225-232. Additional Declarations No competing interests reported. Supplementary Files rawwbofgapdhmaleinfigure2.tif rawwbofpsd95femaleinfigure3.tif rawwbofpsd95maleinfigure3.tif rawwbofgapdhfemaleinfigure2.tif rawwbofAPPmaleinfigure3.tif rawwbofAPPfemaleinfigure3.tif rawwbofAPPctfmaleinfigure3.tif rawwbofapoemaleinfigure2.tif rawwbofAPPctffemaleinfigure3.tif rawwbofapoefemaleinfigure2.tif originalgelofapoe3FIG1PCRs.tif originalgelofapoe4FIG1PCRs.tif originalgelofapoe2FIG1PCRs.tif SupplementalTable1CorrelationCoefficients.xlsx Cite Share Download PDF Status: Published Journal Publication published 27 Sep, 2024 Read the published version in Cell Communication and Signaling → Version 1 posted Editorial decision: Revision requested 08 Aug, 2024 Reviews received at journal 08 Aug, 2024 Reviews received at journal 06 Aug, 2024 Reviewers agreed at journal 02 Aug, 2024 Reviewers agreed at journal 31 Jul, 2024 Reviewers agreed at journal 29 Jul, 2024 Reviewers invited by journal 29 Jul, 2024 Editor assigned by journal 24 Jul, 2024 Submission checks completed at journal 24 Jul, 2024 First submitted to journal 23 Jul, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4789248","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":337878388,"identity":"6a957d82-a4d7-4e8f-af26-cf9aeaa466fe","order_by":0,"name":"Metin Yesiltepe","email":"","orcid":"","institution":"New Jersey Medical School, Rutgers, The State University of New Jersey","correspondingAuthor":false,"prefix":"","firstName":"Metin","middleName":"","lastName":"Yesiltepe","suffix":""},{"id":337878389,"identity":"6edb5834-43d2-40aa-93ed-b6bded11b21e","order_by":1,"name":"Tao Yin","email":"","orcid":"","institution":"New Jersey Medical School, Rutgers, The State University of New Jersey","correspondingAuthor":false,"prefix":"","firstName":"Tao","middleName":"","lastName":"Yin","suffix":""},{"id":337878390,"identity":"25b7fc93-db5b-46d5-b847-172c5da97b24","order_by":2,"name":"Marc Tambini","email":"","orcid":"","institution":"New Jersey Medical School, Rutgers, The State University of New Jersey","correspondingAuthor":false,"prefix":"","firstName":"Marc","middleName":"","lastName":"Tambini","suffix":""},{"id":337878391,"identity":"09fe5c5f-dbff-4b71-9a95-6f11f78f47be","order_by":3,"name":"Hanmei Bao","email":"","orcid":"","institution":"Barshop Institute for Longevity and Aging Studies, University of Texas Health San Antonio, San Antonio, TX, 78229","correspondingAuthor":false,"prefix":"","firstName":"Hanmei","middleName":"","lastName":"Bao","suffix":""},{"id":337878392,"identity":"b39d0767-88fc-407f-ae9d-11b34180b616","order_by":4,"name":"Meixia Pan","email":"","orcid":"","institution":"Barshop Institute for Longevity and Aging Studies, University of Texas Health San Antonio, San Antonio, TX, 78229","correspondingAuthor":false,"prefix":"","firstName":"Meixia","middleName":"","lastName":"Pan","suffix":""},{"id":337878393,"identity":"8c990742-f5cc-4b08-b339-276b0dcd7e08","order_by":5,"name":"Xianlin Han","email":"","orcid":"","institution":"Barshop Institute for Longevity and Aging Studies, University of Texas Health San Antonio, San Antonio, TX, 78229","correspondingAuthor":false,"prefix":"","firstName":"Xianlin","middleName":"","lastName":"Han","suffix":""},{"id":337878394,"identity":"d90dfaf6-0c87-48ee-916b-9ef982e153ee","order_by":6,"name":"Luciano D'Adamio","email":"data:image/png;base64,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","orcid":"","institution":"New Jersey Medical School, Rutgers, The State University of New Jersey","correspondingAuthor":true,"prefix":"","firstName":"Luciano","middleName":"","lastName":"D'Adamio","suffix":""}],"badges":[],"createdAt":"2024-07-23 13:51:23","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4789248/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4789248/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s12964-024-01832-2","type":"published","date":"2024-09-27T15:56:59+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":62944830,"identity":"7d50c3cb-451e-4ca6-aa28-e1928c90be49","added_by":"auto","created_at":"2024-08-21 10:13:25","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1369447,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGeneration of humanized \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e rats.\u003c/strong\u003e \u003cstrong\u003ea.\u003c/strong\u003e The schematic representation of the rat \u003cem\u003eApoe\u003c/em\u003e allele depicts the four exons with the 5' UTR sequences in black, the coding sequences in orange, and the 3' UTR in white. The regions utilized for the homology arms of the KI construct are indicated in blue. The Cas9 targeting site is also highlighted. Below is a schematic representation of the human \u003cem\u003eAPOE KI\u003c/em\u003eallele, with the sites of the oligonucleotides used for PCR analysis indicated. \u003cstrong\u003eb.\u003c/strong\u003e PCR analysis using primer pairs SPF1/SPR1, which detects correct insertion at the 5' region, and primer pairs SPF2/SPR2, which detects correct insertion at the 3' region. The data show that the rats designed as \u003cem\u003eAPOE2, APOE3, \u003c/em\u003eand\u003cem\u003e APOE4\u003c/em\u003e F0 and F1 have correct 5’ and 3’ insertions. \u003cstrong\u003ec.\u003c/strong\u003eThe schematic representation of the Southern blotting technique used for genotyping the \u003cem\u003eAPOE2, APOE3, \u003c/em\u003eand\u003cem\u003e APOE4\u003c/em\u003e rats is presented. For the 5’ arm probe Southern blot, genomic DNA was digested with AflII. The expected fragment size for the wild-type rat \u003cem\u003eApoe\u003c/em\u003e allele was 2.68 kb, while for the human \u003cem\u003eAPOE\u003c/em\u003e \u003cem\u003eKI\u003c/em\u003e allele, it was 7.61 kb. For the 3’ arm, genomic DNA was digested with KpnI plus BstEII. The expected fragment size for the wild-type rat \u003cem\u003eApoe\u003c/em\u003e allele was 4.05 kb, and for the human \u003cem\u003eAPOE KI\u003c/em\u003eallele, it was 6.55 kb. \u003cstrong\u003ed.\u003c/strong\u003e Southern blot analysis shows that the wild-type sample displayed the expected rat \u003cem\u003eApoe\u003c/em\u003e bands of 2.68 kb and 4.05 kb for the 5’ arm probe and 3’ arm probe, respectively. In contrast, the samples identified as \u003cem\u003eAPOE2, APOE3, \u003c/em\u003eand\u003cem\u003e APOE4\u003c/em\u003e F1s by the PCR analysis in part b, exhibited both the wild-type bands and the \u003cem\u003eAPOE\u003c/em\u003e KI bands of 7.61 kb and 6.55 kb for the 5’ arm probe and 3’ arm probe, respectively. No other bands that would indicate off-target, random integration are detected.\u003c/p\u003e","description":"","filename":"Figure1final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/dc423f51d3f9cd7700ceb98b.png"},{"id":62944284,"identity":"0e468e25-4da0-4f32-bbdc-b77c05983a77","added_by":"auto","created_at":"2024-08-21 10:05:25","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1658857,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eLevels of human APOE in 80 days old \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eApoe\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e, \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE2\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e, \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE3\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e and \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE4\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e \u003cstrong\u003erats. a.\u003c/strong\u003e ELISA measurements of human APOE in blood serum (n=4 per sex per genotype) showed significantly higher levels in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e compared to \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e in both males and females. \u003cstrong\u003eb\u003c/strong\u003e. ELISA analysis of human APOE in brain homogenates (\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, females n=6, males n=5; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, females n=6, males n=6; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, females n=4, males n=3; \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, females n=6, males n=6) revealed higher levels in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e compared to \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e in both sexes. Moreover, brain APOE levels were higher in male \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats compared to females. Rat Apoe (\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats) is not detected by the ELISA demonstrating specificity. Therefore, \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats were excluded from the statistical analysis\u003cstrong\u003e. c.\u003c/strong\u003e WB analysis of human APOE in the same brains analyzed by ELISA in panel b confirms higher levels of APOE in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e brains compared to \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e in both sexes.\u0026nbsp; d.\u0026nbsp; Quantitative RT-PCR analysis of rat and human \u003cem\u003eAPOE\u003c/em\u003e mRNA expression in the same brains analyzed by ELISA and WB in panels b and c confirms that \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats express only rat \u003cem\u003eApoe\u003c/em\u003e mRNA, while \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e express exclusively human \u003cem\u003eAPOE\u003c/em\u003e mRNA. Human \u003cem\u003eAPOE\u003c/em\u003e mRNA expression levels were comparable among \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, except for a reduction observed in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males compared to \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males. The WB analysis for GAPDH confirms equal loading of the samples. Data are presented as mean ± SEM and were analyzed by two-way ANOVA followed by post hoc Tukey’s multiple comparisons test when significant differences were detected. Statistical significance is denoted as ** p\u0026lt;0.01, *** p\u0026lt;0.001, **** p\u0026lt;0.0001.\u003c/p\u003e","description":"","filename":"Figure2final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/d99f81a5bcd6786047961011.png"},{"id":62944290,"identity":"5e544aad-b65f-48ec-bfef-0ddb7c98414f","added_by":"auto","created_at":"2024-08-21 10:05:25","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":2513082,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eAnalysis of APP metabolites in brains of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eApoe\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e, \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE2\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e, \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE3\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e and \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE4\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e rats. a-f\u003c/strong\u003e. ELISA measurements for Aβ38 (\u003cstrong\u003ea\u003c/strong\u003e), Aβ40 (\u003cstrong\u003eb\u003c/strong\u003e), Aβ42 (\u003cstrong\u003ec\u003c/strong\u003e), Aβ43 (\u003cstrong\u003ed\u003c/strong\u003e), sAPPα (\u003cstrong\u003ee\u003c/strong\u003e), and sAPPβ (\u003cstrong\u003ef\u003c/strong\u003e) were conducted on the same brain homogenates used in Figure 2b. g. WB analysis of APP, βCTF, and αCTF in brain lysates used in the WBs shown Figure 2c. PSD95 WB was used as a loading control. \u003cstrong\u003eh\u003c/strong\u003e. Quantification of the APP, βCTF, and αCTF signals detected in panel g. Longer exposures of βCTF and αCTF signals, which were used to quantify βCTF and αCTF, are shown below the main WBs. Data are represented as mean ± SEM and were analyzed by two‐way ANOVA followed by post-hoc Tukey's multiple comparisons tests when ANOVA showed significant differences. Statistical significance is denoted as * p\u0026lt;0.05, ** p\u0026lt;0.01, *** p\u0026lt;0.001, **** p\u0026lt;0.0001.\u003c/p\u003e","description":"","filename":"Figure3final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/82a3ab28b7f649680aaf7967.png"},{"id":62944832,"identity":"97c07f46-a2b7-4f2e-9f20-6544e98d2277","added_by":"auto","created_at":"2024-08-21 10:13:25","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":374743,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eMetabolic profiles in 80-day-old rats: blood glucose, lipids, and LDL/HDL ratio after fasting\u003c/strong\u003e. \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats\u003cstrong\u003e \u003c/strong\u003e(n=4 per sex per genotype) were analyzed for blood levels of glucose (\u003cstrong\u003ea\u003c/strong\u003e), triglycerides (\u003cstrong\u003eb\u003c/strong\u003e), total cholesterol (\u003cstrong\u003ec\u003c/strong\u003e), HDL (\u003cstrong\u003ed\u003c/strong\u003e), LDL (\u003cstrong\u003ee\u003c/strong\u003e) and for LDL/HDL ratio (\u003cstrong\u003ef\u003c/strong\u003e).\u0026nbsp; \u003cstrong\u003eg\u003c/strong\u003e. The serum of \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats exhibited a turbid appearance reminiscent of human cases of type III HLP. Data are represented as mean ± SEM and were analyzed by two-way ANOVA followed by post hoc Tukey’s multiple comparisons test when ANOVA showed a significant difference. When the measurements were discovered to exceed the range, the nearest integer beyond the range was assigned. Statistical significance is denoted as * p\u0026lt;0.05, ** p\u0026lt;0.01, *** p\u0026lt;0.001, **** p\u0026lt;0.0001.\u003c/p\u003e","description":"","filename":"Figure4final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/1af86d821f19709270b0c1a8.png"},{"id":62944297,"identity":"cb4ba83c-3506-4711-bdae-1dfe2d533e04","added_by":"auto","created_at":"2024-08-21 10:05:26","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":897384,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSerum lipid profile of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eApoe\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e, \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE2\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e, \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE3\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e and \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eAPOE4\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e \u003cstrong\u003erats. \u003c/strong\u003eLevels of PG (\u003cstrong\u003ea\u003c/strong\u003e), PS (\u003cstrong\u003eb\u003c/strong\u003e), PE (\u003cstrong\u003ec\u003c/strong\u003e), pPE (\u003cstrong\u003ed\u003c/strong\u003e), LPE (\u003cstrong\u003ee\u003c/strong\u003e), CAR (\u003cstrong\u003ef\u003c/strong\u003e), PI (\u003cstrong\u003eg\u003c/strong\u003e), SM (\u003cstrong\u003eh\u003c/strong\u003e), PC (\u003cstrong\u003ei\u003c/strong\u003e), pPC (\u003cstrong\u003ej\u003c/strong\u003e), LPC (\u003cstrong\u003ek\u003c/strong\u003e), TAG (\u003cstrong\u003el\u003c/strong\u003e), FA (\u003cstrong\u003em\u003c/strong\u003e), total Cholesterol (\u003cstrong\u003en\u003c/strong\u003e), free Cholesterol (\u003cstrong\u003eo\u003c/strong\u003e) and Cholesterol esters (\u003cstrong\u003ep\u003c/strong\u003e) and relative ratios of TAG/PC (\u003cstrong\u003eq\u003c/strong\u003e) and FA18:1/FA 18:2 (\u003cstrong\u003er\u003c/strong\u003e)\u003cstrong\u003e \u003c/strong\u003ein serum of 80 days old \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e,\u003cem\u003e APOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e,\u003cem\u003e APOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003eand\u003cem\u003e APOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats are shown (n=4 per sex per genotype). Data are represented as mean ± SEM and were analyzed by two-way ANOVA followed by post hoc Tukey’s multiple comparisons test when ANOVA showed a significant difference. Post hoc Tukey’s Analysis is shown in Table 1 and Table 2. \u003cem\u003e* p\u0026lt;0.05, ** p\u0026lt;0.01, *** p\u0026lt;0.001, **** p\u0026lt;0.0001\u003c/em\u003e\u003c/p\u003e","description":"","filename":"Figure5final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/a3b76adf61536061a4355020.png"},{"id":62944293,"identity":"014db407-0e91-4132-bec6-68dc28154d57","added_by":"auto","created_at":"2024-08-21 10:05:25","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":663330,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSerum lipid profile of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eApoe\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e,\u003c/strong\u003e\u003cem\u003e\u003cstrong\u003e APOE3\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e\u003cstrong\u003e \u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003eand\u003c/strong\u003e\u003cem\u003e\u003cstrong\u003e APOE4\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e rats. \u003c/strong\u003eLevels of PG (\u003cstrong\u003ea\u003c/strong\u003e), PS (\u003cstrong\u003eb\u003c/strong\u003e), PE (\u003cstrong\u003ec\u003c/strong\u003e), pPE (\u003cstrong\u003ed\u003c/strong\u003e), LPE (\u003cstrong\u003ee\u003c/strong\u003e), CAR (\u003cstrong\u003ef\u003c/strong\u003e), PI (\u003cstrong\u003eg\u003c/strong\u003e), SM (\u003cstrong\u003eh\u003c/strong\u003e), PC (\u003cstrong\u003ei\u003c/strong\u003e), pPC (\u003cstrong\u003ej\u003c/strong\u003e), LPC (\u003cstrong\u003ek\u003c/strong\u003e), TAG (\u003cstrong\u003el\u003c/strong\u003e), FA (\u003cstrong\u003em\u003c/strong\u003e), total Cholesterol (\u003cstrong\u003en\u003c/strong\u003e), free Cholesterol (\u003cstrong\u003eo\u003c/strong\u003e) and Cholesterol esters (\u003cstrong\u003ep\u003c/strong\u003e) and relative ratios of TAG/PC (\u003cstrong\u003eq\u003c/strong\u003e) and FA18:1/FA 18:2 (\u003cstrong\u003er\u003c/strong\u003e)\u003cstrong\u003e \u003c/strong\u003ein serum of 80 days old \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003eand\u003cem\u003e APOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e \u003c/strong\u003erats (n=4 per sex per genotype). Data are represented as mean ± SEM and were analyzed by two-way ANOVA followed by post hoc Tukey’s multiple comparisons tests when ANOVA showed a significant difference. \u003cem\u003e* p\u0026lt;0.05, ** p\u0026lt;0.01, *** p\u0026lt;0.001, **** p\u0026lt;0.0001\u003c/em\u003e\u003c/p\u003e","description":"","filename":"Figure6final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/95f633abc25536a67c9a4602.png"},{"id":62945836,"identity":"9539cba2-3458-4ae0-b0c2-0f82f8c9c647","added_by":"auto","created_at":"2024-08-21 10:29:25","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":2966186,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eHeatmap analysis of FA, PI, PG, PS, and CAR in serum of individual rats. \u003c/strong\u003eHeatmap represents the relative levels of various lipids. Each row corresponds to a specific lipid, and each column corresponds to an individual rat. The color of each cell indicates the relative change from the mean of the same row (lipid) across all individuals. The color gradient ranges from dark blue (lowest) to red (highest). Blue colors represent lower relative levels of a particular lipid in an individual, while red colors indicate higher relative levels.\u003c/p\u003e","description":"","filename":"Figure7final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/77cc10b15ab3b59a56334516.png"},{"id":62945331,"identity":"270ea241-a65c-4ea5-a75c-b080b686586d","added_by":"auto","created_at":"2024-08-21 10:21:26","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":2875057,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eHeatmap analysis of PE and related lipid species, SM, PC and related lipid species, and TAG in serum of individual rats. \u003c/strong\u003eHeatmap represents the relative levels of various lipids. Each row corresponds to a specific lipid, and each column corresponds to an individual rat. The color of each cell indicates the relative change from the mean of the same row (lipid) across all individuals. The color gradient ranges from dark blue (lowest) to red (highest). Blue colors represent lower relative levels of a particular lipid in an individual, while red colors indicate higher relative levels.\u003c/p\u003e","description":"","filename":"Figure8final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/d19e87ef9d1d8d991c734e98.png"},{"id":62944307,"identity":"442e5865-57dd-4c99-9d7e-391b1ccf7b6e","added_by":"auto","created_at":"2024-08-21 10:05:29","extension":"png","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":2880906,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCorrelation heatmaps of top 20 correlations with p\u0026lt;0.05 for \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eApoe\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e,\u003c/strong\u003e\u003cem\u003e\u003cstrong\u003e APOE2\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e,\u003c/strong\u003e\u003cem\u003e\u003cstrong\u003e APOE3\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e\u003cstrong\u003e \u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003eand\u003c/strong\u003e\u003cem\u003e\u003cstrong\u003e APOE4\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cstrong\u003ehAβ\u003c/strong\u003e\u003c/em\u003e\u003c/sup\u003e\u003cstrong\u003e rats. \u003c/strong\u003eAll correlation coefficients reported in Supplemental Table 1.\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e","description":"","filename":"Figure9final.png","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/bdea4d39498c4c5bedc27132.png"},{"id":65627076,"identity":"b74eb009-2b4f-4ae0-a944-1cca795f32ca","added_by":"auto","created_at":"2024-09-30 16:11:15","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":16979880,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/67593dfb-a9e4-4646-8a12-917596fb7c23.pdf"},{"id":62944285,"identity":"bed33159-f165-49a7-8fac-b8910b212c7b","added_by":"auto","created_at":"2024-08-21 10:05:25","extension":"tif","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofgapdhmaleinfigure2.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/38555b1a71947276b4bdb65d.tif"},{"id":62944829,"identity":"bbbb1c93-7111-407e-a7a6-45515802292c","added_by":"auto","created_at":"2024-08-21 10:13:25","extension":"tif","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofpsd95femaleinfigure3.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/8e1315c018a9450db3b33a1c.tif"},{"id":62945346,"identity":"782028e7-5669-4f29-82c9-102fac3e6e67","added_by":"auto","created_at":"2024-08-21 10:21:26","extension":"tif","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofpsd95maleinfigure3.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/0e1fad6d97b77632cc7e6b6c.tif"},{"id":62944288,"identity":"565f061d-8215-4be3-bc50-b5510201a42d","added_by":"auto","created_at":"2024-08-21 10:05:25","extension":"tif","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofgapdhfemaleinfigure2.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/e3f3d27f953b97ac741fd422.tif"},{"id":62944836,"identity":"e2d47b84-c58f-4c29-958d-c05d47f14798","added_by":"auto","created_at":"2024-08-21 10:13:27","extension":"tif","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofAPPmaleinfigure3.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/69c91b37464bf98e44fd3429.tif"},{"id":62945325,"identity":"cd8feaef-b9cd-487d-ba79-d117c1d4a496","added_by":"auto","created_at":"2024-08-21 10:21:25","extension":"tif","order_by":6,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofAPPfemaleinfigure3.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/cc62a646173232f8e6910511.tif"},{"id":62944303,"identity":"979633a5-89b8-4999-8696-172d3c8005eb","added_by":"auto","created_at":"2024-08-21 10:05:27","extension":"tif","order_by":7,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofAPPctfmaleinfigure3.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/fe92df1553e190380bab013e.tif"},{"id":62944296,"identity":"e91da5d5-879d-490d-90ea-1e3cc353ec4b","added_by":"auto","created_at":"2024-08-21 10:05:26","extension":"tif","order_by":8,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofapoemaleinfigure2.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/1e51e69563a1b9ae70e1c353.tif"},{"id":62944301,"identity":"e68b729a-ada4-4d54-aa79-e5850d065a26","added_by":"auto","created_at":"2024-08-21 10:05:26","extension":"tif","order_by":9,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofAPPctffemaleinfigure3.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/54f1bab37aa67f496ab9bafd.tif"},{"id":62944306,"identity":"9c680a3b-0487-4cac-9e80-412b96678d72","added_by":"auto","created_at":"2024-08-21 10:05:28","extension":"tif","order_by":10,"title":"","display":"","copyAsset":false,"role":"supplement","size":14761152,"visible":true,"origin":"","legend":"","description":"","filename":"rawwbofapoefemaleinfigure2.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/84c4ed99e0c1f3858ceed44f.tif"},{"id":62944299,"identity":"7bd30c13-48ba-48fd-96a0-8643be5ad849","added_by":"auto","created_at":"2024-08-21 10:05:26","extension":"tif","order_by":11,"title":"","display":"","copyAsset":false,"role":"supplement","size":437516,"visible":true,"origin":"","legend":"","description":"","filename":"originalgelofapoe3FIG1PCRs.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/7bc83e430e5006fe1c8a88d8.tif"},{"id":62944305,"identity":"83184221-b8fe-4122-8578-e1d900351ca5","added_by":"auto","created_at":"2024-08-21 10:05:28","extension":"tif","order_by":12,"title":"","display":"","copyAsset":false,"role":"supplement","size":263160,"visible":true,"origin":"","legend":"","description":"","filename":"originalgelofapoe4FIG1PCRs.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/d9c29f36420c06d9ddfee9b2.tif"},{"id":62944300,"identity":"66818046-7653-4458-b70c-95e3ff1215c1","added_by":"auto","created_at":"2024-08-21 10:05:26","extension":"tif","order_by":13,"title":"","display":"","copyAsset":false,"role":"supplement","size":502588,"visible":true,"origin":"","legend":"","description":"","filename":"originalgelofapoe2FIG1PCRs.tif","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/ffba46427b1ce7876319b660.tif"},{"id":62944302,"identity":"54bf7564-f5af-4efa-9172-04e75aeb8910","added_by":"auto","created_at":"2024-08-21 10:05:26","extension":"xlsx","order_by":14,"title":"","display":"","copyAsset":false,"role":"supplement","size":62928,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementalTable1CorrelationCoefficients.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-4789248/v1/217b31e125358027d69710e7.xlsx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Human APOE variants in Alzheimer’s Disease and type III hyperlipoproteinemia: insights from Long-Evans knock-in rat models with humanized App and APOE","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eThe \u003cem\u003eApolipoprotein E\u003c/em\u003e (\u003cem\u003eAPOE\u003c/em\u003e) gene plays an important role in both lipid metabolism and neurological functions. In humans, there are three forms of \u003cem\u003eAPOE\u003c/em\u003e: \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e. These forms are distinguished by the presence of either arginine or cysteine residues at positions 112 and 158. \u003cem\u003eAPOE4\u003c/em\u003e, the ancestral form of \u003cem\u003eAPOE\u003c/em\u003e, is has arginine residues at positions 112 and 158 [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e] and is present in about 12% of the population. \u003cem\u003eAPOE3\u003c/em\u003e is the most common variant in contemporary human populations with a prevalence of 69\u0026ndash;82% depending on geolocation [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. \u003cem\u003eAPOE3\u003c/em\u003e emerged later through an arginine-to-cysteine substitution at position 112 of \u003cem\u003eAPOE4\u003c/em\u003e, dating back approximately 200,000 years based on time-depth analysis and natural selection assumptions [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. The fact that \u003cem\u003eAPOE3\u003c/em\u003e is more recent compared to \u003cem\u003eAPOE4\u003c/em\u003e, yet significantly more prevalent, implies a positive selection pressure favoring \u003cem\u003eAPOE3\u003c/em\u003e. \u003cem\u003eAPOE2\u003c/em\u003e, the least common variant, originated around 80,000 years ago from an arginine-to-cysteine substitution at position 158 of the \u003cem\u003eAPOE3\u003c/em\u003e gene. \u003cem\u003eAPOE\u003c/em\u003e is predominantly expressed in hepatocytes, macrophages, and astrocytes, a type of glial cell in the brain [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003cem\u003eAPOE\u003c/em\u003e is genetically linked to late-onset sporadic Alzheimer\u0026rsquo;s disease (LOAD), with \u003cem\u003eAPOE4\u003c/em\u003e being a significant risk factor, \u003cem\u003eAPOE3\u003c/em\u003e considered the \"neutral\" allele, and \u003cem\u003eAPOE2\u003c/em\u003e exerting a protective role against LOAD [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. The prevailing hypothesis linking APOE to LOAD suggests that Aβ production and deposition vary with APOE isoforms. Aβ, the primary component of amyloid plaques that characterize AD pathology, originates from sequential cleavage of the Amyloid-β Precursor Protein (APP) by β- and γ-secretases (known as the amyloidogenic processing pathway). APOE isoforms likely influence Aβ production by modulating lipid metabolism and the lipid composition of cellular membranes, with APOE4 promoting amyloidogenic APP cleavage and Aβ production more than APOE3 [\u003cspan additionalcitationids=\"CR9\" citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Moreover, APOE facilitates Aβ clearance and inhibits its aggregation through the formation of APOE-Aβ complexes, with APOE3-Aβ complexes being approximately 20 times more prevalent than APOE4-Aβ complexes [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Additionally, APOE isoforms may impact neurite growth differently [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e] and affect neuronal cell survival in an isoform-dependent manner [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eTo investigate the impact of human APOE isoforms on LOAD, we developed Long-Evans rats with the rat \u003cem\u003eApoe\u003c/em\u003e gene replaced by human \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, or \u003cem\u003eAPOE4\u003c/em\u003e variants. Given that the pathogenic mechanisms linked to \u003cem\u003eAPOE4\u003c/em\u003e may significantly influence LOAD pathogenesis through its effects on Aβ production and metabolism via direct interaction with Aβ, we crossed these human \u003cem\u003eAPOE\u003c/em\u003e replacement rats with rats carrying a humanized \u003cem\u003eApp\u003c/em\u003e allele (\u003cem\u003eApp\u003c/em\u003e\u003csup\u003e\u003cem\u003eh\u003c/em\u003e\u003c/sup\u003e allele) [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. Humanization of \u003cem\u003eApp\u003c/em\u003e targeted the Aβ region via humanization of the three amino acid differences between rodent and human Aβ. Therefore, these rat models, designated as \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, are expected to physiologically express both human \u003cem\u003eAPOE\u003c/em\u003e isoforms and human Aβ.\u003c/p\u003e \u003cp\u003eThese amino acids differences in Aβ may be crucial as human Aβ is more prone to aggregation compared to its rodent counterpart. In addition, rodent Aβ may not interact as effectively with human APOE as human Aβ does, which could reduce the ability to accurately study pathogenic and physiological mechanisms based on APOE-Aβ interactions. This setup enables the investigation of mechanisms underlying the interaction between human APOE and human Aβ. These new rat models represent an advancement compared to earlier rodent models carrying human \u003cem\u003eAPOE\u003c/em\u003e variants, as they now allow for a comprehensive exploration of the interplay between human \u003cem\u003eAPOE\u003c/em\u003e and human Aβ that was not possible in these earlier models [\u003cspan additionalcitationids=\"CR15\" citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eAPOE also plays a crucial role in lipid metabolism, both peripherally and in the central nervous system. Plasma APOE circulates in the bloodstream and is associated with chylomicron, very low-density lipoprotein (VLDL), and high-density lipoprotein (HDL) particles, playing a crucial role in their metabolism. Chylomicrons, which are derived from the intestine, and VLDL particles, which come from the liver, are lipolyzed in the bloodstream by an enzyme called lipoprotein lipase (LPL). APOE on the remnant lipoprotein particles binds to low-density lipoprotein (LDL) receptors, LDL receptor-related proteins (LRP), and heparan sulfate proteoglycans (HSPG) on the surface of liver cells [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. These remnant particles are then endocytosed by the liver cells and removed from the bloodstream. Some VLDL remnants are cleared quickly, while others undergo further lipolysis and are gradually converted into intermediate-density lipoprotein (IDL) and eventually into LDL [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. LDL particles do not contain APOE, and their removal from the bloodstream is facilitated by the binding of another protein, APOB, to the LDL receptor (LDLR) [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. APOE is also crucial for the production of VLDL particles. Its expression within liver cells promotes the assembly and secretion of VLDL particles. Optimal expression of APOE is essential for the normal metabolism of triglyceride (TG)-rich lipoproteins. However, overexpression or accumulation of APOE stimulates the production of VLDL triglycerides [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e], leading to hypertriglyceridemia. Additionally, an excess of APOE on VLDL particles can hinder their lipolysis [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e], resulting in elevated plasma triglyceride levels.\u003c/p\u003e \u003cp\u003eThe critical role of APOE in lipid metabolism is underscored by evidence showing that \u003cem\u003eAPOE2\u003c/em\u003e homozygosity can lead to Hyperlipoproteinemia type III (HLP), characterized by mixed hypercholesterolemia and hypertriglyceridemia [\u003cspan additionalcitationids=\"CR23 CR24 CR25\" citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]. This is attributed to the fact that the cysteine change at position 158 in APOE2, near the LDLR binding region, hinders \u003cem\u003eAPOE2\u003c/em\u003e binding to the LDLR [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e, \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. In addition, \u003cem\u003eAPOE4\u003c/em\u003e is linked to hypercholesterolemia and increased risk of cardiovascular disorders (CVD) [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e], although the specific underlying mechanism remains unclear [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn this study, we provide an initial characterization of \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, focusing on early effects of these \u003cem\u003eAPOE\u003c/em\u003e isoforms on APP processing and plasma lipidomics as an indicator of their influence on peripheral lipid metabolism. While the lipidomics studies do not directly focus on the LOAD-\u003cem\u003eAPOE\u003c/em\u003e link, our findings may shed light on how APOE isoforms modulate LOAD risk, potentially through effects on lipid metabolism pathways.\u003c/p\u003e"},{"header":"MATERIALS AND METHODS","content":"\u003cp\u003e\u003cb\u003eAnimals.\u003c/b\u003e All experiments were done according to policies on the care and use of laboratory animals of the Ethical Guidelines for Treatment of Laboratory Animals of the NIH. Relevant protocols were approved by the Rutgers Institutional Animal Care and Use Committee (Protocol #201702513). All efforts were made to minimize animal suffering and reduce the number of rats used.\u003c/p\u003e \u003cp\u003e \u003cb\u003eGeneration of Long-Evans Rat Models Expressing Human APOE Variants with Human APP Gene\u003c/b\u003e. gRNAs targeting vectors and the donor vector, which is flanked by homologous arms, were constructed and confirmed by sequencing (designed as shown below). The vectors\u0026rsquo; sequences are available at the indicated links.\u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003egRNA designed:\u003c/h2\u003e \u003cp\u003egRNA1 (matches forward strand of gene): AATCACAACTGGGAAGATGAAGG\u003c/p\u003e \u003cp\u003egRNA2 (Cas9_D10A) (matches reverse strand of gene): TTCATCTTCCCAGTTGTGATTGG\u003c/p\u003e \u003cp\u003egRNA3 (Cas9_D10A) (matches forward strand of gene): AATCACAACTGGGAAGATGAAGG\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003eLinks the the gRNA and donor vectors on VectorBuilder:\u003c/h2\u003e \u003cp\u003egRNA1: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.vectorbuilder.com/vector/VB171212-1056smp.html\u003c/span\u003e\u003cspan address=\"https://www.vectorbuilder.com/vector/VB171212-1056smp.html\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/p\u003e \u003cp\u003egRNA2 (Cas9_D10A): \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.vectorbuilder.com/vector/VB171212-1329ath.html\u003c/span\u003e\u003cspan address=\"https://www.vectorbuilder.com/vector/VB171212-1329ath.html\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/p\u003e \u003cp\u003egRNA3 (Cas9_D10A): \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.vectorbuilder.com/vector/VB171212-1058ubb.html\u003c/span\u003e\u003cspan address=\"https://www.vectorbuilder.com/vector/VB171212-1058ubb.html\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/p\u003e \u003cp\u003eDonor vector for \u003cem\u003eAPOE2\u003c/em\u003e plus SV40 late polyadenylation site flanked by homologous arms: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.vectorbuilder.com/vector/VB171225-1186ayg.html\u003c/span\u003e\u003cspan address=\"https://www.vectorbuilder.com/vector/VB171225-1186ayg.html\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/p\u003e \u003cp\u003eDonor vector for \u003cem\u003eAPOE3\u003c/em\u003e plus SV40 late polyadenylation site flanked by homologous arms: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.vectorbuilder.com/vector/VB171225-1189pmt.html\u003c/span\u003e\u003cspan address=\"https://www.vectorbuilder.com/vector/VB171225-1189pmt.html\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/p\u003e \u003cp\u003eDonor vector for \u003cem\u003eAPOE4\u003c/em\u003e plus SV40 late polyadenylation site flanked by homologous arms: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.vectorbuilder.com/vector/VB171225-1191dnt.html\u003c/span\u003e\u003cspan address=\"https://www.vectorbuilder.com/vector/VB171225-1191dnt.html\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/p\u003e \u003cp\u003eCas9 mRNA, gRNA generated by in vitro transcription, and oligo donor were co-injected into fertilized eggs to generate knock-in (KI) rats. The PCR primer pairs used to determine correct KI insertion and the integrity of the APOE sequences' insertions were as follows (5' to 3'):\u003c/p\u003e \u003cp\u003eSPF1: CACCCGTGGCAGAGGAATCAAC x SPR1: TTCTAGCGGGTCGGGTCGTCT\u003c/p\u003e \u003cp\u003eSPF2: CCAACCCCCTTCATCTGGATTTC x SPR2: AAAGGTCAGAATTAGGGTGGGAGG\u0026rsquo;\u003c/p\u003e \u003cp\u003eKI-1-F:TGCTCTATTGTGGAGATGTTTGTGATG x KI-1-R: GTGTGGGGGTGATGGAGAATAAAGATC\u003c/p\u003e \u003cp\u003eKI-2-F: CCACACCCGACTAACTTTTTTGTATTTTC x KI-2-R: TCAACTCCTTCATGGTCTCGTCCATC\u003c/p\u003e \u003cp\u003eKI-3-F: GCCTCCTAGCTCCTTCTTCGTCTCTG x KI-3-R: CAGGCGTATCTGCTGGGCCTG\u003c/p\u003e \u003cp\u003eKI-4-F: TAAGCGGCTCCTCCGCGATG x KI-4-R: AGCAGAATCGCTTGAACCCAAGAG\u003c/p\u003e \u003cp\u003eKI-5-F: CCTCAGTTTCTCTTTCTGCCCACATA x KI-5-R: TATTATGGATAGGGAAAGACAAGGCC\u003c/p\u003e \u003cp\u003eThe primers used for the Southern blot analysis were:\u003c/p\u003e \u003cp\u003e5\u0026rsquo; arm Probe: F: CCAAGATTATACATCCGGCAACCG x R: GGCTGGAGGCTTAAATGGAAATAGG\u003c/p\u003e \u003cp\u003e3\u0026rsquo; arm Probe: F: TGTTGGTCCCATTGCTGACAGGTA x R: AAGCAACAGTGCGTCTGGAAGTCAG\u003c/p\u003e \u003cp\u003eTo generate \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, we doubly crossed humanized \u003cem\u003eAPOE\u003c/em\u003e rats described above with \u003cem\u003eApp\u003c/em\u003e\u003csup\u003e\u003cem\u003eh/h\u003c/em\u003e\u003c/sup\u003e rats that carries \u003cem\u003eApp\u003c/em\u003e genes with the humanized Aβ sequence. The \u003cem\u003eApp\u003c/em\u003e\u003csup\u003e\u003cem\u003eh\u003c/em\u003e\u003c/sup\u003e allele enables the physiological production of human Aβ instead of rodent Aβ from the endogenous rat \u003cem\u003eApp\u003c/em\u003e gene [\u003cspan additionalcitationids=\"CR31 CR32 CR33 CR34 CR35\" citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003cb\u003eProtein preparation.\u003c/b\u003e These procedures were performed as previously described [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. Briefly, the rats were first put under anesthesia using isoflurane, followed by perfusion through intracardiac catheterization using ice-cold PBS. Brains were extracted and homogenized with a glass-teflon homogenizer in 250 mM Sucrose, 20 mM Tris-base pH 7.4, 1 mM EDTA, 1 mM EGTA plus protease and phosphatase inhibitors (Thermo Scientific). All steps were carried out on the ice. Homogenates were solubilized with 1% NP-40 for 30 min rotating and spun at 20,000 g for 10 min. Supernatants were collected and protein content was quantified using the Bradford method.\u003c/p\u003e \u003cp\u003e \u003cb\u003eQuantitative RT-PCR\u003c/b\u003e. Total brain RNA was extracted using the RNeasy RNA Isolation kit (Qiagen) and converted to cDNA using the High-Capacity cDNA Reverse Transcription Kit (Thermo Fisher) with oligo dT priming. For each reaction, 50 ng of cDNA, TaqMan\u0026trade; Fast Advanced Master Mix (Thermo Fisher 4444556), and the appropriate TaqMan probes (Thermo Fisher) were used. Real-time PCR was conducted on a QuantStudio 6 Flex Real-Time PCR System (Thermo Fisher). Relative RNA quantification was performed using LinRegPCR software (hartfaalcentrum.nl). The rat \u003cem\u003eApoe\u003c/em\u003e transcript was detected using probe Rn00593680_m1 targeting exon junction 3\u0026ndash;4, while human \u003cem\u003eAPOE\u003c/em\u003e was detected using probe Hs00171168_m1 for the same exon junction. Amplification data were normalized to rat \u003cem\u003eGapdh\u003c/em\u003e expression, assessed using probe Rn01775763_g1.\u003c/p\u003e \u003cp\u003e \u003cb\u003eELISA.\u003c/b\u003e For analysis of human APOE, Aβ38, Aβ40, Aβ42, sAPPα and sAPPβSw, the following Meso Scale Discovery kits were used: levels of APOE were measured with R-PLEX Human ApoE Assay (K151AMLR) (serum samples were diluted 1:20,000), Aβ38, Aβ40, and Aβ42 were measured with V-PLEX Plus Aβ Peptide Panel 1 6E10 (K15200G); sAPPα and sAPPβ were measured with sAPPα/sAPPβ kit (K15120E). Measurements were performed according to the manufacturer\u0026rsquo;s recommendations. Plates were read on a MESO QuickPlex SQ 120. For analysis of Aβ43, IBL Human Amyloidβ (1\u0026ndash;43) (FL) Assay Kit (27710) was used according to the manufacturer\u0026rsquo;s recommendations.\u003c/p\u003e \u003cp\u003e \u003cb\u003eWestern blots (WB).\u003c/b\u003e WB were performed as follows: proteins were diluted with PBS and LDS Sample Buffer (Invitrogen NP0007) containing 10% β-mercaptoethanol, and 4.5M urea to a concentration of 1 \u0026micro;g/\u0026micro;l. Samples were loaded onto a 4\u0026ndash;12% Bis-Tris polyacrylamide gel (Biorad 3450125) and transferred onto nitrocellulose membranes at 25 V for 7 minutes using the Trans-Blot Turbo system (Biorad). Blotting efficiency was confirmed by red Ponceau staining of the membranes.\u003c/p\u003e \u003cp\u003eMembranes were blocked for 45 minutes in 5% milk (Biorad 1706404), followed by extensive washing in PBS/Tween20-0.05%. Primary antibodies (anti-APOE Rabbit mAb, Cell Signaling Technology, 10197SF; anti-APP (Y188) Rabbit mAb, Abcam, Ab32136; anti-GAPDH Rabbit mAb, Sigma, g9545; anti-PSD95, Cell Signaling Technology, 3450) were applied overnight at 4\u0026deg;C.\u003c/p\u003e \u003cp\u003eAfter washing three times for 10 minutes each with PBS/Tween20-0.05%, membranes were incubated with a mixture of HRP-conjugated anti-rabbit secondary antibodies (Southern Biotech, OB405005 and Cell Signaling Technology, 7074) diluted 1:1,000 in 5% milk for 45 minutes at room temperature with shaking. Blots were developed using Clarity Western ECL reagent (Bio-rad 1705061) and visualized on a ChemiDoc MP Imaging System (Bio-Rad). For WB quantifications, signal intensities were analyzed with Image Lab software (Bio-Rad).\u003c/p\u003e \u003cp\u003e \u003cb\u003eBlood glucose and lipid profile measurements.\u003c/b\u003e Blood glucose levels were measured using the ACCU-CHEK Guide Me system (Roche, mg/dL). For the blood lipid profile, including total cholesterol (TC, range 100\u0026ndash;450 mg/dL), HDL (range 25\u0026ndash;95 mg/dL), LDL (calculated), and TG (range 45\u0026ndash;450 mg/dL), a CURO L7 Lipid Analyzer (CUROfit, CA) cholesterol home test kit was utilized. Briefly, blood was collected via cardiac puncture from 80-day-old rats (with a sample size of 4 per sex per genotype). 35 \u0026micro;L of fresh blood was applied to a lipid profile test strip and measured after 3 minutes. Measurements falling below the specified range were rounded up to the closest integer within the lower end, while those above the range were rounded down to the closest integer within the upper end. LDL levels were calculated using the Friedewald equation: LDL (mg/dL)\u0026thinsp;=\u0026thinsp;TC - HDL - (TG/5).\u003c/p\u003e \u003cp\u003e \u003cb\u003eLipidomic analysis.\u003c/b\u003e Serum for lipidomic analysis was collected from the same rats used for blood glucose and lipid profile measurements. Blood was drawn into serum separator tubes (BD Becton Dickinson vacutainers, SSTTM) and left to incubate at room temperature for 30 minutes. The tubes were then centrifuged at 2000\u0026times;g for 10 minutes to separate the serum, which was subsequently stored at -80\u0026deg;C.\u003c/p\u003e \u003cp\u003eFor lipidomic analysis, the protein content of the thawed serum samples was quantified using a BCA protein assay kit (Pierce, Rockford, IL, USA). A mixture of approximately 20 internal standards was added to serum samples based on the protein content for quantification of individual lipid molecular species as previously described [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e]. The extraction of lipids was carried out using a modified Bligh and Dyer extraction method, as previously described [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e]. Multi-dimensional mass spectrometry (MDMS)-based shotgun lipidomics (MDMS-SL) was performed using electrospray ionization mass spectrometry (ESI/MS) to measure individual lipid molecular species [\u003cspan additionalcitationids=\"CR41\" citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]. Instrumentation utilized was a triple-quadruple mass spectrometer (Thermo Scientific TSQ Altis, San Jose, CA, USA) equipped with a NanoMate device (Advion Bioscience Ltd., Ithaca, NY, USA). Xcalibur system software was utilized for this process. Data processing included several steps such as ion peak selection, baseline correction, data transfer, peak intensity comparison, 13C deisotoping, and quantitation. These steps were conducted using a custom-programmed Microsoft Excel macro, as previously described [\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e]. The concentrations of the total lipid class populations were calculated by summing the individually detected analytes that belonged to the class.\u003c/p\u003e \u003cp\u003e \u003cb\u003eStatistical analysis.\u003c/b\u003e Data were analyzed using GraphPad Prism software and expressed as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SEM. Statistical tests used to evaluate significance and statistical data are shown in Figure legends and \u003cspan refid=\"Sec5\" class=\"InternalRef\"\u003eResults\u003c/span\u003e section. Significant differences were accepted at P\u0026thinsp;\u0026lt;\u0026thinsp;0.05.\u003c/p\u003e \u003c/div\u003e"},{"header":"RESULTS","content":"\u003cp\u003e \u003cb\u003eGeneration of Long-Evans rat models expressing either human\u003c/b\u003e \u003cb\u003eAPOE2\u003c/b\u003e, \u003cb\u003eAPOE3\u003c/b\u003e \u003cb\u003eor\u003c/b\u003e \u003cb\u003eAPOE4\u003c/b\u003e \u003cb\u003evariant genes and the humanized\u003c/b\u003e \u003cb\u003eApp\u003c/b\u003e \u003cb\u003erat gene.\u003c/b\u003e The rat \u003cem\u003eApoe\u003c/em\u003e gene, with GenBank accession number NM_138828.3 and Ensembl ID ENSRNOG00000018454, is on rat chromosome 1. The gene comprises 4 exons, with the ATG start codon located in exon 2 and the TGA stop codon located in exon 4. Long-Evans rat models expressing human \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e variants were separately generated using CRISPR/Cas-mediated genome engineering. To achieve this, the ATG start codon in exon 2 of the rat \u003cem\u003eApoe\u003c/em\u003e gene was replaced with coding sequences for human \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, or \u003cem\u003eAPOE4\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea). These human APOE coding sequences were linked to the SV40 late polyadenylation site. By employing the gene manipulation strategy as described above, the expression of human \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e variants, which replaces the expression of the rat \u003cem\u003eApoe\u003c/em\u003e gene, is controlled using the regulatory elements of the rat \u003cem\u003eApoe\u003c/em\u003e gene. This allows for the accurate and specific regulation of the human \u003cem\u003eAPOE\u003c/em\u003e variants' expression in the Long-Evans rat models. To confirm CRISPR-induced mutations, the resulting pups underwent genotyping by PCR, followed by sequence analysis. PCR was initially performed using primers SPF1 x SPR1 and SPF2 x SPR2 to identify founder rats (F0). Based on this screening, rat 10 was selected as an F0 rat for \u003cem\u003eAPOE2\u003c/em\u003e, rat 26 as an F0 for \u003cem\u003eAPOE3\u003c/em\u003e, and rats 6 and 7 as F0s for \u003cem\u003eAPOE4\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb). Subsequently, the same PCR method was used to identify and designate certain offspring of these F0 rats as \u003cem\u003eAPOE2\u003c/em\u003e F1s (rats 1, 5, 6, 7, and 9), \u003cem\u003eAPOE3\u003c/em\u003e F1s (rats 58, 62, and 63), and \u003cem\u003eAPOE4\u003c/em\u003e F1s (rats 59, 67, 76, and 87) based on their respective parentage (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb). The selection of F0 and F1 rats was further validated by PCR using 5 primer pairs (KI-1-F x KI-1-R, KI-2-F x KI-2-R, KI-3-F x KI-3-R, KI-4-F x KI-4-R, and KI-5-F x KI-5-R) followed by sequencing to confirm the genotyping results. The accurate gene targeting in F1 animals was verified through Southern blot analysis of the tail DNA samples. The Southern blot analysis strategy is depicted in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec. The results demonstrated that all F1 rats analyzed expressed both the rat \u003cem\u003eApoe\u003c/em\u003e allele and the human \u003cem\u003eAPOE\u003c/em\u003e knock-in (KI) allele in a 1:1 ratio (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed). F1 rats were crossed to Long Evans for 5 generations. The probability that F5 rats carry unidentified off-target insertions/mutations (except those that may be on Chr. 1) is ~\u0026thinsp;1.5625%.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cem\u003eAPOE\u003c/em\u003e isoforms have been associated with the risk of LOAD, with \u003cem\u003eAPOE4\u003c/em\u003e increasing the risk and \u003cem\u003eAPOE2\u003c/em\u003e reducing it. Considering the important role of \u003cem\u003eAPP\u003c/em\u003e in LOAD pathogenesis and the interplay between APP, its metabolic product Aβ, and APOE isoforms, we crossed \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e rats with animals carrying a rat \u003cem\u003eApp\u003c/em\u003e allele humanized specifically in the Aβ region (\u003cem\u003eApp\u003c/em\u003e\u003csup\u003e\u003cem\u003eh/h\u003c/em\u003e\u003c/sup\u003e rats). The resulting progeny, double heterozygous for \u003cem\u003eAPOE\u003c/em\u003e and \u003cem\u003eApp\u003c/em\u003e\u003csup\u003e\u003cem\u003eh\u003c/em\u003e\u003c/sup\u003e alleles, were further bred to generate rats homozygous for humanized \u003cem\u003eAPOE\u003c/em\u003e and \u003cem\u003eApp\u003c/em\u003e\u003csup\u003e\u003cem\u003eh\u003c/em\u003e\u003c/sup\u003e, thereby exclusively producing human Aβ species in a physiological manner. These models were designated as \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e. \u003cem\u003eAPP\u003c/em\u003e is expressed in virtually all types of cells, suggesting broader functions beyond the central nervous system (CNS). Studying these double humanized rats could offer significant advantages, particularly in exploring how human APOE's systemic functions related to lipid metabolism are influenced by its interactions with human Aβ. Utilizing \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats for this lipid metabolism study may yield valuable insights into their potential interactions without apparent drawbacks.\u003c/p\u003e \u003cp\u003e \u003cb\u003eSerum and brain APOE levels are highest in\u003c/b\u003e \u003cb\u003eAPOE2\u003c/b\u003e\u003csup\u003e\u003cb\u003ehAβ\u003c/b\u003e\u003c/sup\u003e \u003cb\u003erats.\u003c/b\u003e To characterize these models, we first measured human APOE levels in blood serum and brain. Since \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats have rat \u003cem\u003eApoe\u003c/em\u003e, we excluded them from this analysis and used them as negative control. Serum APOE levels were highest in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats in both males and females compared to \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.2930, P\u0026thinsp;=\u0026thinsp;0.5949; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;95.68, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.5054, P\u0026thinsp;=\u0026thinsp;0.6116. For males \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.6589. For females \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.9979). Remarkably, this pattern of APOE isoform levels mirrors that found in human serum [\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e]. Brain APOE levels measured by ELISA were also the highest in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats compared to \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats in both sexes (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;15.13, P\u0026thinsp;=\u0026thinsp;0.0007; genotype factor, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;68.7, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.765, P\u0026thinsp;=\u0026thinsp;0.0087. For males \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0022**; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.8481. For females \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.4170.). Additionally, in the brains of \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e females, APOE level was higher than male \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (female vs male p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****). The ELISA results were confirmed by WB analysis of brain APOE levels (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ec) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.6498, P\u0026thinsp;=\u0026thinsp;0.4278; genotype factor, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;75.87, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.743, P\u0026thinsp;=\u0026thinsp;0.0837. For males \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0013**; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.2056. For females \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.90380).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTo investigate whether the increase in APOE2 protein levels is due to enhanced transcription, we measured mRNA levels of human \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats. Overall, these mRNA levels were comparable, with the notable exception of a decrease in \u003cem\u003eAPOE2\u003c/em\u003e mRNA levels in male \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e compared to male \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ed), despite APOE2 protein levels being significantly higher than APOE3 levels in male rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea and \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb). Therefore, differences in protein expression are likely attributable to translational and/or protein stability variances.\u003c/p\u003e \u003cp\u003e \u003cb\u003eSex- and APOE isoform-dependent variations in brain APP metabolites\u003c/b\u003e. APP is a substrate of several proteases. α-Secretase cleaves APP to produce a soluble ectodomain (sAPPα) and a membrane-bound C-terminal fragment (αCTF). Alternatively, β-secretase cleaves APP to generate the soluble sAPPβ ectodomain and the membrane-tethered βCTF stub. Subsequent cleavage of βCTF by γ-secretase leads to the production of Aβ peptides and the short intracellular domain of APP [\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e]. APOE modulates APP metabolism in an isoform-dependent manner through distinct mechanisms: 1) influencing APP processing via effects on lipid metabolism [\u003cspan additionalcitationids=\"CR9\" citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e], and 2) modulating Aβ clearance and aggregation via the formation of APOE-Aβ complexes [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Consequently, we investigated APP metabolites in the CNS of \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats. ELISA experiments showed no significant differences in Aβ38 levels among humanized \u003cem\u003eAPOE\u003c/em\u003e variants (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea; \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e-way ANOVA summary; sex factor, F\u003csub\u003e(1, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.717, P\u0026thinsp;=\u0026thinsp;0.1989; genotype factor, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.978, P\u0026thinsp;=\u0026thinsp;0.1358; sex x genotype interaction, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.978, P\u0026thinsp;=\u0026thinsp;0.1358). In female rats, Aβ40 levels were higher in \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e compared to \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;10.53, P\u0026thinsp;=\u0026thinsp;0.0026; genotype factor, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.083, P\u0026thinsp;=\u0026thinsp;0.0051; sex x genotype interaction, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.677, P\u0026thinsp;=\u0026thinsp;0.1902. For females \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0238*). There was no significant difference in Aβ42 levels across the humanized \u003cem\u003eAPOE\u003c/em\u003e variants (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec; \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e-way ANOVA summary; sex factor, F\u003csub\u003e(1, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.449, P\u0026thinsp;=\u0026thinsp;0.5073; genotype factor, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.009, P\u0026thinsp;=\u0026thinsp;0.1311; sex x genotype interaction, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.901, P\u0026thinsp;=\u0026thinsp;0.1480). However, the levels of Aβ43, a determining factor in the onset of pathological amyloid deposition [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e], were higher in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males compared to \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ed) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.840, P\u0026thinsp;=\u0026thinsp;0.1838; genotype factor, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.781, P\u0026thinsp;=\u0026thinsp;0.0558; sex x genotype interaction, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.735, P\u0026thinsp;=\u0026thinsp;0.0587. For males \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0146*). The levels of sAPPα showed gender-specific variations (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ee; \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e-way ANOVA summary; sex factor, F\u003csub\u003e(1, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;39.75, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; genotype factor, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.152, P\u0026thinsp;=\u0026thinsp;0.3424; sex x genotype interaction, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;3.222, P\u0026thinsp;=\u0026thinsp;0.0347). \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e female rats exhibited higher sAPPα levels compared to males of the same genotype (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ee) (for \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e male vs female p\u0026thinsp;=\u0026thinsp;0.0195*; For \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e male vs female p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****). Finally, sAPPβ levels were higher in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e females compared to all other female groups and \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e male rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ef) (2-way ANOVA summary; : sex factor, F\u003csub\u003e(1, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;25.04, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; genotype factor, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;6.863, P\u0026thinsp;=\u0026thinsp;0.0010; sex x genotype interaction, F\u003csub\u003e(3, 34)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;7.099, P\u0026thinsp;=\u0026thinsp;0.0008. For females \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0059**; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0023**; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****. For \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e male vs. female p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eLevels of APP, αCTF and βCTF were quantified by WB, as shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eg. APP levels were similar except for \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males which showed significant lower levels compared to \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e females (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eh)(2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;12.08, P\u0026thinsp;=\u0026thinsp;0.0019; genotype factor, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.0983, P\u0026thinsp;=\u0026thinsp;0.9067; sex x genotype interaction, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.2287, P\u0026thinsp;=\u0026thinsp;0.7972. Males \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. females \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0106*). In contrast, αCTF levels were significantly higher in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males compared to \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e females (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eh)(2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;8.588, P\u0026thinsp;=\u0026thinsp;0.0071; genotype factor, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.7281, P\u0026thinsp;=\u0026thinsp;0.4928; sex x genotype interaction, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.612, P\u0026thinsp;=\u0026thinsp;0.0933. Males \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. females \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0029**). Finally, βCTF showed a more complex pattern (Figure h) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.6420, P\u0026thinsp;=\u0026thinsp;0.4306; genotype factor, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.157, P\u0026thinsp;=\u0026thinsp;0.1368; sex x genotype interaction, F\u003csub\u003e(2, 25)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;8.787, P\u0026thinsp;=\u0026thinsp;0.0013). \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males showed significantly lower levels compared to \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males (males, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0006***) and \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e females (\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e male vs. female p\u0026thinsp;=\u0026thinsp;0.0092**), and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males showed significantly higher levels compared to \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e females (\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e male vs. female p\u0026thinsp;=\u0026thinsp;0.0065**),\u003c/p\u003e \u003cp\u003eOverall, these findings demonstrated distinct regulation of APP metabolites in relation to \u003cem\u003eAPOE\u003c/em\u003e variants, with notable sex differences.\u003c/p\u003e \u003cp\u003e \u003cb\u003eIncreased triglyceride and cholesterol levels in\u003c/b\u003e \u003cb\u003eAPOE2\u003c/b\u003e\u003csup\u003e\u003cb\u003ehAβ\u003c/b\u003e\u003c/sup\u003e \u003cb\u003erats.\u003c/b\u003e Next, we measured blood glucose, triglyceride, cholesterol, LDL and HDL levels using home test kits while collecting blood via cardiac puncture, to assess the metabolic health of our rats after a 16-hour fasting period. No significant variance was observed in the levels of blood glucose (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea; \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e-way ANOVA summary; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;10.40, P\u0026thinsp;=\u0026thinsp;0.0036; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.6423, P\u0026thinsp;=\u0026thinsp;0.5953; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.389, P\u0026thinsp;=\u0026thinsp;0.2701). In contrast, both blood triglyceride (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb; \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e-way ANOVA summary; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.697, P\u0026thinsp;=\u0026thinsp;0.2051; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;35.47, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.1583, P\u0026thinsp;=\u0026thinsp;0.9233. For males \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0013**; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0014**; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026gt;\u0026thinsp;0.9999. For females \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0019**; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0002***; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.9848) and cholesterol levels (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec; \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e-way ANOVA summary; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.798, P\u0026thinsp;=\u0026thinsp;0.1925; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;35.47, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.5164, P\u0026thinsp;=\u0026thinsp;0.6749. For males \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0006***; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0037**; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0102*; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.9998. For females \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0173*; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0074**; \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026gt;\u0026thinsp;0.9999) were significantly higher in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e compared to \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats. In addition, the serum of \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats exhibited a turbid appearance (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eg) reminiscent of human cases of type III HLP [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. This observation, coupled with evident mixed hypercholesterolemia and hypertriglyceridemia, supports the characterization of \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats as a model for type III HLP, a condition prevalent in a significant fraction of individuals homozygous for APOE2 [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e, \u003cspan additionalcitationids=\"CR24 CR25\" citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eHDL levels were similar in all humanized \u003cem\u003eAPOE\u003c/em\u003e groups, but lower compared to \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e group (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ed) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.8298, P\u0026thinsp;=\u0026thinsp;0.3714; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;24.69, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.178, P\u0026thinsp;=\u0026thinsp;0.1168. For males \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001****; \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0090**; \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0020**. For females \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0076**; \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0065**; \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0010***). Moreover, the level of LDL is similar in all humanized \u003cem\u003eAPOE\u003c/em\u003e groups. But LDL levels were higher in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats compared to \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ee) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.4476, P\u0026thinsp;=\u0026thinsp;0.5098; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;11.07, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.4133, P\u0026thinsp;=\u0026thinsp;0.7450. For males \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0163*. For females \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0058**). As a result, the LDL/HDL ratio was much higher in male \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e compared to male \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, but there was no difference between groups in females (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ef) (2-way ANOVA summary; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.0244, P\u0026thinsp;=\u0026thinsp;0.8771; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.385, P\u0026thinsp;=\u0026thinsp;0.0003; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.372, P\u0026thinsp;=\u0026thinsp;0.2752. For males \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0040**; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0288*; \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0796). The LDL/HDL ratio is used to predict Coronary Heart Disease (CHD), with a higher ratio indicating a higher risk of CHD [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003cb\u003eSex- and APOE isoform-dependent variations in serum lipidomics.\u003c/b\u003e A comprehensive lipidomic analysis was conducted to measure 222 lipid molecular species in serum samples of \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, after scanning thousands of lipid molecular species. The lipid species included 3 phosphatidylglycerol (PG) species, 3 phosphatidylserine (PS) species, 21 phosphatidylethanolamine (PE) species, 13 plasmalogen PE (pPE) species, 12 lyso PE (LPE) species, 11 acylcarnitine (CAR) species, 6 phosphatidylinositol (PI) species, 20 sphingomyelin (SM) species, 31 phosphatidylcholine (PC) species, 4 plasmalogen PC (pPC) species, 12 lyso PC (LPC) species, 59 types of triacylglycerol (TAG) without deconvolution of individual molecular species, 11 types of fatty acyl chains in TAG (FA), total cholesterol, free cholesterol, and 14 types of cholesterol esters.\u003c/p\u003e \u003cp\u003eThe results revealed significant differences in levels of various lipid molecular species between the different \u003cem\u003eAPOE\u003c/em\u003e genotypes. Notably, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats displayed markedly elevated levels of serum PG, PS, PE, pPE, SM, LPC, TAG, FA, total cholesterol, free cholesterol, and cholesterol esters compared to \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats in both males and females(PG is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ea; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.492, P\u0026thinsp;=\u0026thinsp;0.1275; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;125.3, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.1789, P\u0026thinsp;=\u0026thinsp;0.9096. PS is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eb; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.3257, P\u0026thinsp;=\u0026thinsp;0.5735; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;260.1, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.486, P\u0026thinsp;=\u0026thinsp;0.0849. PE is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ec; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.382, P\u0026thinsp;=\u0026thinsp;0.2512; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;63.46, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.978, P\u0026thinsp;=\u0026thinsp;0.0515. pPE is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ed; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.117, P\u0026thinsp;=\u0026thinsp;0.1586; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;69.44, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.014, P\u0026thinsp;=\u0026thinsp;0.1388 SM is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eh; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.0049, P\u0026thinsp;=\u0026thinsp;0.9444; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;82.17, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.08844, P\u0026thinsp;=\u0026thinsp;0.9657. LPC is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ek; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.024, P\u0026thinsp;=\u0026thinsp;0.3217; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;28.38, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.421, P\u0026thinsp;=\u0026thinsp;0.261. TAG is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003el; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.519, P\u0026thinsp;=\u0026thinsp;0.1256; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;70.46, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.2196, P\u0026thinsp;=\u0026thinsp;0.8818. FA is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003em; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.318, P\u0026thinsp;=\u0026thinsp;0.141; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;71.29, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.2084, P\u0026thinsp;=\u0026thinsp;0.8896. Cholesterol-Total is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003en; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.158, P\u0026thinsp;=\u0026thinsp;0.2927; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;74.78, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.8491, P\u0026thinsp;=\u0026thinsp;0.4807. Cholesterol-Free is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eo; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.0239, P\u0026thinsp;=\u0026thinsp;0.3460; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;127.6, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.2969, P\u0026thinsp;=\u0026thinsp;0.8273. Cholesterol-Ester is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ep; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.103, P\u0026thinsp;=\u0026thinsp;0.3041; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;45.94, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.105, P\u0026thinsp;=\u0026thinsp;0.3666.). This indicates a distinct lipid profile associated with the \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e genotype (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e) (see Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e for multiple comparisons).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePost hoc Tukey\u0026rsquo;s Analysis: p-Values for Multiple Comparisons of Various Lipids.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"14\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c12\" colnum=\"12\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c13\" colnum=\"13\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c14\" colnum=\"14\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePG\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePS\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePE\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003epPE\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eLPE\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eCAR\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003ePI\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eSM\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003ePC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c11\"\u003e \u003cp\u003epPC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c12\"\u003e \u003cp\u003eLPC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c13\"\u003e \u003cp\u003eTAG\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c14\"\u003e \u003cp\u003eFA\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003evs.♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0031\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.5878\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.4875\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0235\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.2004\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9901\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.6910\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.9265\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.5522\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.9434\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.7490\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.7492\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.8712\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.8751\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9388\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.9998\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.9696\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.1423\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.0392\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.7406\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.9667\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.3953\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0893\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.9918\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.9928\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.8432\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9904\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.4333\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.5589\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0424\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0050\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.0033\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.9526\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.9630\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0007\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.0004\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.6346\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.0740\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9901\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9561\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.9989\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.9679\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.9998\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.7642\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.9941\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.2140\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.3535\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.1697\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.1800\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9988\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9991\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.9352\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.9734\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.9656\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.7112\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.6869\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.3542\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0285\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.8855\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.9061\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003evs. ♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.1396\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0861\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.0120\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0005\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0003\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0021\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.0003\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.0002\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.0006\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.9473\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.2633\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.0002\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.1521\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.0002\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.2001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9963\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.1967\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.6931\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.5118\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.6229\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0574\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.8368\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.8801\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.5854\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.9721\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.9817\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0339\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.8409\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.6325\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0003\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.3461\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.0867\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.1008\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.1720\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.7425\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.7973\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.9997\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.9968\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.9987\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.8292\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.9991\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.9991\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9244\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9998\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.8919\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.3764\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.6909\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0003\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.8510\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.0014\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.1753\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.1923\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.9644\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.9669\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0024\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0431\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9995\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9998\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.9968\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.7248\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.4121\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.9998\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.9733\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.9972\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.8679\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.8772\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.2661\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.8567\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.9877\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.1797\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.9928\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.9840\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.9993\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.8652\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.9972\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.5188\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.6409\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9998\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9748\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.3066\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0041\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.5802\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.0003\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.0542\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0088\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.9997\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.9997\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0014\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.0338\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9998\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.4931\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0580\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.0232\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.9930\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.5586\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.5678\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9993\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.6820\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.9957\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.2929\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.2865\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.9968\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.9993\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.9994\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.9997\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.1126\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.9917\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.0067\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.0058\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.9944\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.6781\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9629\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.2485\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.2972\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0143\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.6064\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.0005\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.0229\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0512\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.2909\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.3083\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9099\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.1781\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.0833\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0044\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0018\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e0.5488\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e0.0460\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0024\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.9704\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.9791\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.0005\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0021\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0094\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.0003\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.0004\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.1039\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.7253\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.9988\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.5078\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.9872\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.9045\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c9\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c10\"\u003e \u003cp\u003e0.9774\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e \u003cp\u003e0.8879\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e \u003cp\u003e0.8524\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e \u003cp\u003e0.8416\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePost hoc Tukey\u0026rsquo;s Analysis: p-Values for Multiple Comparisons of Cholesterol Species\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCholesterol-Total\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCholesterol-Free\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eCholesterol-Esters\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.0005\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9996\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9929\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9994\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9889\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9966\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9870\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9991\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9987\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9950\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9803\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9996\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9592\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.0019\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.6360\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9638\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.5012\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9484\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9602\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9606\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.9995\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9991\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.8189\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.8556\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.8525\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♂:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.8983\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.9594\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.8956\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.7383\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.8685\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.7378\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.0001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e♀:\u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. ♀:\u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;0.9999\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eThe levels of LPE, CAR, PI, and PC showed gender-dependent variations in \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (LPE is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ee; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.143, P\u0026thinsp;=\u0026thinsp;0.0326; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;27.96, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;3.351, P\u0026thinsp;=\u0026thinsp;0.0357. CAR is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ef; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;33.52, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;21.55, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.325, P\u0026thinsp;=\u0026thinsp;0.2894. PI is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eg; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;82.76, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;49.39, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;7.462, P\u0026thinsp;=\u0026thinsp;0.0011. PC is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ei; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.798, P\u0026thinsp;=\u0026thinsp;0.0241; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;71.09, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.659, P\u0026thinsp;=\u0026thinsp;0.0044. pPC is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ej; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.4159, P\u0026thinsp;=\u0026thinsp;0.5251; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;59.09, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.165, P\u0026thinsp;=\u0026thinsp;0.3436.). In male rats, the levels of these lipid species were lower compared to female rats. Moreover, within the \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, males exhibited lower levels of PI compared to females (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e) (see Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e for multiple comparisons).\u003c/p\u003e \u003cp\u003eFurthermore, a comparison among the humanized \u003cem\u003eAPOE\u003c/em\u003e genotypes revealed additional insights. Female \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats exhibited lower levels of LPE and CAR compared to female \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats. Additionally, male \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats displayed higher levels of LPE compared to male \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, and higher levels of CAR compared to male \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, females showed reduced levels of PI, PC, and pPC compared to \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats of the same sex. Additionally, male \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats had lower levels of PI compared to male \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, with male \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats also displaying lower levels of PI compared to male \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e. Moreover, male \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats exhibited higher levels of PC, pPC, and LPC compared to male \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eWe also analyzed the ratios of TAG to PC and FA 18:1 to FA 18:2 which are indicators of the size of lipoproteins and FA profiles in TAG pools, respectively, since FA 18:1 largely represents the de novo synthesized pool and FA 18:2 represents the portion from dietary uptake. \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats have much higher TAG/PC ratio compared to all other variants in male animals, indicating a relatively larger size of lipoprotein particles in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e male rats compared to the other counterparts (TAG/PC is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eq; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.472, P\u0026thinsp;=\u0026thinsp;0.1290; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;20.62, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.071, P\u0026thinsp;=\u0026thinsp;0.3798.). But, in females TAG/PC ratio was higher in both \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e compared to \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eq). Moreover, FA18:1/FA 18:2 ratio was lower in male \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats compared to females (FA18:1/ FA18:2 is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003er; sex factor, F\u003csub\u003e(1, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;23.33, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; genotype factor, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;20.65, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(3, 24)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.608, P\u0026thinsp;=\u0026thinsp;0.0749\u003cb\u003e)\u003c/b\u003e. Also, FA18:1/FA 18:2 ratio was lower in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e compared to \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e in both sexes. A lower FA18:1/FA 18:2 ratio was manifest in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e females compared to \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003er). These different FA ratios clearly indicate the different lipid metabolism at the FA levels.\u003c/p\u003e \u003cp\u003eGiven the markedly elevated lipid levels in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e relative to other variants, it becomes impractical to compare lipid changes among \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e. Consequently, in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e, we have selectively excluded the \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, thereby replicating the analytical approach employed in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e without incorporating the \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e dataset. First, we observed that both sexes of \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats had lower levels of PG compared to \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ea) (PG is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ea; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.437, P\u0026thinsp;=\u0026thinsp;0.1359; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;11.39, P\u0026thinsp;=\u0026thinsp;0.0006; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.3179, P\u0026thinsp;=\u0026thinsp;0.7317.) (For male \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0415*. For females \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e vs. \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e p\u0026thinsp;=\u0026thinsp;0.0429*).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIn male \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals, the levels of PE, pPE, LPE, CAR, PI, PC, total cholesterol, and free cholesterol were found to be lower compared to their female counterparts. In \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals, females exhibited lower levels of PE, pPE, LPE, CAR, PI, SM, PC, pPC, LPC, total cholesterol, free cholesterol, cholesterol esters, and the ratio of FA18:1/FA18:2 when compared to female \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals (PE is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ec; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.3314, P\u0026thinsp;=\u0026thinsp;0.5719; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.552, P\u0026thinsp;=\u0026thinsp;0.2388; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;8.314, P\u0026thinsp;=\u0026thinsp;0.0028. pPE is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ed; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.776, P\u0026thinsp;=\u0026thinsp;0.1993; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;21.43, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;12.23, P\u0026thinsp;=\u0026thinsp;0.0004. LPE is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ee; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.997, P\u0026thinsp;=\u0026thinsp;0.1005; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;10.30, P\u0026thinsp;=\u0026thinsp;0.0010; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.70, P\u0026thinsp;=\u0026thinsp;0.0121. CAR is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ef; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;20.87, P\u0026thinsp;=\u0026thinsp;0.0002; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;13.27, P\u0026thinsp;=\u0026thinsp;0.0003; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.718, P\u0026thinsp;=\u0026thinsp;0.2076. PI is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eg; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;64.51, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;52.32, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;12.77, P\u0026thinsp;=\u0026thinsp;0.0004. SM is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eh; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.2447, P\u0026thinsp;=\u0026thinsp;0.6268; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;52.96, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.818, P\u0026thinsp;=\u0026thinsp;0.1910. PC is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ei; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;7.169, P\u0026thinsp;=\u0026thinsp;0.0154; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;26.70, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;15.82, P\u0026thinsp;=\u0026thinsp;0.0001. pPC is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ej; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.0014, P\u0026thinsp;=\u0026thinsp;0.9701; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;32.94, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;3.648, P\u0026thinsp;=\u0026thinsp;0.0468. LPC is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ek; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.01269, P\u0026thinsp;=\u0026thinsp;0.9116; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;22.77, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.368, P\u0026thinsp;=\u0026thinsp;0.2799. Cholesterol-Total is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003en; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.041, P\u0026thinsp;=\u0026thinsp;0.3212; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;34.85, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.507, P\u0026thinsp;=\u0026thinsp;0.0136. Cholesterol-Free is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eo; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;6.462, P\u0026thinsp;=\u0026thinsp;0.0204; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;22.53, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.915, P\u0026thinsp;=\u0026thinsp;0.0106. Cholesterol-Ester is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ep; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.0808, P\u0026thinsp;=\u0026thinsp;0.7794; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;39.83, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.055, P\u0026thinsp;=\u0026thinsp;0.0181. FA18:1/FA18:2 is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003er; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;19.17, P\u0026thinsp;=\u0026thinsp;0.0004; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;25.58, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.596, P\u0026thinsp;=\u0026thinsp;0.1022). Similarly, male \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats had lower levels of PI, SM, pPC, LPC, total cholesterol, and cholesterol esters compared to male \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats. Additionally, female \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats showed lower levels of pPE, CAR, PI, SM, PC, pPC, LPC, total cholesterol, free cholesterol, cholesterol esters and ratio of FA18:1/FA18:2 compared to female \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats. Conversely, female \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats showed a higher ratio of TAG to PC (TAG/PC is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eq; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.756, P\u0026thinsp;=\u0026thinsp;0.1140; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;8.524, P\u0026thinsp;=\u0026thinsp;0.0025; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.102, P\u0026thinsp;=\u0026thinsp;0.3537). Male \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, on the other hand, only had lower levels of SM compared to male \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals. PS, TAG and FA levels were comparable in \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e (PS is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eb; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;3.876, P\u0026thinsp;=\u0026thinsp;0.0646; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.1970, P\u0026thinsp;=\u0026thinsp;0.8230; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.4870, P\u0026thinsp;=\u0026thinsp;0.6223. TAG is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003el; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.321, P\u0026thinsp;=\u0026thinsp;0.1450; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.081, P\u0026thinsp;=\u0026thinsp;0178; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.4433, P\u0026thinsp;=\u0026thinsp;0.6488. FA is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003em; sex factor, F\u003csub\u003e(1, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.233, P\u0026thinsp;=\u0026thinsp;0.1524; genotype factor, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.030, P\u0026thinsp;=\u0026thinsp;0184; sex \u0026times; genotype interaction, F\u003csub\u003e(2, 18)\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.4383, P\u0026thinsp;=\u0026thinsp;0.6519).\u003c/p\u003e \u003cp\u003eThis comprehensive lipidomic analysis across nine lipid classes (CAR, FA, PC, PE, PI, PG, PS, TAG, and SM) revealed distinct patterns of lipid abundance in different rat samples, identified through the generation of heatmaps. Each heatmap provided a color-coded representation of the relative abundance of each lipid type within each sample. Our major finding was that \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e male rats consistently displayed unique lipidomic profiles across all lipid classes. This pattern underscores the substantial influence of genotype on lipid metabolism and suggests that specific genotypes and sex, such as \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e males, might uniquely influence lipid metabolism. This insight has potential implications for understanding disease susceptibilities or responses to treatments in these genetic models.\u003c/p\u003e \u003cp\u003eWe observed substantial heterogeneity in lipid composition across samples and lipid species, which underlined the diversity of the lipidomic landscape in the studied rat models. Certain lipid species like PS P-18:0/22:6 and PS O-16:0/22:6 in the PS lipid data, and TAG 48:0 and TAG 50:1 in the TAG lipid data, consistently exhibited lower relative abundances across all samples. These patterns suggest that these lipids might have less dominant role in the overall lipid metabolism, or their functions might be conserved across different rat models. Conversely, certain lipids were found to be consistently present at higher levels across all samples, indicating their prominent role in lipid metabolism across different rat genotypes and sexes. For instance, in the PC lipid data, lipid species such as PC 32:0 and PC 34:1 were observed to have higher relative abundances in most samples (Figs.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e and \u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eCorrelation heatmaps (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e9\u003c/span\u003e) depicted relationships among lipid species in \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats. We used the Pearson correlation coefficient to calculate all correlations and p-values between 222 lipid species in each group. We identified the top 20 correlations based on absolute values of correlation coefficients with p\u0026thinsp;\u0026lt;\u0026thinsp;0.05. The heatmaps reveal strong positive correlations between different types of lipid species across all groups. The strongest positive correlations were between TAG C52:2/C53:9 and PE D18:0\u0026ndash;20:4/D16:0\u0026ndash;22:4 in \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.97), between PE P18:1\u0026ndash;20:4/P16:0\u0026ndash;22:5 and PE A20:0\u0026ndash;20:4/P18:0\u0026ndash;22:3 in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.98), between CAR 16:2 and CAR 18:2 in \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.97), between CAR 18:0 and CAR 18:1 in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.97). There are strong positive correlations between LPE 20:4 and pPE P18:0\u0026ndash;22:6/P18:1\u0026ndash;22:5 with correlation coefficients of 0.97, 0.45 and 0.33 in \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, respectively. Conversely, there are strong negative correlations between CAR 7:0 and PE P16:0\u0026ndash;20:3/P18:1\u0026ndash;18:2 in \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= -0.91) and between SM N18:0 and PE D16:0\u0026ndash;20:4/D18:2\u0026ndash;18:2 in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= -0.94). The heatmaps also show that there are several opposite correlations between the different lipid types in the different groups of animals. A strong positive correlation exists between SM N20:0 and PC D18:0\u0026ndash;22:5 with correlation coefficients of 0.76 and 0.32 in \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, respectively but in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals there is negative correlation (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= -0.55). Similarly, there is a strong negative correlation between PC D18:2\u0026ndash;18:2/D16:0\u0026ndash;20:4 and SM N24:2 in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats with correlation coefficients of -0.68, while there is positive correlation in \u003cem\u003eApoe\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.32) and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.36). The correlation between CAR 7:0 and CAR 18:1 is more robust in \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.91) than in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.29), suggesting a more pronounced relationship between these two lipid species in \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats. Another intriguing observation is that the correlation between PE D16:0\u0026ndash;22:6 and SM N18:0, while there is strong negative correlation in \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= -0.56), there is slightly positive correlation in \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:r\\)\u003c/span\u003e\u003c/span\u003e= 0.018). All other correlation coefficients are reported in Supplemental Table\u0026nbsp;1. In conclusion, our findings offer a complex picture of lipid homeostasis across \u003cem\u003eAPOE\u003c/em\u003e variants of rat models. The substantial variations in lipid composition could have significant implications for understanding lipid metabolism in these models, potentially serving as a basis for the identification of lipidomic biomarkers for different physiological or pathological states.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eIn this study, we have characterized Long-Evans rat models that express human \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e variant genes, as well as humanized \u003cem\u003eApp\u003c/em\u003e\u003csup\u003e\u003cem\u003eh\u003c/em\u003e\u003c/sup\u003e alleles. We observed that \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats exhibited the highest levels of APOE in both serum and brain compared to \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, which showed comparable levels of APOE. These changes were unlikely due to transcriptional differences, as mRNA levels of human \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e were comparable. Therefore, differences in protein expression are likely attributed to translational and/or protein stability variances. While further investigation into these mechanisms is warranted, these findings closely mirror observations from human studies, underscoring the validity and translatability of these rat models [\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e, \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e, \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe humanization of both APP/Aβ and APOE in model organisms that express these human proteins in a physiological manner enables a comprehensive study of the multifaceted APOE-Aβ interaction. Our findings revealed a significant elevation in Aβ43 levels in male \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats compared to their \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e counterparts. This observation is particularly noteworthy as Aβ43 has high propensity for oligomerization and is a primary determinant of amyloid pathology [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e], while APOE4 is the most prominent genetic risk factor for LOAD, increasing the risk up to 12-fold, whereas APOE2 is a protective factor [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e]. This finding also aligns with evidence that Aβ oligomer levels in APOE4 AD patients' brains are 2.7 times higher than those in APOE3 AD patients [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e] and is consistent with findings from mouse models expressing APOE4 and APOE2 [\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003cem\u003eAPOE2\u003c/em\u003e homozygosity can lead to type III HLP in humans. Our findings suggest that \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats serve as a suitable model for studying this condition, showing elevated triglyceride and cholesterol levels. This led us to investigate how different human \u003cem\u003eAPOE\u003c/em\u003e isoforms affect lipid metabolism in models expressing humanized APOE variants and humanized APP/Aβ in a more comprehensive manner. Our findings confirm the elevated lipid levels previously observed in humanized \u003cem\u003eAPOE2\u003c/em\u003e mice compared to those carrying \u003cem\u003eAPOE3\u003c/em\u003e and \u003cem\u003eAPOE4\u003c/em\u003e [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e]. Moreover, we identified significant differences with the mouse models, including notably higher serum levels of total PG, pPE, LPE, CAR, PI, PC, pPC, LPC, FA in TAG, free-cholesterol, and cholesterol-esters. These observations are consistent with data showing that brain samples from \u003cem\u003eAPOE2\u003c/em\u003e LOAD patients exhibit elevated levels of CAR, SM, PC, and LPE compared to \u003cem\u003eAPOE3\u003c/em\u003e and \u003cem\u003eAPOE4\u003c/em\u003e LOAD patients [\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e]. Thus, rats, known for their more human-like metabolism compared to mice [\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e], and engineered to carry human \u003cem\u003eAPOE2\u003c/em\u003e along with the humanized \u003cem\u003eApp\u003c/em\u003e gene, may offer enhanced modeling capabilities for type III HLP compared to their mouse counterparts.\u003c/p\u003e \u003cp\u003eA follow-up analysis aimed at discerning differences between \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats revealed that only PG levels were lower in both male and female \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats compared to \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e animals. All other lipid categories showed no significant variation between these two groups. This serum lipidomic similarity between \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats mirrors the lipid homogeneity observed in synaptosomes isolated from 2-month-old mice expressing human \u003cem\u003eAPOE3\u003c/em\u003e and \u003cem\u003eAPOE4\u003c/em\u003e [\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e].\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eIn this study, we utilized Long-Evans knock-in rats with humanized \u003cem\u003eApp\u003c/em\u003e and \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, and \u003cem\u003eAPOE4\u003c/em\u003e isoforms to investigate their roles in LOAD type III HLP. Our findings reveal distinct profiles of APOE expression and lipid metabolism associated with each \u003cem\u003eAPOE\u003c/em\u003e isoform. Specifically, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats exhibited higher APOE levels in both serum and brain compared to \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, suggesting isoform-specific differences in protein expression likely due to translational or stability factors rather than transcriptional differences.\u003c/p\u003e \u003cp\u003eNotably, elevated Aβ43 levels were observed in male \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats, which could be instrumental in studying amyloid pathology due to Aβ43's high aggregation propensity. Furthermore, \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats showed elevated triglyceride and cholesterol levels, supporting their utility as a model for type III HLP. Minimal lipid profile differences between \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats reflect previously observed patterns in mouse models\u003c/p\u003e \u003cp\u003eOverall, these Long-Evans rat models provide a valuable platform for exploring the interactions between human \u003cem\u003eAPOE\u003c/em\u003e isoforms and Aβ in the context of LOAD and lipid metabolism. Future studies will expand lipidomic analyses to various CNS regions, offering deeper insights into the effects of human APOE isoforms on lipid metabolism and Alzheimer\u0026rsquo;s disease pathology. Comparing the brain lipidomic of \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e, \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats with lipidomic profiles from postmortem AD brains will further validate the translatability of the models described here [\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e].\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMY: \u003cstrong\u003eAlzheimer\u0026apos;s Association\u003c/strong\u003e,\u0026nbsp;24AARFD-1243865\u003c/p\u003e\n\u003cp\u003eXH: National Institute on Aging, R01AG061729; National Institutes of Health, P30 AG013319\u003c/p\u003e\n\u003cp\u003eLD:\u0026nbsp;\u003cstrong\u003eNational Institute on Aging\u003c/strong\u003e,\u0026nbsp;R01AG073182;\u0026nbsp;\u003cstrong\u003eNational Institute on Aging\u003c/strong\u003e,\u0026nbsp;R01AG063407;\u0026nbsp;\u003cstrong\u003eNational Institute on Aging\u003c/strong\u003e, RF1AG064821\u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eMY and LD wrote the first draft of the manuscript. MY, TY, MT, HB, MP, and LD contributed to the material preparation, data collection, and figure drawing. MY, TY, MT, XH, and LD contributed to the critical review of the manuscript. All authors reviewed and approved the final version.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eYamazaki, Y., et al., \u003cem\u003eApolipoprotein E and Alzheimer disease: pathobiology and targeting strategies.\u003c/em\u003e Nature Reviews Neurology, 2019. \u003cstrong\u003e15\u003c/strong\u003e(9): p. 501-518.\u003c/li\u003e\n\u003cli\u003eHuebbe, P. and G. Rimbach, \u003cem\u003eEvolution of human apolipoprotein E (APOE) isoforms: Gene structure, protein function and interaction with dietary factors.\u003c/em\u003e Ageing Research Reviews, 2017. \u003cstrong\u003e37\u003c/strong\u003e: p. 146-161.\u003c/li\u003e\n\u003cli\u003eFullerton, S.M., et al., \u003cem\u003eApolipoprotein E variation at the sequence haplotype level: implications for the origin and maintenance of a major human polymorphism.\u003c/em\u003e The American journal of human genetics, 2000. \u003cstrong\u003e67\u003c/strong\u003e(4): p. 881-900.\u003c/li\u003e\n\u003cli\u003eKayden, H.J., F. Maschio, and M.G. Traber, \u003cem\u003eThe secretion of apolipoprotein E by human monocytederived macrophages.\u003c/em\u003e Archives of biochemistry and biophysics, 1985. \u003cstrong\u003e239\u003c/strong\u003e(2): p. 388-395.\u003c/li\u003e\n\u003cli\u003eGee, J.R. and J.N. Keller, \u003cem\u003eAstrocytes: regulation of brain homeostasis via apolipoprotein E.\u003c/em\u003e The International Journal of Biochemistry \u0026amp; Cell Biology, 2005. \u003cstrong\u003e37\u003c/strong\u003e(6): p. 1145-1150.\u003c/li\u003e\n\u003cli\u003eWest, H.L., G.W. Rebeck, and B.T. Hyman, \u003cem\u003eFrequency of the apolipoprotein E \u0026epsilon;2 allele is diminished in sporadic Alzheimer disease.\u003c/em\u003e Neuroscience letters, 1994. \u003cstrong\u003e175\u003c/strong\u003e(1-2): p. 46-48.\u003c/li\u003e\n\u003cli\u003eFarrer, L.A., et al., \u003cem\u003eEffects of Age, Sex, and Ethnicity on the Association Between Apolipoprotein E Genotype and Alzheimer Disease: A Meta-analysis.\u003c/em\u003e JAMA, 1997. \u003cstrong\u003e278\u003c/strong\u003e(16): p. 1349-1356.\u003c/li\u003e\n\u003cli\u003eKojro, E., et al., \u003cem\u003eLow cholesterol stimulates the nonamyloidogenic pathway by its effect on the \u0026alpha;-secretase ADAM 10.\u003c/em\u003e Proceedings of the National Academy of Sciences, 2001. \u003cstrong\u003e98\u003c/strong\u003e(10): p. 5815-5820.\u003c/li\u003e\n\u003cli\u003eGr\u0026ouml;sgen, S., et al., \u003cem\u003eRole of amyloid beta in lipid homeostasis.\u003c/em\u003e Biochimica et Biophysica Acta (BBA)-Molecular and Cell Biology of Lipids, 2010. \u003cstrong\u003e1801\u003c/strong\u003e(8): p. 966-974.\u003c/li\u003e\n\u003cli\u003eCordy, J.M., et al., \u003cem\u003eExclusively targeting \u0026beta;-secretase to lipid rafts by GPI-anchor addition up-regulates \u0026beta;-site processing of the amyloid precursor protein.\u003c/em\u003e Proceedings of the National Academy of Sciences, 2003. \u003cstrong\u003e100\u003c/strong\u003e(20): p. 11735-11740.\u003c/li\u003e\n\u003cli\u003eLaDu, M.J., et al., \u003cem\u003eIsoform-specific binding of apolipoprotein E to beta-amyloid.\u003c/em\u003e J Biol Chem, 1994. \u003cstrong\u003e269\u003c/strong\u003e(38): p. 23403-6.\u003c/li\u003e\n\u003cli\u003eSharman, M.J., et al., \u003cem\u003eAPOE genotype results in differential effects on the peripheral clearance of amyloid-\u0026beta; 42 in APOE knock-in and knock-out mice.\u003c/em\u003e Journal of Alzheimer\u0026apos;s Disease, 2010. \u003cstrong\u003e21\u003c/strong\u003e(2): p. 403-409.\u003c/li\u003e\n\u003cli\u003eTambini, M.D., W. Yao, and L. D\u0026apos;Adamio, \u003cem\u003eFacilitation of glutamate, but not GABA, release in Familial Alzheimer\u0026apos;s APP mutant Knock-in rats with increased beta-cleavage of APP.\u003c/em\u003e Aging Cell, 2019. \u003cstrong\u003e18\u003c/strong\u003e(6): p. e13033.\u003c/li\u003e\n\u003cli\u003ePiedrahita, J.A., et al., \u003cem\u003eGeneration of mice carrying a mutant apolipoprotein E gene inactivated by gene targeting in embryonic stem cells.\u003c/em\u003e Proceedings of the National Academy of Sciences, 1992. \u003cstrong\u003e89\u003c/strong\u003e(10): p. 4471-4475.\u003c/li\u003e\n\u003cli\u003eSullivan, P.M., et al., \u003cem\u003eTargeted replacement of the mouse apolipoprotein E gene with the common human APOE3 allele enhances diet-induced hypercholesterolemia and atherosclerosis.\u003c/em\u003e J Biol Chem, 1997. \u003cstrong\u003e272\u003c/strong\u003e(29): p. 17972-80.\u003c/li\u003e\n\u003cli\u003eWu, Y., et al., \u003cem\u003eFeatures of Lipid Metabolism in Humanized ApoE Knockin Rat Models.\u003c/em\u003e International Journal of Molecular Sciences, 2021. \u003cstrong\u003e22\u003c/strong\u003e(15): p. 8262.\u003c/li\u003e\n\u003cli\u003eMahley, R.W., Y. Huang, and S.C. Rall, \u003cem\u003ePathogenesis of type III hyperlipoproteinemia (dysbetalipoproteinemia): questions, quandaries, and paradoxes.\u003c/em\u003e Journal of lipid research, 1999. \u003cstrong\u003e40\u003c/strong\u003e(11): p. 1933-1949.\u003c/li\u003e\n\u003cli\u003eMahley, R.W. and Z.-S. Ji, \u003cem\u003eRemnant lipoprotein metabolism: key pathways involving cell-surface heparan sulfate proteoglycans and apolipoprotein E.\u003c/em\u003e Journal of Lipid Research, 1999. \u003cstrong\u003e40\u003c/strong\u003e(1): p. 1-16.\u003c/li\u003e\n\u003cli\u003eLund-Katz, S., et al., \u003cem\u003eApolipoprotein B-100 Conformation and Particle Surface Charge in Human LDL Subspecies:\u0026thinsp; Implication for LDL Receptor Interaction.\u003c/em\u003e Biochemistry, 1998. \u003cstrong\u003e37\u003c/strong\u003e(37): p. 12867-12874.\u003c/li\u003e\n\u003cli\u003eHuang, Y., et al., \u003cem\u003eOverexpression and Accumulation of Apolipoprotein E as a Cause of Hypertriglyceridemia *.\u003c/em\u003e Journal of Biological Chemistry, 1998. \u003cstrong\u003e273\u003c/strong\u003e(41): p. 26388-26393.\u003c/li\u003e\n\u003cli\u003eRensen, P.C.N. and T.J.C. van Berkel, \u003cem\u003eApolipoprotein E Effectively Inhibits Lipoprotein Lipase-mediated Lipolysis of Chylomicron-like Triglyceride-rich Lipid Emulsions \u0026lt;strong\u0026gt;\u0026lt;em\u0026gt;in Vitro\u0026lt;/em\u0026gt;\u0026lt;/strong\u0026gt; and \u0026lt;strong\u0026gt;\u0026lt;em\u0026gt;in Vivo\u0026lt;/em\u0026gt;\u0026lt;/strong\u0026gt;*.\u003c/em\u003e Journal of Biological Chemistry, 1996. \u003cstrong\u003e271\u003c/strong\u003e(25): p. 14791-14799.\u003c/li\u003e\n\u003cli\u003eFredrickson, D.S., R.I. Levy, and R.S. Lees, \u003cem\u003eFat Transport in Lipoproteins \u0026mdash; An Integrated Approach to Mechanisms and Disorders.\u003c/em\u003e New England Journal of Medicine, 1967. \u003cstrong\u003e276\u003c/strong\u003e(3): p. 148-156.\u003c/li\u003e\n\u003cli\u003eUtermann, G., M. Jaeschke, and J. Menzel, \u003cem\u003eFamilial hyperlipoproteinemia type III: deficiency of a specific apolipoprotein (apo E-III) in the very-low-density lipoproteins.\u003c/em\u003e FEBS lett, 1975. \u003cstrong\u003e56\u003c/strong\u003e(2): p. 352-355.\u003c/li\u003e\n\u003cli\u003eZannis, V.I. and J.L. Breslow, \u003cem\u003eHuman very low density lipoprotein apolipoprotein E isoprotein polymorphism is explained by genetic variation and posttranslational modification.\u003c/em\u003e Biochemistry, 1981. \u003cstrong\u003e20\u003c/strong\u003e(4): p. 1033-1041.\u003c/li\u003e\n\u003cli\u003eMahley, R.W., \u003cem\u003eApolipoprotein E: Cholesterol Transport Protein with Expanding Role in Cell Biology.\u003c/em\u003e Science, 1988. \u003cstrong\u003e240\u003c/strong\u003e(4852): p. 622-630.\u003c/li\u003e\n\u003cli\u003eDavignon, J., R.E. Gregg, and C.F. Sing, \u003cem\u003eApolipoprotein E polymorphism and atherosclerosis.\u003c/em\u003e Arteriosclerosis: An Official Journal of the American Heart Association, Inc., 1988. \u003cstrong\u003e8\u003c/strong\u003e(1): p. 1-21.\u003c/li\u003e\n\u003cli\u003ePhillips, M.C., \u003cem\u003eApolipoprotein E isoforms and lipoprotein metabolism.\u003c/em\u003e IUBMB Life, 2014. \u003cstrong\u003e66\u003c/strong\u003e(9): p. 616-623.\u003c/li\u003e\n\u003cli\u003eGustavsson, J., et al., \u003cem\u003eInteraction of apolipoprotein E genotype with smoking and physical inactivity on coronary heart disease risk in men and women.\u003c/em\u003e Atherosclerosis, 2012. \u003cstrong\u003e220\u003c/strong\u003e(2): p. 486-492.\u003c/li\u003e\n\u003cli\u003eWeisgraber, K.H., T.L. Innerarity, and R.W. Mahley, \u003cem\u003eAbnormal lipoprotein receptor-binding activity of the human E apoprotein due to cysteine-arginine interchange at a single site.\u003c/em\u003e Journal of Biological Chemistry, 1982. \u003cstrong\u003e257\u003c/strong\u003e(5): p. 2518-2521.\u003c/li\u003e\n\u003cli\u003ePham, H., T. Yin, and L. D\u0026apos;Adamio, \u003cem\u003eInitial assessment of the spatial learning, reversal, and sequencing task capabilities of knock-in rats with humanizing mutations in the A\u0026beta;-coding region of App.\u003c/em\u003e PLoS One, 2022. \u003cstrong\u003e17\u003c/strong\u003e(5): p. e0263546.\u003c/li\u003e\n\u003cli\u003eTambini, M.D., W. Yao, and L. D\u0026apos;Adamio, \u003cem\u003eFacilitation of glutamate, but not GABA, release in Familial Alzheimer\u0026apos;s APP mutant Knock-in rats with increased \u0026beta;-cleavage of APP.\u003c/em\u003e Aging Cell, 2019. \u003cstrong\u003e18\u003c/strong\u003e(6): p. e13033.\u003c/li\u003e\n\u003cli\u003eYesiltepe, M., et al., \u003cem\u003eLate-long-term potentiation magnitude, but not Abeta levels and amyloid pathology, is associated with behavioral performance in a rat knock-in model of Alzheimer disease.\u003c/em\u003e Front Aging Neurosci, 2022. \u003cstrong\u003e14\u003c/strong\u003e: p. 1040576.\u003c/li\u003e\n\u003cli\u003eTambini, M.D., et al., \u003cem\u003eAbeta43 levels determine the onset of pathological amyloid deposition.\u003c/em\u003e J Biol Chem, 2023. \u003cstrong\u003e299\u003c/strong\u003e(7): p. 104868.\u003c/li\u003e\n\u003cli\u003eYin, T., et al., \u003cem\u003eA familial Danish dementia rat shows impaired presynaptic and postsynaptic glutamatergic transmission.\u003c/em\u003e J Biol Chem, 2021. \u003cstrong\u003e297\u003c/strong\u003e(3): p. 101089.\u003c/li\u003e\n\u003cli\u003eRen, S., et al., \u003cem\u003eTNF-alpha-mediated reduction in inhibitory neurotransmission precedes sporadic Alzheimer\u0026apos;s disease pathology in young Trem2(R47H) rats.\u003c/em\u003e J Biol Chem, 2021. \u003cstrong\u003e296\u003c/strong\u003e: p. 100089.\u003c/li\u003e\n\u003cli\u003eRen, S., et al., \u003cem\u003eMicroglia TREM2(R47H) Alzheimer-linked variant enhances excitatory transmission and reduces LTP via increased TNF-alpha levels.\u003c/em\u003e Elife, 2020. \u003cstrong\u003e9\u003c/strong\u003e.\u003c/li\u003e\n\u003cli\u003eTambini, M.D., et al., \u003cem\u003eA\u0026beta;43 levels determine the onset of pathological amyloid deposition.\u003c/em\u003e Journal of Biological Chemistry, 2023. \u003cstrong\u003e299\u003c/strong\u003e(7).\u003c/li\u003e\n\u003cli\u003eHan, X., \u003cem\u003eLipidomics: Comprehensive mass spectrometry of lipids\u003c/em\u003e. 2016: John Wiley \u0026amp; Sons.\u003c/li\u003e\n\u003cli\u003ePalavicini, J.P., et al., \u003cem\u003eNovel molecular insights into the critical role of sulfatide in myelin maintenance/function.\u003c/em\u003e Journal of neurochemistry, 2016. \u003cstrong\u003e139\u003c/strong\u003e(1): p. 40-54.\u003c/li\u003e\n\u003cli\u003eHan, X., K. Yang, and R.W. Gross, \u003cem\u003eMicrofluidics-based electrospray ionization enhances the intrasource separation of lipid classes and extends identification of individual molecular species through multi-dimensional mass spectrometry: development of an automated high-throughput platform for shotgun lipidomics.\u003c/em\u003e Rapid Commun Mass Spectrom, 2008. \u003cstrong\u003e22\u003c/strong\u003e(13): p. 2115-24.\u003c/li\u003e\n\u003cli\u003eYang, K., et al., \u003cem\u003eAutomated lipid identification and quantification by multidimensional mass spectrometry-based shotgun lipidomics.\u003c/em\u003e Anal Chem, 2009. \u003cstrong\u003e81\u003c/strong\u003e(11): p. 4356-68.\u003c/li\u003e\n\u003cli\u003eWang, M. and X. Han, \u003cem\u003eMultidimensional mass spectrometry-based shotgun lipidomics.\u003c/em\u003e Methods Mol Biol, 2014. \u003cstrong\u003e1198\u003c/strong\u003e: p. 203-20.\u003c/li\u003e\n\u003cli\u003eYang, K., et al., \u003cem\u003eAutomated lipid identification and quantification by multidimensional mass spectrometry-based shotgun lipidomics.\u003c/em\u003e Analytical chemistry, 2009. \u003cstrong\u003e81\u003c/strong\u003e(11): p. 4356-4368.\u003c/li\u003e\n\u003cli\u003eCai, C., Z. Wen, and L. Li, \u003cem\u003eThe relationship between ApoE gene polymorphism and the efficacy of statins controlling hyperlipidemia.\u003c/em\u003e Am J Transl Res, 2021. \u003cstrong\u003e13\u003c/strong\u003e(6): p. 6772-6777.\u003c/li\u003e\n\u003cli\u003ePasser, B., et al., \u003cem\u003eGeneration of an apoptotic intracellular peptide by gamma-secretase cleavage of Alzheimer\u0026apos;s amyloid beta protein precursor.\u003c/em\u003e J Alzheimers Dis, 2000. \u003cstrong\u003e2\u003c/strong\u003e(3-4): p. 289-301.\u003c/li\u003e\n\u003cli\u003eKinosian, B., H. Glick, and G. Garland, \u003cem\u003eCholesterol and coronary heart disease: predicting risks by levels and ratios.\u003c/em\u003e Annals of internal medicine, 1994. \u003cstrong\u003e121\u003c/strong\u003e(9): p. 641-647.\u003c/li\u003e\n\u003cli\u003eMart\u0026iacute;nez-Morillo, E., et al., \u003cem\u003eTotal apolipoprotein E levels and specific isoform composition in cerebrospinal fluid and plasma from Alzheimer\u0026rsquo;s disease patients and controls.\u003c/em\u003e Acta Neuropathologica, 2014. \u003cstrong\u003e127\u003c/strong\u003e(5): p. 633-643.\u003c/li\u003e\n\u003cli\u003eGiannisis, A., et al., \u003cem\u003ePlasma apolipoprotein E levels, isoform composition, and dimer profile in relation to plasma lipids in racially diverse patients with Alzheimer\u0026rsquo;s disease and mild cognitive impairment.\u003c/em\u003e Alzheimer\u0026apos;s Research \u0026amp; Therapy, 2023. \u003cstrong\u003e15\u003c/strong\u003e(1): p. 119.\u003c/li\u003e\n\u003cli\u003eMichaelson, D.M., \u003cem\u003eAPOE \u0026epsilon;4: The most prevalent yet understudied risk factor for Alzheimer\u0026apos;s disease.\u003c/em\u003e Alzheimer\u0026apos;s \u0026amp; Dementia, 2014. \u003cstrong\u003e10\u003c/strong\u003e(6): p. 861-868.\u003c/li\u003e\n\u003cli\u003eHashimoto, T., et al., \u003cem\u003eApolipoprotein E, Especially Apolipoprotein E4, Increases the Oligomerization of Amyloid \u0026beta; Peptide.\u003c/em\u003e The Journal of Neuroscience, 2012. \u003cstrong\u003e32\u003c/strong\u003e(43): p. 15181-15192.\u003c/li\u003e\n\u003cli\u003eHudry, E., et al., \u003cem\u003eGene Transfer of Human \u0026lt;i\u0026gt;Apoe\u0026lt;/i\u0026gt; Isoforms Results in Differential Modulation of Amyloid Deposition and Neurotoxicity in Mouse Brain.\u003c/em\u003e Science Translational Medicine, 2013. \u003cstrong\u003e5\u003c/strong\u003e(212): p. 212ra161-212ra161.\u003c/li\u003e\n\u003cli\u003eSharman, M.J., et al., \u003cem\u003eProfiling brain and plasma lipids in human apoe \u0026epsilon;2, \u0026epsilon;3, and \u0026epsilon;4 knock-in mice using electrospray ionization mass spectrometry.\u003c/em\u003e Journal of Alzheimer\u0026apos;s Disease, 2010. \u003cstrong\u003e20\u003c/strong\u003e(1): p. 105-111.\u003c/li\u003e\n\u003cli\u003eLefterov, I., et al., \u003cem\u003eAPOE2 orchestrated differences in transcriptomic and lipidomic profiles of postmortem AD brain.\u003c/em\u003e Alzheimer\u0026apos;s Research \u0026amp; Therapy, 2019. \u003cstrong\u003e11\u003c/strong\u003e(1): p. 113.\u003c/li\u003e\n\u003cli\u003eIgbavboa, U., et al., \u003cem\u003eMurine synaptosomal lipid raft protein and lipid composition are altered by expression of human apoE 3 and 4 and by increasing age.\u003c/em\u003e Journal of the Neurological Sciences, 2005. \u003cstrong\u003e229\u003c/strong\u003e: p. 225-232.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"cell-communication-and-signaling","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"ccas","sideBox":"Learn more about [Cell Communication and Signaling](http://biosignaling.biomedcentral.com/)","snPcode":"12964","submissionUrl":"https://submission.nature.com/new-submission/12964/3","title":"Cell Communication and Signaling","twitterHandle":"@bmc","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-4789248/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4789248/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eAPOE is a major genetic factor in late-onset Alzheimer's disease (LOAD), with APOE4 significantly increasing risk, APOE3 acting as a neutral isoform, and APOE2 offering protective effects. The primary hypothesis links APOE isoforms to LOAD through their impact on Aβ production and deposition, which is thought to be related to their effects on lipid metabolism. Specifically, APOE4 enhances Aβ production and accumulation in amyloid plaques more than APOE3. In contrast, APOE3-Aβ complexes, which promote Aβ clearance and reduce Aβ aggregation, are approximately 20 times more prevalent than APOE4-Aβ complexes, highlighting differences in their functional interactions. APOE is also important in lipid metabolism, affecting both peripheral and central systems. It is involved in the metabolism of lipoproteins and plays a key role in triglyceride and cholesterol regulation. APOE2 is notably associated with Hyperlipoproteinemia type III (HLP), which is characterized by mixed hypercholesterolemia and hypertriglyceridemia due to impaired binding to Low Density Lipoproteins receptors. To explore the impact of human APOE isoforms on lipid metabolism and LOAD, we developed Long-Evans rats with the rat \u003cem\u003eApoe\u003c/em\u003e gene replaced by human \u003cem\u003eAPOE2\u003c/em\u003e, \u003cem\u003eAPOE3\u003c/em\u003e, or \u003cem\u003eAPOE4\u003c/em\u003e. These rats were crossed with those carrying a humanized \u003cem\u003eApp\u003c/em\u003e allele, enabling the expression of human Aβ, which is more aggregation-prone than rodent Aβ. This model offers a significant advancement for studying APOE-Aβ interactions. We found that \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats had the highest levels of APOE in serum and brain, with no significant transcriptional differences among isoforms, suggesting variations in protein translation or stability. Elevated Aβ43 levels in male \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats compared to \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats highlight the model\u0026rsquo;s utility for amyloid pathology studies. Additionally, a lipidomic analysis of 222 lipid molecular species in serum samples showed that \u003cem\u003eAPOE2\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats displayed elevated triglycerides and cholesterol, making them a valuable model for studying HLP. These rats also exhibited elevated levels of phosphatidylglycerol, phosphatidylserine, phosphatidylethanolamine, sphingomyelin, and lysophosphatidylcholine. Minimal differences in lipid profiles between \u003cem\u003eAPOE3\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eAPOE4\u003c/em\u003e\u003csup\u003e\u003cem\u003ehAβ\u003c/em\u003e\u003c/sup\u003e rats reflect findings from mouse models. Future studies will include comprehensive lipidomic analyses in various CNS regions to further validate these models and explore the effects of \u003cem\u003eAPOE\u003c/em\u003e isoforms on lipid metabolism in relation to AD pathology.\u003c/p\u003e","manuscriptTitle":"Human APOE variants in Alzheimer’s Disease and type III hyperlipoproteinemia: insights from Long-Evans knock-in rat models with humanized App and APOE","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-08-21 10:05:20","doi":"10.21203/rs.3.rs-4789248/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-08-08T22:31:18+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-08-08T19:40:15+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-08-06T18:12:36+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"261124197749722705111591355632210059742","date":"2024-08-02T15:05:04+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"143191342837468183099422748319966345338","date":"2024-07-31T07:40:44+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"129395177974386967147869364577602183453","date":"2024-07-29T14:29:39+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-07-29T11:54:10+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-07-25T03:50:28+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-07-25T03:50:03+00:00","index":"","fulltext":""},{"type":"submitted","content":"Cell Communication and Signaling","date":"2024-07-23T13:47:57+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"cell-communication-and-signaling","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"ccas","sideBox":"Learn more about [Cell Communication and Signaling](http://biosignaling.biomedcentral.com/)","snPcode":"12964","submissionUrl":"https://submission.nature.com/new-submission/12964/3","title":"Cell Communication and Signaling","twitterHandle":"@bmc","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"5a2c5f5a-e4fc-43bf-b3cf-7c1781348821","owner":[],"postedDate":"August 21st, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2024-09-30T15:59:28+00:00","versionOfRecord":{"articleIdentity":"rs-4789248","link":"https://doi.org/10.1186/s12964-024-01832-2","journal":{"identity":"cell-communication-and-signaling","isVorOnly":false,"title":"Cell Communication and Signaling"},"publishedOn":"2024-09-27 15:56:59","publishedOnDateReadable":"September 27th, 2024"},"versionCreatedAt":"2024-08-21 10:05:20","video":"","vorDoi":"10.1186/s12964-024-01832-2","vorDoiUrl":"https://doi.org/10.1186/s12964-024-01832-2","workflowStages":[]},"version":"v1","identity":"rs-4789248","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4789248","identity":"rs-4789248","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}