SCARF1 Deficiency Exacerbates Gut Inflammation and Autoimmune Pathology

SCARF1 Deficiency Exacerbates Gut Inflammation and Autoimmune Pathology | 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 Article SCARF1 Deficiency Exacerbates Gut Inflammation and Autoimmune Pathology Dominique M Shepard, Sabine Hahn, Monika Chitre, Haley Neff, Doyle V Ward, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7023221/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 11 Feb, 2026 Read the published version in Scientific Reports → Version 1 posted 16 You are reading this latest preprint version Abstract Systemic lupus erythematosus (SLE) is a complex autoimmune disease known for its heterogeneity in both manifestation and presentation. Recent evidence has increasingly implicated the gut microbiome within immunomodulation and autoimmunity. This study aims to characterize the intestinal inflammation and microbial profile associated with autoimmune diseases, particularly SLE, and to identify unique biomarkers and shared microbial signatures for potential therapeutic measures. Our lab identified scavenger receptor class F, member 1 (SCARF1, SREC-1) as an efferocytosis receptor essential for the clearance of apoptotic debris, and its deficiency results in the development of lupus-like disease. SCARF1 is crucial in immune homeostasis, and defects in efferocytosis lead to inflammation. However, the role of SCARF1 in homeostasis in the gut remains to be elucidated. To answer our question, we analyzed and compared the metagenomic datasets generated through whole genome shotgun sequencing between our Scarf1 −/− lupus-prone mouse model and healthy counterparts. We found that Scarf1 −/− mice had significantly lengthened intestines, elevated immune cell infiltration, and structural changes in the colon. Microbiome analysis revealed gut dysbiosis, including reduced alpha diversity and increased F/B ratio. Notably, beneficial taxa such as Akkermansia muciniphila was absent in Scarf1 −/− mice. Linear regression analysis identified positive associations between lupus disease severity and increased abundances of Bacillota, Alistipes, Lachnospiraceae , and Hominisplanchenecus . Function analysis of the gut microbiome in Scarf1 −/− mice indicated downregulation of multiple pathways related to cell proliferation. These findings highlight the role of SCARF1 involvement in the gut microbiome and immune regulation in the context of inflammation and SLE. Biological sciences/Computational biology and bioinformatics Health sciences/Diseases Biological sciences/Immunology Biological sciences/Microbiology lupus microbiome autoimmune inflammation gut SCARF1 Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Introduction The innate immune system is the first line of defense following a pathological insult 1 . During infection, pattern recognition receptors (PRRs) identify pathogen associated molecular patterns (PAMPs) to initiate an immune response 2 . This leads to an intracellular signaling cascade with the end results of eliminating the pathogen. The gut contains 70–80% of the body’s immune cells due to its susceptibility towards antigen exposure through pathogens, diet, and the gut microbiome. Nonetheless, humans have a variety of microbes that reside in our bodies that contribute to homeostasis 3 . The mammalian immune system co-evolved with commensal microbes. Comprising the gut microbiota, these commensal microbes not only contribute towards shaping the immune system during neonatal development, but also directly participate to protect us from infection 4 . Recent research increasingly highlights the gut microbiome’s complex and multifaceted role in the immune system. This includes, but is not limited to, providing colonization resistance through competition between commensal and pathogenic bacteria; interacting with the intestinal epithelium and mucosal membranes to support the regulation of mucus production, immune signaling, and epithelial barrier integrity; and producing microbial metabolites that act as immune protective signals, influencing T cell differentiation within the gut and activating innate immune cells, such as neutrophils and macrophages, beyond the gut 5 . Dysbiosis or the imbalance of the microbiota, will result in the disruption of the immune homeostasis and can induce an inflammatory response. Gut dysbiosis has been linked to multiple diseases, such as inflammatory bowel disease, cancer and diabetes 6 , 7 . Recent evidence has increasingly suggested that gut microbiota dysbiosis has a role within systemic lupus erythematosus (SLE) development and pathogenesis 8 . SLE is a complex and heterogeneous disease with a wide range of manifestations known to impact multiple organ systems, such as the gastrointestinal (GI) tract 9 . Side effects of medications and increased susceptibility to infection may disrupt the balance of immune activation, ultimately impacting immune homeostasis 10 . Nonetheless, host genetics also play a significant role in shaping the composition and function of the microbiome 11 . Scavenger receptors (SR) are considered a subset of PRRs, as they can identify a broad spectrum of ligands including apoptotic cells and microbial components 12 , 13 . One member of the SR family, SCARF1 (scavenger receptor class F member 1, also known as SR-F1 or SREC1) was identified as a non-redundant efferocytosis receptor and as a lupus-prone mouse model 14 . This is the result of dying cells accumulating in tissues leading to a lupus-like disease with spontaneous generation of autoantibodies to chromatin, cell activation, dermatitis and nephritis. Our initial studies filled some gaps within SCARF1-mediated regulation of autoimmunity. In this study, we plan to expand our knowledgeabout the role of SCARF1 and investigate the gut microbiome within gut inflammation in Scarf1 −/− lupus-prone mice. Using shotgun sequencing we compared fecal pellets from Scarf1 −/− and Scarf1 +/+ , where we observed changes in certain bacterial species. Together, our findings provide insights in bacterial species associated with inflammation and SLE. Results Scarf1 deficiency results in enhanced gut inflammation and defects in efferocytosis We previously showed that Scarf1 is responsible for the removal of apoptotic debris, and mice deficient in this receptor develop lupus-like disease 14 . Our group observed Scarf1 −/− mice exhibit a significantly longer gut when compared to wild-type mice, including colon length (Fig. 1 A-B). Gut microbiome dysbiosis has often been linked to not only inflammation, but also anatomical and structural changes in the gastrointestinal organ system 15 . We aimed to assess whether Scarf1 +/+ and Scarf1 −/− mice exhibited notable changes in gut inflammation and microbiome. We assessed the development of autoimmunity through the presence of antinuclear autoantibodies (Sup Fig. 1-BA). In confirmation with earlier work, Scarf1 −/− mice develop autoimmunity through multiple symptom manifestations, including alopecia, loss of whiskers, (Sup Fig. 1C-D) and nephritis (Sup Fig E) at 20-weeks of age. Histological analysis of Scarf1-deficient murine colons indicated increased cell infiltration and structural alterations (Fig. 1 C, bottom panels), while pathological analysis found increased numbers of neutrophils and apoptotic cells (Fig. 1 D). Our previous work has shown the scavenger receptor, SCARF1, mediates the efferocytosis of apoptotic cells (ACs) in mice in a non-redundant manner and dysregulation of SCARF1 leads to the accumulation of ACs 14 , 16 . We next aimed to investigate whether Scarf1-deficiency leads to an accumulation of apoptotic debris in the gut. Using fluorescent microscopy, we detected a significant increase in cellular debris in the Scarf1 −/− mice (Fig. 1 E-F). Our data confirms that Scarf1 is essential for the removal of apoptotic debris and suggests a potential role for Scarf1 in the mediation of tissue homeostasis. The gut microbiome in Scarf1 −/− mice exhibits reduced alpha diversity and an increased Firmicutes/Bacteroidetes (F/B) ratio The gut microbiota is sensitive to changes in the tissue homeostasis 17 . We compared the fecal microbiome of 20-week-old Scarf1 +/+ and Scarf1 −/− (Fig. 2 ). Our data shows reduced alpha diversity in the Scarf1 −/− mice in both female and male animals when compared to Scarf1 +/+ mice (Fig. 2 A-B). Changes in the Firmicutes/ Bacteroidota (F/B) ratio are a biomarker for gut dysbiosis. Assessing F/B ratio across strains and sexes, we uncovered a significant dysbiosis in the Scarf1 −/− female mice, through a marked increase in Firmicutes and decrease in Bacteroidota (Fig. 2 C). Interestingly, we noted that Scarf1 -deficient males reveal an F/B ratio similar to wild-type mice, although still trending higher (Fig. 2 C). Our data corroborate the large sex difference observed in the development of autoimmune disease. In SLE, females face a 9:1 incidence ratio in comparison to males 18 . Nonetheless, while a significant difference between wild-type and Scarf1 -deficient mice was present, we noted that the clusters within our Principal Coordinates Analysis (PCoA) of bacterial beta-diversity as analyzed by Bray-Curtis dissimilarity were independent of sex (Fig. 2 D). Due to the coprophagous behaviors of mice, we next questioned whether co-housed wild-type and Scarf1 −/− mice would lead to fecal microbial self-reinoculation and subsequent alterations in the microbiome 19 (Supplemental Fig. 2). No significant difference in the alpha diversity between wild-type and Scarf1 -deficient mice within control and co-housed groups was found (Supp Fig. 2 A-B). However, PCoA data and Bray-Curtis dissimilarity analysis indicated the co-housed mice cluster together, suggesting microbiome composition is impacted (Supp Fig. 2 C). Taxonomic analysis revealed a reduction in Akkermansia and Porphyromenadaceae , and an increase in Alistipes in wild-type mice (Supp Fig. 2 D). No significant difference in Firmicutes (also known as Bacillota ) or F/B ratio between co-housed and control mice (Supp Fig. 2 E-F). Deficiency in SCARF1 will affect immune homeostasis and mutations of the receptor will affect ligand-receptor interactions 20 . Therefore, based on our data we can conclude that the observed dysbiosis in Scarf1 -deficient mice is likely attributed to the absence of the SCARF1 receptor itself and the associated inflammation. Wild-type littermates exhibit greater relative bacterial abundance and higher alpha diversity Since co-housing mice for 2 weeks did indeed have an impact on the microbiome, we next asked whether a longer timeline would have a clear impact on the clinical development of autoimmune disease. To address this question, F2 littermate mice were co-housed for a minimum of 20-weeks. 50% of heterozygous mice ( Scarf1 −/+ ) developed low levels of autoimmunity, as examined by ANA immunofluorescence study (Fig. 3 A and data not shown). Scarf1 −/+ mice positive for ANA staining had a higher F/B ratio (Fig. 3 B); however, no difference in diversity between Scarf1 −/− and Scarf1 −/+ was identified through Shannon nor Simpson index measurements (Fig. 3 C-D). Wild-type ( Scarf1 +/+ ) mice display a more diverse microbiota compared to Scarf1 −/− or Scarf1 −/+ (Fig. 3 C-D), suggesting even a partial deficiency of Scarf1 could alter gut microbial homeostasis. Nonetheless, Bray-Curtis dissimilarity analysis of Scarf1 −/− , Scarf1 −/+ and Scarf1 +/+ mice showed no clustering, suggesting shared microbial taxa across the three mouse strains (Fig. 3 E). Relative abundance data comparing Scarf1 −/− , Scarf1 −/+ and Scarf1 +/+ mice indicates a higher relative abundance for wild-type mice when compared to Scarf1 −/− and Scarf1 −/+ (Fig. 4 F). We observed that boths littermates wild-type and het mice faced a reduction in Akkermasia (Fig. 3 F); however, wild-type mice experienced an increase in other beneficial bacteria such as Ligilactobacillus murinus , a species known for maintaining gut health. Healthy mice express higher levels of Akkermansia We hypothesized that beneficial bacteria are reduced or absent in Scarf1 -deficient mice. Overall, bacterial species abundance was lower in the Scarf1 −/− mice when compared to wild-type controls (Fig. 4 A-B). Examining the top 20 species, we observed higher abundance of Akkermansia muciniciphila , Dubosiella sp004793885 and Bacteriodales bacterium M2 in wild type animals compared to Scarf1 −/− mice (Fig. 4 ). As expected, we observed an increase in Firmicutes (Bacillota) in Scarf1 −/− mice (Fig. 4 C). Our data also show decreased levels of Akkermansia (Fig. 4 D), Dubosiella (Fig. 4 E), Bacteroidales (Fig. 4 F) along with increased abundance of Alistipes (Fig. 4 E) and Duncaniella (Fig. 4 F) in Scarf1 −/− mice. Akkermansia muciniciphila is associated with a healthy gut by stimulating metabolic and immune responses 21 . Interestingly, wild-type mice that were co-housed with Scarf1 −/− mice for 2 weeks had decreased levels of Akkermansia (Supp Fig. 2 D); however, the co-housed wild-type mice did not develop autoimmune disease, as assessed by ANA analysis (data not shown). Loss of specific bacterial species is associated with autoimmunity An increasing number of studies suggest that the gut microbiota is involved in the initiation and progression of inflammatory and autoimmune diseases 22 . To assess this, we investigated whether we could identify specific species potentially associated with disease development. We developed a disease score (Fig. 5 A and Supp Fig. 1 ) based on the levels of ANA staining, alopecia, nephritis (as measured by glomerular inflammation), and the average number of apoptotic cells. Using a simple linear regression, we analyzed the relationship between bacterial species abundance of species and disease score (Fig. 5 B-J). Healthy mice exhibit significantly higher levels of Akkermansia (Fig. 5 B) and Dubosiella (Fig. 5 G) in contrast to Scarf1 −/− mice which experience significantly increased in Alistipes (Fig. 5 B), Bacteroides (Fig. 5 D), Lachnospiraceae (Fig. 5 E) and Hominisplanchenecus (Fig. 5 F). While not statistically significant, we noticed a trend toward increased abundance of Lepagella (Fig. 5 H), Porphyromonadaceae (Fig. 5 I) and Heminphilus (Fig. 5 J) in Scarf1 −/− mice. Together, these findings suggest that SCARF1 deficiency decreases beneficial bacterial populations, leading to an altered gut microbiome and potentially contributing to the gut inflammation and autoimmune pathology. Gut functional analysis suggests that a decrease in “internal component of membrane pathway” and an increase in “regulation of cell proliferation pathway” in Scarf1 -/- mice Microbiome changes affect both the metabolic and immune systems 23 . To gain more insight into the microbiome-host interaction, we performed a functional gene pathway analysis on the bacterial species (Fig. 6 ). We noticed a decrease in the “integral component of membrane” in Scarf1 −/− mice (Fig. 6 A, Supp Fig. 3 A). In turn, we also observed an increase in the “positive regulation of cell proliferation” (Fig. 6 A, Supp Fig. 3 B). Despite statistical insignificance, increased regulation of cell proliferation in Scarf1 −/− mice may provide a potential explanation for increased gut size observed in Scarf1 −/− mice compared to wild-type controls. Pathway analysis further showed that Scarf1 −/− mice have decreased levels of L-valine biosynthesis, L-isoleucine biosynthesis and adenosine ribonucleotide de novo biosynthesis compared to wild-type (Fig. 6 B). These compounds are essential in the ability to produce branched-chain fatty acids (BCFA), which are essential lipid membrane components of the gut bacteria 24 . Furthermore, these compounds have been shown to protect against intestinal damage 25 . Alterations in the biosynthesis of BCFA have been linked to dysbiosis 26 , further suggesting that changes in lipids can affect homeostasis. As described above, we found that Akkermansia is present only in the microbiota of Scarf1 +/+ mice (Fig. 6 C-D, refer Fig. 4 D and Fig. 5 B). Studies have shown that A. muciniphila is involved in regulating lipid metabolism and modulating the immune response by reducing inflammation 27 . Taken together, our data shows a different microbiome and inflammation profile between Scarf1 +/+ and Scarf1 −/− . Discussion Dysbiosis of the gut microbiome has been linked to the development of SLE 28 , 29 . Numerous factors have been studied associating dysbiosis with the development of SLE. Side effects of medications, increased susceptibility to infection and tissue damage are a few examples that lead to the impairment of gut homeostasis 10 , 30 . However, the impact of genetic SLE-associated risk factors on gut dysbiosis is less understood. Here we characterize the gut microbiome in a mouse model of spontaneous lupus. Using the lupus-prone mouse Scarf1 −/− , we observed a significant size increase in the colon size when compared to wild-type mice. This observation was striking, as the inflamed gut is shorter in size 31 , 32 . SCARF1’s role as an efferocytosis receptor 14 , 33 and the accumulation of uncleared apoptotic cells and subsequent inflammatory immune and epithelial stimulation within diseased mice suggest a possible explanation for the increase in colon size. The absence of SCARF1 disrupts immune homeostasis 14 , leading us to assess inherent changes in composition and diversity of gut microbiota between control and disease mice. Wild-type mice express a higher diversity and a richer bacterial composition when compared to Scarf1 −/− mice. Data also shows differences that are sex specific, with diseased female mice exhibiting low diversity with increased F/B ratio. Similar observations indicating significant sex-specific differences in gut mucosa have been shown using SWR × NZB F1 (SNF1) mice 34 . Sex hormones might be influencing the microbiome, as this was indicated in a type 1 diabetes study in which male mice were castrated resulting in disease progression 35 , 36 . However, we observed differences in the gut microbiome that were strain specific as Scarf1 −/− mice showed similar microbiota trends regardless of being cohoused for 2 or 20 weeks. SCARF1 plays a significant role in maintaining lipid homeostasis. Initially, SCARF1 on endothelial cells as a receptor for modified lipoproteins 33 , 37 . De novo biosynthesis of L-valine, L-isoleucine and adenosine ribonucleotide is essential to produce BCFA and dampen inflammation 38 , 39 . Consistent with maintaining lipid homeostasis, Scarf1 −/− exhibit decreased beneficial bacteria that synthesize BCFA and short-chain fatty acids (SCFA). Although additional work is needed, we can propose that the development of autoimmunity in Scarf1 -deficient mice isdriven in part by the dysbiosis and defects in efferocytosis. Akkermansia muciniphila is a gram-negative anaerobic bacterium that colonizes the intestinal tract early in life 40 . This beneficial intestinal commensal is known for colonizing the mucosal layer, where it plays an essential role in host metabolism and immune response 27 . Control mice express high levels of A. muciniphila , however this bacterium is completely absent in Scarf1 −/− mice. A. muciniphila is associated with health and maintaining the mucosal barrier. Furthermore, disease mice express high levels of Alistipes , Bacteroides , Lachnospiraceae which are associated with dysbiosis and metabolic diseases 41 . Altogether, our data shows various lupus-associated changes in the gut microbiome. Intestinal colonization of A. muciniphila was found to negatively correlate with disease development. A. muciniphila probiotics and derived postbiotics have already been identified as promising therapeutics within multiple inflammatory diseases, including SLE 42 . Our data corroborate the potential use of A. muciniphila as a probiotic to decrease lupus-like symptoms and lower inflammation; however, further research is required to understand the impact of A. municiphila postbiotics, or metabolites, directly. Assessing metabolic changes in control and disease murine models would be an interesting avenue given previous studies implicate SCARF1 in lipid metabolism and homeostasis 20 . The work presented here are the first steps in understanding the role of SCARF1 in gut inflammation and homeostasis. Future work will focus on the mechanistic interaction of apoptotic debris and dysbiosis. Material and Methods Mice : All mice were maintained under micro isolation in specific pathogen–free conditions at the animal facility of UMass Chan Medical School under a protocol approved by the Institutional Animal Care and Use Committee. In addition, experimental design shows a comparative study between Scarf1 +/+ (wild-type) or Scarf1 −/− following ARRIVE guidelines ( https://arriveguidelines.org/ ). This includes the use of control animals, inclusion criteria for male and female mice, blinding of samples for data analysis and the use of statistical methods are described below. Wild-type (WT, Scarf1 +/+ ) C57BL/6 mice were obtained from Jackson Laboratories and bred in-house. Scarf1 −/− mice were transferred from Massachusetts General Hospital and bred in-house for at least 10 generations. All mice were used after 20-weeks of age to allow for disease development, as previously described 14 . Offspring of Scarf1 −/− and WT mice were produced at normal Mendelian ratios. Mice were not randomized or placed in specific groups for these studies. In accordance with ALAAS learning library and IACUC, for end-point studies mice were euthanize using a two-step euthanasia protocol. Mice were anesthetized with isoflurane using saturated vapor, then we performed cardiac puncture for blood collection. To ensure death, cervical dislocation was performed before harvesting organs for analysis. Fecal collection for microbiome preparation : Mice were selected for analysis from a cohort of Scarf1 −/− mice and their healthy littermate controls. Following euthanasia, fecal samples were collected and immediately frozen at -80C until processing. Samples were then placed in individual tubes containing DNA stabilization buffers (Transnetyx Microbiome Kits, Cordova, TN) to preserve sample integrity and stability during shipment. All DNA extraction, library preparation, and sequencing were performed by Transnetyx (Cordova, TN). DNA extraction and metagenomic sequencing : DNA extraction and metagenomic sequencing was perform by Transnetyx Inc. Briefly, stool DNA was extracted using a robust method that ensures reproducible extraction that captures the accurate microbial diversity. DNA quality control was performed to confirm sample integrity. Genomic DNA was converted into sequencing libraries and sequenced using shotgun metagenomic sequencing, generating approximately 2 million 2x150 bp read pairs to obtain microbial species and strain-level taxonomic resolution. Raw sequencing data were uploaded to the OneCodex platform for analysis and aligned against a database of ~ 148K complete microbial genomes, including 71K bacterial, 72K viral, and thousands of archaeal and eukaryotic genomes. To reduce false positives, classification results underwent further analysis to group the samples in their required experimental approach. Microbial taxonomy and diversity metrics were computed within OneCodex to compare Scarf1 −/− mice and healthy littermate controls. Samples were also compared with Transnetyx’s global diversity averages derived from historic datasets for quality control. Low read counts were normalized relative to the total number of identifiable reads within each host sample. Finally, sequencing data were aligned against the Gene Ontology (GO) and KEGG Orthology databases via the OneCodex platform for downstream functional analysis of microbial communities. Autoantibody profiles : Antinuclear autoantibodies (ANA) were measured as previously described 43 . ANA assays from mouse serum were performed using immunofluorescence assays according to the manufacturer’s instructions (Bio-Rad). Mouse serum was diluted at 1:200 and incubated with Hep-2 cells, followed by Alexa Fluor 488 secondary to detect bound ANAs (Cat #A11001, Invitrogen). Staining was scored by three independent observers ‘blinded’ to the genotypes of the mice. Histology : The small and large intestines were dissected from 20-week-old Scarf1 −/− and C57BL/6J (B6) wild-type mice. To assess histological signs of gut inflammation, the intestines were fixed in 10% phosphate-formalin and embedded in paraffin. Sections were prepared and stained in hematoxylin-eosin. For all other studies, the intestines were flash frozen on Optimal Cutting Temperature (OCT) Compound (Sciegen Scientific, Gardena CA) embedding media. Sections from frozen samples were prepared (7um) and stained as described below. All slides were imaged using ECHO fluorescent microscope equipped with a high-resolution (Discover Echo, San Diego, CA, USA) and analyzed using Discover ECHO App (iOS 16 + ) (Discover Echo, San Diego, CA, USA) and Adobe Photoshop (Adobe, San Jose CA). TUNEL assay : Frozen sections (7um) were allowed to warm to 25°C. A TUNEL assay (terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling) for the detection of apoptotic cells was done according to manufacturer’s instructions (Invitrogen). Paraformaldehyde-fixed tissue was made permeable with 60 minutes 0.25% Triton X-100 in PBS at room temperature and was washed twice with PBS. Slides were incubated with reaction buffer (25 mM Tris-HCl, pH 6.6, 200 mM sodium cacodylate, 0.25 mg/ml BSA and 1 mM cobalt) containing terminal deoxynucleotidyl transferase, then were washed twice and were incubated for 45 min at 25°C in the dark (protected from light) with reaction buffer containing the fluorescent label Alexa Fluor 488. To identify tissue structures, sections we stained with Phalloidin-Rhodamin for 25 min at RT and EPCAM1 cells were stained for 30 min at 25°C with Alexa Fluor 647–anti-EPCAM1 (1:50 dilution; Invitrogen A22283) in PBS. Samples were washed twice with PBS. Finally, DNA was stained for 10 min at 25°C with Hoechst 33342 (1:1,000 dilution; Molecular Probes, Invitrogen). Slides were mounted with Prolong Gold antifade reagent (Invitrogen P36935) and were visualized with ECHO fluorescent microscope equipped with a high-resolution. Data were analyzed with Discover ECHO App (iOS 16 + ) (Discover Echo, San Diego, CA, USA) and Adobe Photoshop (Adobe, San Jose CA). Histological assessment and disease scoring : Histological evaluation of the intestinal H&E sections was performed in a blinded manner by a pathologist (Jadhav, Nupur). Inflammation and cellular infiltration were scored, and cell types were identified. Tissue inflammation severity was scored on a scale from 0–5, where grade of inflammation 1- minimal, 2- mild, 3-moderate, 4-severe, 5-severe with ulceration. For presence of apoptosis in the tissue, 1- present and 2- absent. A composite disease score was calculated by averaging the individual scores for antinuclear autoantibodies (ANA), alopecia, nephritis, and the number of apoptotic cells. Statistical analysis: Statistical calculations were done with a statistical software package GraphPad Prism, version 10.4.2 (GraphPad Software, San Diego, CA). For comparisons between two or more groups, the mean ± s.e.m was analyzed by unpaired two-tailed Student’s t test or ANOVA, respectively. Statistical analysis of the microbiota profiling data was performed on the proportional representation of the taxa using Shapiro-Wilk normality test. Parametric test with Welch’s corrections or nonparametric test with Mann-Whitney corrections were used depending on if passed the normality test. Multivariate analyses of disease score versus relative abundance of bacteria were done. The investigators were not blinded to the genotype of the mice except where indicated. Values of P < 0.05 were considered statistically significant. Declarations Funding This work was funded by the Department of Defense LRP-Impact Award (W81XWH-21-1-0803) (ZGRO), Lupus Research Alliance Innovation Award (ZGRO) and UMass Chan Medical School Start-up funds (ZGRO). Acknowledgements We would like to acknowledge lead microbiome researchers, Dr. Beth McCormick and Dr. Ana Maldonado-Contreras, for their great advice and expertise. We also thank Dr. Stuart Levitz and his lab for providing additional advice in the preparation of the manuscript. Conflict of Interest JMR is an inventor on patent application #62489191 and #15/851,651 which covers IL-15 and CXCR3 for the treatment of vitiligo, respectively; and on patent #63/478,900 filed for “Diagnosis of skin diseases in veterinary and human patients” for CTCL. The other authors have no conflicts of interest to disclose. Author Contribution DMS and ZGRO designed and performed the experimental, analyzed data and wrote the manuscript. ZGRO developed the mouse model. DMS and ZGRO figures 1-5ZGRO and DVJ figure 6NJ Figure 1 and all pathologySH, MC, JMR, HN provided advice in the experimental design and edited the manuscript. JMR, DVW and NF validated the results. NJ analyzed pathology samples. JMR, DVW, HN provided advice on data analysis.All authors reviewed the manuscript Acknowledgement We would like to acknowledge lead microbiome researchers, Dr. Beth McCormick and Dr. Ana Maldonado-Contreras, for their great advice and expertise. We also thank Dr. Stuart Levitz and his lab for providing additional advice in the preparation of the manuscript. Data Availability Data is provided within the manuscript or supplementary information files. The datasets for the current study are available through OneCodex upon request to the corresponding author. References Keogh, C. E., Rude, K. M. & Gareau, M. G. 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A) \u003cem\u003eScarf1-\u003c/em\u003e/- gut is enlarged when compared to wild-type mice. Representative image of gut measurement. B) Quantification of gut length (cm) in wild-type vs. \u003cem\u003eScarf1-\u003c/em\u003e/- mice. ** p \u0026lt; 0.05, ns = not significant C) Increased inflammation in \u003cem\u003eScarf1-\u003c/em\u003e/- gut tissue. Hematoxylin-and-eosin staining of fixed cryosections of duodenum, Ileum, and colon of 20-week-old female \u003cem\u003eScarf+/+\u003c/em\u003e and \u003cem\u003eScarf1\u003c/em\u003e-/- mice (n = 3 per group). Original magnification, ×20. D) Pathology score. Slides were scored blinded, and the grade of inflammation 1- minimal 2- mild, 3-moderate, 4-severe, 5-severe with ulceration. (n = 3 mice per group). Data was analyzed using 2-Way ANOVA. \u0026nbsp;E) Fluorescence microscopy of Duodenum, Ileum and Colon sections from \u0026gt;20-week-old \u003cem\u003eScarf1\u003c/em\u003e+/+ and \u003cem\u003eScarf1\u003c/em\u003e-/-mice, showing apoptotic cells stained by TUNEL (green) and phalloidin (red), EPCAM-1 (white) and DAPI (blue). Representative image of 3 independent experiments. 20x magnification\u0026nbsp; F) Quantification of images at left by automated analysis of staining with the nuclear dye DAPI and by TUNEL. *P \u0026lt; 0.001, 1; Student’s t-test.\u003c/p\u003e","description":"","filename":"OnlineMicroFig1.png","url":"https://assets-eu.researchsquare.com/files/rs-7023221/v1/b43e685caf8893804b18cf96.png"},{"id":87506157,"identity":"1b5d591b-6613-479f-bf61-a1973f865e78","added_by":"auto","created_at":"2025-07-24 14:48:45","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":870719,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cu\u003e\u003cstrong\u003eLower alpha diversity and increased Firmicute/Bacteroidetes (F/B) ratio in female \u003c/strong\u003e\u003c/u\u003e\u003cu\u003e\u003cem\u003e\u003cstrong\u003eScarf1-\u003c/strong\u003e\u003c/em\u003e\u003c/u\u003e\u003cu\u003e\u003cstrong\u003e/- mice.\u003c/strong\u003e\u003c/u\u003e\u0026nbsp; A-B) Alpha diversity of fecal microbiota from wild-type and \u003cem\u003eScarf1-\u003c/em\u003e/- mice analyzed by A) Shannon and B) Simpson diversity indices. N= 20 mice, 5 per group. Samples show trends but are not statistically significant according to the non-parametric Wilcoxon rank statistical test. C) F/B ratio calculated from normalized read counts. N = 24 mice (\u003cem\u003eScarf1-\u003c/em\u003e/- female 7, male 7, \u003cem\u003eWild-type\u003c/em\u003e female 6, male 4) ** p \u0026lt; 0.03 when comparing female \u003cem\u003eScarf1-\u003c/em\u003e/- to all other groups. Data is not significant between wild-type and male \u003cem\u003eScarf1-\u003c/em\u003e/- D) Principal Coordinates Analysis (PCoA) plot of bacterial beta-diversity based on Bray–Curtis dissimilarities. Data represent two combined independent experiments.\u003c/p\u003e","description":"","filename":"OnlineMicroFig2.png","url":"https://assets-eu.researchsquare.com/files/rs-7023221/v1/0bd0a6be66dd4bdfa464ca5d.png"},{"id":87506160,"identity":"58201c79-8c08-488f-9a9c-510f6aa0f0a2","added_by":"auto","created_at":"2025-07-24 14:48:45","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":5402337,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cu\u003e\u003cstrong\u003eWild-type mice have a higher alpha diversity and reduced F/B ratio compared to Scarf1-deficient littermates\u003c/strong\u003e\u003c/u\u003e. F2 littermates were co-housed for 20 weeks. A) \u003cem\u003eScarf1-/+ develop autoimmunity\u003c/em\u003e.\u0026nbsp; Antinuclear antibody (ANA) immunofluorescence of HEp-2 cells using serum from 20-week-old female \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e, \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e-/-\u003c/sup\u003e mice (\u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e n = 4 female mice per group; \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/+\u003c/sup\u003e n=9 female mice per group). Original magnification, ×20. Representative images from two independent experiments. B) Wild-type mice have a lower F/B ratio. F/B ratio calculated from normalized read counts. C-D) Alpha diversity of fecal microbiota assessed by C) Shannon and D) Simpson diversity indices\u003cem\u003e Scarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e n = 4 female mice per group; \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/+\u003c/sup\u003e n=9 female mice per group. E) Principal Coordinates Analysis (PCoA) plot of bacterial beta-diversity based on Bray–Curtis dissimilarities. Data represent two independent experiments combined. F) Comparison of gut microbiota profiles between 20-week-old female wild-type, \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/+\u003c/sup\u003e\u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e mice. Whole genome shotgun sequencing was performed by Transnetyx and analyzed using OneCodex.\u003c/p\u003e","description":"","filename":"OnlineMIcroFig3.png","url":"https://assets-eu.researchsquare.com/files/rs-7023221/v1/4d0fd308b63e96b5de58179a.png"},{"id":87506193,"identity":"8dccf724-b53a-4071-880e-e330fcfeb5d4","added_by":"auto","created_at":"2025-07-24 14:48:45","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":3469228,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cu\u003e\u003cstrong\u003eHealthy species are present in wild-type mice but absent in \u003c/strong\u003e\u003c/u\u003e\u003cu\u003e\u003cem\u003e\u003cstrong\u003eScarf1\u003c/strong\u003e\u003c/em\u003e\u003c/u\u003e\u003csup\u003e\u003cu\u003e\u003cem\u003e\u003cstrong\u003e-\u003c/strong\u003e\u003c/em\u003e\u003c/u\u003e\u003c/sup\u003e\u003csup\u003e\u003cu\u003e\u003cstrong\u003e/- \u003c/strong\u003e\u003c/u\u003e\u003c/sup\u003e\u003cu\u003e\u003cstrong\u003emice.\u003c/strong\u003e\u003c/u\u003e\u003c/p\u003e\n\u003cp\u003eA-C) Comparison of gut microbiota profiles between 20-week-old female wild-type and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e mice. Whole genome shotgun sequencing was performed by Transnetyx, and data was analyzed using OneCodex. N = 21 female mice per group. (n= 9 wild-type; n=12 \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e) A) Relative abundance of the top 15 species. B)\u0026nbsp; Heat map of the top 15 genera. C) Mean relative abundance of the top 10 phyla. D-H) Levels of specific microbial communities measured by shotgun sequencing as described above. Statistical significance: **** p \u0026lt;0.0001,*** p = 0.0004, * p = 0.04, NS = not significant.\u0026nbsp; D) \u003cem\u003eAkkermansia muciniphila, \u003c/em\u003eE) \u003cem\u003eDubosiella\u003c/em\u003e sp004793885, F) \u003cem\u003eBacteroidales bacterium \u003c/em\u003eM2, G)\u003cem\u003e Alistipes\u003c/em\u003e MGBC116833, H) \u003cem\u003eDuncaniella\u003c/em\u003e MGBC142302. Data represent two independent experiments, N= 13 mice per group.\u003c/p\u003e","description":"","filename":"Onlinemicrofig4.png","url":"https://assets-eu.researchsquare.com/files/rs-7023221/v1/068b7098c73c074f2cf3ca6a.png"},{"id":87507696,"identity":"bfd27fee-17a9-49fd-b5f0-fc3e0b7ac1bd","added_by":"auto","created_at":"2025-07-24 15:04:45","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":1275875,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cu\u003e\u003cstrong\u003eCorrelation between SLE-like disease score and relative microbiota abundance\u003c/strong\u003e\u003c/u\u003e. A) Heat map of disease scores in 20-week-old female \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-/-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e \u003c/sup\u003emice (n= 3 mice per group). The disease score represents the average of blinded assessments for ANA immunofluorescence on Hep-2 cells, nephritis, alopecia, and apoptotic cell counts in the duodenum, ileum, and colon. B-J) Simple linear regression analyses between individual bacterial species and disease score. N=3 mice per group. Data are from a representative experiment.\u003c/p\u003e","description":"","filename":"Onlinemicrofig5.png","url":"https://assets-eu.researchsquare.com/files/rs-7023221/v1/9ccf3eaa0104c435a34e5147.png"},{"id":87506162,"identity":"98d5d1e1-81b2-4966-9728-0fde7e54da7b","added_by":"auto","created_at":"2025-07-24 14:48:45","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":4487087,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cu\u003e\u003cstrong\u003eGut functional analysis shows a decrease in the internal component of the membrane in \u003c/strong\u003e\u003c/u\u003e\u003cu\u003e\u003cem\u003e\u003cstrong\u003eScarf1\u003c/strong\u003e\u003c/em\u003e\u003c/u\u003e\u003csup\u003e\u003cu\u003e\u003cem\u003e\u003cstrong\u003e-\u003c/strong\u003e\u003c/em\u003e\u003c/u\u003e\u003c/sup\u003e\u003csup\u003e\u003cu\u003e\u003cstrong\u003e/-\u003c/strong\u003e\u003c/u\u003e\u003c/sup\u003e\u003cu\u003e\u003cstrong\u003e mice.\u003c/strong\u003e\u003c/u\u003e A) Top 20 GO Term analysis in gut sample as copies per million comparing \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e mice B) Pathway analysis of gut samples as copies per million comparing \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e mice C-D) Mapped to functional group runs using both nucleotide and translated protein alignment comparing C) \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and D) \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e Representative analysis of n = 6 \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and n=10 \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e-\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e/-\u003c/sup\u003e\u003c/p\u003e","description":"","filename":"Onlinemicrofig6.png","url":"https://assets-eu.researchsquare.com/files/rs-7023221/v1/87c56b48ec9c466d72ce29ae.png"},{"id":102785175,"identity":"5044cfa1-b52e-4e1e-97d4-4c9ab528c214","added_by":"auto","created_at":"2026-02-16 16:01:04","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":4587570,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7023221/v1/4e5dd3d2-1659-4e64-a6dd-bc467dc5a533.pdf"},{"id":87508944,"identity":"6a498c3e-e2ef-41ab-ada4-677910e5db12","added_by":"auto","created_at":"2025-07-24 15:12:45","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":10586094,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementalFigureSCARF1.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7023221/v1/2e887c49dba57ccc9cbfff60.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"SCARF1 Deficiency Exacerbates Gut Inflammation and Autoimmune Pathology","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThe innate immune system is the first line of defense following a pathological insult\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. During infection, pattern recognition receptors (PRRs) identify pathogen associated molecular patterns (PAMPs) to initiate an immune response\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e. This leads to an intracellular signaling cascade with the end results of eliminating the pathogen. The gut contains 70\u0026ndash;80% of the body\u0026rsquo;s immune cells due to its susceptibility towards antigen exposure through pathogens, diet, and the gut microbiome. Nonetheless, humans have a variety of microbes that reside in our bodies that contribute to homeostasis\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eThe mammalian immune system co-evolved with commensal microbes. Comprising the gut microbiota, these commensal microbes not only contribute towards shaping the immune system during neonatal development, but also directly participate to protect us from infection\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. Recent research increasingly highlights the gut microbiome\u0026rsquo;s complex and multifaceted role in the immune system. This includes, but is not limited to, providing colonization resistance through competition between commensal and pathogenic bacteria; interacting with the intestinal epithelium and mucosal membranes to support the regulation of mucus production, immune signaling, and epithelial barrier integrity; and producing microbial metabolites that act as immune protective signals, influencing T cell differentiation within the gut and activating innate immune cells, such as neutrophils and macrophages, beyond the gut\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eDysbiosis or the imbalance of the microbiota, will result in the disruption of the immune homeostasis and can induce an inflammatory response. Gut dysbiosis has been linked to multiple diseases, such as inflammatory bowel disease, cancer and diabetes\u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e,\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e. Recent evidence has increasingly suggested that gut microbiota dysbiosis has a role within systemic lupus erythematosus (SLE) development and pathogenesis\u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e. SLE is a complex and heterogeneous disease with a wide range of manifestations known to impact multiple organ systems, such as the gastrointestinal (GI) tract\u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e. Side effects of medications and increased susceptibility to infection may disrupt the balance of immune activation, ultimately impacting immune homeostasis\u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e. Nonetheless, host genetics also play a significant role in shaping the composition and function of the microbiome\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eScavenger receptors (SR) are considered a subset of PRRs, as they can identify a broad spectrum of ligands including apoptotic cells and microbial components\u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e,\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e. One member of the SR family, SCARF1 (scavenger receptor class F member 1, also known as SR-F1 or SREC1) was identified as a non-redundant efferocytosis receptor and as a lupus-prone mouse model\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e. This is the result of dying cells accumulating in tissues leading to a lupus-like disease with spontaneous generation of autoantibodies to chromatin, cell activation, dermatitis and nephritis. Our initial studies filled some gaps within SCARF1-mediated regulation of autoimmunity. In this study, we plan to expand our knowledgeabout the role of SCARF1 and investigate the gut microbiome within gut inflammation in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e lupus-prone mice. Using shotgun sequencing we compared fecal pellets from \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e, where we observed changes in certain bacterial species. Together, our findings provide insights in bacterial species associated with inflammation and SLE.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eScarf1\u003c/span\u003e \u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003edeficiency results in enhanced gut inflammation and defects in efferocytosis\u003c/span\u003e\u003c/p\u003e\u003cp\u003eWe previously showed that \u003cem\u003eScarf1\u003c/em\u003e is responsible for the removal of apoptotic debris, and mice deficient in this receptor develop lupus-like disease\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e. Our group observed \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice exhibit a significantly longer gut when compared to wild-type mice, including colon length (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA-B). Gut microbiome dysbiosis has often been linked to not only inflammation, but also anatomical and structural changes in the gastrointestinal organ system\u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e. We aimed to assess whether \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e+/+\u003c/em\u003e\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice exhibited notable changes in gut inflammation and microbiome. We assessed the development of autoimmunity through the presence of antinuclear autoantibodies (Sup Fig.\u0026nbsp;1-BA). In confirmation with earlier work, \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice develop autoimmunity through multiple symptom manifestations, including alopecia, loss of whiskers, (Sup Fig.\u0026nbsp;1C-D) and nephritis (Sup Fig E) at 20-weeks of age. Histological analysis of Scarf1-deficient murine colons indicated increased cell infiltration and structural alterations (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC, bottom panels), while pathological analysis found increased numbers of neutrophils and apoptotic cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eOur previous work has shown the scavenger receptor, SCARF1, mediates the efferocytosis of apoptotic cells (ACs) in mice in a non-redundant manner and dysregulation of SCARF1 leads to the accumulation of ACs\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e,\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e. We next aimed to investigate whether Scarf1-deficiency leads to an accumulation of apoptotic debris in the gut. Using fluorescent microscopy, we detected a significant increase in cellular debris in the \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eE-F). Our data confirms that \u003cem\u003eScarf1\u003c/em\u003e is essential for the removal of apoptotic debris and suggests a potential role for Scarf1 in the mediation of tissue homeostasis.\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eThe gut microbiome in\u003c/span\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eScarf1\u003c/span\u003e\u003csup\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003e\u0026minus;/\u0026minus;\u003c/span\u003e\u003c/sup\u003e \u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003emice exhibits reduced alpha diversity and an increased Firmicutes/Bacteroidetes (F/B) ratio\u003c/span\u003e\u003c/p\u003e\u003cp\u003eThe gut microbiota is sensitive to changes in the tissue homeostasis\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e. We compared the fecal microbiome of 20-week-old \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Our data shows reduced alpha diversity in the \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice in both female and male animals when compared to \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA-B). Changes in the \u003cem\u003eFirmicutes/ Bacteroidota\u003c/em\u003e (F/B) ratio are a biomarker for gut dysbiosis. Assessing F/B ratio across strains and sexes, we uncovered a significant dysbiosis in the \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e female mice, through a marked increase in \u003cem\u003eFirmicutes\u003c/em\u003e and decrease in \u003cem\u003eBacteroidota\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC). Interestingly, we noted that \u003cem\u003eScarf1\u003c/em\u003e-deficient males reveal an F/B ratio similar to wild-type mice, although still trending higher (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC). Our data corroborate the large sex difference observed in the development of autoimmune disease. In SLE, females face a 9:1 incidence ratio in comparison to males\u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e. Nonetheless, while a significant difference between wild-type and \u003cem\u003eScarf1\u003c/em\u003e-deficient mice was present, we noted that the clusters within our Principal Coordinates Analysis (PCoA) of bacterial beta-diversity as analyzed by Bray-Curtis dissimilarity were independent of sex (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eDue to the coprophagous behaviors of mice, we next questioned whether co-housed wild-type and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice would lead to fecal microbial self-reinoculation and subsequent alterations in the microbiome\u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e (Supplemental Fig.\u0026nbsp;2). No significant difference in the alpha diversity between wild-type and \u003cem\u003eScarf1\u003c/em\u003e-deficient mice within control and co-housed groups was found (Supp Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA-B). However, PCoA data and Bray-Curtis dissimilarity analysis indicated the co-housed mice cluster together, suggesting microbiome composition is impacted (Supp Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC). Taxonomic analysis revealed a reduction in \u003cem\u003eAkkermansia\u003c/em\u003e and \u003cem\u003ePorphyromenadaceae\u003c/em\u003e, and an increase in \u003cem\u003eAlistipes\u003c/em\u003e in wild-type mice (Supp Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD). No significant difference in \u003cem\u003eFirmicutes\u003c/em\u003e (also known as \u003cem\u003eBacillota\u003c/em\u003e) or F/B ratio between co-housed and control mice (Supp Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eE-F). Deficiency in SCARF1 will affect immune homeostasis and mutations of the receptor will affect ligand-receptor interactions\u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e. Therefore, based on our data we can conclude that the observed dysbiosis in \u003cem\u003eScarf1\u003c/em\u003e-deficient mice is likely attributed to the absence of the SCARF1 receptor itself and the associated inflammation.\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eWild-type littermates exhibit greater relative bacterial abundance and higher alpha diversity\u003c/span\u003e\u003c/p\u003e\u003cp\u003eSince co-housing mice for 2 weeks did indeed have an impact on the microbiome, we next asked whether a longer timeline would have a clear impact on the clinical development of autoimmune disease. To address this question, F2 littermate mice were co-housed for a minimum of 20-weeks. 50% of heterozygous mice (\u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/+\u003c/sup\u003e) developed low levels of autoimmunity, as examined by ANA immunofluorescence study (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA and data not shown). \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/+\u003c/sup\u003e mice positive for ANA staining had a higher F/B ratio (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB); however, no difference in diversity between \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/+\u003c/sup\u003e was identified through Shannon nor Simpson index measurements (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eC-D). Wild-type (\u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e) mice display a more diverse microbiota compared to \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e or \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/+\u003c/sup\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eC-D), suggesting even a partial deficiency of \u003cem\u003eScarf1\u003c/em\u003e could alter gut microbial homeostasis. Nonetheless, Bray-Curtis dissimilarity analysis of \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e, \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e mice showed no clustering, suggesting shared microbial taxa across the three mouse strains (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eE). Relative abundance data comparing \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e, \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e mice indicates a higher relative abundance for wild-type mice when compared to \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/+\u003c/sup\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eF). We observed that boths littermates wild-type and het mice faced a reduction in \u003cem\u003eAkkermasia\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eF); however, wild-type mice experienced an increase in other beneficial bacteria such as \u003cem\u003eLigilactobacillus murinus\u003c/em\u003e, a species known for maintaining gut health.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eHealthy mice express higher levels of\u003c/span\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eAkkermansia\u003c/span\u003e\u003c/p\u003e\u003cp\u003eWe hypothesized that beneficial bacteria are reduced or absent in \u003cem\u003eScarf1\u003c/em\u003e-deficient mice. Overall, bacterial species abundance was lower in the \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice when compared to wild-type controls (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA-B). Examining the top 20 species, we observed higher abundance of \u003cem\u003eAkkermansia muciniciphila\u003c/em\u003e, \u003cem\u003eDubosiella sp004793885\u003c/em\u003e and \u003cem\u003eBacteriodales bacterium M2\u003c/em\u003e in wild type animals compared to \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). As expected, we observed an increase in \u003cem\u003eFirmicutes (Bacillota)\u003c/em\u003e in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eC). Our data also show decreased levels of \u003cem\u003eAkkermansia\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eD), \u003cem\u003eDubosiella\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eE), \u003cem\u003eBacteroidales\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eF) along with increased abundance of Alistipes (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eE) and \u003cem\u003eDuncaniella\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eF) in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice. \u003cem\u003eAkkermansia muciniciphila\u003c/em\u003e is associated with a healthy gut by stimulating metabolic and immune responses\u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e. Interestingly, wild-type mice that were co-housed with \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice for 2 weeks had decreased levels of \u003cem\u003eAkkermansia\u003c/em\u003e (Supp Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD); however, the co-housed wild-type mice did not develop autoimmune disease, as assessed by ANA analysis (data not shown).\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eLoss of specific bacterial species\u003c/span\u003e \u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eis associated with autoimmunity\u003c/span\u003e\u003c/p\u003e\u003cp\u003eAn increasing number of studies suggest that the gut microbiota is involved in the initiation and progression of inflammatory and autoimmune diseases\u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e. To assess this, we investigated whether we could identify specific species potentially associated with disease development. We developed a disease score (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eA and Supp Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) based on the levels of ANA staining, alopecia, nephritis (as measured by glomerular inflammation), and the average number of apoptotic cells. Using a simple linear regression, we analyzed the relationship between bacterial species abundance of species and disease score (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eB-J). Healthy mice exhibit significantly higher levels of \u003cem\u003eAkkermansia\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eB) and \u003cem\u003eDubosiella\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eG) in contrast to \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice which experience significantly increased in \u003cem\u003eAlistipes\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eB), \u003cem\u003eBacteroides\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eD), \u003cem\u003eLachnospiraceae\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eE) and \u003cem\u003eHominisplanchenecus\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eF). While not statistically significant, we noticed a trend toward increased abundance of \u003cem\u003eLepagella\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eH), \u003cem\u003ePorphyromonadaceae\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eI) and \u003cem\u003eHeminphilus\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eJ) in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice. Together, these findings suggest that SCARF1 deficiency decreases beneficial bacterial populations, leading to an altered gut microbiome and potentially contributing to the gut inflammation and autoimmune pathology.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eGut functional analysis suggests that a decrease in \u0026ldquo;internal component of membrane pathway\u0026rdquo; and an increase in \u0026ldquo;regulation of cell proliferation pathway\u0026rdquo; in\u003c/span\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eScarf1\u003c/span\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003e-/- mice\u003c/span\u003e\u003c/p\u003e\u003cp\u003eMicrobiome changes affect both the metabolic and immune systems\u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e. To gain more insight into the microbiome-host interaction, we performed a functional gene pathway analysis on the bacterial species (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e). We noticed a decrease in the \u0026ldquo;integral component of membrane\u0026rdquo; in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eA, Supp Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA). In turn, we also observed an increase in the \u0026ldquo;positive regulation of cell proliferation\u0026rdquo; (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eA, Supp Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB). Despite statistical insignificance, increased regulation of cell proliferation in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice may provide a potential explanation for increased gut size observed in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice compared to wild-type controls. Pathway analysis further showed that \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice have decreased levels of L-valine biosynthesis, L-isoleucine biosynthesis and adenosine ribonucleotide de novo biosynthesis compared to wild-type (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eB). These compounds are essential in the ability to produce branched-chain fatty acids (BCFA), which are essential lipid membrane components of the gut bacteria\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e. Furthermore, these compounds have been shown to protect against intestinal damage\u003csup\u003e\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e. Alterations in the biosynthesis of BCFA have been linked to dysbiosis\u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e, further suggesting that changes in lipids can affect homeostasis. As described above, we found that \u003cem\u003eAkkermansia\u003c/em\u003e is present only in the microbiota of \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eC-D, refer Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eD and Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eB). Studies have shown that \u003cem\u003eA. muciniphila\u003c/em\u003e is involved in regulating lipid metabolism and modulating the immune response by reducing inflammation\u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e. Taken together, our data shows a different microbiome and inflammation profile between \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e and \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eDysbiosis of the gut microbiome has been linked to the development of SLE\u003csup\u003e\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e,\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e. Numerous factors have been studied associating dysbiosis with the development of SLE. Side effects of medications, increased susceptibility to infection and tissue damage are a few examples that lead to the impairment of gut homeostasis\u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e,\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e. However, the impact of genetic SLE-associated risk factors on gut dysbiosis is less understood.\u003c/p\u003e\u003cp\u003eHere we characterize the gut microbiome in a mouse model of spontaneous lupus. Using the lupus-prone mouse \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e, we observed a significant size increase in the colon size when compared to wild-type mice. This observation was striking, as the inflamed gut is shorter in size\u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e,\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e. SCARF1\u0026rsquo;s role as an efferocytosis receptor\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e,\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e\u003c/sup\u003e and the accumulation of uncleared apoptotic cells and subsequent inflammatory immune and epithelial stimulation within diseased mice suggest a possible explanation for the increase in colon size.\u003c/p\u003e\u003cp\u003eThe absence of SCARF1 disrupts immune homeostasis\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e, leading us to assess inherent changes in composition and diversity of gut microbiota between control and disease mice. Wild-type mice express a higher diversity and a richer bacterial composition when compared to \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice. Data also shows differences that are sex specific, with diseased female mice exhibiting low diversity with increased F/B ratio. Similar observations indicating significant sex-specific differences in gut mucosa have been shown using SWR \u0026times; NZB F1 (SNF1) mice\u003csup\u003e\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e. Sex hormones might be influencing the microbiome, as this was indicated in a type 1 diabetes study in which male mice were castrated resulting in disease progression\u003csup\u003e\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e,\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e. However, we observed differences in the gut microbiome that were strain specific as \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice showed similar microbiota trends regardless of being cohoused for 2 or 20 weeks.\u003c/p\u003e\u003cp\u003eSCARF1 plays a significant role in maintaining lipid homeostasis. Initially, SCARF1 on endothelial cells as a receptor for modified lipoproteins\u003csup\u003e\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e,\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e. De novo biosynthesis of L-valine, L-isoleucine and adenosine ribonucleotide is essential to produce BCFA and dampen inflammation\u003csup\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e,\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/sup\u003e. Consistent with maintaining lipid homeostasis, \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e exhibit decreased beneficial bacteria that synthesize BCFA and short-chain fatty acids (SCFA). Although additional work is needed, we can propose that the development of autoimmunity in \u003cem\u003eScarf1\u003c/em\u003e-deficient mice isdriven in part by the dysbiosis and defects in efferocytosis.\u003c/p\u003e\u003cp\u003e\u003cem\u003eAkkermansia muciniphila\u003c/em\u003e is a gram-negative anaerobic bacterium that colonizes the intestinal tract early in life\u003csup\u003e\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e. This beneficial intestinal commensal is known for colonizing the mucosal layer, where it plays an essential role in host metabolism and immune response\u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e. Control mice express high levels of \u003cem\u003eA. muciniphila\u003c/em\u003e, however this bacterium is completely absent in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice. \u003cem\u003eA. muciniphila\u003c/em\u003e is associated with health and maintaining the mucosal barrier. Furthermore, disease mice express high levels of \u003cem\u003eAlistipes\u003c/em\u003e, \u003cem\u003eBacteroides\u003c/em\u003e, \u003cem\u003eLachnospiraceae\u003c/em\u003e which are associated with dysbiosis and metabolic diseases\u003csup\u003e\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e\u003cp\u003eAltogether, our data shows various lupus-associated changes in the gut microbiome. Intestinal colonization of \u003cem\u003eA. muciniphila\u003c/em\u003e was found to negatively correlate with disease development. \u003cem\u003eA. muciniphila\u003c/em\u003e probiotics and derived postbiotics have already been identified as promising therapeutics within multiple inflammatory diseases, including SLE\u003csup\u003e\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e\u003c/sup\u003e. Our data corroborate the potential use of \u003cem\u003eA. muciniphila\u003c/em\u003e as a probiotic to decrease lupus-like symptoms and lower inflammation; however, further research is required to understand the impact of \u003cem\u003eA. municiphila\u003c/em\u003e postbiotics, or metabolites, directly. Assessing metabolic changes in control and disease murine models would be an interesting avenue given previous studies implicate SCARF1 in lipid metabolism and homeostasis\u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e. The work presented here are the first steps in understanding the role of SCARF1 in gut inflammation and homeostasis. Future work will focus on the mechanistic interaction of apoptotic debris and dysbiosis.\u003c/p\u003e"},{"header":"Material and Methods","content":"\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eMice\u003c/span\u003e:\u003c/p\u003e\u003cp\u003e All mice were maintained under micro isolation in specific pathogen\u0026ndash;free conditions at the animal facility of UMass Chan Medical School under a protocol approved by the Institutional Animal Care and Use Committee. In addition, experimental design shows a comparative study between \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e (wild-type) or \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e following ARRIVE guidelines (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://arriveguidelines.org/\u003c/span\u003e\u003cspan address=\"https://arriveguidelines.org/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e). This includes the use of control animals, inclusion criteria for male and female mice, blinding of samples for data analysis and the use of statistical methods are described below.\u003c/p\u003e\u003cp\u003eWild-type (WT, \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e+/+\u003c/sup\u003e) C57BL/6 mice were obtained from Jackson Laboratories and bred in-house. \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice were transferred from Massachusetts General Hospital and bred in-house for at least 10 generations. All mice were used after 20-weeks of age to allow for disease development, as previously described\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e. Offspring of \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e and WT mice were produced at normal Mendelian ratios. Mice were not randomized or placed in specific groups for these studies.\u003c/p\u003e\u003cp\u003e In accordance with ALAAS learning library and IACUC, for end-point studies mice were euthanize using a two-step euthanasia protocol. Mice were anesthetized with isoflurane using saturated vapor, then we performed cardiac puncture for blood collection. To ensure death, cervical dislocation was performed before harvesting organs for analysis.\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eFecal collection for microbiome preparation\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eMice were selected for analysis from a cohort of \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice and their healthy littermate controls. Following euthanasia, fecal samples were collected and immediately frozen at -80C until processing. Samples were then placed in individual tubes containing DNA stabilization buffers (Transnetyx Microbiome Kits, Cordova, TN) to preserve sample integrity and stability during shipment. All DNA extraction, library preparation, and sequencing were performed by Transnetyx (Cordova, TN).\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eDNA extraction and metagenomic sequencing\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eDNA extraction and metagenomic sequencing was perform by Transnetyx Inc. Briefly, stool DNA was extracted using a robust method that ensures reproducible extraction that captures the accurate microbial diversity. DNA quality control was performed to confirm sample integrity. Genomic DNA was converted into sequencing libraries and sequenced using shotgun metagenomic sequencing, generating approximately 2\u0026nbsp;million 2x150 bp read pairs to obtain microbial species and strain-level taxonomic resolution.\u003c/p\u003e\u003cp\u003eRaw sequencing data were uploaded to the OneCodex platform for analysis and aligned against a database of ~\u0026thinsp;148K complete microbial genomes, including 71K bacterial, 72K viral, and thousands of archaeal and eukaryotic genomes. To reduce false positives, classification results underwent further analysis to group the samples in their required experimental approach.\u003c/p\u003e\u003cp\u003eMicrobial taxonomy and diversity metrics were computed within OneCodex to compare \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e mice and healthy littermate controls. Samples were also compared with Transnetyx\u0026rsquo;s global diversity averages derived from historic datasets for quality control. Low read counts were normalized relative to the total number of identifiable reads within each host sample. Finally, sequencing data were aligned against the Gene Ontology (GO) and KEGG Orthology databases via the OneCodex platform for downstream functional analysis of microbial communities.\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eAutoantibody profiles\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eAntinuclear autoantibodies (ANA) were measured as previously described\u003csup\u003e\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e\u003c/sup\u003e. ANA assays from mouse serum were performed using immunofluorescence assays according to the manufacturer\u0026rsquo;s instructions (Bio-Rad). Mouse serum was diluted at 1:200 and incubated with Hep-2 cells, followed by Alexa Fluor 488 secondary to detect bound ANAs (Cat #A11001, Invitrogen). Staining was scored by three independent observers \u0026lsquo;blinded\u0026rsquo; to the genotypes of the mice.\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eHistology\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eThe small and large intestines were dissected from 20-week-old \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u0026minus;/\u0026minus;\u003c/sup\u003e and C57BL/6J (B6) wild-type mice. To assess histological signs of gut inflammation, the intestines were fixed in 10% phosphate-formalin and embedded in paraffin. Sections were prepared and stained in hematoxylin-eosin. For all other studies, the intestines were flash frozen on Optimal Cutting Temperature (OCT) Compound (Sciegen Scientific, Gardena CA) embedding media. Sections from frozen samples were prepared (7um) and stained as described below. All slides were imaged using ECHO fluorescent microscope equipped with a high-resolution (Discover Echo, San Diego, CA, USA) and analyzed using Discover ECHO App (iOS 16\u003cem\u003e+\u003c/em\u003e) (Discover Echo, San Diego, CA, USA) and Adobe Photoshop (Adobe, San Jose CA).\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eTUNEL assay\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eFrozen sections (7um) were allowed to warm to 25\u0026deg;C. A TUNEL assay (terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling) for the detection of apoptotic cells was done according to manufacturer\u0026rsquo;s instructions (Invitrogen). Paraformaldehyde-fixed tissue was made permeable with 60 minutes 0.25% Triton X-100 in PBS at room temperature and was washed twice with PBS. Slides were incubated with reaction buffer (25 mM Tris-HCl, pH 6.6, 200 mM sodium cacodylate, 0.25 mg/ml BSA and 1 mM cobalt) containing terminal deoxynucleotidyl transferase, then were washed twice and were incubated for 45 min at 25\u0026deg;C in the dark (protected from light) with reaction buffer containing the fluorescent label Alexa Fluor 488. To identify tissue structures, sections we stained with Phalloidin-Rhodamin for 25 min at RT and EPCAM1 cells were stained for 30 min at 25\u0026deg;C with Alexa Fluor 647\u0026ndash;anti-EPCAM1 (1:50 dilution; Invitrogen A22283) in PBS. Samples were washed twice with PBS. Finally, DNA was stained for 10 min at 25\u0026deg;C with Hoechst 33342 (1:1,000 dilution; Molecular Probes, Invitrogen). Slides were mounted with Prolong Gold antifade reagent (Invitrogen P36935) and were visualized with ECHO fluorescent microscope equipped with a high-resolution. Data were analyzed with Discover ECHO App (iOS 16\u003cem\u003e+\u003c/em\u003e) (Discover Echo, San Diego, CA, USA) and Adobe Photoshop (Adobe, San Jose CA).\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eHistological assessment and disease scoring\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eHistological evaluation of the intestinal H\u0026amp;E sections was performed in a blinded manner by a pathologist (Jadhav, Nupur). Inflammation and cellular infiltration were scored, and cell types were identified. Tissue inflammation severity was scored on a scale from 0\u0026ndash;5, where grade of inflammation 1- minimal, 2- mild, 3-moderate, 4-severe, 5-severe with ulceration. For presence of apoptosis in the tissue, 1- present and 2- absent. A composite disease score was calculated by averaging the individual scores for antinuclear autoantibodies (ANA), alopecia, nephritis, and the number of apoptotic cells.\u003c/p\u003e\u003cdiv id=\"Sec5\" class=\"Section2\"\u003e\u003ch2\u003eStatistical analysis:\u003c/h2\u003e\u003cp\u003eStatistical calculations were done with a statistical software package GraphPad Prism, version 10.4.2 (GraphPad Software, San Diego, CA). For comparisons between two or more groups, the mean\u0026thinsp;\u0026plusmn;\u0026thinsp;s.e.m was analyzed by unpaired two-tailed Student\u0026rsquo;s t test or ANOVA, respectively. Statistical analysis of the microbiota profiling data was performed on the proportional representation of the taxa using Shapiro-Wilk normality test. Parametric test with Welch\u0026rsquo;s corrections or nonparametric test with Mann-Whitney corrections were used depending on if passed the normality test. Multivariate analyses of disease score versus relative abundance of bacteria were done. The investigators were not blinded to the genotype of the mice except where indicated. Values of P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 were considered statistically significant.\u003c/p\u003e\u003c/div\u003e"},{"header":"Declarations","content":"\u003ch2\u003eFunding\u003c/h2\u003e\n\u003cp\u003eThis work was funded by the Department of Defense LRP-Impact Award (W81XWH-21-1-0803) (ZGRO), Lupus Research Alliance Innovation Award (ZGRO) and UMass Chan Medical School Start-up funds (ZGRO).\u003c/p\u003e\n\u003ch2\u003eAcknowledgements\u003c/h2\u003e\n\u003cp\u003eWe would like to acknowledge lead microbiome researchers, Dr. Beth McCormick and Dr. Ana Maldonado-Contreras, for their great advice and expertise. We also thank Dr. Stuart Levitz and his lab for providing additional advice in the preparation of the manuscript.\u003c/p\u003e\n\u003ch2\u003eConflict of Interest\u003c/h2\u003e\n\u003cp\u003eJMR is an inventor on patent application #62489191 and #15/851,651 which covers IL-15 and CXCR3 for the treatment of vitiligo, respectively; and on patent #63/478,900 filed for \u0026ldquo;Diagnosis of skin diseases in veterinary and human patients\u0026rdquo; for CTCL. The other authors have no conflicts of interest to disclose.\u003c/p\u003e\n\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\n\u003cp\u003eDMS and ZGRO designed and performed the experimental, analyzed data and wrote the manuscript. ZGRO developed the mouse model. DMS and ZGRO figures 1-5ZGRO and DVJ figure 6NJ Figure 1 and all pathologySH, MC, JMR, HN provided advice in the experimental design and edited the manuscript. JMR, DVW and NF validated the results. NJ analyzed pathology samples. JMR, DVW, HN provided advice on data analysis.All authors reviewed the manuscript\u003c/p\u003e\n\u003ch2\u003eAcknowledgement\u003c/h2\u003e\n\u003cp\u003eWe would like to acknowledge lead microbiome researchers, Dr. Beth McCormick and Dr. Ana Maldonado-Contreras, for their great advice and expertise. We also thank Dr. Stuart Levitz and his lab for providing additional advice in the preparation of the manuscript.\u003c/p\u003e\n\u003ch2\u003eData Availability\u003c/h2\u003e\n\u003cp\u003eData is provided within the manuscript or supplementary information files. The datasets for the current study are available through OneCodex upon request to the corresponding author.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eKeogh, C. E., Rude, K. M. \u0026amp; Gareau, M. G. Role of pattern recognition receptors and the microbiota in neurological disorders. \u003cem\u003eJ. 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Immunol.\u003c/em\u003e \u003cb\u003e16\u003c/b\u003e, 495\u0026ndash;504. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1038/ni.3143\u003c/span\u003e\u003cspan address=\"10.1038/ni.3143\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2015).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"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":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"lupus, microbiome, autoimmune, inflammation, gut, SCARF1","lastPublishedDoi":"10.21203/rs.3.rs-7023221/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7023221/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eSystemic lupus erythematosus (SLE) is a complex autoimmune disease known for its heterogeneity in both manifestation and presentation. Recent evidence has increasingly implicated the gut microbiome within immunomodulation and autoimmunity. This study aims to characterize the intestinal inflammation and microbial profile associated with autoimmune diseases, particularly SLE, and to identify unique biomarkers and shared microbial signatures for potential therapeutic measures. Our lab identified scavenger receptor class F, member 1 (SCARF1, SREC-1) as an efferocytosis receptor essential for the clearance of apoptotic debris, and its deficiency results in the development of lupus-like disease. SCARF1 is crucial in immune homeostasis, and defects in efferocytosis lead to inflammation. However, the role of SCARF1 in homeostasis in the gut remains to be elucidated. To answer our question, we analyzed and compared the metagenomic datasets generated through whole genome shotgun sequencing between our \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e lupus-prone mouse model and healthy counterparts. We found that \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice had significantly lengthened intestines, elevated immune cell infiltration, and structural changes in the colon. Microbiome analysis revealed gut dysbiosis, including reduced alpha diversity and increased F/B ratio. Notably, beneficial taxa such as \u003cem\u003eAkkermansia muciniphila\u003c/em\u003e was absent in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice. Linear regression analysis identified positive associations between lupus disease severity and increased abundances of \u003cem\u003eBacillota, Alistipes, Lachnospiraceae\u003c/em\u003e, and \u003cem\u003eHominisplanchenecus\u003c/em\u003e. Function analysis of the gut microbiome in \u003cem\u003eScarf1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice indicated downregulation of multiple pathways related to cell proliferation. These findings highlight the role of SCARF1 involvement in the gut microbiome and immune regulation in the context of inflammation and SLE.\u003c/p\u003e","manuscriptTitle":"SCARF1 Deficiency Exacerbates Gut Inflammation and Autoimmune Pathology","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-07-24 14:48:40","doi":"10.21203/rs.3.rs-7023221/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-10-13T11:00:43+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-12T21:40:38+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-11T17:48:22+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-09T17:15:37+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-09-29T08:30:19+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"265146168181982998329141005073773150419","date":"2025-09-15T02:51:26+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"314656208384331339550630086938959500913","date":"2025-09-11T11:55:31+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"229230169536573905499878186417267447738","date":"2025-09-11T10:48:03+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"43149688477214099895569078727352218973","date":"2025-09-11T09:16:47+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-08-29T19:37:37+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"208962673594807378933921976475335764373","date":"2025-08-03T15:47:32+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"186744415397667292314917978550266207738","date":"2025-07-29T20:43:13+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-07-29T13:51:10+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-07-10T14:49:50+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-07-07T13:22:50+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2025-07-07T13:19:24+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ef95c470-0c1f-4e64-a0a3-3a6cfe3fe9ef","owner":[],"postedDate":"July 24th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":51348147,"name":"Biological sciences/Computational biology and bioinformatics"},{"id":51348148,"name":"Health sciences/Diseases"},{"id":51348149,"name":"Biological sciences/Immunology"},{"id":51348150,"name":"Biological sciences/Microbiology"}],"tags":[],"updatedAt":"2026-02-16T15:59:56+00:00","versionOfRecord":{"articleIdentity":"rs-7023221","link":"https://doi.org/10.1038/s41598-026-39378-7","journal":{"identity":"scientific-reports","isVorOnly":false,"title":"Scientific Reports"},"publishedOn":"2026-02-11 15:57:11","publishedOnDateReadable":"February 11th, 2026"},"versionCreatedAt":"2025-07-24 14:48:40","video":"","vorDoi":"10.1038/s41598-026-39378-7","vorDoiUrl":"https://doi.org/10.1038/s41598-026-39378-7","workflowStages":[]},"version":"v1","identity":"rs-7023221","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7023221","identity":"rs-7023221","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}