{"paper_id":"41b000b9-253e-4301-b9be-ecc38c8ee29c","body_text":"Managing diabetes with nanomedicine: nanoMIL-89 as a promising drug delivery system for metformin. | 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 Managing diabetes with nanomedicine: nanoMIL-89 as a promising drug delivery system for metformin. Hana Mohamed, Nura Mohamed, Shantelle Macasa, Hamda Basha, Adna Adan, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-3893992/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 28 Dec, 2024 Read the published version in Scientific Reports → Version 1 posted 11 You are reading this latest preprint version Abstract Diabetes Mellitus is a chronic disease characterized by metabolic defects, including insulin deficiency and resistance. Individuals with diabetes are at increased risk of developing cardiovascular complications, such as atherosclerosis, coronary artery disease, and hypertension. Conventional treatment methods, though effective, are often challenging, costly, and may lead to systemic side effects. This study explores the potential of nanomedicine applications, specifically Metal-Organic Frameworks (MOFs), as drug carriers to overcome these limitations. The Materials Institute Lavoisier-89 nanoparticles (nanoMIL-89) have previously demonstrated promise as a drug delivery vehicle for chronic diseases due to their anti-oxidant and cardio-protective properties. In this investigation, nanoMIL-89 was loaded with the anti-diabetic drug metformin (MET), creating MET@nanoMIL-89 formulation. We examined the drug release kinetics of MET@nanoMIL-89 over 96 hours and assessed its impact on the viability of various endothelial cells. Furthermore, we investigated the nanoformulation effect on inflammatory markers in these cells and explored its influence on phosphorylated eNOS, total eNOS, and AKT levels. Our findings indicate that nanoMIL-89 effectively released metformin over 96 hours and caused a concentration-dependent reduction in CXCL-8 release from endothelial cells. Notably, MET@nanoMIL-89 reduced dihydroethidium levels and increased phosphorylated eNOS, total eNOS, and AKT levels. Our results underscore the potential of nanoMIL-89 as a versatile potential drug delivery platform for anti-diabetic drugs, offering a prospective therapeutic approach for diabetic patients with associated cardiovascular complications. Biological sciences/Biotechnology Health sciences/Diseases Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 1. Introduction Diabetes mellitus (DM) stands as one of the most prevalent chronic metabolic disorders globally, associated with decreased life expectancy and heightened mortality rates [ 1 ]. The escalating prevalence of DM is alarming, with a projected rise from approximately 424.9 million cases worldwide in 2019 to an anticipated 700 million cases by 2045 [ 2 ]. Notably, Type-2 DM constitutes approximately 90% of all DM cases globally [ 3 ]. Achieving glycemic control in type-2 DM involves lifestyle modifications alongside pharmacological interventions, including oral antidiabetic drugs such as biguanides (e.g., metformin)[ 4 ], sulfonylureas[ 4 ], dipeptidyl peptidase IV (DPP-4) inhibitors (gliptins)[ 4 ], glucagon-like peptide-1 (GLP-1) receptor agonists[ 4 ], sodium-glucose cotransporter (SGLT-2) inhibitors[ 4 ], alpha-glucosidase inhibitors[ 4 ], thiazolidinediones[ 4 ], and amylin analogs[ 4 ]. However, drug resistance, side effects, and individual variability responses may impede the effectiveness of these conventional therapies [ 1 ]. DM is frequently accompanied by complications such as vision, gastric, renal, and notably cardiovascular issues; with the latter being the most common even with intensive glycemic control. [ 5 ]. Recent studies indicate that DM-associated cardiovascular complications may arise from dysfunctional endothelium, observable even in early DM stages, making endothelial dysfunction a potential early biomarker and risk factor for cardiovascular diseases (CVDs) [ 6 , 7 ]. Understanding the mechanisms through which DM induces endothelial cell dysfunction is crucial for preventing the onset of CVDs, such as atherosclerosis, hypertension, and stroke [ 8 ]. Despite technological advancements in DM treatments, management remains suboptimal, marked by side effects and limitations [ 4 ]. Nanomedicine applications emerge as promising solutions to address these challenges. Nanotherapies tailored for DM management offer effective blood glucose regulation and gradual release of antidiabetic drugs [ 1 ]. This extension of drug duration reduces the likelihood of acute and chronic complications associated with antidiabetic drugs [ 1 ]. The exploration of various nanoparticles (NPs) loaded with antidiabetic drugs, including liposomes, niosomes, polymers, dendrimers, micelles, and metal-based NPs, has been ongoing [ 9 – 22 ]. However, safety, tolerance, and stability challenges have been encountered [ 9 – 22 ]. To overcome these limitations, we propose the utilization of iron oxide-based NPs, known for their anti-inflammatory and hypoglycemic properties [ 23 , 24 ]. They also possess antioxidant activities that can be easily detected via magnetic resonance imagining (MRI) [ 25 ]. Among these, nanoMIL-89, a metal-organic framework (MOF), demonstrates suitability as a drug carrier. NanoMIL-89 has exhibited selective uptake by endothelial cells, passage to daughter cells, and reduced inflammatory markers in dysfunctional endothelial cells [ 26 ]. Additionally, it has proven to be a suitable carrier for the drug sildenafil, used to treat pulmonary arterial hypertension (PAH) [ 27 ]. Moreover, it has been demonstrated that nanoMIL-89 exhibits non-toxic effects both in vitro and in vivo , as evidenced by studies utilizing zebrafish embryos and rodent models [ 24 , 26 ]. Our selection of nanoMIL-89 as a carrier for the antidiabetic drug metformin (MET) is grounded in its cardioprotective activity and ability to carry small drugs with a molecular weight under 500 Da [ 27 ]. Moreover, MET's positive effects on blood glucose levels, insulin sensitivity, antioxidant activity, and endothelial protection align with our goal of mitigating hyperglycemia's impact on the endothelium [ 28 , 29 ]. MET's association with a lower risk of CVDs in type-2 DM patients and its compatibility with nanoMIL-89 due to its size further reinforce our choice [ 30 ]. Despite MET’s low bioavailability and short half-life, loading it into nanoMIL-89 presents an opportunity to reduce dosage, improve patient compliance, and potentially minimize gastrointestinal side effects [ 31 – 33 ]. Therefore, our objective is to load MET into nanoMIL-89, investigate its release kinetics, and evaluate its impact on endothelial cell function and oxidative stress under both normoglycemic and hyperglycemic conditions in vitro . 2. Results 2.1. Pharmacokinetics of metformin-loaded nanoMIL-89 (MET@nanoMIL-89): The pharmacokinetics of MET@nanoMIL-89 were investigated and compared to MET alone. MET@nanoMIL-89 demonstrated a time-dependent release of MET, persisting for up to 96 hours with an approximate concentration of ~ 96 µg/ml. Notably, the initial hours (0.5, 1, 3, 6, and 24 hrs) showed a gradual release of MET (~ 50 µg/ml), reaching a peak concentration of ~ 100 µg/ml after 48 hours, maintaining stability until the end of the experiment (Fig. 1 ). In contrast, MET alone exhibited consistent concentrations of 100–180 µg/ml throughout the experiment, while the nanoMIL-89 (used as a control) showed no detectable release. 2.2. Effect of the metformin-Loaded nanoMIL-89 on endothelial cells proliferation, anti-inflammatory and antioxidant effect: The impact of nanoMIL-89 and MET@nanoMIL-89 on endothelial cell proliferation was assessed using different concentrations [0, 1, 3, 10, 30, and 100 µg/ml]. The cell respiration rate, measured by the AlamarBlue® assay, indicated that nanoMIL-89 and MET@nanoMIL-89 had no significant effect on cell viability (respiration) at concentrations ≤ 30 µg/ml in all endothelial cells (Fig. 2 a: HUVECs, PAECs; and Fig. 3 a: HAECs, and HAECs-Type-2 DM). The cytotoxicity of the nanoformulations used for antidiabetic drug delivery was investigated using the LDH Assay. Results from the LDH release assay (Fig. 2 b: HUVECs, PAECs; and Fig. 3 b: HAECs, and HAECs-Type-2 DM) demonstrated that nanoMIL-89 and MET@nanoMIL-89 did not induce significant cytotoxic effects in any of the four cell lines. Furthermore, the anti-inflammatory effect of nanoMIL-89 and MET@nanoMIL-89 was examined by measuring the levels of CXCL-8 in the conditioned media. Results (Fig. 2 c: HUVECs, PAECs; and Fig. 3 c: HAECs and HAECs-Type-2 DM) revealed that both nanoformulations exhibited a concentration-dependent decrease in CXCL-8 release from endothelial cells. This effect was more pronounced in HUVECs and HAECs, where the reduction in CXCL8 release was more significant in the MET@nanoMIL-89 group compared to the group treated with nanoMIL-89 alone. 2.3. Effect of metformin-loaded nanoMIL-89 on reducing reactive oxygen Species production by endothelial cells: Hyperglycemic conditions are known to induce endothelial dysfunction, reducing nitric oxide (NO) production, and increasing reactive oxygen species (ROS) release and oxidative stress. As MET is acknowledged for its in vitro antioxidant effects independently [ 39 ], this experiment delves into exploring whether MET-loaded nanoMIL-89 can sustain or amplify these antioxidant properties. Results demonstrated that hyperglycemia increased ROS levels, evident by higher DHE levels in cells seeded under hyperglycemic media for the control groups. The use of nanoMIL-89 and MET@nanoMIL-89 induced a concentration-dependent reduction in DHE levels, starting at concentrations ≥ 1µg/ml in the hyperglycemic environment (Fig. 4 ). This indicates that the nanoformulation possesses antioxidant activity, reducing DHE release under hyperglycemic conditions. 3.4. Effect of metformin-loaded nanoMIL-89 on endothelial cells eNOS and AKT levels: Given that hyperglycemic conditions induce a reduction in eNOS phosphorylation levels in endothelial cells, our results showed a decrease in eNOS levels under hyperglycemic conditions (Fig. 5 ). Furthermore, the use of MET restored the reduction in eNOS levels in the treated group compared to the untreated group (Fig. 5 b). Additionally, both nanoMIL-89 and MET@nanoMIL-89 could induce a concentration-dependent increase in eNOS levels, with MET@nanoMIL-89 causing a more significant increase. This could be attributed to the individual increases in eNOS levels by the nanoparticle and metformin, and when combined, MET@nanoMIL-89 further enhanced the efficacy of metformin, thus promoting eNOS activity. Moreover, we measured the levels of AKT, with hyperglycemic conditions generally reducing AKT levels in endothelial cells. However, when cells were incubated in hyperglycemic media, both nanoMIL-89 and MET@nanoMIL-89 induced an increase in AKT levels. Nevertheless, MET@nanoMIL-89 caused a more significant increase compared to nanoMIL-89 alone (Fig. 6 ). This correlates with the finding that both nanoMIL-89 and metformin have antioxidant activity, and when conjugated, the efficacy of the treatment increases. 3. Discussion Metformin, renowned for its potent antidiabetic and cardioprotective properties, has been recognized to induce eNOS and Akt phosphorylation, as demonstrated in previous studies by our collaborators and others [ 29 ]. Additionally, it exhibits the ability to decrease DHE levels, thereby mitigating oxidative stress associated with diabetes mellitus (DM) [ 39 ]. Despite its efficacy, Metformin is burdened by gastrointestinal side effects and a short half-life (~ 4 hrs), limiting its therapeutic impact [ 33 ]. Harnessing nanomedicine to encapsulate metformin in nanoparticles (NPs) offers a promising avenue to overcome these challenges [ 33 ]. This study focused on exploring the potential of nanoMIL-89 as a carrier for metformin, and the observed results suggest that nanoMIL-89 indeed possesses promising therapeutic features for delivering metformin. The evaluation of metformin release from nanoMIL-89 at different time points revealed a successful and controlled release, reaching approximately 100 µg/ml at 96 hours. This gradual release pattern is crucial in evading premature drug deactivation by the immune system, preventing rapid immune tolerance, and ensuring sustained therapeutic efficacy [ 40 ]. These findings align with a previous study on nanoMIL-89, which demonstrated its ability to circulate, and release loaded drugs in plasma for up to 4 days [ 27 ], potentially addressing metformin's short plasma half-life [ 31 , 32 ]. The in vitro assessments were performed using endothelial cells, considering the well-documented impact of DM on endothelial cell dysfunction. Hyperglycemia-induced dysfunction is characterized by reduced eNOS phosphorylation, altered total eNOS function, decreased Akt phosphorylation [ 29 ], and increased oxidative stress represented by elevated DHE levels [ 39 ]. Results from this study, utilizing endothelial cells from various sources, revealed no detectable cytotoxic effects of MET@nanoMIL-89 and nanoMIL-89. Metabolic dysfunction, rather than toxicity from nanoformulation, was responsible for the observed decrease in cell proliferation. Moreover, loading metformin into nanoMIL-89 effectively countered hyperglycemia-induced oxidative stress, reducing DHE levels. Additionally, it elevated phosphorylated eNOS, total eNOS, and AKT levels in endothelial cells under hyperglycemic conditions, mimicking the diabetic environment. These findings imply that nanoMIL-89 may possess cardioprotective activity by mitigating endothelial cell dysfunction, a property often associated with metallic NPs like iron oxide NPs [ 3 ]. 4. Methodology 4.1. NPs synthesis, characterizations, and metformin loading: NanoMIL-89 was synthesized according to previously published procedures [ 24 , 34 ]. In brief, 10 mmol of iron (III) chloride hexahydrate (FeCl 3 .6H 2 O) and 10 mmol of trans-trans muconic acid were dissolved in 100 ml of absolute ethanol (99.8%). After subjecting the reaction mixture to sonication for 15 minutes, the subsequent step involved adding 20 ml of glacial acetic acid (99.8%) before heating it at 90°C for 24 hours within a Parr reactor. Precipitated nanoparticles were recovered by centrifugation at 7000 rpm for 15 minutes, followed by several washes in distilled water. The precipitate was then vacuum-dried to obtain nanoMIL-89, which presented as a brown precipitate (100–200 mg/reaction). Subsequently, nanoMIL-89 was loaded with metformin as follows: Solutions of 1000, 300, 100, 30, and 10 µg/ml metformin in Phosphate Buffer Saline (PBS; pH 7.4) were prepared. To each 5 ml of metformin concentration, 200 mg of nanoMIL-89 was added. The mixture was incubated overnight (16–18 hrs.) at room temperature, and metformin-loaded nanoMIL-89 was harvested by centrifugation at 7000 rpm for 15 minutes at room temperature. The harvested metformin-loaded nanoMIL-89 was characterized by Infrared Imaging (IR), Powder X-ray Diffraction (PXRD), and Scanning Electron Microscope (SEM), with results compared to the unloaded nanoMIL-89. Finally, the metformin nanoMIL-89 prototype was utilized for pharmacokinetic and in vitro experiments. 4.2. Metformin release pharmacokinetics: As described above, nanoMIL-89 was loaded with MET, and the release kinetics were measured over 96 hours. NanoMIL-89 loaded with MET was precipitated at 7000 rpm at room temperature for 15 minutes. The resulting precipitate was dissolved in 5 ml of PBS and incubated at room temperature. At various time points (0, 0.5, 1, 3, 6, 24, 48, 72, and 96 hours), 1.5 ml of the supernatant was collected. The amount of released MET into PBS was determined using a previously described colorimetric method [ 35 ], which relies on the interaction of MET with ninhydrin reagent at pH 5.6, resulting in a red color. A MET standard curve was incorporated. Each collected 1.5 ml aliquot was mixed with 10.0 ml of Phthalate buffer (pH = 5.6) and 5.0 ml of 0.1% Ninhydrin. The mixture underwent heating in a water bath for 1 hour at 90ºC. Subsequently, the solution was cooled to room temperature and topped up with distilled water to a final volume of 25 ml, and the optical density was measured at 567 nm. Readouts were corrected for a blank, and concentrations were estimated using the MET standard curve [ 35 ]. 4.3. Effect of metformin-loaded nanoMIL-89 on endothelial cell proliferation, anti-inflammatory, and antioxidant effects: 4.3.1. Cell line culture: Standard culture techniques were employed for seeding and maintaining the following endothelial cell lines: (i) Human Pulmonary Artery Endothelial Cells (PAECs) [ 36 ] and (ii) Human Umbilical Vein Endothelial Cells (HUVECs) [ 37 ], both cultured according to the suppliers' recommendations. The responses of healthy cells cultured under hyperglycemic conditions were compared to those of a diabetic endothelial cell line: Human Primary Diabetic Aortic Endothelial Cells (HDAOEC; Cellbiologics), cultured according to the manufacturer's instructions [ 38 ]. 4.3.2. Cell viability, cytotoxicity, and cytokine release: Endothelial cells were plated in 96-well plates at a seeding density of 10,000 cells/well under hyperglycemic conditions (25 mM glucose). After overnight incubation (16–18 hrs) in humidified conditions (37 ºC, 5% CO 2 ), cells were treated with either nanoMIL-89 alone (0, 1, 3, 10, 30, and 100 µg/ml), metformin alone (50 µM), or metformin-loaded nanoMIL-89 (MET@nanoMIL-89; 0, 1, 3, 10, 30, and 100 µg/ml) for 24 hrs under the same conditions. Adherent cells were used to determine the effects of different treatments on cell viability using the AlamarBlue® assay following the manufacturer’s instructions. Conditioned media were collected to assess cell cytotoxicity using the LDH Kit (Abcam, UK) and anti-inflammatory effects using CXCL8-ELISA Kits (R&D systems, USA). AlamarBlue, LDH, and ELISA assays were performed following manufacturer instructions. 4.3.3. Effect of metformin-loaded nanoMIL-89 on reactive oxygen species, eNOS, and AKT levels: Endothelial cells were seeded into a black 96-well plate and treated as described above. After 24 hrs in humidified conditions (37 ºC, 5% CO 2 ), the Dihydroethidium (DHE, Abcam, UK) assay kit was used to measure Reactive Oxygen Species (ROS) release according to the manufacturer's instructions. For Endothelial Nitric Oxide Synthase (eNOS) and Phospho-AKT, cells were seeded into 6-well plates at a density of 100,000 cells/well and treated as described above for 24 hours. Following treatment, proteins were extracted, and the levels of phosphorylated eNOS and total eNOS were measured using the Human Phospho-eNOS and Total eNOS ELISA kit (RayBio®, USA) as instructed. Additionally, the expression of total and phosphorylated AKT was measured using the Phospho-AKT (RayBio®, USA) ELISA kit following the manufacturer's instructions. Statistical analysis : Results from drug release studies and in vitro culture studies were analyzed and presented as mean ± SEM for n experiments with well-defined figure legends. Statistical tests were performed using GraphPad Prism v5. Two-way ANOVA followed by Bonferroni Posttests were used for comparing effects between different groups (i.e., metformin-loaded nanoMIL-89, metformin alone, and unloaded nanoMIL-89). One-way ANOVA followed by Dunnett’s Multiple Comparison Tests was used for comparing the effect of different nanoformulation concentrations within each group compared to the control. Statistical significance was noted at p < 0.05. 5. Conclusion This study highlights nanoMIL-89 as a biocompatible nanoparticle endowed with cardioprotective properties, as evidenced by its antioxidant and anti-inflammatory effects, along with its capacity to enhance the nitric oxide pathway. The observed potential suggests that this nanoparticle could have versatile applications in diverse diseases, such as cardiovascular diseases (CVDs) and diabetes mellitus (DM). The experimental findings underscore nanoMIL-89 as a promising carrier for delivering antidiabetic drugs, specifically metformin. However, to gain a comprehensive understanding of its mechanism of action, pharmacodynamics, and long-term biological effects, further in vivo studies utilizing diabetic models are imperative. The success of such studies could propel this nanoparticle into pre-clinical stages, unveiling promising therapeutic opportunities not only for diabetes but also for other challenging diseases, including cancer. The ongoing exploration of nanoMIL-89's capabilities holds great potential for advancing our understanding of nanomedicine applications and their broader impact on medical treatments. Declarations Acknowledgement: We express our appreciation for the Open Access funding provided by the Qatar National Library. Furthermore, we convey our profound gratitude to the Qatar National Research Fund for offering financial support for this research through grant # UREP26-019-3-006 awarded to H.A.S. We also acknowledge the British Pharmacological Society, United Kingdom, for the generous assistance provided through the Pickford Award, granted to N.A.M. This support played a crucial role in facilitating the development of the nanoparticles employed in this study. The graphical abstract utilized in this research was generated using BioRender.com. Authors Contributions: Conceptualization: [N.A.M., I.M., H.A.S.], Methodology: [N.A.M., I.M., H.A.S.], Formal analysis and investigation: [N.A.M., I.M., H.A.S., H.A.M., S.S.M., H.K.B., A.M.A.]; Writing - original draft preparation: [H.A.M, N.A.M.]; Writing - review and editing: [H.A.S., S.C., N.A.M., I.M., C.R.T., H.D.]; Funding acquisition: [H.A.S., N.A.M., I.M.]; Resources: [N.A.M., I.M., H.A.S.]; Supervision: [N.A.M., H.A.S.]. All authors have read and agreed to the published version of the manuscript. Conflict of interest: The authors declare that they have no conflict of interest regarding the publication of this article. Funding: This research was supported by Qatar National Research Fund under the Undergraduate Research experience Program Grant no. [UREP26-019-3-006] (to H.A.S.), and the Pickford Award from the British Pharmacological Society (to N.A.M). Ethical Approval: This article does not contain any studies with human participants or animals performed by any of the authors. References Kesharwani, P.; Gorain, B.; Low, S.Y.; Tan, S.A.; Ling, E.C.S.; Lim, Y.K.; Chin, C.M.; Lee, P.Y.; Lee, C.M.; Ooi, C.H.; et al. Nanotechnology based approaches for antidiabetic drugs delivery. Diabetes Res Clin Pract 2018 , 136 , 52-77, doi:10.1016/j.diabres.2017.11.018. Saeedi P, P.I., Salpea P, Malanda B, Karuranga S, Unwin N, Colagiuri S, Guariguata L, Motala AA, Ogurtsova K, Shaw JE, Bright D, Williams R. IDF Diabetes Atlas Committee. Global and regional diabetes prevalence estimates for 2019 and projections for 2030 and 2045: Results from the International Diabetes Federation Diabetes Atlas, 9th edition. . 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Metformin kinetics in healthy subjects and in patients with diabetes mellitus. Br J Clin Pharmacol 1981 , 12 , 235-246, doi:10.1111/j.1365-2125.1981.tb01206.x. Padwal, R.S.; Gabr, R.Q.; Sharma, A.M.; Langkaas, L.A.; Birch, D.W.; Karmali, S.; Brocks, D.R. Effect of gastric bypass surgery on the absorption and bioavailability of metformin. Diabetes Care 2011 , 34 , 1295-1300, doi:10.2337/dc10-2140. Cetin, M.; Sahin, S. Microparticulate and nanoparticulate drug delivery systems for metformin hydrochloride. Drug Deliv 2016 , 23 , 2796-2805, doi:10.3109/10717544.2015.1089957. Horcajada, P.; Chalati, T.; Serre, C.; Gillet, B.; Sebrie, C.; Baati, T.; Eubank, J.F.; Heurtaux, D.; Clayette, P.; Kreuz, C.; et al. Porous metal-organic-framework nanoscale carriers as a potential platform for drug delivery and imaging. Nat Mater 2010 , 9 , 172-178, doi:10.1038/nmat2608. Vijay Vikram Singh, A.K., G.T.Kulkarni, R. Hemalatha. Estimation and validation of metformin by colorimetry method. Analytical CHEMISTRY An Indian Journal 2010 , ACAIJ, 9(1) 2010 145-150. Sakao, S.; Tatsumi, K.; Voelkel, N.F. Endothelial cells and pulmonary arterial hypertension: apoptosis, proliferation, interaction and transdifferentiation. Respir Res 2009 , 10 , 95, doi:10.1186/1465-9921-10-95. Bala, K.; Ambwani, K.; Gohil, N.K. Effect of different mitogens and serum concentration on HUVEC morphology and characteristics: implication on use of higher passage cells. Tissue Cell 2011 , 43 , 216-222, doi:10.1016/j.tice.2011.03.004. Cutiongco, M.F.A.; Chua, B.M.X.; Neo, DJH; Rizwan, M.; Yim, E.K.F. Functional differences between healthy and diabetic endothelial cells on topographical cues. Biomaterials 2018 , 153 , 70-84, doi:10.1016/j.biomaterials.2017.10.037. Ouslimani, N.; Peynet, J.; Bonnefont-Rousselot, D.; Therond, P.; Legrand, A.; Beaudeux, J.L. Metformin decreases intracellular production of reactive oxygen species in aortic endothelial cells. Metabolism 2005 , 54 , 829-834, doi:10.1016/j.metabol.2005.01.029. Li, H., Yang, Y. G., & Sun, T. (2022). Nanoparticle-Based Drug Delivery Systems for Induction of Tolerance and Treatment of Autoimmune Diseases. Frontiers in bioengineering and biotechnology , 10 , 889291. https://doi.org/10.3389/fbioe.2022.889291 Additional Declarations No competing interests reported. Supplementary Files GraphicalAbstract.jpeg Cite Share Download PDF Status: Published Journal Publication published 28 Dec, 2024 Read the published version in Scientific Reports → Version 1 posted Editorial decision: Revision requested 26 Apr, 2024 Reviews received at journal 25 Apr, 2024 Reviewers agreed at journal 24 Mar, 2024 Reviews received at journal 23 Feb, 2024 Reviewers agreed at journal 07 Feb, 2024 Reviewers agreed at journal 07 Feb, 2024 Reviewers invited by journal 05 Feb, 2024 Editor assigned by journal 04 Feb, 2024 Editor invited by journal 04 Feb, 2024 Submission checks completed at journal 04 Feb, 2024 First submitted to journal 24 Jan, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {\"props\":{\"pageProps\":{\"initialData\":{\"identity\":\"rs-3893992\",\"acceptedTermsAndConditions\":true,\"allowDirectSubmit\":false,\"archivedVersions\":[],\"articleType\":\"Article\",\"associatedPublications\":[],\"authors\":[{\"id\":271066201,\"identity\":\"208a7b3a-269b-4d68-9a62-9b8781d48f43\",\"order_by\":0,\"name\":\"Hana Mohamed\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Qatar University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Hana\",\"middleName\":\"\",\"lastName\":\"Mohamed\",\"suffix\":\"\"},{\"id\":271066202,\"identity\":\"dc930ea8-dc86-4708-8389-24e707aefe6b\",\"order_by\":1,\"name\":\"Nura Mohamed\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Qatar University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Nura\",\"middleName\":\"\",\"lastName\":\"Mohamed\",\"suffix\":\"\"},{\"id\":271066203,\"identity\":\"20ad853a-9aaa-46fa-af5f-af93f77adc54\",\"order_by\":2,\"name\":\"Shantelle Macasa\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Qatar University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Shantelle\",\"middleName\":\"\",\"lastName\":\"Macasa\",\"suffix\":\"\"},{\"id\":271066204,\"identity\":\"c9c73494-24e8-42b2-baa3-e326a3180ebc\",\"order_by\":3,\"name\":\"Hamda Basha\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Qatar University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Hamda\",\"middleName\":\"\",\"lastName\":\"Basha\",\"suffix\":\"\"},{\"id\":271066205,\"identity\":\"f3674a6e-be94-4393-97f4-92ebc407be1e\",\"order_by\":4,\"name\":\"Adna Adan\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Qatar University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Adna\",\"middleName\":\"\",\"lastName\":\"Adan\",\"suffix\":\"\"},{\"id\":271066206,\"identity\":\"370bf4cf-75a3-44d1-9c45-ea6b68c14ce0\",\"order_by\":5,\"name\":\"Isra Marei\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Imperial College London\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Isra\",\"middleName\":\"\",\"lastName\":\"Marei\",\"suffix\":\"\"},{\"id\":271066207,\"identity\":\"6095f539-e4cf-4174-94a2-049dfa155a6d\",\"order_by\":6,\"name\":\"Hong Ding\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Weill Cornell Medicine - Qatar\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Hong\",\"middleName\":\"\",\"lastName\":\"Ding\",\"suffix\":\"\"},{\"id\":271066208,\"identity\":\"065dbad1-9250-46b1-815a-8cde5ae5faa8\",\"order_by\":7,\"name\":\"Christopher Triggle\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Weill Cornell Medicine - Qatar\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Christopher\",\"middleName\":\"\",\"lastName\":\"Triggle\",\"suffix\":\"\"},{\"id\":271066209,\"identity\":\"dbc6d370-1855-4bbb-81e4-f8a6cb966eb3\",\"order_by\":8,\"name\":\"Sergio Crovella\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Qatar University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Sergio\",\"middleName\":\"\",\"lastName\":\"Crovella\",\"suffix\":\"\"},{\"id\":271066210,\"identity\":\"4224a90d-6697-462a-b370-3ff4eec1fabb\",\"order_by\":9,\"name\":\"Haissam Abou-Saleh\",\"email\":\"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAz0lEQVRIiWNgGAWjYBACAxDxgOGAHJjHQ7SWBIYDxqRrSWwgWos5+/GHHxIq7qT33UhgfPC2jUG2v4GAFsueHGOJhDPPcmfeSGA2nNvGYDzjACGHHchhkEhsO5y74UYCmzRvG0NiA0Et558//pH473C6wY0E9t8gLfMJarmRYCaR2HA4AchgYwZp2UBYyxszi4Rjhw1nnnnYLDnnnITxRsIOS39840PNYXm+48kHP7wps5GdR0gLAhxgbACSEmCSWC0QihQto2AUjIJRMEIAAFV0TBoAE0Y7AAAAAElFTkSuQmCC\",\"orcid\":\"\",\"institution\":\"Qatar University\",\"correspondingAuthor\":true,\"prefix\":\"\",\"firstName\":\"Haissam\",\"middleName\":\"\",\"lastName\":\"Abou-Saleh\",\"suffix\":\"\"}],\"badges\":[],\"createdAt\":\"2024-01-24 12:05:18\",\"currentVersionCode\":1,\"declarations\":\"\",\"doi\":\"10.21203/rs.3.rs-3893992/v1\",\"doiUrl\":\"https://doi.org/10.21203/rs.3.rs-3893992/v1\",\"draftVersion\":[],\"editorialEvents\":[{\"content\":\"https://doi.org/10.1038/s41598-024-81427-6\",\"type\":\"published\",\"date\":\"2024-12-28T15:57:41+00:00\"}],\"editorialNote\":\"\",\"failedWorkflow\":false,\"files\":[{\"id\":50752336,\"identity\":\"7f6a283f-488d-42e2-9aa0-729c7a6612ec\",\"added_by\":\"auto\",\"created_at\":\"2024-02-06 17:48:07\",\"extension\":\"jpg\",\"order_by\":1,\"title\":\"Figure 1\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":37380,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eCharacterization of nanoMIL-89 and MET@nanoMIL-89 using \\u003cstrong\\u003ea)\\u003c/strong\\u003e Scanning Electron Microscope (SEM), \\u003cstrong\\u003eb\\u003c/strong\\u003e) Powder X-ray Diffraction (PXRD), and \\u003cstrong\\u003ec\\u003c/strong\\u003e) MET release from nanoMIL-89 over a 96-hour period compared to MET alone and nanoMIL-89 alone.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"1.jpg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-3893992/v1/583be14b002f628fd3779490.jpg\"},{\"id\":50752337,\"identity\":\"e9d807a9-21df-4af7-b9e6-33da7bd99571\",\"added_by\":\"auto\",\"created_at\":\"2024-02-06 17:48:07\",\"extension\":\"jpg\",\"order_by\":2,\"title\":\"Figure 2\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":69840,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eEffect of nanoMIL-89 and metformin-loaded nanoMIL-89 (MET@nanoMIL-89) on \\u003cstrong\\u003ea)\\u003c/strong\\u003e Cell viability, \\u003cstrong\\u003eb)\\u003c/strong\\u003e Cell cytotoxicity, and \\u003cstrong\\u003ec)\\u003c/strong\\u003e CXCL-8 release by HUVECs and PAECs in hyperglycemic media.\\u003cstrong\\u003e \\u003c/strong\\u003eData are presented as mean ± SEM for n=3. Statistical analysis for comparing effects between different experimental groups was conducted using two-way ANOVA (#P\\u0026lt;0.05) followed by Bonferroni Posttests. Each group was compared to the control through one-way ANOVA, followed by Dunnett’s Multiple Comparison Tests (*P\\u0026lt;0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"2.jpg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-3893992/v1/9fa3b48d2eb8526f928d8ba5.jpg\"},{\"id\":50752340,\"identity\":\"0118ceef-8ca1-4762-8337-3847ce21b6ed\",\"added_by\":\"auto\",\"created_at\":\"2024-02-06 17:48:07\",\"extension\":\"jpg\",\"order_by\":3,\"title\":\"Figure 3\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":69054,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eEffect of nanoMIL-89 and metformin-loaded nanoMIL-89 (MET@nanoMIL-89) on \\u003cstrong\\u003ea)\\u003c/strong\\u003e Cell viability, \\u003cstrong\\u003eb)\\u003c/strong\\u003e Cell cytotoxicity, and \\u003cstrong\\u003ec)\\u003c/strong\\u003e CXCL-8 release by HAECs and HAECs-Type 2 DM.\\u003cstrong\\u003e \\u003c/strong\\u003eData are presented as mean ± SEM for n=3. Statistical analysis for comparing effects between different experimental groups was conducted using two-way ANOVA (#P\\u0026lt;0.05) followed by Bonferroni Posttests. Each group was compared to the control through one-way ANOVA, followed by Dunnett’s Multiple Comparison Tests (*P\\u0026lt;0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"3.jpg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-3893992/v1/1bf5f2069ce05f5de0f65467.jpg\"},{\"id\":50752338,\"identity\":\"01982197-0c3a-426e-b91d-34215392b413\",\"added_by\":\"auto\",\"created_at\":\"2024-02-06 17:48:07\",\"extension\":\"jpg\",\"order_by\":4,\"title\":\"Figure 4\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":15272,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eEffect of nanoMIL-89 and metformin-loaded nanoMIL-89 (MET@nanoMIL-89) on DHE levels in PAECs treated with nanoMIL-89 and MET@nanoMIL-89 in hyperglycemic media. Data are presented as mean ± SEM for n=3. Statistical analysis for comparing effects between different experimental groups was conducted using two-way ANOVA (#P\\u0026lt;0.05) followed by Bonferroni Posttests. Each group was compared to the control through one-way ANOVA, followed by Dunnett’s Multiple Comparison Tests (*P\\u0026lt;0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"4.jpg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-3893992/v1/8346838102dd316f1966e154.jpg\"},{\"id\":50752787,\"identity\":\"922872e6-096b-407f-ad4e-f393d0a0b38e\",\"added_by\":\"auto\",\"created_at\":\"2024-02-06 17:56:07\",\"extension\":\"jpg\",\"order_by\":5,\"title\":\"Figure 5\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":26675,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eEffect of nanoMIL-89 and metformin-loaded nanoMIL-89 (MET@nanoMIL-89) on \\u003cstrong\\u003ea)\\u003c/strong\\u003ePhosphorylated eNOS and \\u003cstrong\\u003eb)\\u003c/strong\\u003e Total eNOS levels in PAECs treated under hyperglycemia.\\u003cstrong\\u003e \\u003c/strong\\u003eData are presented as mean ± SEM for n=3. Statistical analysis for comparing effects between different experimental groups was conducted using one-way ANOVA, followed by Dunnett’s Multiple Comparison Tests (P\\u0026lt;0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"5.jpg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-3893992/v1/caa3a8bfe6b8322154af64c0.jpg\"},{\"id\":50752339,\"identity\":\"0585823c-981f-4ce2-aa39-0ff2bc8c0469\",\"added_by\":\"auto\",\"created_at\":\"2024-02-06 17:48:07\",\"extension\":\"jpg\",\"order_by\":6,\"title\":\"Figure 6\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":13624,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eEffect of nanoMIL-89 and metformin-loaded nanoMIL-89 (MET@nanoMIL-89) on AKT levels in PAECs treated under hyperglycemia. Data are presented as mean ± SEM for n=3. Statistical analysis for comparing effects between different experimental groups was conducted using one-way ANOVA, followed by Dunnett’s Multiple Comparison Tests (P\\u0026lt;0.05).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"6.jpg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-3893992/v1/06a5f4d0081abadb67fc289e.jpg\"},{\"id\":72640843,\"identity\":\"8207cbe3-9522-488f-bb27-74c1aceade7b\",\"added_by\":\"auto\",\"created_at\":\"2024-12-30 16:10:20\",\"extension\":\"pdf\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"manuscript-pdf\",\"size\":899895,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"manuscript.pdf\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-3893992/v1/86cdbf2c-10e5-47d2-a8dd-2f91b4be2ced.pdf\"},{\"id\":50752343,\"identity\":\"3f9ce188-5058-4ad9-8394-f513030b8dac\",\"added_by\":\"auto\",\"created_at\":\"2024-02-06 17:48:08\",\"extension\":\"jpeg\",\"order_by\":1,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"supplement\",\"size\":8841079,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"GraphicalAbstract.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-3893992/v1/8f7fbf9885702b4bc3901b8c.jpeg\"}],\"financialInterests\":\"No competing interests reported.\",\"formattedTitle\":\"Managing diabetes with nanomedicine: nanoMIL-89 as a promising drug delivery system for metformin.\",\"fulltext\":[{\"header\":\"1. Introduction\",\"content\":\"\\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eDiabetes mellitus (DM) stands as one of the most prevalent chronic metabolic disorders globally, associated with decreased life expectancy and heightened mortality rates [\\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e1\\u003c/span\\u003e]. The escalating prevalence of DM is alarming, with a projected rise from approximately 424.9\\u0026nbsp;million cases worldwide in 2019 to an anticipated 700\\u0026nbsp;million cases by 2045 [\\u003cspan citationid=\\\"CR2\\\" class=\\\"CitationRef\\\"\\u003e2\\u003c/span\\u003e]. Notably, Type-2 DM constitutes approximately 90% of all DM cases globally [\\u003cspan citationid=\\\"CR3\\\" class=\\\"CitationRef\\\"\\u003e3\\u003c/span\\u003e]. Achieving glycemic control in type-2 DM involves lifestyle modifications alongside pharmacological interventions, including oral antidiabetic drugs such as biguanides (e.g., metformin)[\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e], sulfonylureas[\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e], dipeptidyl peptidase IV (DPP-4) inhibitors (gliptins)[\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e], glucagon-like peptide-1 (GLP-1) receptor agonists[\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e], sodium-glucose cotransporter (SGLT-2) inhibitors[\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e], alpha-glucosidase inhibitors[\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e], thiazolidinediones[\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e], and amylin analogs[\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e]. However, drug resistance, side effects, and individual variability responses may impede the effectiveness of these conventional therapies [\\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e1\\u003c/span\\u003e]. DM is frequently accompanied by complications such as vision, gastric, renal, and notably cardiovascular issues; with the latter being the most common even with intensive glycemic control. [\\u003cspan citationid=\\\"CR5\\\" class=\\\"CitationRef\\\"\\u003e5\\u003c/span\\u003e]. Recent studies indicate that DM-associated cardiovascular complications may arise from dysfunctional endothelium, observable even in early DM stages, making endothelial dysfunction a potential early biomarker and risk factor for cardiovascular diseases (CVDs) [\\u003cspan citationid=\\\"CR6\\\" class=\\\"CitationRef\\\"\\u003e6\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e7\\u003c/span\\u003e]. Understanding the mechanisms through which DM induces endothelial cell dysfunction is crucial for preventing the onset of CVDs, such as atherosclerosis, hypertension, and stroke [\\u003cspan citationid=\\\"CR8\\\" class=\\\"CitationRef\\\"\\u003e8\\u003c/span\\u003e].\\u003c/p\\u003e \\u003cp\\u003eDespite technological advancements in DM treatments, management remains suboptimal, marked by side effects and limitations [\\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e]. Nanomedicine applications emerge as promising solutions to address these challenges. Nanotherapies tailored for DM management offer effective blood glucose regulation and gradual release of antidiabetic drugs [\\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e1\\u003c/span\\u003e]. This extension of drug duration reduces the likelihood of acute and chronic complications associated with antidiabetic drugs [\\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e1\\u003c/span\\u003e]. The exploration of various nanoparticles (NPs) loaded with antidiabetic drugs, including liposomes, niosomes, polymers, dendrimers, micelles, and metal-based NPs, has been ongoing [\\u003cspan additionalcitationids=\\\"CR10 CR11 CR12 CR13 CR14 CR15 CR16 CR17 CR18 CR19 CR20 CR21\\\" citationid=\\\"CR9\\\" class=\\\"CitationRef\\\"\\u003e9\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR22\\\" class=\\\"CitationRef\\\"\\u003e22\\u003c/span\\u003e]. However, safety, tolerance, and stability challenges have been encountered [\\u003cspan additionalcitationids=\\\"CR10 CR11 CR12 CR13 CR14 CR15 CR16 CR17 CR18 CR19 CR20 CR21\\\" citationid=\\\"CR9\\\" class=\\\"CitationRef\\\"\\u003e9\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR22\\\" class=\\\"CitationRef\\\"\\u003e22\\u003c/span\\u003e]. To overcome these limitations, we propose the utilization of iron oxide-based NPs, known for their anti-inflammatory and hypoglycemic properties [\\u003cspan citationid=\\\"CR23\\\" class=\\\"CitationRef\\\"\\u003e23\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR24\\\" class=\\\"CitationRef\\\"\\u003e24\\u003c/span\\u003e]. They also possess antioxidant activities that can be easily detected via magnetic resonance imagining (MRI) [\\u003cspan citationid=\\\"CR25\\\" class=\\\"CitationRef\\\"\\u003e25\\u003c/span\\u003e]. Among these, nanoMIL-89, a metal-organic framework (MOF), demonstrates suitability as a drug carrier. NanoMIL-89 has exhibited selective uptake by endothelial cells, passage to daughter cells, and reduced inflammatory markers in dysfunctional endothelial cells [\\u003cspan citationid=\\\"CR26\\\" class=\\\"CitationRef\\\"\\u003e26\\u003c/span\\u003e]. Additionally, it has proven to be a suitable carrier for the drug sildenafil, used to treat pulmonary arterial hypertension (PAH) [\\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e27\\u003c/span\\u003e]. Moreover, it has been demonstrated that nanoMIL-89 exhibits non-toxic effects both \\u003cem\\u003ein vitro\\u003c/em\\u003e and \\u003cem\\u003ein vivo\\u003c/em\\u003e, as evidenced by studies utilizing zebrafish embryos and rodent models [\\u003cspan citationid=\\\"CR24\\\" class=\\\"CitationRef\\\"\\u003e24\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR26\\\" class=\\\"CitationRef\\\"\\u003e26\\u003c/span\\u003e].\\u003c/p\\u003e \\u003cp\\u003eOur selection of nanoMIL-89 as a carrier for the antidiabetic drug metformin (MET) is grounded in its cardioprotective activity and ability to carry small drugs with a molecular weight under 500 Da [\\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e27\\u003c/span\\u003e]. Moreover, MET's positive effects on blood glucose levels, insulin sensitivity, antioxidant activity, and endothelial protection align with our goal of mitigating hyperglycemia's impact on the endothelium [\\u003cspan citationid=\\\"CR28\\\" class=\\\"CitationRef\\\"\\u003e28\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR29\\\" class=\\\"CitationRef\\\"\\u003e29\\u003c/span\\u003e]. MET's association with a lower risk of CVDs in type-2 DM patients and its compatibility with nanoMIL-89 due to its size further reinforce our choice [\\u003cspan citationid=\\\"CR30\\\" class=\\\"CitationRef\\\"\\u003e30\\u003c/span\\u003e].\\u003c/p\\u003e \\u003cp\\u003eDespite MET\\u0026rsquo;s low bioavailability and short half-life, loading it into nanoMIL-89 presents an opportunity to reduce dosage, improve patient compliance, and potentially minimize gastrointestinal side effects [\\u003cspan additionalcitationids=\\\"CR32\\\" citationid=\\\"CR31\\\" class=\\\"CitationRef\\\"\\u003e31\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR33\\\" class=\\\"CitationRef\\\"\\u003e33\\u003c/span\\u003e]. Therefore, our objective is to load MET into nanoMIL-89, investigate its release kinetics, and evaluate its impact on endothelial cell function and oxidative stress under both normoglycemic and hyperglycemic conditions \\u003cem\\u003ein vitro\\u003c/em\\u003e.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e\"},{\"header\":\"2. Results\",\"content\":\"\\u003cdiv id=\\\"Sec3\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003e2.1. Pharmacokinetics of metformin-loaded nanoMIL-89 (MET@nanoMIL-89):\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eThe pharmacokinetics of MET@nanoMIL-89 were investigated and compared to MET alone. MET@nanoMIL-89 demonstrated a time-dependent release of MET, persisting for up to 96 hours with an approximate concentration of ~\\u0026thinsp;96 \\u0026micro;g/ml. Notably, the initial hours (0.5, 1, 3, 6, and 24 hrs) showed a gradual release of MET (~\\u0026thinsp;50 \\u0026micro;g/ml), reaching a peak concentration of ~\\u0026thinsp;100 \\u0026micro;g/ml after 48 hours, maintaining stability until the end of the experiment (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e). In contrast, MET alone exhibited consistent concentrations of 100\\u0026ndash;180 \\u0026micro;g/ml throughout the experiment, while the nanoMIL-89 (used as a control) showed no detectable release.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec4\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003e2.2. Effect of the metformin-Loaded nanoMIL-89 on endothelial cells proliferation, anti-inflammatory and antioxidant effect:\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eThe impact of nanoMIL-89 and MET@nanoMIL-89 on endothelial cell proliferation was assessed using different concentrations [0, 1, 3, 10, 30, and 100 \\u0026micro;g/ml]. The cell respiration rate, measured by the AlamarBlue\\u0026reg; assay, indicated that nanoMIL-89 and MET@nanoMIL-89 had no significant effect on cell viability (respiration) at concentrations\\u0026thinsp;\\u0026le;\\u0026thinsp;30 \\u0026micro;g/ml in all endothelial cells (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003ea: HUVECs, PAECs; and Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003ea: HAECs, and HAECs-Type-2 DM). The cytotoxicity of the nanoformulations used for antidiabetic drug delivery was investigated using the LDH Assay. Results from the LDH release assay (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003eb: HUVECs, PAECs; and Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003eb: HAECs, and HAECs-Type-2 DM) demonstrated that nanoMIL-89 and MET@nanoMIL-89 did not induce significant cytotoxic effects in any of the four cell lines. Furthermore, the anti-inflammatory effect of nanoMIL-89 and MET@nanoMIL-89 was examined by measuring the levels of CXCL-8 in the conditioned media. Results (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003ec: HUVECs, PAECs; and Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003ec: HAECs and HAECs-Type-2 DM) revealed that both nanoformulations exhibited a concentration-dependent decrease in CXCL-8 release from endothelial cells. This effect was more pronounced in HUVECs and HAECs, where the reduction in CXCL8 release was more significant in the MET@nanoMIL-89 group compared to the group treated with nanoMIL-89 alone.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec5\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003e2.3. Effect of metformin-loaded nanoMIL-89 on reducing reactive oxygen Species production by endothelial cells:\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eHyperglycemic conditions are known to induce endothelial dysfunction, reducing nitric oxide (NO) production, and increasing reactive oxygen species (ROS) release and oxidative stress. As MET is acknowledged for its in vitro antioxidant effects independently [\\u003cspan citationid=\\\"CR39\\\" class=\\\"CitationRef\\\"\\u003e39\\u003c/span\\u003e], this experiment delves into exploring whether MET-loaded nanoMIL-89 can sustain or amplify these antioxidant properties. Results demonstrated that hyperglycemia increased ROS levels, evident by higher DHE levels in cells seeded under hyperglycemic media for the control groups. The use of nanoMIL-89 and MET@nanoMIL-89 induced a concentration-dependent reduction in DHE levels, starting at concentrations\\u0026thinsp;\\u0026ge;\\u0026thinsp;1\\u0026micro;g/ml in the hyperglycemic environment (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig4\\\" class=\\\"InternalRef\\\"\\u003e4\\u003c/span\\u003e). This indicates that the nanoformulation possesses antioxidant activity, reducing DHE release under hyperglycemic conditions.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec6\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003e3.4. Effect of metformin-loaded nanoMIL-89 on endothelial cells eNOS and AKT levels:\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eGiven that hyperglycemic conditions induce a reduction in eNOS phosphorylation levels in endothelial cells, our results showed a decrease in eNOS levels under hyperglycemic conditions (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig5\\\" class=\\\"InternalRef\\\"\\u003e5\\u003c/span\\u003e). Furthermore, the use of MET restored the reduction in eNOS levels in the treated group compared to the untreated group (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig5\\\" class=\\\"InternalRef\\\"\\u003e5\\u003c/span\\u003eb). Additionally, both nanoMIL-89 and MET@nanoMIL-89 could induce a concentration-dependent increase in eNOS levels, with MET@nanoMIL-89 causing a more significant increase. This could be attributed to the individual increases in eNOS levels by the nanoparticle and metformin, and when combined, MET@nanoMIL-89 further enhanced the efficacy of metformin, thus promoting eNOS activity.\\u003c/p\\u003e \\u003cp\\u003eMoreover, we measured the levels of AKT, with hyperglycemic conditions generally reducing AKT levels in endothelial cells. However, when cells were incubated in hyperglycemic media, both nanoMIL-89 and MET@nanoMIL-89 induced an increase in AKT levels. Nevertheless, MET@nanoMIL-89 caused a more significant increase compared to nanoMIL-89 alone (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig6\\\" class=\\\"InternalRef\\\"\\u003e6\\u003c/span\\u003e). This correlates with the finding that both nanoMIL-89 and metformin have antioxidant activity, and when conjugated, the efficacy of the treatment increases.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e \\u003c/div\\u003e\"},{\"header\":\"3. Discussion\",\"content\":\"\\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eMetformin, renowned for its potent antidiabetic and cardioprotective properties, has been recognized to induce eNOS and Akt phosphorylation, as demonstrated in previous studies by our collaborators and others [\\u003cspan citationid=\\\"CR29\\\" class=\\\"CitationRef\\\"\\u003e29\\u003c/span\\u003e]. Additionally, it exhibits the ability to decrease DHE levels, thereby mitigating oxidative stress associated with diabetes mellitus (DM) [\\u003cspan citationid=\\\"CR39\\\" class=\\\"CitationRef\\\"\\u003e39\\u003c/span\\u003e]. Despite its efficacy, Metformin is burdened by gastrointestinal side effects and a short half-life (~\\u0026thinsp;4 hrs), limiting its therapeutic impact [\\u003cspan citationid=\\\"CR33\\\" class=\\\"CitationRef\\\"\\u003e33\\u003c/span\\u003e]. Harnessing nanomedicine to encapsulate metformin in nanoparticles (NPs) offers a promising avenue to overcome these challenges [\\u003cspan citationid=\\\"CR33\\\" class=\\\"CitationRef\\\"\\u003e33\\u003c/span\\u003e]. This study focused on exploring the potential of nanoMIL-89 as a carrier for metformin, and the observed results suggest that nanoMIL-89 indeed possesses promising therapeutic features for delivering metformin. The evaluation of metformin release from nanoMIL-89 at different time points revealed a successful and controlled release, reaching approximately 100 \\u0026micro;g/ml at 96 hours. This gradual release pattern is crucial in evading premature drug deactivation by the immune system, preventing rapid immune tolerance, and ensuring sustained therapeutic efficacy [\\u003cspan citationid=\\\"CR40\\\" class=\\\"CitationRef\\\"\\u003e40\\u003c/span\\u003e]. These findings align with a previous study on nanoMIL-89, which demonstrated its ability to circulate, and release loaded drugs in plasma for up to 4 days [\\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e27\\u003c/span\\u003e], potentially addressing metformin's short plasma half-life [\\u003cspan citationid=\\\"CR31\\\" class=\\\"CitationRef\\\"\\u003e31\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR32\\\" class=\\\"CitationRef\\\"\\u003e32\\u003c/span\\u003e].\\u003c/p\\u003e \\u003cp\\u003eThe \\u003cem\\u003ein vitro\\u003c/em\\u003e assessments were performed using endothelial cells, considering the well-documented impact of DM on endothelial cell dysfunction. Hyperglycemia-induced dysfunction is characterized by reduced eNOS phosphorylation, altered total eNOS function, decreased Akt phosphorylation [\\u003cspan citationid=\\\"CR29\\\" class=\\\"CitationRef\\\"\\u003e29\\u003c/span\\u003e], and increased oxidative stress represented by elevated DHE levels [\\u003cspan citationid=\\\"CR39\\\" class=\\\"CitationRef\\\"\\u003e39\\u003c/span\\u003e]. Results from this study, utilizing endothelial cells from various sources, revealed no detectable cytotoxic effects of MET@nanoMIL-89 and nanoMIL-89. Metabolic dysfunction, rather than toxicity from nanoformulation, was responsible for the observed decrease in cell proliferation. Moreover, loading metformin into nanoMIL-89 effectively countered hyperglycemia-induced oxidative stress, reducing DHE levels. Additionally, it elevated phosphorylated eNOS, total eNOS, and AKT levels in endothelial cells under hyperglycemic conditions, mimicking the diabetic environment.\\u003c/p\\u003e \\u003cp\\u003eThese findings imply that nanoMIL-89 may possess cardioprotective activity by mitigating endothelial cell dysfunction, a property often associated with metallic NPs like iron oxide NPs [\\u003cspan citationid=\\\"CR3\\\" class=\\\"CitationRef\\\"\\u003e3\\u003c/span\\u003e].\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e\"},{\"header\":\"4. Methodology\",\"content\":\"\\u003cdiv id=\\\"Sec9\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003e4.1. NPs synthesis, characterizations, and metformin loading:\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eNanoMIL-89 was synthesized according to previously published procedures [\\u003cspan citationid=\\\"CR24\\\" class=\\\"CitationRef\\\"\\u003e24\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR34\\\" class=\\\"CitationRef\\\"\\u003e34\\u003c/span\\u003e]. In brief, 10 mmol of iron (III) chloride hexahydrate (FeCl\\u003csub\\u003e3\\u003c/sub\\u003e.6H\\u003csub\\u003e2\\u003c/sub\\u003eO) and 10 mmol of trans-trans muconic acid were dissolved in 100 ml of absolute ethanol (99.8%). After subjecting the reaction mixture to sonication for 15 minutes, the subsequent step involved adding 20 ml of glacial acetic acid (99.8%) before heating it at 90\\u0026deg;C for 24 hours within a Parr reactor. Precipitated nanoparticles were recovered by centrifugation at 7000 rpm for 15 minutes, followed by several washes in distilled water. The precipitate was then vacuum-dried to obtain nanoMIL-89, which presented as a brown precipitate (100\\u0026ndash;200 mg/reaction).\\u003c/p\\u003e \\u003cp\\u003eSubsequently, nanoMIL-89 was loaded with metformin as follows: Solutions of 1000, 300, 100, 30, and 10 \\u0026micro;g/ml metformin in Phosphate Buffer Saline (PBS; pH 7.4) were prepared. To each 5 ml of metformin concentration, 200 mg of nanoMIL-89 was added. The mixture was incubated overnight (16\\u0026ndash;18 hrs.) at room temperature, and metformin-loaded nanoMIL-89 was harvested by centrifugation at 7000 rpm for 15 minutes at room temperature. The harvested metformin-loaded nanoMIL-89 was characterized by Infrared Imaging (IR), Powder X-ray Diffraction (PXRD), and Scanning Electron Microscope (SEM), with results compared to the unloaded nanoMIL-89. Finally, the metformin nanoMIL-89 prototype was utilized for pharmacokinetic and \\u003cem\\u003ein vitro\\u003c/em\\u003e experiments.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec10\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003e4.2. Metformin release pharmacokinetics:\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eAs described above, nanoMIL-89 was loaded with MET, and the release kinetics were measured over 96 hours. NanoMIL-89 loaded with MET was precipitated at 7000 rpm at room temperature for 15 minutes. The resulting precipitate was dissolved in 5 ml of PBS and incubated at room temperature. At various time points (0, 0.5, 1, 3, 6, 24, 48, 72, and 96 hours), 1.5 ml of the supernatant was collected. The amount of released MET into PBS was determined using a previously described colorimetric method [\\u003cspan citationid=\\\"CR35\\\" class=\\\"CitationRef\\\"\\u003e35\\u003c/span\\u003e], which relies on the interaction of MET with ninhydrin reagent at pH 5.6, resulting in a red color. A MET standard curve was incorporated. Each collected 1.5 ml aliquot was mixed with 10.0 ml of Phthalate buffer (pH\\u0026thinsp;=\\u0026thinsp;5.6) and 5.0 ml of 0.1% Ninhydrin. The mixture underwent heating in a water bath for 1 hour at 90\\u0026ordm;C. Subsequently, the solution was cooled to room temperature and topped up with distilled water to a final volume of 25 ml, and the optical density was measured at 567 nm. Readouts were corrected for a blank, and concentrations were estimated using the MET standard curve [\\u003cspan citationid=\\\"CR35\\\" class=\\\"CitationRef\\\"\\u003e35\\u003c/span\\u003e].\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec11\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003e4.3. Effect of metformin-loaded nanoMIL-89 on endothelial cell proliferation, anti-inflammatory, and antioxidant effects:\\u003c/h2\\u003e \\u003cdiv id=\\\"Sec12\\\" class=\\\"Section3\\\"\\u003e \\u003ch2\\u003e4.3.1. Cell line culture:\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eStandard culture techniques were employed for seeding and maintaining the following endothelial cell lines: (i) Human Pulmonary Artery Endothelial Cells (PAECs) [\\u003cspan citationid=\\\"CR36\\\" class=\\\"CitationRef\\\"\\u003e36\\u003c/span\\u003e] and (ii) Human Umbilical Vein Endothelial Cells (HUVECs) [\\u003cspan citationid=\\\"CR37\\\" class=\\\"CitationRef\\\"\\u003e37\\u003c/span\\u003e], both cultured according to the suppliers' recommendations. The responses of healthy cells cultured under hyperglycemic conditions were compared to those of a diabetic endothelial cell line: Human Primary Diabetic Aortic Endothelial Cells (HDAOEC; Cellbiologics), cultured according to the manufacturer's instructions [\\u003cspan citationid=\\\"CR38\\\" class=\\\"CitationRef\\\"\\u003e38\\u003c/span\\u003e].\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec13\\\" class=\\\"Section3\\\"\\u003e \\u003ch2\\u003e4.3.2. Cell viability, cytotoxicity, and cytokine release:\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eEndothelial cells were plated in 96-well plates at a seeding density of 10,000 cells/well under hyperglycemic conditions (25 mM glucose). After overnight incubation (16\\u0026ndash;18 hrs) in humidified conditions (37 \\u0026ordm;C, 5% CO\\u003csub\\u003e2\\u003c/sub\\u003e), cells were treated with either nanoMIL-89 alone (0, 1, 3, 10, 30, and 100 \\u0026micro;g/ml), metformin alone (50 \\u0026micro;M), or metformin-loaded nanoMIL-89 (MET@nanoMIL-89; 0, 1, 3, 10, 30, and 100 \\u0026micro;g/ml) for 24 hrs under the same conditions. Adherent cells were used to determine the effects of different treatments on cell viability using the AlamarBlue\\u0026reg; assay following the manufacturer\\u0026rsquo;s instructions. Conditioned media were collected to assess cell cytotoxicity using the LDH Kit (Abcam, UK) and anti-inflammatory effects using CXCL8-ELISA Kits (R\\u0026amp;D systems, USA). AlamarBlue, LDH, and ELISA assays were performed following manufacturer instructions.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003cdiv id=\\\"Sec14\\\" class=\\\"Section3\\\"\\u003e \\u003ch2\\u003e4.3.3. Effect of metformin-loaded nanoMIL-89 on reactive oxygen species, eNOS, and AKT levels:\\u003c/h2\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eEndothelial cells were seeded into a black 96-well plate and treated as described above. After 24 hrs in humidified conditions (37 \\u0026ordm;C, 5% CO\\u003csub\\u003e2\\u003c/sub\\u003e), the Dihydroethidium (DHE, Abcam, UK) assay kit was used to measure Reactive Oxygen Species (ROS) release according to the manufacturer's instructions. For Endothelial Nitric Oxide Synthase (eNOS) and Phospho-AKT, cells were seeded into 6-well plates at a density of 100,000 cells/well and treated as described above for 24 hours. Following treatment, proteins were extracted, and the levels of phosphorylated eNOS and total eNOS were measured using the Human Phospho-eNOS and Total eNOS ELISA kit (RayBio\\u0026reg;, USA) as instructed. Additionally, the expression of total and phosphorylated AKT was measured using the Phospho-AKT (RayBio\\u0026reg;, USA) ELISA kit following the manufacturer's instructions.\\u003c/p\\u003e \\u003cp\\u003e \\u003cb\\u003eStatistical analysis\\u003c/b\\u003e:\\u003c/p\\u003e \\u003cp\\u003eResults from drug release studies and \\u003cem\\u003ein vitro\\u003c/em\\u003e culture studies were analyzed and presented as mean\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;SEM for n experiments with well-defined figure legends. Statistical tests were performed using GraphPad Prism v5. Two-way ANOVA followed by Bonferroni Posttests were used for comparing effects between different groups (i.e., metformin-loaded nanoMIL-89, metformin alone, and unloaded nanoMIL-89). One-way ANOVA followed by Dunnett\\u0026rsquo;s Multiple Comparison Tests was used for comparing the effect of different nanoformulation concentrations within each group compared to the control. Statistical significance was noted at p\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.05.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e \\u003c/div\\u003e\"},{\"header\":\"5. Conclusion\",\"content\":\"\\u003cp\\u003e \\u003cdiv class=\\\"BlockQuote\\\"\\u003e \\u003cp\\u003eThis study highlights nanoMIL-89 as a biocompatible nanoparticle endowed with cardioprotective properties, as evidenced by its antioxidant and anti-inflammatory effects, along with its capacity to enhance the nitric oxide pathway. The observed potential suggests that this nanoparticle could have versatile applications in diverse diseases, such as cardiovascular diseases (CVDs) and diabetes mellitus (DM). The experimental findings underscore nanoMIL-89 as a promising carrier for delivering antidiabetic drugs, specifically metformin. However, to gain a comprehensive understanding of its mechanism of action, pharmacodynamics, and long-term biological effects, further in vivo studies utilizing diabetic models are imperative. The success of such studies could propel this nanoparticle into pre-clinical stages, unveiling promising therapeutic opportunities not only for diabetes but also for other challenging diseases, including cancer. The ongoing exploration of nanoMIL-89's capabilities holds great potential for advancing our understanding of nanomedicine applications and their broader impact on medical treatments.\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/p\\u003e\"},{\"header\":\"Declarations\",\"content\":\"\\u003cp\\u003e\\u003cstrong\\u003eAcknowledgement:\\u0026nbsp;\\u003c/strong\\u003eWe express our appreciation for the Open Access funding provided by the Qatar National Library. Furthermore, we convey our profound gratitude to the Qatar National Research Fund for offering financial support for this research through grant # UREP26-019-3-006 awarded to H.A.S. We also acknowledge the British Pharmacological Society, United Kingdom, for the generous assistance provided through the Pickford Award, granted to N.A.M. This support played a crucial role in facilitating the development of the nanoparticles employed in this study. The graphical abstract utilized in this research was generated using BioRender.com.\\u003c/p\\u003e\\n\\u003cp\\u003e\\u003cstrong\\u003eAuthors Contributions:\\u0026nbsp;\\u003c/strong\\u003eConceptualization: [N.A.M., I.M., H.A.S.], \\u0026nbsp;Methodology: [N.A.M., I.M., H.A.S.], Formal analysis and investigation: [N.A.M., I.M., H.A.S., H.A.M., S.S.M., H.K.B., A.M.A.]; Writing - original draft preparation: [H.A.M, N.A.M.]; Writing - review and editing: [H.A.S., S.C., N.A.M., I.M., C.R.T., H.D.]; Funding acquisition: [H.A.S., N.A.M., I.M.]; Resources: [N.A.M., I.M., H.A.S.]; Supervision: [N.A.M., H.A.S.]. All authors have read and agreed to the published version of the manuscript.\\u003c/p\\u003e\\n\\u003cp\\u003e\\u003cstrong\\u003eConflict of interest:\\u0026nbsp;\\u003c/strong\\u003eThe authors declare that they have no conflict of interest regarding the publication of this article.\\u0026nbsp;\\u003c/p\\u003e\\n\\u003cp\\u003e\\u003cstrong\\u003eFunding:\\u0026nbsp;\\u003c/strong\\u003eThis research was supported by Qatar National Research Fund under the Undergraduate Research experience Program Grant no. [UREP26-019-3-006] (to H.A.S.), and the Pickford Award from the British Pharmacological Society (to N.A.M). \\u0026nbsp;\\u003c/p\\u003e\\n\\u003cp\\u003e\\u003cstrong\\u003eEthical Approval:\\u0026nbsp;\\u003c/strong\\u003eThis article does not contain any studies with human participants or animals performed by any of the authors.\\u0026nbsp;\\u003c/p\\u003e\"},{\"header\":\"References\",\"content\":\"\\u003col\\u003e\\n\\u003cli\\u003eKesharwani, P.; Gorain, B.; Low, S.Y.; Tan, S.A.; Ling, E.C.S.; Lim, Y.K.; Chin, C.M.; Lee, P.Y.; Lee, C.M.; Ooi, C.H.; et al. Nanotechnology based approaches for antidiabetic drugs delivery. \\u003cem\\u003eDiabetes Res Clin Pract \\u003c/em\\u003e\\u003cstrong\\u003e2018\\u003c/strong\\u003e, \\u003cem\\u003e136\\u003c/em\\u003e, 52-77, doi:10.1016/j.diabres.2017.11.018.\\u003c/li\\u003e\\n\\u003cli\\u003eSaeedi P, P.I., Salpea P, Malanda B, Karuranga S, Unwin N, Colagiuri S, Guariguata L, Motala AA, Ogurtsova K, Shaw JE, Bright D, Williams R. 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Metformin decreases intracellular production of reactive oxygen species in aortic endothelial cells. \\u003cem\\u003eMetabolism \\u003c/em\\u003e\\u003cstrong\\u003e2005\\u003c/strong\\u003e, \\u003cem\\u003e54\\u003c/em\\u003e, 829-834, doi:10.1016/j.metabol.2005.01.029.\\u003c/li\\u003e\\n\\u003cli\\u003eLi, H., Yang, Y. G., \\u0026amp; Sun, T. (2022). Nanoparticle-Based Drug Delivery Systems for Induction of Tolerance and Treatment of Autoimmune Diseases. \\u003cem\\u003eFrontiers in bioengineering and biotechnology\\u003c/em\\u003e, \\u003cem\\u003e10\\u003c/em\\u003e, 889291. https://doi.org/10.3389/fbioe.2022.889291\\u003c/li\\u003e\\n\\u003c/ol\\u003e\"}],\"fulltextSource\":\"\",\"fullText\":\"\",\"funders\":[],\"hasAdminPriorityOnWorkflow\":false,\"hasManuscriptDocX\":true,\"hasOptedInToPreprint\":true,\"hasPassedJournalQc\":\"\",\"hasAnyPriority\":false,\"hideJournal\":false,\"highlight\":\"\",\"institution\":\"\",\"isAcceptedByJournal\":true,\"isAuthorSuppliedPdf\":false,\"isDeskRejected\":\"\",\"isHiddenFromSearch\":false,\"isInQc\":false,\"isInWorkflow\":false,\"isPdf\":false,\"isPdfUpToDate\":true,\"isWithdrawnOrRetracted\":false,\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"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\":\"\",\"lastPublishedDoi\":\"10.21203/rs.3.rs-3893992/v1\",\"lastPublishedDoiUrl\":\"https://doi.org/10.21203/rs.3.rs-3893992/v1\",\"license\":{\"name\":\"CC BY 4.0\",\"url\":\"https://creativecommons.org/licenses/by/4.0/\"},\"manuscriptAbstract\":\"\\u003cp\\u003eDiabetes Mellitus is a chronic disease characterized by metabolic defects, including insulin deficiency and resistance. Individuals with diabetes are at increased risk of developing cardiovascular complications, such as atherosclerosis, coronary artery disease, and hypertension. Conventional treatment methods, though effective, are often challenging, costly, and may lead to systemic side effects. This study explores the potential of nanomedicine applications, specifically Metal-Organic Frameworks (MOFs), as drug carriers to overcome these limitations. The Materials Institute Lavoisier-89 nanoparticles (nanoMIL-89) have previously demonstrated promise as a drug delivery vehicle for chronic diseases due to their anti-oxidant and cardio-protective properties. In this investigation, nanoMIL-89 was loaded with the anti-diabetic drug metformin (MET), creating MET@nanoMIL-89 formulation.\\u003c/p\\u003e\\n\\u003cp\\u003eWe examined the drug release kinetics of MET@nanoMIL-89 over 96 hours and assessed its impact on the viability of various endothelial cells. Furthermore, we investigated the nanoformulation effect on inflammatory markers in these cells and explored its influence on phosphorylated eNOS, total eNOS, and AKT levels. Our findings indicate that nanoMIL-89 effectively released metformin over 96 hours and caused a concentration-dependent reduction in CXCL-8 release from endothelial cells. Notably, MET@nanoMIL-89 reduced dihydroethidium levels and increased phosphorylated eNOS, total eNOS, and AKT levels.\\u003c/p\\u003e\\n\\u003cp\\u003eOur results underscore the potential of nanoMIL-89 as a versatile potential drug delivery platform for anti-diabetic drugs, offering a prospective therapeutic approach for diabetic patients with associated cardiovascular complications.\\u003c/p\\u003e\",\"manuscriptTitle\":\"Managing diabetes with nanomedicine: nanoMIL-89 as a promising drug delivery system for metformin.\",\"msid\":\"\",\"msnumber\":\"\",\"nonDraftVersions\":[{\"code\":1,\"date\":\"2024-02-06 17:48:02\",\"doi\":\"10.21203/rs.3.rs-3893992/v1\",\"editorialEvents\":[{\"type\":\"communityComments\",\"content\":0},{\"type\":\"decision\",\"content\":\"Revision requested\",\"date\":\"2024-04-26T12:13:53+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"editorInvitedReview\",\"content\":\"\",\"date\":\"2024-04-25T05:06:01+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"b2a74a30-773c-43bd-9b79-90717c6acdd4\",\"date\":\"2024-03-24T16:42:38+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"editorInvitedReview\",\"content\":\"\",\"date\":\"2024-02-23T07:29:06+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"2504902b-5457-4e28-9e4f-ee3084efcd6c\",\"date\":\"2024-02-07T16:44:27+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"11f9052d-9f76-4391-a7f2-07ff00963027\",\"date\":\"2024-02-07T10:21:05+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewersInvited\",\"content\":\"\",\"date\":\"2024-02-05T07:39:09+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"editorAssigned\",\"content\":\"\",\"date\":\"2024-02-04T17:21:18+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"editorInvited\",\"content\":\"\",\"date\":\"2024-02-04T17:07:58+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"checksComplete\",\"content\":\"\",\"date\":\"2024-02-04T12:34:03+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"submitted\",\"content\":\"Scientific Reports\",\"date\":\"2024-01-24T12:03:05+00:00\",\"index\":\"\",\"fulltext\":\"\"}],\"status\":\"published\",\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"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\":\"efc50885-dc28-4a98-a879-a6f133eb3179\",\"owner\":[],\"postedDate\":\"February 6th, 2024\",\"published\":true,\"recentEditorialEvents\":[],\"rejectedJournal\":[],\"revision\":\"\",\"amendment\":\"\",\"status\":\"published-in-journal\",\"subjectAreas\":[{\"id\":28565251,\"name\":\"Biological sciences/Biotechnology\"},{\"id\":28565252,\"name\":\"Health sciences/Diseases\"}],\"tags\":[],\"updatedAt\":\"2024-12-30T16:05:24+00:00\",\"versionOfRecord\":{\"articleIdentity\":\"rs-3893992\",\"link\":\"https://doi.org/10.1038/s41598-024-81427-6\",\"journal\":{\"identity\":\"scientific-reports\",\"isVorOnly\":false,\"title\":\"Scientific Reports\"},\"publishedOn\":\"2024-12-28 15:57:41\",\"publishedOnDateReadable\":\"December 28th, 2024\"},\"versionCreatedAt\":\"2024-02-06 17:48:02\",\"video\":\"\",\"vorDoi\":\"10.1038/s41598-024-81427-6\",\"vorDoiUrl\":\"https://doi.org/10.1038/s41598-024-81427-6\",\"workflowStages\":[]},\"version\":\"v1\",\"identity\":\"rs-3893992\",\"journalConfig\":\"researchsquare\"},\"__N_SSP\":true},\"page\":\"/article/[identity]/[[...version]]\",\"query\":{\"redirect\":\"/article/rs-3893992\",\"identity\":\"rs-3893992\",\"version\":[\"v1\"]},\"buildId\":\"qtupq5eGEP_6zYnWcrvyt\",\"isFallback\":false,\"isExperimentalCompile\":false,\"dynamicIds\":[84888],\"gssp\":true,\"scriptLoader\":[]}","source_license":"CC-BY-4.0","license_restricted":false}