Abstract 4368701: Microvascular Endothelial Heterogeneity in Canonical Tissues of Human Insulin Resistance Identifies Depot Specific ER Stress as a Novel Therapeutic Target

Introduction: Endothelial cell (EC) heterogeneity across vascular beds and disease states is increasingly recognized. However, tissue-specific EC heterogeneity in individuals with type 2 diabetes mellitus (T2DM) is poorly characterized. Hypothesis: We hypothesized that ECs from different tissue beds exhibit distinct transcriptional responses to T2DM. Specifically, we investigated whether subcutaneous adipose tissue ECs (ATECs) and skeletal muscle ECs (SkMECs) from individuals with T2DM show differential activation of endoplasmic reticulum (ER) stress. Materials and Methods: We prospectively recruited patients undergoing cardiac implantable electronic device implantation at Leeds Teaching Hospitals and obtained paired biopsies of pectoralis major and adipose tissue. ECs were isolated using CD31 coated beads and purity confirmed using flow cytometry. Bulk RNA sequencing (RNAseq) of cultured EC isolates was performed. Differentially expressed genes (DEGs) were identified between people with T2DM versus no T2DM (ND) using DESeq2. Gene ontology (GO) enrichment analysis was used to identify differential pathways between tissue beds. The presence of ER stress markers was validated by immunoblotting and correlated with clinical data. Results: We recruited 16 participants (mean age 72 years, 56% male, mean BMI 28). Among individuals with T2DM, mean HbA1c was 57 mmol/mol. RNAseq revealed 63 DEG in SkMEC and 1085 in ATEC in people with T2DM vs ND, with key stress-response genes (e.g. ATF6B, EIF2AK2, ERP29) differentially expressed (Panel A). GO analysis identified response to ER stress as an enriched biological process in ATEC but not SkMECs in people with T2DM. Immunoblotting confirmed increased protein expression of ER stress markers IRE1α and CHOP in ATECs but not in SkMECs (Panel C). Serum HbA1c showed a strong positive correlation with IRE1α (r = 0.78, p = 0.02) and CHOP (r = 0.75, p = 0.02) in ATECs, but not in SkMECs (Panel D). Regression analysis further identified HbA1c as an independent predictor of ER stress in ATECs, after adjustment for age, sex, and BMI (Panel E). Conclusion: Our study reveals a striking tissue-specific heterogeneity in EC response to T2DM, with selective activation of ER stress via the IRE1α pathway in ATECs. These findings suggest that ATECs are uniquely susceptible to hyperglycemia-induced stress and highlight the unfolded protein response as a potential therapeutic target in adipose-specific microvascular dysfunction in T2DM.