#1656 Ezetimibe improves diabetic nephropathy by upregulating Nrf2 to alleviate endoplasmic reticulum stress

Abstract Background and Aims Diabetic nephropathy (DN), a prevalent complication of diabetes mellitus, stands as the primary contributor to chronic kidney disease and end-stage renal disease. The progression of diabetic nephropathy is facilitated by endoplasmic reticulum stress and the excessive generation of reactive oxygen species (ROS). In addition to its cholesterol-lowering properties, recent research has revealed that ezetimibe also ameliorates insulin resistance and glucose intolerance abnormalities, and is also beneficial in chronic kidney disease. However, the specific impact of ezetimibe on diabetic nephropathy and the underlying mechanism remain unknown. Therefore, the objective of this research is to investigate the effect of ezetimibe on diabetic nephropathy and elucidate its mechanism. Method In vivo experiments were conducted using C57BL/6J mice, with a total of 24 mice randomly assigned to three groups: NC, DM, and EZE (10 mg/kg/day). In vitro experiments involved the treatment of human renal proximal tubular (HK-2) cells with ezetimibe (50 μM) and high glucose (30 mmol/L) for 48 hours. Then, combined with histopathological examination, biochemical evaluation, TUNEL stain, western blot analysis, Si-RNA transfection, transmission electron microscopy and ROS probe were used to detect the renal morphology and function, oxidative stress and endoplasmic reticulum homeostasis level. Results The results demonstrated that a 10-week treatment with ezetimibe led to significant improvements in urine volume, beta-N-acetylglucosaminidase (NAG), and the urinary albumin/creatinine ratio (ACR). The administration of ezetimibe in the treatment exhibited enhancements in glomerular hypertrophy and renal tubular vacuolization, as well as mesangial matrix expansion and glomerular basement membrane thickness, in diabetic mice. Additionally, ezetimibe treatment resulted in a reduction in ROS formation by upregulating the expression of antioxidant enzymes, such as superoxide dismutase (SOD) and catalase (CAT), through the activation of NF-E2-related factor-2 (Nrf2). Furthermore, ezetimibe administration partially reversed the increased expression of endoplasmic reticulum stress-related proteins, including activating transcription factor 6 (ATF6), glucose-regulated protein 78 (GRP78), and C/EBP homologous protein (CHOP), in DN mice. Ezetimibe demonstrated inhibitory effects on the accumulation of ROS and alleviated endoplasmic reticulum stress in HK-2 cells treated with high glucose. Furthermore, electron microscopy analysis revealed that ezetimibe effectively reduced the distension and dilation of the endoplasmic reticulum. However, the anti-endoplasmic reticulum stress effect of ezetimibe was significantly diminished following transfection with si-Nrf2. These findings suggest that ezetimibe exerts its anti-endoplasmic reticulum stress effect, at least in part, by enhancing antioxidant capacity and reducing ROS levels through the upregulation of Nrf2. Conclusion In summary, our research findings demonstrate the notable contribution of endoplasmic reticulum stress to the progression of DN. Through the up-regulation of Nrf2, ezetimibe effectively mitigates ROS generation in the context of DN, consequently ameliorating the adverse consequences of ERS, restoring equilibrium within the endoplasmic reticulum, and ultimately safeguarding against renal cell demise. Ezetimibe is an attractive potential therapy for the treatment of DN.