SIRT3 involves high glucose-induced endothelial impairment in diabetic retinopathy through OPA1-mediated mitochondrial dynamics imbalance

Abstract Background The oxidative stress of endothelial cells plays a key role in diabetic retinopathy (DR), and mitochondrial homeostasis is crucial for oxidative stress. Aim To explore the potential roles of mitochondrial sirtuins (SIRTs) in oxidative stress-related endothelial impairment in DR and of mitochondrial homeostasis damage. Methods The activity of retinal microvascular endothelial cells was detected via the cell counting kit-8 experiment. The dead or alive cells were assessed by calcein-AM/PI staining. The reactive oxygen species (ROS) in cells were labeled using the DCFH-DA fluorescence probe method. The expression of mitochondrial SIRTs (SIRT3–5) and key molecules of mitochondrial homeostasis regulation pathway was detected using qPCR and further verified by western blot. Mitochondria morphology was labeled with an electron microscope and MitoTracker fluorescence probe, and quantitative analysis was performed with Image J software. Results Cell viability significantly decreased after 48 h of high glucose (HG), and HG led to a significant increase in the proportion of dead cells and the ROS level, accompanied by a decrease in SIRT3 and the level of mitochondrial dynamics-related molecules. SIRT3 overexpression reversed the proportion of dead cells and ROS levels and had a significant effect on the expression of mitochondrial dynamics-related molecules, especially the mitochondrial fusion-related molecule OPA1. Mitochondrial morphological changes increased mitochondrial fusion after SIRT3 overexpression under HG. However, MYLS22, the OPA1 inhibitor, blocked the protective effect of SIRT3 overexpression, which was manifested by increasing the number of dead cells, the ROS level, and the intensification of mitochondrial fragmentation. Conclusion This study found that SIRT3 involved an imbalance of mitochondrial dynamics caused by HG-induced endothelial impairment in DR, possibly through the OPA1 pathway.