High-sugar high-fat treatment induces autophagy of retinal microvascular endothelial cells

• Published in: Biochemical and biophysical research communications 2022-04, Vol.600, p.22-28
• Main Authors: Mao, Xinbang, Wan, Yuwen, Huang, Sidan, Wang, Yan, Wu, Yunfei, Zhou, Shenghong, Feng, Xia, Gao, Caixia, Wu, Chen
• Format: Article
• Language: English
• Subjects: Animals; Autophagy; Cathepsin B - metabolism; Cathepsin D - metabolism; Chloroquine; Chloroquine - pharmacology; Endothelial Cells - metabolism; Glucose - metabolism; Lipoproteins, LDL - metabolism; Lipoproteins, LDL - pharmacology; MITO-Tempol; Oxidative Stress; Rat retinal microvascular endothelial cells; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species - metabolism; Resveratrol; Sugars - metabolism
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2022.02.032

To investigate the role of high-sugar high-fat treatment in inducing autophagy of rat retinal microvascular endothelial cells. The optimal concentrations and time points of glucose and oxidized low-density lipoprotein (ox-LDL) in inducing rat retinal microvascular endothelial cells were determined by examining the proliferate rate by CCK-8 assay. They were divided into control group (blank control), model group (treatment of 50 mM glucose and 10 μg/ml ox-LDL for 24 h), chloroquine group (treatment of 20 μM chloroquine, 50 mM glucose and 10 μg/ml ox-LDL for 24 h), resveratrol group (treatment of 50 μM resveratrol, 50 mM glucose and 10 μg/ml ox-LDL for 24 h) and MITO-Tempol group (treatment of 20 μM MITO-Tempol, 50 mM glucose and 10 μg/ml ox-LDL for 24 h). Reactive oxygen species (ROS) level in rat retinal microvascular endothelial cells induced with high sugar high-fat treatment was measured by flow cytometry. In addition, protein levels of cathepsin B and cathepsin D in rat retinal microvascular endothelial cells induced with high sugar high-fat treatment were examined by immunofluorescence, and protein levels of LC3 A/B and the autophagy substrate P62 were detected by Western blot. Primary retinal microvascular endothelial cells were isolated from neonatal Sprague-Dawley (SD) rats. ROS level was significantly higher in model group than that of control group (P