miR-16-5p Regulates Proliferation and Apoptosis in High Glucose-Treated Human Retinal Microvascular Endothelial Cells by Targeting VEGFA and TGFBR1.

Abstract

Diabetic retinopathy (DR) is a common complication of diabetes and the main cause of vision loss in the middle-aged and elderly people. miRNAs play vital roles in the development of DR. This study aimed to explore the effects of miR-16-5p on high glucose (HG)-stimulated human retinal microvascular endothelial cells (HRECs) by modulating vascular endothelial growth factor A (VEGFA) and transforming growth factor beta receptor 1 (TGFBR1). HRECs were treated with 5 mM, 10 mM, 20 mM, and 30 mM of HG to induce the DR cell model. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression of miR-16-5p and mRNAs of VEGFA and TGFBR1. Western blot was used to examine VEGFA and TGFBR1 protein levels. The 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide assay was conducted to test cell proliferation. Flow cytometry with Annexin V-FITC/PI double staining was carried out to assess cell apoptosis ratio. Dual-luciferase assay was used to identify the target relationship between miR-16-5p and VEGFA and TGFBR1. Results found that the expression of miR-16-5p in HG-treated HRECs was reduced, and VEGFA and TGFBR1 expressions were upregulated. Knockdown of miR-16-5p increased VEGFA and TGFBR1 mRNA and protein levels, promoted cell proliferation, and inhibited apoptosis in HG-treated HRECs. VEGFA and TGFBR1 inhibition reversed the effect of knocking down miR-16-5p on HRECs. Dual-luciferase reporter assay revealed that VEGFA and TGFBR1 were the target of miR-16-5p. Overall, knockdown of miR-16-5p enhances proliferation and inhibits apoptosis of HRECs by upregulating VEGFA and TGFBR1 expression.