Rong Luo1, Fan Xiao2, Pei Wang2, Yu-Xiang Hu2. 1. Department of Ophthalmology, Jiangxi Provincial People's Hospital Affiliated to Nanchang University, No. 92, Aiguo Road, Donghu District, Nanchang, 330006, Jiangx, People's Republic of China. luorongs57@163.com. 2. Department of Ophthalmology, Jiangxi Provincial People's Hospital Affiliated to Nanchang University, No. 92, Aiguo Road, Donghu District, Nanchang, 330006, Jiangx, People's Republic of China.
Abstract
OBJECTIVE: Diabetic retinopathy (DR) is a major complication of both type 1 and type 2 diabetes. Recently, inflammation was found to play an important role in DR pathogenesis. But the mechanism has not been fully understood. METHODS: ARPE-19 cells were cultured under normal condition and high-glucose condition, then the expressions of miR-93, XBP1s and lncRNA H19 were measured using RT-qPCR or western blots. Besides, the mRNA level of eIF2α and GRP78 and protein level of p-eIF2α and GRP78 were measured by RT-qPCR or western blots. In addition, RT-qPCR and ELISA were adopted to examine the expression and secretion of cytokine factors in these conditions. Dual-luciferase reporter gene assay was used to elucidate the binding and regulation among XBP1s, miR-93 and H19. RNA immunoprecipitation was also performed to verify the interaction between H19 and miR-93. The expressions of DNAJC3 and DNAJB9, the downstream targets of XBP1s, were detected by RT-qPCR. RESULTS: We identified that H19 and XBP1s were down-regulated in ARPE-19 cells under high-glucose condition, while miR-93 was up-regulated. ER stress inducers TM and IRE1 inhibitor STF-083010 were adopted and data suggest that ER stress could be induced during high-glucose treatment. In addition, the altered expressions of miR-93, XBP1s and H19 might mediate the increased level of pro-inflammatory cytokines. Furthermore, miR-93 interacted with either lncRNA H19 or XBP1s then modulating the inflammatory processes. CONCLUSIONS: H19 played an important role in regulating inflammatory processes in retinal endothelial cells under high-glucose condition through modulating miR-93/XBP1s axis.
OBJECTIVE:Diabetic retinopathy (DR) is a major complication of both type 1 and type 2 diabetes. Recently, inflammation was found to play an important role in DR pathogenesis. But the mechanism has not been fully understood. METHODS:ARPE-19 cells were cultured under normal condition and high-glucose condition, then the expressions of miR-93, XBP1s and lncRNA H19 were measured using RT-qPCR or western blots. Besides, the mRNA level of eIF2α and GRP78 and protein level of p-eIF2α and GRP78 were measured by RT-qPCR or western blots. In addition, RT-qPCR and ELISA were adopted to examine the expression and secretion of cytokine factors in these conditions. Dual-luciferase reporter gene assay was used to elucidate the binding and regulation among XBP1s, miR-93 and H19. RNA immunoprecipitation was also performed to verify the interaction between H19 and miR-93. The expressions of DNAJC3 and DNAJB9, the downstream targets of XBP1s, were detected by RT-qPCR. RESULTS: We identified that H19 and XBP1s were down-regulated in ARPE-19 cells under high-glucose condition, while miR-93 was up-regulated. ER stress inducers TM and IRE1 inhibitor STF-083010 were adopted and data suggest that ER stress could be induced during high-glucose treatment. In addition, the altered expressions of miR-93, XBP1s and H19 might mediate the increased level of pro-inflammatory cytokines. Furthermore, miR-93 interacted with either lncRNA H19 or XBP1s then modulating the inflammatory processes. CONCLUSIONS:H19 played an important role in regulating inflammatory processes in retinal endothelial cells under high-glucose condition through modulating miR-93/XBP1s axis.
Entities:
Keywords:
Diabetic retinopathy; H19; Inflammation; Long non-coding RNA; MicroRNA; miR-93
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