芝麻素通过lncRNA WEE2-AS1/miR-515-5p通路影响高糖诱导的血管内皮细胞损伤的机制研究

孙珂; 周慧超; 曹云玲; 井庆平

doi:10.13898/j.cnki.issn.1000-2200.2023.02.003

芝麻素通过lncRNA WEE2-AS1/miR-515-5p通路影响高糖诱导的血管内皮细胞损伤的机制研究

Effect of sesamin on high glucose-induced vascular endothelial cell injury through lncRNA WEE2-AS1/miR-515-5p pathway

摘要

摘要:
目的探讨芝麻素对高糖诱导的血管内皮细胞（HUVEC）损伤的影响及其可能作用机制。
方法高糖诱导HUVEC建立细胞损伤模型，用不同浓度的芝麻素处理细胞；qRT-PCR法检测LncRNA WEE2-AS1与miR-515-5p的表达量；sh-NC、sh-WEE2-AS1转染至HUVEC后加入30 mmol/L葡萄糖处理细胞（HG+sh-NC组、HG+sh-WEE2-AS1组）；构建WEE2-AS1稳定过表达HUVEC细胞，用30 mmol/L葡萄糖处理细胞（HG+WEE2-AS1-LV组），用40 μmol/L芝麻素与30 mmol/L葡萄糖共同处理细胞（HG+SES+WEE2-AS1-LV组）；MTT、流式细胞术分别检测细胞增殖及凋亡率；试剂盒检测超氧化物歧化酶（SOD）、乳酸脱氢酶（LDH）、丙二醛（MDA）的水平；双荧光素酶报告基因实验检测WEE2-AS1与miR-515-5p的靶向关系；Western blotting法检测cleaved-caspase3蛋白表达量。
结果芝麻素可降低高糖诱导的HUVEC中WEE2-AS1的表达量（P < 0.05），可降低凋亡率和cleaved-caspase3蛋白水平（P < 0.05），并可降低LDH的活性和MDA的水平，还可促进miR-515-5p的表达以及增强细胞活力和SOD的活性（P < 0.05），呈剂量依赖性；与HG+sh-NC组比较，HG+sh-WEE2-AS1组miR-515-5p的表达量升高，细胞活力和SOD的活性升高，凋亡率和cleaved-caspase3蛋白水平降低，LDH的活性和MDA的水平降低，差异均有统计学意义（P < 0.05）；WEE2-AS1可靶向调控miR-515-5p；与HG+SES组比较，HG+SES+WEE2-AS1-LV组miR-515-5p的表达量降低，细胞活力和SOD的活性降低，凋亡率和cleaved-caspase3蛋白水平升高，LDH的活性和MDA的水平升高，差异均有统计学意义（P < 0.05）。
结论芝麻素可通过调控WEE2-AS1/miR-515-5p而促进细胞增殖及抑制细胞凋亡、氧化应激进而减轻高糖诱导的血管内皮细胞损伤。

Abstract:
ObjectiveTo explore the effect of sesamin on human umbilical vein endothelial cells (HUVEC) injury induced by high glucose and its possible mechanism.
MethodsHigh glucose stimulated the HUVEC to establish the cell damage model, and the cells were treated with different concentrations of sesamin.qRT-PCR was used to detect the expressions of LncRNA WEE2-AS1 and miR-515-5p.After transfection of sh-NC and sh-WEE2-AS1 into HUVEC, the cells were treated with 30 mmol/L glucose(HG+sh-NC group, HG+sh-WEE2-AS1 group).HUVEC cells stably overexpressing WEE2-AS1 were constructed.HG+WEE2-AS1-LV group was treated with 30 mmol/L glucose.HG+SES+WEE2 -AS1-LV group was treated with 40 μmol/L sesamin and 30 mmol/L glucose.MTT and flow cytometry were used to detect cell proliferation and apoptosis rates.The levels of SOD, LDH, and MDA were tested according to the kit instructions.The dual luciferase reporter gene experiment was used to detect the targeting relationship between WEE2-AS1 and miR-515-5p.Western blotting was used to detect the expression of cleaved-caspase3 protein.
ResultsSesamin reduced the expression of WEE2-AS1, the rate of apoptosis, the protein level of cleaved-caspase3, the activity of LDH and the level of MDA in HUVEC after stimulated with high glucose (P < 0.05).It could promote the expression of miR-515-5p and enhance the cell viability and the activity of SOD (P < 0.05) in a dose-dependent manner.Compared with the HG+sh-NC group, the expression of miR-515-5p in the HG+sh-WEE2-AS1 group was increased (P < 0.05), the cell viability and the activity of SOD were increased (P < 0.05), and apoptosis rate and the protein level of cleaved-caspase3 were decreased (P < 0.05), and the activity of LDH and the level of MDA were decreased (P < 0.05).WEE2-AS1 could targetedly regulate miR-515-5p.Compared with the HG+SES group, the expression of miR-515-5p in the HG+SES+WEE2-AS1-LV group was decreased (P < 0.05), the cell viability and the activity of SOD were decreased (P < 0.05), the apoptosis rate and the protein level of cleaved-caspase3 were increased (P < 0.05), the activity of LDH and the level of MDA were increased (P < 0.05).
ConclusionsSesamin can promote cell proliferation and inhibit apoptosis and oxidative stress by regulating WEE2-AS1/miR-515-5p, thereby reducing the damage of vascular endothelial induced by high glucose.

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