microRNA-497在肾小球系膜细胞焦亡中的作用及机制

发布时间：2018-08-06 12:56

【摘要】：目的:探讨microRNA-497(miR-497)对高糖和胰岛素诱导的肾小球系膜细胞焦亡的调控作用。方法:高糖和胰岛素处理人肾小球系膜细胞(HRMC),在不同时间运用流式细胞仪检测细胞焦亡,实时定量PCR检测HRMC细胞中miR-497表达水平;转染不同浓度miR-497 mimic,检测细胞焦亡通路关键基因mRNA表达水平,运用Western印记检测NLRP1蛋白的表达水平;运用生物信息学和萤光素酶报告基因技术预测并验证miR-497对炎性小体NLRP1基因的靶向结合作用;miR-497 mimic和pc DNA-NLRP1分别单独转染或共转染后检测细胞增殖。结果:高糖和胰岛素处理48 h以上HRMC细胞焦亡水平显著增加(P0.05),miR-497表达水平显著降低(P0.05);miR-497 mimic转染显著抑制HRMC细胞焦亡,和焦亡通路关键基因IL-1β、TNFα和caspase-1表达水平下降(P0.05);miR-497可直接靶向作用于NLRP1基因并抑制NLRP1蛋白的表达;NLRP1过表达可消除miR-497对细胞焦亡的抑制作用。结论:miR-497 mimic抑制高糖和胰岛素诱导的肾小球系膜细胞焦亡。
[Abstract]:Objective: To investigate the regulatory effect of microRNA-497 (miR-497) on the death of glomerular mesangial cells induced by high glucose and insulin. Methods: high glucose and insulin were used to treat human glomerular mesangial cells (HRMC). The expression of miR-497 in HRMC cells was detected by flow cytometry at different time. The expression of miR-497 in HRMC cells was detected by real-time quantitative PCR, and miR was transfected with different concentrations of miR. -497 mimic was used to detect the expression level of key gene mRNA in the cell focal pathway, and to detect the expression level of NLRP1 protein by Western imprint. The target binding effect of miR-497 on the NLRP1 gene of inflammatory corpuscle was predicted and verified by bioinformatics and luciferase reporter gene technique, and miR-497 mimic and PC DNA-NLRP1 were transfected or co transfected separately. Results: the results showed that the level of HRMC cells with high glucose and insulin treatment more than 48 h increased significantly (P0.05), and the expression level of miR-497 decreased significantly (P0.05); miR-497 mimic transfection significantly inhibited the death of HRMC cells, and the key genes of IL-1 beta, TNF A and caspase-1 decreased (P0.05). The expression of the NLRP1 gene and the inhibition of the expression of NLRP1 protein; NLRP1 overexpression can eliminate the inhibitory effect of miR-497 on the apoptosis of cells. Conclusion: miR-497 mimic inhibits the death of glomerular mesangial cells induced by high glucose and insulin.
【作者单位】：第四军医大学西京医院肾内科;第四军医大学学员旅三营九连;
【基金】：国家自然科学基金(81400699) 陕西省自然科学基金重点项目(2014JZ007)
【分类号】：R692

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