MiR-26b通过靶基因GSK3β调控高糖诱导的系膜细胞肥大

MiR-26b Regulates High Glucose-induced Mesangial Cell Hypertrophy Via Targeting GSK3β

目的:研究miR-26b调控高糖诱导系膜细胞(MCs)肥大的机制,为其在糖尿病肾病(DN)早期发病机制研究提供新线索。方法:采用生物信息学方法预测调控GSK3β的候选miRNAs;在db/db小鼠肾皮质中验证所有候选miRNAs的表达并分别与GSK3β做pearson相关性分析;以高糖诱导的MCs为载体,采用real-time PCR,观察与GSK3β显著负相关的miRNAs和GSK3β表达水平;转染miR-26b抑制子后,检测高糖诱导的GSK3β以及α-SMA的表达改变。结果:预测到调控GSK3β的候选miRNAs共9个:分别为miR-199a-5p,miR-128,miR-23b,miR-23a,miR-29a,miR-29c,29b,miR-26b,miR-26a;与对照组相比,miR-128,miR-23b,miR-26b,miR-29a,26b在db/db小鼠肾皮质中表达显著上调;其中miR-26b与GSK3β表达水平负相关最显著(r=-0.470 29);高糖诱导的MCs中miR-26b与GSK3β亦呈现显著负相关,miR-26b抑制子部分逆转GSK3β表达水平,同时α-SMA、Ⅳ型胶原表达水平上调。结论:miR-26b可能通过其靶基因GSK3β参与高糖诱导的MCs肥大机制的调控,为DN早期发病机制研究提供了新的思路。

Objective：To investigate mechanism by which miR -26b regulates high glucose-induced mesangial cells （MCs） hypertrophy, providing a novel clue for the insight into diabetic nephropathy （DN） at early stage. Methods：Applying bioinformatics to predict the candidates of miRNAs potentially targeting glycogen synthase kinase -3β （GSK313）. To verify the expressions of all of miRNAs candidates in renal cortex of db/db mice and perform pearson relavant analysis between miRNAs and GSK3βrespectively as well. The selected miRNA and GSK313 were detected in MCs exposure to high glucose condition by using real - time PCR . Furthermore, GSK313, hypertrophic gene α - SMA and Collagen IV were detected in MCs cultured in high glucose 72 hours after transfection of selected miRNA inhibitor by real - time PCR and western blot. Results： A total of 9 miRNAs candidates potentially targeting GSK313were screened by bioinformatics, which were miR- 199a- 5p, miR- 128, miR- 23b, miR- 23a, miR- 29a, miR-29c, 29b, miR-26b, miR-26a. Among these candidates, miR-128, miR-23b, miR-26b, miR-29a upregulated remarkably in renal cortex in db/db mice compare with db/m controls （P 〈 0.05 ）. One of these four miRNAs, namely miR - 26b showing significantly negative relevance with GSK3 [3by person analysis （r = -0.470 29）. The similar results were found in high glucose -induced MCs. The levels of GSK3β were restored partly by transfection of miR -26b inhibitor, meanwhile α-SMA and Collagen IV ameliorated accordingly in cultured MCs in high glucose conditions. Conclusion：Taken together, miR -26b might involve in the regulation of high glucose - induced MCs hypertrophy via targeting GSK3β. The current study provides a novel insight into the pathogenesis of DN at early stage.