miR-17-5p诱导小鼠肾足细胞系凋亡

miR-17-5p induces apoptosis of glomerular podocytes in mice

目的探讨miR-17-5p在小儿肾病综合征(NS)发病中的作用及其机制。方法将miR-17-5p mimic转染入小鼠肾足细胞系MPC5 24 h后收集细胞。通过microRNA.org、Target Scan和PicTar在线预测miR-17-5p对蛋白酪氨酸磷酸酶受体(PTPRO)-3′UTR区的调控作用。用RT-PCR和Western blot检测PTPRO mRNA以及蛋白的表达,Fluo-3-Am特异性Ca2+荧光指示剂检测足细胞内游离钙离子浓度([Ca2+]),annexin V/PI双染色流式细胞计量术检测足细胞凋亡率。结果生物信息学技术预测,小鼠源及人源miR-17-5p都有多个位点结合于相应的PTPRO-3′UTR区,并抑制PTPRO蛋白及mRNA的表达。PTPRO过表达抑制miR-17-5p诱导的足细胞[Ca2+]升高,PTPRO过表达减缓miR-17-5p mimic诱导的肾小球足细胞凋亡。结论miR-17-5p抑制小鼠肾足细胞内PTPRO表达,并激发肾小球足细胞[Ca2+]升高,诱导肾小球足细胞凋亡。

Objective To explore the role and mechanisms of miR-17-5p in the pathogenesis of childhood nephrotic syndrome.Methods miR-17-5p mimic were transfected into podocytes in kidney of mice(MPC5)for 24 h and then harvested the cells.The potential targets of miR-117-5P were searched using the prediction programs microRNA.org,TargetScan and PicTar.RT-PCR and Western blot were used to detect expression of protein tyrosine phosphatase receptor type O(PTPRO)mRNA and protein.Ca 2+fluorescence indicator with Fluo-3-Amspecificity was used to measure intracellular free calcium concentration([Ca 2+])in podocytes.Annexin V/PI flow cytomeutry was used to test apoptosis rate of podocytes.Results According to the bioinformatics technology prediction,miR-17-5p derived from human or mouse can all bind to multiple sites of the corresponding PTPRO-3′UTR region and inhibit the expression of PTPRO protein and mRNA.PTPRO overexpression inhibited miR-17-5p-induced increase of[Ca 2+]in podocytes in kidney of mice.PTPRO over-expression alleviated miR-17-5p-induced apoptosis of podocytes in kidney of mice.Conclusions miR-17-5p inhibits expression of PTPRO in podocytes in kidney of mice and stimulates increase of intracellular free calcium concentration([Ca 2+])in podocytes,inducing apoptosis of podocytes in kidney of mice.