miR-204和miR-211下调Arx基因表达影响胰岛素分泌细胞诱导分化进程  

miR-204 and miR-211 down-regulate Arx expression during induction of insulin-producing cells

作　　者：王涛 李晨 穆长征 WANG Tao;LI Chen;MU Changzheng(Medical Treatment College,Jinzhou 121000,Liaoning Province,China;Department of Histology and Embryology Jinzhou Medical University,Jinzhou 121000,Liaoning Province,China)

机构地区：[1]锦州医科大学医疗学院,辽宁锦州121013 [2]锦州医科大学组织学与胚胎学教研室,辽宁锦州121013

出　　处：《解放军医学院学报》2020年第11期1113-1117,共5页Academic Journal of Chinese PLA Medical School

基　　金：辽宁省自然科学基金指导计划项目资助(20180551187,20170540341)。

摘　　要：目的探讨骨髓间充质干细胞(bone marrow mesenchymal stem cells,bMSCs)向胰岛素分泌细胞(insulin-producing cells,IPCs)诱导分化过程中调控Arx(Aristaless-related homeobox)基因表达的miRNAs的鉴定。方法应用Conophylline和尼克酰胺诱导bMSCs向IPCs分化,双硫腙染色观察诱导后的细胞,细胞免疫荧光化学检测细胞内C肽的表达。生物信息学方法预测能够与Arx基因匹配的miRNAs,Real-time PCR检测诱导分化过程中预测的miRNAs以及靶基因Arx的表达。转染miRNAs模拟物进入细胞,实现过表达miRNAs后,Western blotting检测Arx蛋白以及胰岛细胞重要转录因子Pdx1蛋白的表达。结果细胞实施诱导分化后,双硫腙染色呈阳性,细胞着猩红色,免疫荧光化学发现细胞内C肽表达阳性。miRanda和TargetScan两个靶基因预测数据库显示miR-204和miR-211能够与Arx基因靶向匹配,Real-time PCR表明诱导过程中miR-204和miR-211的表达量逐渐降低,而Arx mRNA的表达量逐渐升高。Western blotting发现miR-204和miR-211的过表达能够下调靶蛋白Arx以及Pdx1蛋白的表达。结论miR-204和miR-211能够下调IPCs诱导分化过程中Arx的表达,影响分化进程。Objective To illustrate miRNA regulating aristaless-related homeobox(Arx)expression during differentiation of bone marrow mesenchymal stem cells(bMSCs)into insulin-producing cells.Methods bMSCs were isolated and induced into insulinproducing cells using conophylline and nicotinamide.Induced cells were detected by Dithizone staining and expression of C-peptide was tested by immunofluorescence.The miRNA regulating Arx expression was predicted by bioinformatics and expressions of predicted miRNA and Arx mRNA were measured by real-time PCR during induction of insulin-producing cells.Expressions of Arx and Pdx1 protein were also tested by western blotting after transfection of the predicted miRNA.Results Induced cells could be stained scarlet and the expressions of C-peptide were positive.miR-204 and miR-211 which might regulate Arx expression were obtained using miRanda and TargetScan.Results of real-time PCR showed that expressions of miR-204 and miR-211 were almost reciprocal to the expression of Arx mRNA.Western blotting explored that transfection of miR-204 and miR-211 declined the amount of Arx and Pdx1 protein.Conclusion Therefore,miR-204 and miR-211 can down-regulate the Arx expression and then affect the induction of insulin-producing cells.

关 键 词：微小RNA 骨髓间充质干细胞 胰岛素分泌细胞 细胞分化 小鼠 

分 类 号：R342.2[医药卫生—基础医学]