miR-141基因干扰靶向PTEN影响小鼠BMSCs成骨分化的实验研究  

Effect of miR-141 gene interference targeting PTEN on osteogenic differentiation of mouse BMSCs

作　　者：鲍运平[1] 朱正荣[2] 田丰 易洋[2] 程帆 BAO Yunping;ZHU Zhengrong;TIAN Feng;YI Yang;CHENG Fan(the First Clinical Medical College of Yangtze University,Jingzhou 434000,China;Department of Joint Surgery,Jingzhou First People’s Hospital,Jingzhou 434000)

机构地区：[1]长江大学第一临床医学院,湖北荆州434000 [2]荆州市第一人民医院关节外科,湖北荆州434000

出　　处：《中国比较医学杂志》2023年第4期96-103,共8页Chinese Journal of Comparative Medicine

摘　　要：目的 探讨微小核糖核酸-141(micro-RNA-141,miR-141)基因干扰靶向蛋白酪氨酸磷酸酶基因(PTEN)对小鼠骨髓间充质干细胞(bone mesenchymal stem cells, BMSCs)成骨分化的影响。方法 取小鼠BMSCs并鉴定;构建miR-141 mimics、miR-141 inhibitor、miR-141 mimics-NC、miR-141 inhibitor-NC质粒,分别转染小鼠BMSCs,并记为过表达组、沉默组、过表达对照组、沉默对照组,另取小鼠BMSCs常规培养记为空白对照组。碱性磷酸酶(alkaline phosphatase, ALP)染色、茜素红染色检测各组成骨分化能力;检测各组miR-141、PTEN通路相关基因及蛋白表达;验证miR-141是否可靶向调控PTEN。结果 与空白对照组、过表达对照组相比,过表达组ALP定量,AKT、GSK3β mRNA及蛋白表达,Runx2、Osterix蛋白表达,p-AKT、p-GSK3β均下降(P<0.05),钙化结节大小、数量、密度均显著减少,miR-141 mRNA表达及PTEN mRNA和蛋白表达均升高(P<0.05);与空白对照组、沉默对照组相比,沉默组ALP定量,AKT、GSK3β mRNA及蛋白表达,Runx2、Osterix蛋白表达,p-AKT、p-GSK3β均升高(P<0.05),钙化结节也显著增多,miR-141 mRNA表达及PTEN mRNA和蛋白表达低于其余4组(P<0.05);荧光素酶活性实验验证miR-141可靶向调控PTEN。结论 miR-141过表达可激活PTEN信号通路抑制小鼠BMSCs成骨分化,而miR-141基因沉默可下调PTEN,上调AKT、GSK3β表达,增加p-AKT、p-GSK3β水平,并促进成骨标志蛋白表达,促进小鼠BMSCs成骨分化。Objective To investigate the effect of microRNA-141(miR-141)gene interference targeting the protein tyrosine phosphatase gene(PTEN)on the osteogenic differentiation of mouse bone marrow mesenchymal stem cells(BMSCs).Methods miR-141 mimics,an miR-141 inhibitor,miR-141 mimics-NC,and an miR-141 inhibitor-NC plasmids were constructed and transfected into mouse BMSCs to form an over expression group,silent group,over expression control group,and silent control group,respectively.Mouse BMSCs in routine culture were used as a blank control group.Alkaline phosphatase(ALP)staining and alizarin red staining were used to detect the bone differentiation ability of each component.The expression of miR-141 and PTEN pathway-related genes and proteins was detected in each group to verify whether miR-141 can target PTEN.Results Compared with the blank control and over expression control groups,the over expression group had lower expression of ALP,AKT,and GSK3βmRNA and protein;Runx2,Osterix protein,p-AKT and p-GSK3β;and significantly decreased size,number and density of calcified nodules but higher expression of miR-141 mRNA and PTEN mRNA and protein.Compared with the blank control and silent control groups,the silent group had higher expression of ALP,AKT,and GSK3βmRNA and protein;Runx2,Osterix protein,p-AKT,and p-GSK3β;and significantly increased calcified nodules but lower expression of miR-141 mRNA and PTEN mRNA and protein.The luciferase activity experiment verified that miR-141 could target and regulate PTEN.Conclusions miR-141 overexpression activated the PTEN signaling pathway and inhibited the osteogenic differentiation of mouse BMSCs,while miR-141 gene silencing downregulated PTEN and upregulated AKT and GSK3βexpression,increased p-AKT and p-GSK3β,promoted the expression of osteogenic marker protein,and promoted the osteogenic differentiation of mouse BMSCs.

关 键 词：微小核糖核酸-141 蛋白酪氨酸磷酸酶基因 骨髓间充质干细胞 成骨分化 

分 类 号：R-33[医药卫生]