Wnt/β-catenin通路在FSHβ增强BMP9诱导间充质干细胞成骨分化的作用研究  

The study of the Wnt/β-catenin on FSHβenhanced BMP9-induced mesenchymal stem cells osteogenesis

作　　者：成亚琳 赵怡心 张任飞[1] 刘宏[2] CHENG Yalin;ZHAO Yixin;ZHANG Renfei;LIU Hong(Department of Clinical Laboratory, Mianyang Municipal Third People’s Hospital/ Sichuan Provincial Mental Health Center, Mianyang 621000, China;Department of Reproduction Health and Infertility, the First Affiliated Hospital of Chongqing Medical University,Chongqing 400016, China)

机构地区：[1]绵阳市第三人民医院/四川省精神卫生中心,四川绵阳621000 [2]重庆医科大学附属第一医院生殖健康与不孕症中心,重庆400016

出　　处：《中国骨质疏松杂志》2020年第12期1773-1778,共6页Chinese Journal of Osteoporosis

摘　　要：目的探讨Wnt/β-catenin通路在促卵泡激素β亚基(follicle-stimulating hormoneβ-subunit,FSHβ)增强骨形成蛋白9(bone morphogenetic protein 9,BMP9)诱导间充质干细胞(mesenchymal stem cells,MSCs)成骨分化的作用。方法C3H10T1/2细胞为研究对象,通过碱性磷酸酶(alkaline phosphatase,ALP)染色方法检测Ad-GFP对照组、Ad-FSHβ实验组、Ad-BMP9实验组、Ad-BMP9+Ad-FSHβ实验组或Ad-BMP9+Ad-FSHβ+XAV-939实验组等各研究组ALP表达;茜素红S染色(alizarin red s,ARS)方法检测各研究组钙盐小节沉积;实时荧光定量多聚核苷酸链式反应(real-time quantitative polymerase chain reaction,qPCR)技术检测各研究组cyclin D1转录。结果单独的Ad-FSHβ实验组骨形成早、晚期标志物(ALP和钙盐小结)与Ad-GFP对照组相比无显著差异;单独的Ad-BMP9实验组中,两者的表达均高于Ad-GFP对照组;Ad-FSHβ与Ad-BMP9联合实验组的表达量较Ad-BMP9实验组显著增加。检测各研究组Wnt/β-catenin通路的靶基因cyclin D1的转录,与Ad-GFP对照组相比,Ad-FSHβ实验组的表达无显著变化,Ad-BMP9实验组的表达水平显著升高(P<0.01),Ad-FSHβ与Ad-BMP9联合实验组cyclin D1 mRNA表达量显著高于Ad-BMP9实验组(P<0.01)。Wnt/β-catenin通路抑制剂XAV-939处理后,Ad-BMP9与XAV-939联合实验组cyclin D1 mRNA的水平较Ad-BMP9实验组下降(P<0.01),Ad-FSHβ、Ad-BMP9与XAV-939三者联合实验组水平也较Ad-FSHβ与Ad-BMP9联合实验组下降(P<0.05)。ALP的表达呈现出与cyclin D1转录相似的变化趋势。结论Wnt/β-catenin通路可能在促卵泡激素β亚基增强骨形成蛋白9诱导MSCs成骨分化中起重要作用。Objective To investigate the influence of Wnt/β-catenin pathway on follicle-stimulating hormoneβ-subunit(FSHβ)enhanced bone morphogenetic protein 9(BMP9)induced mesenchymal stem cells(MSCs)osteogenesis.Methods C3H10T1/2 cells were the research objects,Alkaline phosphatase(ALP)chromogenic reaction tested the ALP expression in each group,contained:Ad-GFP control group,Ad-FSHβexperimental group,Ad-BMP9 experimental group and Ad-BMP9+Ad-FSHβexperimental group.Alizarin Res S(ARS)staining detected the calcium salt nodules in all groups.Real-time quantitative polymerase chain reaction(qPCR)technology explored the cyclin D1 transcription in each group.Results There were no significant differences in the early and late markers expression of bone formation(ALP and calcium salt deposition)in Ad-FSHβgroup and Ad-GFP control group;in the Ad-BMP9 alone,the expression of both markers were higher than the Ad-GFP control group;in the Ad-FSHβand the Ad-BMP9 combine group,the expression of both markers increased significantly compared with the Ad-BMP9 group.Detected the transcription of Wnt/β-catenin pathway target gene cyclin D1,compared with Ad-GFP control group,there was no significant difference with Ad-FSH experimental group,the level in the Ad-BMP9 experimental group was increased significantly(P<0.01),and the cyclin D1 mRNA expression level in the Ad-FSH and the Ad-BMP9 combined the experimental group was more higher than Ad-BMP9 alone(P<0.01).After treatment with Wnt/β-catenin pathway inhibitor XAV-939,the level of cyclin D1 mRNA in the Ad-BMP9 and the XAV-939 combined group was decreased than the Ad-BMP9 group(P<0.01),and the level of Ad-FSHβ,Ad-BMP9 and XAV-939 combined experimental group was also lower than that of Ad-FSHβand Ad-BMP9 combined experimental group(P<0.05).The change trend of ALP was similar with cyclin D1 transcription.Conclusion Wnt/β-catenin pathway may play an crucial role in follicle-stimulating hormoneβ-subunit enhanced bone morphogenetic protein 9 induced mesenchymal stem cells osteogenes

关 键 词：C3H10T1/2细胞 骨形成蛋白9 促卵泡激素β亚基 WNT信号 XAV-939 

分 类 号：R322.54[医药卫生—人体解剖和组织胚胎学] R322.71[医药卫生—基础医学]