mTORC1信号通路在张应力下对小鼠骨髓间充质细胞成骨分化的作用  

The role of the mTORC1 signaling pathway during osteogenic differentiation of mouse bone marrow mesenchymal cells under tension stress

作　　者：彭海艳 蒋校文 黄华庆 陈金勇 PENG Haiyan;JIANG Xiaowen;HUANG Huaqing;CHEN Jinyong(Depart-ment of Stomatology,The First People's Hospital of Chenzhou City,The South Medical University&Institute of Transla-tion Medicine,University of China South,Chenzhou 423000,China)

机构地区：[1]南方医科大学附属郴州市第一人民院口腔科南华大学转化医学研究所

出　　处：《口腔疾病防治》2020年第4期219-223,共5页Journal of Prevention and Treatment for Stomatological Diseases

基　　金：国家自然基金(81301651);湖南省自然科学基金(2018JJ2015);郴州市第一人民医院重点项目(N2019-003)

摘　　要：目的探讨哺乳动物雷帕霉素复合物1(mammalian target of rapamycin complex 1,m TORC1)信号通路在周期性单轴牵张力作用下对小鼠骨髓间充质细胞(bone marrow mesenchymal cells,BMMSCs)的成骨分化的作用。方法对体外分离培养的小鼠BMMSCs施加形变量10%的单轴动态牵张力,在牵张后0、1、2、4、8 h利用western blot检测内源性mTORC1信号通路主要分子mTOR、Raptor、核糖体蛋白S6激酶(ribosomal proteinS6kinases,S6K)的表达变化,并利用化学比色法检测碱性磷酸酶(alkaline phosphatase,ALP)的活性变化,ELISA法检测检测骨钙素(osteocalcin,OCN)表达量,RT-PCR法检测Runt相关转录因子(Runt-related transcription factor 2,Runx2)的mRNA表达变化。将BMMSCs分为抑制组、激活组及对照组,分别加入工具药PP242、MHY1485及PBS,加力2 h后用上述方法检测S6K与成骨信号相关因子的活性或表达变化。结果Western blot结果显示,mTORC1信号通路主要分子在牵张力作用后的8 h内均有表达,并在加力后2 h表达最高。与对照组相比,抑制mTORC1信号通路表达后,ALP活性、OCN表达下降,Runx2的mRNA水平上升,差异均具有统计学意义(P<0.001);激活mTORC1信号通路表达后,ALP活性、OCN表达上升,而Runx2 mRNA水平下降,差异均具有统计学意义(P<0.001)。结论 mTORC1信号通路参与了牵张力下小鼠BMMSCs成骨分化过程,激活该信号通路可以促进其成骨分化。Objective To investigate the expression of the mTORC1 signaling pathway during the osteogenic differentiation of mouse bone marrow mesenchymal cells(BMMSCs) under cyclic uniaxial tension and explore its possible role.MethodsThe BMMSCs of mice were affected by uniaxial dynamic tensile force. Western blot was used to detect the expression changes of major molecules(mTOR, Raptor, S6 K) in the endogenous mTORC1 signaling pathway at 0, 1,2, 4, and 8 hours after stretching. Chemical colorimetry, ELISA and PCR were used to detect alkaline phosphatase(ALP), osteocalcin(OCN) and Runx2 mRNA, respectively. Then, inhibition, activation and control groups were established by administration of the drugs PP242, MHY1485 and PBS, respectively. Two hours after the stress, the expression of S6 K was detected by western blot, and the expression of the osteogenic signal was continuously detected by the above methods.Results Western blot analysis showed that the main molecules of the mTORC1 signaling pathwaywere all expressed within 8 hours after traction, and the highest expression was 2 hours after the stress. Compared withthose in the control group, the ALP activity and OCN expression decreased and the Runx2 m RNA levels increased afterthe m TORC1 signal pathway was inhibited(P < 0.001);ALP activity and OCN expression increased after the m TORC1 signal pathway was activated, while the Runx2 m RNA levels decreased(P < 0.001).ConclusionThe mTORC1 signal-ing pathway participates in the osteogenic differentiation of mouse BMMSCs under tension. The osteogenesis of BMMSCs under cyclic uniaxial tension would be enhanced if the mTORC1 signaling pathway was activated.

关 键 词：骨髓间充质细胞 哺乳动物雷帕霉素靶蛋白复合体1 周期性单轴牵张力 牵张成骨 碱性磷酸酶 骨钙素 Runt相关转录因子 核糖体蛋白S6激酶 

分 类 号：R78[医药卫生—口腔医学]