mTOR信号通路介导的自噬在镉抑制人骨髓间充质干细胞成骨分化中的作用  

mTOR signaling pathway-mediated autophagy involved in inhibition of osteoblast differentiation induced by cadmium in human bone marrow mesenchymal stem cells

作　　者：张样聪 卫秦芝 胡友坤 吴璐 李冬丽 王照洁 刘起展[3] 杨杏芬 Zhang Yangcong;Wei Qinzhi;Hu Youkun;Wu Lu;Li Dongli;Wang Zhaojie;Liu Qizhan;Yang Xingfen(Food Safety and Health Research Center School of Public Health,Southern Medical University,Guangzhou 510515,China;School of Medicine,Jinan University,Guangzhou 510632,China;Center for Global Health,School of Public Health,Nanjing Medical University,Nanjing 211166,China)

机构地区：[1]南方医科大学公共卫生学院食物安全与健康研究中心,广州510515 [2]暨南大学基础医学院,广州510632 [3]南京医科大学公共卫生学院,全球健康中心,211166

出　　处：《中华预防医学杂志》2021年第9期1123-1128,共6页Chinese Journal of Preventive Medicine

基　　金：国家自然科学基金(81872642);广东省自然科学基金(2018A0303130222);国家重点基础研发计划(2018YFC1603101);广东省科技攻关计划(2019B020210002)。

摘　　要：目的研究mTOR信号通路介导的自噬在镉抑制人骨髓间充质干细胞(hBMSCs)成骨分化中的作用。方法根据氯化镉的染毒剂量,分为0、2.5和5.0μmol/L染毒组,每组细胞均处理1 d、4 d和(或)7 d,检测ALP活力、成骨标志物(ALP,RUNX2和OSTERIX)mRNA和蛋白质表达水平、自噬相关蛋白(LC3和Beclin-1)和mTOR信号通路相关蛋白(mTOR,p-mTOR和p-p70S6K)表达情况,碱性磷酸酶染色和茜素红染色情况。选择MHY 1485为信号通路激活剂,设置对照组、氯化镉组(5.0μmol/L)、MHY 1485组和氯化镉+MHY 1485联合处理组,处理7 d后分别检测各组hBMSCs的自噬相关蛋白和mTOR信号通路相关蛋白表达水平。结果在第1天和第4天,0、2.5和5.0μmol/L组之间的ALP活力差异无统计学意义(P值均>0.05);在第7天,与0μmol/L组相比,2.5、5.0μmol/L组的ALP活力、成骨标志物ALP、RUNX2和OSTERIX表达水平和mTOR信号通路相关蛋白mTOR、p-mTOR、p-p70S6K表达水平均降低(P值均<0.05),5.0μmol/L组的自噬相关蛋白LC3、Beclin-1表达均有所增加(P值均<0.05)。与0μmol/L组相比,2.5、5.0μmol/L组的染色变浅,ALP和矿化结节生成减少。与氯化镉组相比,氯化镉与MHY 1485联合处理组的自噬相关蛋白表达下降,mTOR信号通路相关蛋白表达增加,差异具有统计学意义(P值均<0.05)。结论镉抑制hBMSCs成骨分化可能与mTOR信号通路介导的自噬有关。Objective To investigate the role of autophagy mediated by mTOR signaling pathway in the inhibition of osteogenic differentiation of human bone marrow mesenchymal stem cells(hBMSCs)induced by cadmium.Methods HBMSCs were divided into 0,2.5 or 5.0μmol/L groups according to the exposure dose of cadmium chloride(CdCl2),and each group was treated for 1 day,4 days and(or)7 days.The ALP activity and mRNA and protein expression levels of osteogenesis markers(ALP,RUNX2 and OSTERIX),autophagy-related proteins(LC3 and Beclin-1)and mTOR signaling pathway related proteins(mTOR,p-mTOR and p-p70S6K)expression,alkaline phosphatase staining and alizarin red staining were detected.MHY 1485 was selected as the signaling pathway activator.The control group,CdCl2 group(5.0μmol/L),MHY 1485 group and CdCl2+MHY 1485 combined treatment group were set.After 7 days of treatment,the expression levels of autophagy related proteins and mTOR signaling pathway related proteins of hBMSCs in each group were detected.Results There was no significant difference in ALP activity between 0,2.5 and 5.0μmol/L groups on day 1 and 4(P>0.05);On day 7,compared with the 0μmol/L group,the ALP activity,expression of osteogenic markers(ALP,RUNX2,OSTERIX)and mTOR signaling pathway related proteins(mTOR,p-mTOR,p-p70S6K)expression decreased in the 2.5 and 5.0μmol/L group(P<0.05).Compared with the 0μmol/L group,the staining of the 2.5 and 5.0μmol/L groups became lighter,and the formation of ALP and mineralized nodules was reduced.Compared with the CdCl2 group,the autophagy related protein expression in the CdCl2+MHY 1485 combined treatment group decreased,and the mTOR signaling pathway related protein expression increased.The difference was statistically significant(P<0.05).Conclusion The inhibition of osteogenic differentiation of hBMSCs by cadmium may be related to autophagy mediated by mTOR signaling pathway.

关 键 词：镉 自噬 成骨细胞 人骨髓间充质干细胞 

分 类 号：R99[医药卫生—毒理学]