基于TGF-β_(1)/CTGF信号通路探讨山茱萸总甙对人骨关节炎软骨细胞增殖和凋亡的影响  

The effect of cornus officinalis on TGF-β_(1)/CTGF signaling pathway in the proliferation and apoptosis of human osteoarthritis chondrocytes

作　　者：王冠贤 李宝林 唐爱民 黄永湘 Wang Guanxian;Li Baolin;Tang Aimin;Huang Yongxiang(Department of Orthopedics, Danzhou People's Hospital, Hainan Province, Danzhou 571700,China)

机构地区：[1]海南省儋州市人民医院骨科,571700

出　　处：《疑难病杂志》2021年第3期283-288,共6页Chinese Journal of Difficult and Complicated Cases

基　　金：海南省自然科学基金项目(817395)。

摘　　要：目的基于转化生长因子-β1(TGF-β_(1))/结缔组织生长因子(CTGF)信号通路观察山茱萸总甙对人骨关节炎(OA)软骨细胞增殖和凋亡的影响。方法2020年4—5月于海南省儋州市人民医院实验室进行实验。取软骨组织培养软骨细胞,而后分为OA软骨细胞组、罗非昔布组(200μg/ml)及山茱萸总甙低、高剂量组(200、400μg/ml);各组细胞分别设6个平行样,培养72 h。培养结束后,噻唑蓝(MTT)染色测定细胞活力,吉姆萨染色测定细胞群落数,流式细胞仪测定细胞凋亡水平,实时荧光定量PCR(qRT-PCR)及免疫印迹(WB)法测定OA软骨细胞TGF-β_(1)、CTGF mRNA和蛋白表达水平。结果与OA软骨细胞组比较,罗非昔布组及山茱萸总甙低、高剂量组OD值、存活率、群落数明显升高(F/P=15.635/0.000、54.547/0.000、156.632/0.000),凋亡率、TGF-β_(1)及CTGF的mRNA和蛋白表达水平明显降低(F/P=54.236/0.000、36.214/0.000、39.365/0.000、19.541/0.000、23.547/0.000);与罗非昔布组比较,山茱萸总甙低剂量组OD值、存活率、群落数明显降低(q/P=8.954/0.000、8.321/0.000、149.651/0.000),凋亡率及TGF-β_(1)、CTGF mRNA和蛋白表达水平明显升高(q/P=23.458/0.000、8.654/0.000、14.548/0.000、15.458/0.000、22.154/0.000);山茱萸总甙高剂量组OD值、存活率、群落数、凋亡率及TGF-β_(1)、CTGF mRNA和蛋白表达水平与罗非昔布组比较差异无统计学意义(P>0.05);且山茱萸总甙高剂量组OD值、存活率、群落数高于山茱萸总甙低剂量组(q/P=10.654/0.000、8.324/0.000、89.695/0.000),凋亡率及TGF-β_(1)、CTGF mRNA和蛋白表达水平低于山茱萸总甙低剂量组(q/P=38.419/0.000、16.874/0.000、25.654/0.000、19.545/0.000、26.524/0.000)。结论山茱萸总甙能明显促进人骨关节炎软骨细胞增殖,抑制其凋亡,且呈剂量依赖性;其机制与山茱萸总甙抑制人骨关节炎软骨细胞TGF-β_(1)/CTGF信号通路激活有关。Objective Based on the transforming growth factorβ1(TGF-β_(1))/connective tissue growth factor(CTGF)signaling pathway,the effects of total glycosides of cornus on the proliferation and apoptosis of human osteoarthritis(OA)chondrocytes were explored.Methods From April to May 2020,the experiment was conducted in the Laboratory of the People's Hospital of Danzhou City,Hainan Province.The cartilage tissues were taken to culture chondrocytes,and then divided into OA chondrocyte group,rofecoxib group(200μg/ml),and cornus total glycosides low and high dose groups(200,400μg/ml).There were 6 cells in each group.Two parallel samples were cultured for 72 h.After incubation,thiazolyl blue(MTT)staining was used to determine cell viability,Giemsa staining to determine the number of cell colonies,flow cytometry to determine the level of apoptosis,real-time fluorescent quantitative PCR(qRT-PCR)and western blot(WB)methods to determine OA cartilage Cell TGF-β_(1),CTGF mRNA and protein expression levels.Results Compared with the OA chondrocyte group,the OD value,survival rate and number of colonies in the rofecoxib group and the low and high dose groups of cornus total glycosides increased significantly(F/P=15.635/0.000,54.547/0.000,156.632/0.000),Apoptosis rate,TGF-β_(1),CTGF mRNA and protein expression levels were significantly reduced(F/P=54.236/0.000,36.214/0.000,39.365/0.000,19.541/0.000,23.547/0.000);compared with the rofecoxib group,the total dogwood The OD value,survival rate and number of colonies in the low-dose glycoside group were significantly reduced(q/P=8.954/0.000,8.321/0.000,149.651/0.000),and the apoptosis rate,TGF-β_(1),CTGF mRNA and protein expression levels were significantly increased(q/P=23.458/0.000,8.654/0.000,14.548/0.000,15.458/0.000,22.154/0.000).There was no significant difference in OD value,survival rate,colony number,apoptosis rate,TGF-β_(1),CTGF mRNA and protein expression levels in the high-dose dogwood total glycosides group compared with the rofecoxib group(P>0.05);and the dogwood total g

关 键 词：转化生长因子-β_(1)/结缔组织生长因子信号通路 山茱萸总甙 骨关节炎 软骨细胞 增殖 凋亡 

分 类 号：R681.3[医药卫生—骨科学]