雷公藤多苷通过Wnt/β-catenin缓解IL-1β诱导的软骨细胞损伤

Alleviation of IL-1β-induced chondrocyte injury by tripterygium glycosides via Wnt/β-catenin pathway

目的探究雷公藤多苷对IL-1β诱导的软骨细胞损伤的作用及其潜在的分子机制。方法密度梯度分离法分离SD大鼠软骨细胞。软骨细胞随机分为对照组、模型组、雷公藤多苷组(10、50μg/mL雷公藤多苷)、LiCl组(50μg/mL雷公藤多苷+20 mmol/L LiCl),加入10 ng/mL IL-1β诱导软骨细胞24 h,各给药组加入相应药物继续处理24 h。CCK-8检测细胞存活率,试剂盒检测LDH释放量及GAG水平,流式细胞仪检测细胞凋亡,ELISA检测细胞培养上清液中COX-2、iNOS水平,RT-PCR检测MMP13、Acan、Sox-9 mRNA表达,Western blot检测Wnt通路Wnt3a和β-catenin蛋白的表达。结果雷公藤多苷单独作用对软骨细胞存活率及LDH释放无影响。与对照组比较,模型组细胞存活率降低,LDH释放量增加,细胞凋亡率增加,GAG水平及Acan、Sox-9 mRNA表达降低,COX-2、iNOS水平及MMP13 mRNA表达升高(P<0.05);经雷公藤多苷(50μg/mL)干预后,细胞存活率升高、LDH释放量降低,细胞凋亡率降低,GAG水平及Acan、Sox-9 mRNA表达升高,COX-2、iNOS水平及MMP13 mRNA表达降低(P<0.05);而将Wnt通路激活剂作用之后,雷公藤多苷对IL-1β诱导的软骨细胞上述作用全部被反转。结论雷公藤多苷通过抑制Wnt/β-catenin通路缓解IL-1β诱导的软骨细胞损伤。

AIM To investigate the effects of tripterygium glycosides on IL-1β-induced chondrocyte injury and its potential molecular mechanism.METHODS Chondrocytes isolated from SD rats by density gradient separation were randomly divided into control group,model group,tripterygium glycosides group(10,50μg/mL),and LiCl group(50μg/mL tripterygium glycosides+20 mmol/L LiCl).The chondrocytes exposed to 10 ng/mL IL-1βfor 24 h were treated with corresponding agents for another 24 h.The chondrocytes had their cellular viability detection by CCK-8,their determination of LDH release and GAG level by kit,their assessment of apoptosis by flow cytometry,the detection of COX-2 and iNOS levels in the supernatant of cell culture by ELISA,the detection of expression of MMP13,Acan and Sox-9 mRNA by RT-PCR,and detection of expression of Wnt pathway Wnt3a and-catenin protein by Western blot.RESULTS Tripterygium glycosides alone had no effect on the survival rate and LDH release of chondrocytes.Compared with the control group,the model group displayed decreased survival rate,increased LDH release and apoptosis rate,decreased GAG level and Acan,Sox-9 mRNA expression,and increased COX-2,iNOS and MMP13 mRNA expression(P<0.05).The intervention of tripterygium glycosides(50μg/mL)brought forth increased cell survival rate,decreased LDH release and cell apoptosis rate,increased GAG level and Acan,SARBOX-9 mRNA expression,and decreased COX-2,iNOS and MMP13 mRNA expression(P<0.05).The use of Wnt pathway activator completely reversed tripterygium glycosides’s effects on IL-1β-induced chondrocytes.CONCLUSION Tripterygium glycosides alleviates IL-1β-induced chondrocyte injury via Wnt/β-catenin pathway inhibition.