黄芩苷改善IL-1β诱导的软骨细胞凋亡的机制分析

Mechanism analysis of baicalin ameliorative IL-1β induced chondrocyte apoptosis

目的探讨黄芩苷改善IL-1β诱导的软骨细胞凋亡机制,旨在为临床治疗骨关节炎(Osteoarthritis,OA)提供新的靶点。方法将C28/I12细胞以1×10^(4)个/孔的密度接种于96孔板24 h,然后分别用10 ng/mL IL-1β和/或不同浓度(10、20和40μmol/L)的黄芩苷处理12 h,处理0、24、48和72 h。CCK-8和EdU实验用于评估黄芩苷对IL-1β处理的人软骨细胞增殖的影响;TUNEL和流式细胞术实验用于评估黄芩苷对IL-1β处理的软骨细胞凋亡的影响;Western blot实验用于检测黄芩苷对增殖和凋亡相关蛋白表达的影响;ELISA检测用于评估黄芩苷对IL-1β处理的软骨细胞炎症反应的影响。MDC染色、qRT-PCR、Western blot实验评估黄芩苷对IL-1β处理的软骨细胞自噬的影响。最后通过与雷帕霉素联合处理,验证黄芩苷对IL-1β处理的软骨细胞增殖、凋亡、炎症反应和自噬的影响。结果结果表明,黄芩苷以时间和剂量依赖性方式显著提高了IL-1β刺激下软骨细胞的活力和增殖能力,同时抑制细胞凋亡,改善细胞的炎症反应。此外黄芩苷能够明显抑制IL-1β诱导的软骨细胞的自噬。雷帕霉素可部分逆转黄芩苷对IL-1β刺激的软骨细胞增殖、凋亡和炎症反应的影响。结论黄芩苷通过自噬依赖性激活mTOR信号通路促进IL-1β处理的软骨细胞增殖,抑制其凋亡和炎症,从而对OA具有潜在的治疗作用。

Objective To investigate the effect of Baicalin on IL-1βand mechanism of induced chondrocyte apoptosis aims to provide a new target for the clinical treatment of OA.Methods C28/I12 cells were treated with 1×10^(4)/well density was inoculated on 96 well plates for 24 hours,and then 10 ng/ml IL-1 was used respectively and/or different concentrations(10,20 and 40μmol/L)for 12 h,0,24,48 and 72 h.CCK-8 and EdU experiments were used to evaluate the effect of Baicalin on IL-1β.Effect of treatment on proliferation of human chondrocytes;TUNEL and flow cytometry were used to evaluate the effect of Baicalin on IL-1β.Effect of treatment on chondrocyte apoptosis;Western blot was used to detect the effect of Baicalin on the expression of proliferation and apoptosis related proteins;ELISA was used to evaluate the effect of Baicalin on IL-1β.The effect of treatment on the inflammatory response of chondrocytes.The effect of Baicalin on IL-1 was evaluated by MDC staining,QRT PCR and Western blot.Effect of treatment on autophagy of chondrocytes.Finally,the effect of Baicalin on IL-1 was verified by combined treatment with rapamycin.The effects of treatment on chondrocyte proliferation,apoptosis,inflammatory response and autophagy.Results The results showed that baicalin significantly improved the viability and proliferation of chondrocytes stimulated by IL-1βin a time-and dose-dependent manner,while inhibiting cell apoptosis and improving the inflammatory response of cells.In addition,baicalin can significantly inhibit the autophagy of chondrocytes induced by IL-1β.Rapamycin can partially reverse the effects of baicalin on the proliferation,apoptosis and inflammation of chondrocytes stimulated by IL-1β.Conclusion Baicalin promotes IL-1 through autophagy dependent activation of mTOR signaling pathwayβThe treated chondrocytes proliferate and inhibit their apoptosis and inflammation,which has a potential therapeutic effect on OA.