摘要
目的:探究骨痹合剂对膝关节骨性关节炎(KOA)模型鼠软骨细胞PI3K/AKT信号通路的影响。方法:体外培养小鼠KOA软骨细胞凋亡模型,予骨痹合剂含药血清进行干预,通过MTT检测细胞生长情况,运用免疫荧光染色、RT-PCR、Western Blot法检测软骨细胞Bad、Caspase-3、Caspase-9、NF-κB蛋白水平的表达。结果:与5%和20%含药血清浓度比较,10%含药血清更加促进细胞增殖;与第4、6、8天比较,第2天的时间点各组增殖变化最为明显。与正常对照组比较,KOA模型组软骨细胞Bad、Caspase-3、Caspase-9、NF-κB蛋白表达上调,mRNA表达显著上调(P<0.01);与KOA模型组比较,KOA+骨痹合剂组软骨细胞Bad、Caspase-3、Caspase-9、NF-κB蛋白表达下调,Bad、NF-κB mRNA表达显著下调(P<0.05,P<0.01)。与正常对照组比较,KOA模型组软骨细胞PI3K、AKT、mTOR蛋白表达均显著上调(P<0.05);与KOA模型组比较,KOA+骨痹合剂组软骨细胞PI3K、mTOR蛋白表达显著下调(P<0.01)。结论:骨痹合剂能通过抑制PI3K/AKT/mTOR信号通路,增加软骨细胞的自噬能力,抑制Bad、Caspase-3、Caspase-9、NF-κB相关蛋白及基因的表达,从而减少软骨细胞的凋亡。
Objective:To investigate the effects of Gubi Mixture which has the function of activating blood circulation and dredging collaterals on P13K/AKT signaling pathway in chondrocytes of knee osteoarthritis(KOA)model mice.Methods:The apoptotic model of murine KOA chondrocytes was cultured in vitro,and Gubi Mixture containing drug serum was given for intervention.Cell growth was detected by MTT,immunofluorescence staining,RT-PCR and Western Blot were used to detect the expression of related protein levels.Results:Compared with 5%and 20%drug serum concentration,10%drug serum increased cell proliferation;Compared with the 4th,6th and 8th day,the proliferation of each group was the most significant at the time point of the 2nd day.Compared with the chondrocytes in the normal control group,the protein expressions of Bad,Caspase-3,Caspase-9 and NF-κB in the chondrocytes from the KOA model group were up-regulated,and the mRNA expressions of those were up-regulated(P<0.01);Compared with KOA model group the protein expressions of Bad,Caspase-3,Caspase-9 and NF-κB in KOA+Gubi Mixture group were inhibited,and the mRNA expressions of Bad and NF-κB were inhibited(P<0.05,P<0.01).The protein expressions of PI3K,AKT and mTOR were increased in the KOA model group(P<0.05);Compared with KOA model group,the protein expressions of PI3K and mTOR were decreased(P<0.01).Conclusion:Gubi Mixture can increase autophagy ability of chondrocytes and inhibit expression of Bad,Caspase-3,Caspase-9,NF-κB by inhibiting the PI3K/AKT/mTOR signaling pathway,thereby reducing the apoptosis of chondrocytes.
作者
廖建青
马富文
张英杰
谢邦
李具宝
韦佳佳
何渝煦
王琦
LIAO Jian-qing;MA Fu-wen;ZHANG Ying-jie;XIE Bang;LI Ju-bao;WEI Jia-jia;HE Yu-xu;WANG Qi(Yunnan University of Traditional Chinese Medicine,Kunming 650500,China;Yunnan Provincial Hospital of Traditional Chinese Medicine,Kunming 650021,China)
出处
《中华中医药杂志》
CAS
CSCD
北大核心
2022年第4期2322-2326,共5页
China Journal of Traditional Chinese Medicine and Pharmacy
基金
国家自然科学基金项目(No.81560781)
云南省内设研究研究基金(No.2018NS0038)
云南中医药大学2020年度研究生科学研究基金(No.2020Y09)。
