独活寄生汤对IL-1β诱导软骨细胞TNF-α分泌与PI3K、AKT表达的影响

Effects of Duhuo Jisheng Decoction on TNF-αSecretion and Expression of PI3K,AKT in IL-1β-Induced Chondrocytes

【目的】探讨独活寄生汤对白细胞介素1β(IL-1β)诱导的软骨细胞肿瘤坏死因子α(TNF-α)分泌与磷脂酰肌醇-3-激酶/蛋白激酶B(PI3K/AKT)信号通路中PI3K、AKT蛋白与m RNA表达的影响。【方法】将软骨细胞分为空白对照组,模型对照组,中药低、中、高剂量组和阳性对照组。模型对照组,中药低、中、高剂量组和阳性对照组软骨细胞用IL-1β10 ng·mL-1诱导24 h后,分别加入空白血清,独活寄生汤低、中、高剂量含药血清,鹿瓜多肽注射液含药血清培养12 h。空白对照组不经IL-1β诱导,加入空白血清。应用细胞计数试剂盒8(CCK-8)测定软骨细胞活力的变化,酶联免疫吸附分析(ELISA)检测软细胞培养基中TNF-α含量,蛋白免疫印迹(Western Blot)、荧光实时聚合酶链反应(PCR)法分别检测软骨细胞PI3K、AKT蛋白、mRNA表达水平。【结果】与空白对照组比较,模型对照组IL-1β诱导的软骨细胞活力明显下降(P<0.01),TNF-α的含量明显增高(P<0.01),PI3K、AKT蛋白及mRAN表达水平明显降低(P<0.01);与模型对照组比较,中药中剂量组和阳性对照组软骨细胞活力明显升高(P<0.01),TNF-α含量明显降低(P<0.01),PI3K、AKT蛋白与mRAN表达水平明显升高(P<0.01),且2个治疗组比较,差异无统计学意义(P>0.05)。【结论】独活寄生汤对于IL-1β诱导的软骨细胞损伤有治疗作用,其作用机制可能与调控炎症因子TNF-α的分泌及PI3K/AKT信号通路中PI3K、AKT的表达有关。

Objective To explore the effects of Duhuo Jisheng Decoction on tumor necrosis factorα(TNF-α)secretion and protein and mRNA expression of phosphatidylinositol 3-kinase(PI3 K),protein kinase B(AKT)in PI3 K/AKT signaling pathway from interleukin 1β(IL-1β)-induced chondrocytes.Methods The chondrocytes were divided into blank control group,model control group,low-,middle-and high-dose Chinese medicine groups,and positive control group.The chondrocytes in the model control group,low-,middle-and highChinese medicine groups,and positive control group were induced by IL-1βof 10 ng·mL-1 for 24 h,and then were given blank serum,serum containing low-,middle-and high-dose Duhuo Jisheng Decoction,serum containing Cervus and Cucumis Polypeptide Injection,respectively,for 12 h,while the chondrocytes in the blank control group had no IL-1βinduction,and were only given blank serum.The change of viability of chondrocytes was measured by cell counting kit 8(CCK-8),content of TNF-αin medium of chondrocytes was measured by enzyme-linked immunosorbent assay(ELISA),and protein and mRNA expression levels of PI3 K and AKT were measured by Western blotting assay and real time fluorescence quantitative polymerase chain reaction(PCR),respectively.Results Compared with the blank control group,the viability of IL-1β-induced chondrocytes in the model control group was significantly decreased(P<0.01),content of TNF-αwas significantly increased(P<0.01),and protein and m RNA expression levels of PI3 K and AKT was significantly reduced(P<0.01);Compared with the model control group,the cell viability in the middle-dose Chinese medicine group and positive control group was significantly increased(P<0.01),content of TNF-αwas significantly reduced(P<0.01),and protein and mRNA expression levels of PI3 K and AKT was significantly increased(P<0.01),but there were no differences between the two treatment groups(P>0.05).Conclusion Duhuo Jisheng Decoction has a therapeutic effect on chondrocyte injury induced by IL-1β,and its mechanism may be related to regulating and controlling inflammatory factor TNF-αsecretion and expression of PI3 K and AKT in PI3 K/AKT signaling pathway.