目的:探讨c-Jun氨端激酶(JNK)信号通路在左归丸含药血清调控成骨前体细胞(MC3T3-E1)增殖和成骨特异转录因子核心结合因子(Runx2)mRNA表达中的作用。方法:以MC3T3-E1为研究对象,制备左归丸含药血清,选用JNK特异抑制剂SP 600125,实验分为空白对照组、SP 600125组、左归丸组、左归丸加SP 600125组、倍美力组、倍美力加SP 600125组。孵育48 h后,采用噻唑蓝(MTT)法检测SP600125对左归丸含药血清干预MC3T3-E1成骨前体细胞增殖作用的影响,采用Western blot法分析JNK蛋白磷酸化水平,采用Real Time RT-PCR法检测成骨细胞特异转录因子Runx2 mRNA表达情况。结果:与空白对照组比较,左归丸含药血清组显著促进细胞增殖,明显上调p-JNK蛋白和Runx2 mRNA表达(P〈0.01);SP600125显著抑制左归丸含药血清诱导的增殖和p-JNK蛋白表达(P〈0.01),对Runx2 mRNA表达的影响不显著。结论:JNK信号通路的激活可能参与了左归丸含药血清诱导的MC3T3-E1成骨前体细胞增殖,但左归丸含药血清诱导的Runx2mRNA高表达对JNK信号通路依赖不显著。
Objective:To examine the role of c-Jun N-terminal kinase(JNK)in proliferation and bone-specific transcription factor runt-related transcription factor 2(Runx2) mRNA expression stimulated by the serum of Zuogui pill in MC3T3-E1 osteoblastic cells.Method: We used model of MC3T3-E1 osteoblastic cells cultured in different serums with or without specific inhibitors of JNK SP 600125 within 48 hours.Grouping:control group,SP 600125 group,Zuogui pill group,Zuogui pill plus SP 600125 group,premarin group,premarin plus SP600125 group.We investigated proliferation using thiazolyl blue(MTT)assay.We analyzed JNK,and p-JNK by immunoblotting.We examined Runx2 mRNA expression in MC3T3-E1 osteoblastic cells by Real Time RT-PCR.Result: Compared with the control group,Zuogui pill group was found to stimulate proliferation significantly,the expression of p-JNK protein and Runx2 mRNA of the Zuogui pill group were increased significantly(P0.01).SP 600125 decreased the proliferation and expression of p-JNK induced by Zuogui pill(P0.01),while the expression of Runx2 mRNA was not altered significantly.Conclusion: JNK signaling pathway may be involved in the proliferation induced by Zuogui pill in MC3T3-E1 osteoblastic cells,while Runx2 mRNA expression induced by Zuogui pill was independent of JNK signaling.