苯并(a)芘通过活化蛋白-1-细胞周期蛋白D1/E2F-1信号通路引起细胞周期的改变

目的 探讨活化蛋白-1(activated protein-1,AP-1)在苯并(a)芘benzo(a)pyrene,B(a)P]引起的人胚肺成纤维细胞(HELF)周期改变中的作用以及AP-1与细胞周期蛋白D1(cyclin D1)、细胞周期蛋白依赖激酶4(CDK4)和E2F-1/4的上下游关系.方法 转染AP-1荧光报告质粒的HELF细胞(AP-H)无血清培养48 h后,加入2μmol/L B(a)P作用24 h,用荧光检测法检测AP-1的相对活性.用AP-1的化学抑制剂姜黄素(curcumin)抑制其活性,观察它与cyclin D1/CDK4和E2F-1/4的上下游关系.采用流式细胞仪检测细胞周期的变化,用Western blot检测细胞中cyclin D1、CDK4和E2F-1/4蛋白水平的改变.结果 2 μmol/LB(a)P作用24 h后,G1期细胞比例由(71±2)%减少为(48±3)%,差异有统计学意义(P＜0.05);AP-1的活性增加;cyclin D1/E2F-1的蛋白含量增加;CDK4/E2F-4的蛋白水平没有明显改变.抑制AP-1活性后,B(a)P诱导的细胞周期的改变被逆转,B(a)P诱导的cyclin D1/E2F-1蛋白含量的增加被抑制,CDK4/E2F-4蛋白含量没有明显改变.结论 B(a)P通过AP-1引起HELF周期的改变.AP-1是cyclin D1/E2F-1的上游信号分子,但对CDK4/E2F-4的表达不具有调节作用.

Activated protein 1-cyclin D1/E2F 1 pathways involved in cell cycle changes induced by benzo (a) pyrene

OBJECTIVE: To investigate the roles of activated protein 1 (AP-1) in cell cycle changes on human embryo lung fibroblasts (HELF) induced by benzo (a) pyrene B (a) P], and relationships between AP-1 and cyclin D1/CDK4-E2F-1/4. METHODS: Cells transfected with AP-1 luciferase reporter plasmid (AP-H) were cultured with serum-free RPMI1640 for 48 h, and treated with 2 micromol/L B (a) P for 24 h. AP-1 relative activity was detected by luciferase assay. Changes of cell cycle and the expression of cyclin D1, CDK4 and E2F-1/4 were checked using the flow cytometer and Western blot assay. RESULTS: After B (a) P was treated for 24 h, the ratio of G1 phase cells (71 +/- 2)% was decreased to (48 +/- 3)% (P < 0.05), and an increase was observed in the ratio of S phase. AP-1 activity and cyclin D1/E2F-1 expression were increased significantly, but CDK4/E2F-4 expression did not change after B (a) P treatment. When AP-1 activity was inhibited by curcumin, decreases of G1 phase in response to B (a) P treatment were blocked, and overexpression of cyclin D1/E2F-1 was attenuated, but CDK4/E2F-4 expression was not changed significantly. CONCLUSION: AP-1 is involved in B (a) P induced cell cycle changes, and is the upstream signals of cyclin D1/E2F-1, but not CDK4/E2F-4.