镉对蛋白磷酸酶2A-B55δ高表达L02细胞周期和核因子-κB活化影响  

Effect of cadmium on the cell cycle and activation of nuclear factor-kappaB in L02 cell with overexpression of protein phosphatase 2A-B55δ

作　　者：胡耀明[1] 陈慧峰[1] 罗洁[1] 李晓杰[1] 陈宇曦[1] 林丽娜[1] 麦剑荣[1] 陈雯[1] 林忠宁[1] 林育纯[1] 

机构地区：[1]广东省营养膳食与健康重点实验室 中山大学公共卫生学院,广东广州510080

出　　处：《中国职业医学》2012年第3期181-185,共5页China Occupational Medicine

基　　金：国家自然科学基金资助项目(81172705,81072334);教育部博士点基金项目(20090171110052);广东省自然科学基金重点项目(S2011020002769)

摘　　要：目的探讨镉对蛋白磷酸酶2A(PP2A)的B55δ亚基高表达和不同多态性单体型调控高表达的肝细胞周期的影响及其与核因子-kappaB(NF-κB)活化的关系。方法选择永生化人正常肝L02细胞,采用B55δ编码基因PPP2R2D序列构建高表达的L02-2R2Dc细胞、与不同启动子区单体型调控高表达的L02-2R2D-PC1和L02-2R2D-PC3细胞株,以导入空载体的L02-pBabe为对照细胞。各细胞给予5～80μmol/L的氯化镉(CdCl2)处理建立细胞模型。噻唑蓝法检测各细胞生长的抑制作用;流式细胞术检测细胞周期;蛋白免疫印迹(Western blotting)检测细胞核内NF-κBp65蛋白的活化情况;荧光定量聚合酶链反应(QRT-PCR)检测环氧合酶-2(COX-2)基因mRNA水平的改变。结果与对照组比较,CdCl2组各细胞的增殖抑制率随镉水平增高呈剂量依赖性增高(Pearson’s r均P<0.05);与CdCl2组比较,冈田酸+CdCl2组各细胞的增殖抑制率明显增加(P<0.05);与对照组比较,CdCl2组细胞G1期比例下降、S期比例增加(P<0.05);胞核p65蛋白水平增高,COX-2 mRNA水平上调;这种效应在不同细胞株之间比较,差异有统计学意义(P<0.05)。结论镉可引起PP2A-B55δ高表达的人肝L02细胞增殖功能抑制并导致细胞周期S期阻滞,这种效应与NF-κB信号通路的活化有关。Objective To investigate the effect of overexpression of protein phosphatase 2A （PP2A） -B55δ subunit on the cell cycle and the mechanisms related to the activation of nuclear factor-kappaB （NF-r：B） in human hepatocytes treated by cadmium. Methods The immortalized human normal hepatocyte L02 cell line was used to construct the PPP2R2D overexpression and regulatory overexpression cells, while L02-pBabe was used as the control ceil. The cadmium chloride （CdCl2 ） groups were treated with 5 to 80 umol/L CdCl2. MTT assay was used to test the cell viability. Flow cytometry （FCM） was used to detect the change of cell cycle. Cyclooxygenase-2 （COX-2） mRNA was detected by real-time polymerase chain reaction （QRT-PCR）. The activation of NF-KB p65 was detected by Western blotting. Results The inhibition of viability in CdCl2 group of all the cell lines increased in a dose-dependent manner （ Pearson＇ s r, P 〈 0.05 ） compared with that of the control group. Compared with the CdCl2 group, the inhibition of viability in OA ＋ CdCl2 group of all the cell lines increased significantly （ P 〈 0.05 ）. In CdCl2 groups, the proportion of cells in G1-phase decreased but in S-phase increased, NF-KB p65 was activated and the COX-2 mRNA level was elevated. All of the effects differed from cell to cell （ P 〈 0. 05 ）. Conclusion It was indicated that the overexpression of PP2A-B55δ subunit involved in the inhibition of cell viability and S-phase cell cycle was arrested in human liver cells induced by CdCl2 , and these effects were related to the activation of NF-KB signal pathway.

关 键 词：蛋白磷酸酶2A 氯化镉 细胞周期 核因子-ΚB L02细胞 

分 类 号：R135.1[医药卫生—劳动卫生] R994.3[医药卫生—公共卫生与预防医学]