bFGF诱导人晶状体上皮细胞增殖过程中细胞外信号调节激酶的作用

目的:检测细胞外信号调节激酶(extracellular regulated kinase,ERK)在碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)诱导的人晶状体上皮细胞增殖过程中的表达与活化;检测ERK1/2特异性阻断剂PD98059对人晶状体上皮细胞增殖的影响。方法:人晶状体上皮细胞系SRA01/04体外培养,采用MTT比色法和3H]-TdR掺入法检测ERK阻断剂PD98059对人晶状体上皮细胞活力和DNA合成的影响;Westernblot法测定bFGF对培养的人晶状体上皮细胞ERK,c-Jun氨基末端激酶(JNK),P38蛋白表达及PD98059对环氧合酶-2(COX-2)表达的影响。结果:10μg/LbFGF能显著刺激人晶状体上皮细胞增生,1μmol/LPD98059即能显著抑制bFGF启动的HLEC增殖,其效应呈浓度依赖性(P<0.01)。10μg/LbFGF可激活ERK信号通路,诱导COX-2蛋白表达,其效应在30min内达到高峰,6h后逐渐减弱,此作用可被10μmol/LPD98059阻断;10μg/LbFGF对非磷酸化ERK,磷酸化和非磷酸化JNK及P38表达无明显影响。结论:有丝分裂原活性蛋白激酶信号传导通路中ERK磷酸化对bFGF启动的人晶状体上皮细胞增殖具有重要的调节作用。

Activation of extracellular regulated kinase in the proliferation process of human lens epithelial cells induced by bFGF

AIM:To investigate MAPKs-ERK signal transduction in the proliferation process of human lens epithelial cell(HLEC) induced by basic fibroblast growth factor(bFGF) and to detect the inhibition effect of ERK’s blocker PD98059 on the proliferation of HLEC. ·METHODS:HLEC line SRA01/04 was cultured in vitro. MTT and 3H]-thymidine incorporation were used to observe the inhibition effect of PD98059 on the cultured cells and the synthesis of DNA. Western-blot was employed to detect the expression ERK1/2,JNK,P38 and COX-2 protein in HLEC. ·RESULTS:PD98059 treatment at different concentrations(1,5,10μmol/L) resulted in significantly inhibition on cell proliferation and DNA synthesis of HLEC induced by bFGF in a dose-dependent manner. The results of Western-blot showed that expression of phospho-ERK1/2 protein increased in HLEC after treated with 10μg/L bFGF in a time-dependent manner,and negative effect occured after co-treated with 10μmol/L PD98059. bFGF at 10μg/L concentration had no impact on the expression level of nonphospho-ERK1/2,JNK and P38 protein. ·CONCLUSION:The phosphor-ERK is an important regulator of the signal transdution in the proliferation process of HLEC induced by bFGF.