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目的研究P38有丝分裂素激活蛋白激酶(MAPK)在H2O2诱导的人晶状体上皮细胞热休克蛋白质27(HSP27)表达中的作用。方法使用100和200μmol/LH2O2分别以不同时间刺激培养的人晶状体上皮细胞系B3,在阻断实验中加入特异性P38MAPK阻断剂SB203580阻断P38MAPK信号转导通路。采用逆转录聚合酶链反应检测刺激不同时间的人晶状体上皮细胞HSP27mRNA的表达情况,采用免疫印迹法(Westernblotting)检测细胞中HSP27和磷酸化P38MAPK的表达情况。结果(1)H2O2刺激后的人晶状体上皮细胞中HSP27mRNA及其蛋白的表达显著增加;(2)100和200μmol/LH2O2刺激30min,人晶状体上皮细胞应激活化蛋白激酶的活性(平均灰度值)分别增强至800±081和825±050,6h后恢复至基线水平(P<001);磷酸化P38MAPK的表达随H2O2浓度的增高而增加,且在H2O2刺激后15min达到峰值,随后逐渐下降;(3)加用特异性P38MAPK阻断剂SB203580预处理后,200μmol/LH2O2刺激6h的人晶状体上皮细胞HSP27表达(平均灰度值)从3600±082降至875±126,受到显著抑制(P<001)。结论P38MAPK信号转导途径参与H2O2诱导的人晶状体上皮细胞HSP27的表达。(中华眼科杂志,2005,414751)
Objective To investigate the role of P38 mitogen - activated protein kinase (MAPK) in H2O2 - induced heat shock protein 27 (HSP27) expression in human lens epithelial cells. Methods Cultured human lens epithelial cell line B3 was stimulated with 100 and 200 μmol / LH 2 O 2 at different times, and the specific P38 MAPK inhibitor SB203580 was added to block the P38 MAPK signal transduction pathway. The expression of HSP27 mRNA in human lens epithelial cells stimulated with different time was detected by reverse transcription polymerase chain reaction (RT-PCR). The expression of HSP27 and phosphorylated P38MAPK in cells were detected by Western blotting. Results (1) The expression of HSP27 mRNA and protein in human lens epithelial cells stimulated by H2O2 increased significantly. (2) The activation of protein kinase in human lens epithelial cells stimulated by 100 and 200μmol / LHO2 for 30min (average gray value) (P <0.001). The phosphorylated P38MAPK expression increased with the increase of H2O2 concentration and reached the peak at 15 min after H2O2 stimulation, then decreased gradually. ( 3) After pretreatment with specific P38MAPK inhibitor SB203580, the expression of HSP27 (mean gray value) in human lens epithelial cells stimulated by 200μmol / LH 2 O 2 decreased from 3600 ± 082 to 875 ± 126, and was significantly inhibited (P <0.001 ). Conclusion P38MAPK signal transduction pathway is involved in H2O2-induced HSP27 expression in human lens epithelial cells. (Chinese Journal of Ophthalmology, 2005,414751)