高氧对胎鼠肺成纤维细胞p53和增殖细胞核抗原表达的影响

Effect of hyperoxia on p53 and PCNA expression in fetal rat lung fibroblasts

摘要:

目的 探讨高氧对胎鼠肺成纤维细胞（LFs）p53 和增殖细胞核抗原（PCNA）表达的影响。方法 原代培养胎鼠肺 LFs，待生长至亚汇合状态时，随机分为：空气组和高氧组（95% O₂/5% CO₂）。于培养 12 h 和 24 h 时，采用噻唑蓝（MTT）实验测定细胞增殖状况，半定量 RT-PCR 方法检测 p53 mRNA 表达，Western blot 技术检测 p53 和 PCNA 蛋白的表达。结果（1）与空气组比较，高氧组 12 h 和 24 h 的 LFs 生长抑制率分别为 8% 和 23%；（2）高氧组在 12 h 和 24 h 时 p53 mRNA 表达明显高于空气组（P<0.01）；（3）高氧组在 12 h 和 24 h 时 p53 蛋白表达明显高于空气组（P<0.01），而 PCNA 蛋白表达水平在 24 h 明显低于空气组（P<0.01）。结论 高氧暴露抑制 PCNA 表达、促进 p53 表达，从而抑制 LFs 增殖和 DNA 复制，是导致肺发育异常的重要因素。

Abstract:

Objective To study the effect of hyperoxia exposure on the expression of p53 and proliferating cell nuclear antigen（PCNA) in fetal rat lung fibroblasts（LFs). Methods Primary rat embryonic LFs were cultured in vitro.LFs grew to subconfluence and then were randomly divided into air and hyperoxia exposure（95% O₂, 5% CO₂) groups.After LFs were cultured for 12 and 24 hours, the proliferation was analyzed by MTT. p53 mRNA level was detected by semi-quantitative reverse transcription polymerase chain reaction（RT-PCR). p53 and PCNA protein levels were determined by Western blot. Results After 12 and 24 hours of culture the growth inhibition rate of LFs was 8% and 23% respectively in the hyperoxia exposure group. p53 mRNA and protein levels increased significantly（P<0.01) in the hyperoxia exposure group after 12 and 24 hours of culture compared with the air exposure group. Hyperoxia exposure decreased PCNA expression after 24 hours of culture（P<0.01). Conclusion Hyperoxia exposure increases p53 level and decreases PCNA expression, resulting in inhibitions of LFs proliferation and DNA repair.