高浓度氧通过抑制黏着斑激酶表达损伤早产大鼠肺泡Ⅱ型细胞

Hyperoxia-indueed apoptosis of AEC Ⅱ in premature rats by inhibiting expression of focal adhesion kinase

目的探讨高浓度氧暴露不同时间点早产鼠肺泡Ⅱ型上皮细胞（AECⅡ）凋亡规律及其与黏着斑激酶（FAK）表达的关系。方法原代培养早产鼠AECⅡ，暴露于高氧环境中6、12、24和48h，并以空气组作为对照组，采用Annexin—V和PI双标法经流式细胞仪检测AECⅡ凋亡情况，并采用Western印迹、RT—PCR技术分析AECⅡFAK多肽、磷酸化FAK（FAK—Tyr^397）多肽和FAKmRNA表达变化。结果与空气组比较，高氧暴露12h，Annexin—V^＋／PI^-（早期凋亡）亚群细胞所占比例最高，达（23．83±4．43）％。随高氧暴露时间延长，Annexin—V^＋／PI^-亚群细胞所占比例逐步减低，而Annexin—V^＋／PI^-亚群细胞所占比例逐步增高（P〈0．05或P〈0．01）。AECⅡFAK、FAK—Tyr^397多肽及FAKmRNA表达水平随高氧暴露时间延长呈明显下降趋势（P〈0．05或P〈0．01）。结论高浓度氧抑制FAK表达可能是AECⅡ凋亡和坏死的重要原因之一。

Objective To explore the role of focal adhesion kinase （FAK） in hyperoxia-apopto- sis of type Ⅱ alveolar epithelial cells （AEC Ⅱ s） of premature rats. Methods AEC Ⅱ from prema ture rat lungs were cultured and randomly assigned to air group and hyperoxia group. After exposed to hyperoxia for 6, 12, 24 and 48 h, apoptosis rate of AECⅡ were analyzed by flow cytometry with annexin-V/propidium iodine （PI） double staining. FAK mRNA and FAK and fAK-Tyr397 peptide were detected by RT-PCR and Western blot, respectively. Results Positive cells of Annexin-V^＋/ PI^- in AECⅡ after 6,12,24 and 48 h of hyperoxia exposure were significantly decreased, and the maximal apoptosis rate of AEC Ⅱ （stained of Annexin-V^＋/ PI^- ） was found in the hyperoxia group at 12 h （23.83%±4.43%）. Compared to the air group, the expression of FAK mRNA and of FAK decreased markedly and progressively in hyperoxia groups at 12, 24 and 48 h（P±0.05）. Conclusions Decreased expression of FAK induced by hyperoxia is likely to contribute to the apoptosis and necrosis of AEC Ⅱ.