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

摘要:

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48 h,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-Tyr397)多肽和FAKmRNA表达变化.结果 与空气组比较,高氧暴露12 h,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).AEC Ⅱ FAK、FAK-Tyr397多肽及FAK mRNA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.

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abstracts:

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 AEC Ⅱ were analyzed by flow cytometry with an- nexin-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 AEC Ⅱ 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 de- creased 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 neco-z sis of AEC Ⅱ.

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