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目的研究高体积分数氧(高氧)对人肺腺癌A549细胞硫氧还蛋白-1(Trx1)及硫氧还蛋白还原酶-1(TrxR1)表达的影响,探讨Trx1及TrxR1在高氧肺损伤中的作用。方法体外传代培养A549细胞,待其生长至融合状态时随机分为高氧组和空气组。高氧组置于一饱和湿度密封箱中,通以950 mL.L-1氧气(O2)[含50 mL.L-1二氧化碳(CO2)]15 min;空气组置于同一室空气中15 min;2组同时置于37℃含50 mL.L-1CO2细胞培养箱中,分别于6 h、12 h、24 h提取其A549细胞总RNA和总蛋白。采取半定量反转录(RT)-PCR测定其Trx1 mRNA、TrxR1 mRNA的表达;采用Western blot检测其A549细胞Trx1蛋白的表达。结果 1.与空气组比较,A549细胞高氧暴露6 h、12 h时Trx1 mRNA表达显著增强(P=0.012,0.002),24 h时减弱,与空气组比较差异无统计学意义(P=0.795);与空气组比较,高氧暴露6 h TrxR1 mRNA无明显变化(P=0.878);12 h、24 h时TrxR1 mRNA表达明显增强(P=0.003,0.001)。2.与空气组比较,高氧暴露6 h Trx1蛋白有所增加但差异无统计学意义(P=0.575);12 h、24 h时Trx1蛋白表达均显著增强(P=0.003,0.026)。结论高氧暴露可上调A549细胞中Trx1及TrxR1的表达,提示内源性的Trx1及TrxR1在高氧肺损伤机制中提供了保护作用。
Objective To investigate the effect of high volume fraction of oxygen (hyperoxia) on the expression of Trx1 and TrxR1 in human lung adenocarcinoma A549 cells and to explore the role of Trx1 and TrxR1 in hyperoxia-induced lung injury The role of injury. Methods A549 cells were subcultured in vitro and randomly divided into hyperoxia group and air group when they grew to confluence. The hyperoxia group was placed in a sealed chamber with a saturated humidity of 950 mL.L-1 oxygen (containing 50 mL.L-1 carbon dioxide (CO2)] for 15 min. The air group was placed in the same room air for 15 min ; 2 groups were simultaneously placed in 37 ℃ containing 50 mL.L-1CO2 cell incubator, respectively, at 6 h, 12 h, 24 h were extracted A549 cell total RNA and total protein. Trx1 mRNA and TrxR1 mRNA were detected by semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). The expression of Trx1 protein in A549 cells was detected by Western blot. Compared with the air group, A549 cells exposed to hyperoxia for 6 h showed a significant increase in Trx1 mRNA expression at 12 h (P = 0.012, 0.002), and decreased at 24 h, with no significant difference compared with air group (P = 0.795 ). Compared with the air group, TrxR1 mRNA had no significant change at 6 h (P = 0.878). The expression of TrxR1 mRNA increased significantly at 12 h and 24 h (P = 0.003, 0.001). Compared with the air group, the Trx1 protein increased 6 h after hyperoxia exposure (P = 0.575), but Trx1 protein expression increased significantly at 12 h and 24 h (P = 0.003, 0.026). Conclusion Hyperoxia exposure can up-regulate the expression of Trx1 and TrxR1 in A549 cells, suggesting that endogenous Trx1 and TrxR1 may provide protective effects in the mechanism of hyperoxia-induced lung injury.