ALDH2在高氧引起的肺泡上皮细胞损伤中的作用

目的探讨线粒体乙醛脱氢酶2（mitochondrialaldehydedehydrogenase2，AI．DH2）在高氧引起肺泡上皮细胞损伤中的作用。方法采用pcDNA3．1ALDH2转染A549细胞增加ALDH2的表达，采用ALDH2siRNA转染A549细胞抑制ALDH2的表达，细胞分组如下：①对照组（A549细胞），②高ALDH2表达组（A549细胞＋pcDNA3．ALDH2），③低ALDH2表达组（A549细胞＋ALDH2siRNA），将3组细胞均置于常氧（95％空气＋5％C02）和高氧（95％O2＋5％C02）中培养72h。ALDH2表达量检测采用Westernblot，ALDH2活性通过在酶标仪上测定A34（）nm下NAD＋转化为NADH的含量变化来得到，细胞脂质过氧化程度检测采用硫代巴比妥酸法测定丙二醛（malondialdehyde，MDA）含量进行，细胞损伤情况通过检测细胞培养液和细胞内乳酸脱氢酶活力来计算，细胞凋亡检测采用AnnexinVFITC及PI染色后流式细胞术进行，细胞死亡率检测采用台盼蓝染色法。结果转染pcDNA3．1-ALDH2后A549细胞中ALDH2表达显著增多（P〈0．05），活性显著增强（P〈0．05），转染ALDH2siRNA后A549细胞中ALDH2的表达显著减少（P〈0．05），活性显著减弱（P〈0．05），高氧暴露可显著降低ALDH2的表达及活性（P〈0．05）。转染pcD—NA3．1-ALDH2显著减轻了A549细胞高氧暴露后的脂质过氧化和细胞损伤程度（P〈0．05），降低了细胞凋亡率及死亡率（P〈0．05）；转染ALDH2siRNA则显著加重了A549细胞高氧暴露后的脂质过氧化和细胞损伤程度（P〈0．05），增加了细胞凋亡率及死亡率（P〈0．05）。结论ALDH2可以减轻高氧引起的肺泡上皮细胞损伤，其机制可能与ALDH2清除了细胞脂质过氧化产物有关。

ALDH2 Attenuates Hyperoxia-induced Lung Epithelial Cells Injury

Objective To study the role of mitochondrial aldehyde dehydrogenase（ALDH2 or mtALDH） in hyper- oxia-induced lung epithelial cells injury. Methods The pcDNA 3. 1-ALDH2 was transfected into human lung epithelial cells （A549 cells） to increase the expression of ALDH2, and ALDH2 siRNA was transfected into A549 ceils to reduce the expression of ALDH2 by using lipofectamine 2000 （Invitrogen）. A549 cells were divided into three groups： the con- trol group （A549 cells without transfection）, the more expression of ALDH2 group （A549 cells transfeeted with peDNA 3. 1-ALDH2）, and the fewer expression of ALDH2 group （A549 cells transfected with ALDH2 siRNA）. Three groups of A549 cells were cultured in normoxia （95~ air＋ 50/00 CO_, ） or hyperoxia （95~/00 02 ＋ 5% CO2 ） for 72 hours. The ex- pression of ALDH2 was analysed by Western blot, and the activity of ALDH2 was measured by monitoring the reductive reaction of NAD ＋ to NADH at A340 nm in a spectrophotometer. The lipid peroxidation degree of A549 cells was reflec- ted by the malondialdehyde （MDA） level determined by the thiobarbituric acid reactive substance test. Lactate dehydro- genase method was used to test cell injury, and flow cytometry was used to test cell apoptosis after the ceils were stained with Annexin V-FITC and PI. Trypan blue staining method was used to test cell death. Results The expression and ac- tivity of ALDH2 were increased significantly after pcDNA 3. I-ALDH2 was transfeeted into A549 cells （P~O. 05）, and decreased significantly after ALDH2 siRNA was transfected into A549 cells （P%0. 05）. Exposing to hyperoxia reduced the expression and activity of ALDH2 in A549 cells significantly （P〈0. 05）. The degree of lipid peroxidation and cell in- jury of A549 cells exposed to hyperoxia were alleviated, and the ratio of necrotic and apoptotic cells were decreased by the transfection of pcDNA3. 1-ALDH2 significantly （P〈0. 05）. On the contrary, The degree of lipid peroxidation and cell injury of A549 cells exposed to hyperoxia were aggravated, and the ratio of necrotic and apoptotic cells were increased by the transfection of ALDH2 siRNA sig- nificantly （P 〈0. 05 ）. Conclusion ALDH2 attenuates hyperoxia-induced lung epithelial cells injury through re-moving the product of lipid peroxidation.