高氧暴露胎鼠肺泡Ⅱ型上皮细胞线粒体蛋白质组学初步研究以及ND4动态表达  

Differential expression of mitochondrial proteome and NADH-dehydrogenase subunit 4 in type Ⅱ alveolar epithelial cell of primary fetal rat exposed to hyperoxia

作　　者：李文斌[1] 常立文[1] 卢红艳[1] 容志惠[1] 汪鸿[1] 刘伟[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院儿科,武汉430030

出　　处：《中华围产医学杂志》2008年第4期258-261,共4页Chinese Journal of Perinatal Medicine

基　　金：国家自然科学基金资助项目（30672261和30770944）;志谢：感谢华中科技大学同济医学院生物化学与分子生物学系蛋白质组学平台田俊、李东两位老师在线粒体蛋白质提取以及双向电泳工作中给予的指导.

摘　　要：目的初步分析高氧暴露下肺泡Ⅱ型上皮细胞（alveolar epithelial cell Ⅱ，AECⅡ）线粒体蛋白质表达谱的改变并就NADH脱氢酶亚单位4（NADH—dehydrogenase subunit4，ND4）的动态表达进行研究。方法原代培养胎鼠AECⅡ并随机分为空气组和高氧组，采用双向电泳检测线粒体蛋白差异性表达，运用RT—PCR和Western blot方法分别检测ND4 mRNA及其蛋白表达。结果与空气组比较，高氧暴露24h导致55种AECⅡ线粒体蛋白质出现差异性表达；高氧暴露6、12、24和48h，原代胎鼠AECⅡ ND4 mRNA（1．20±0．12，1．20±0．12，0．69±0．07和1．54±0．10）表达均较对照组（2．3±0．13，4．61±0．43，2．15±0．14和2．21±0．21）显著降低，差异均有统计学意义（P均〈0．01）。高氧暴露12和24h时，原代胎鼠AECⅡ ND4蛋白表达（1．58±0．21和1．33±0．10）较对照组（2．77±0．19和2．23±0．05）下降，差异有统计学意义（P均〈0．01）。结论高氧暴露导致AECⅡ线粒体（包括呼吸链）功能状态改变可能是促使肺损伤发生发展的重要因素。Objective To explore the effect of hyperoxia exposure on the mitochondrial proteome and the expression of NADH-dehydrogenase subunit 4 （ND4） of type Ⅱ alveolar epithelial cells （AEC Ⅱ ） of primary fetal rat. Methods The primary fetal rats （gestation 19-20 d） AEC Ⅱ were cultured in vitro and then were randomly divided into 2 groups： Air-exposed control group and hyperoxia-exposed group. The differential expression of mitochondrial proteome was analyzed by twodimensional electrophoresis. The expression of ND4 mRNA and protein was determined by reverse transcription polymerase chain reaction （RT-PCR） and Western blot, respectively. Results Compared with air control group, we found 55 protein spots with differential expression after 24 h of hyperoxia. After exposure to hyperoxia for 12 h and 24 h, the expression of AEC Ⅱ ND4 mRNA （1.20±0.12 vs 4.61±0.43,0.69±0.07 vs 2.15±0. 14, P〈0.01 for all） and protein （1.58±0.21 vs 2.77±0. 19, 1.33±0.10 vs 2. 23±0. 05, P〈0. 01 for all） were reduced significantly. At 6 h and 48 h, ND4 mRNA was decreased markedly （1.02±0.14 vs 2.3±0.13, 1.54±0.10 vs 2.21±0.21, P〈0.01 for all）, but the expression of its protein did not change （1.04±0.06 vs 1.00±0.00, 0.97 ±0.06 vs 1.05 ± 0.15, P〉 0.05 for all）. Conclusions The changes of AEC Ⅱ mitochondrial （including respiratory chain） function might play a critical role in the development of hyperoxia lung injury.

关 键 词：高氧症 肺泡 上皮细胞 线粒体蛋白质类 蛋白质组 NADH脱氢酶 

分 类 号：R[医药卫生]