雷帕霉素及雷帕霉素靶蛋白小干扰RNA对高体积分数氧诱导的早产鼠肺成纤维细胞增殖、凋亡和细胞外基质的影响  

Effects of Rapamycin and mammalian target of rapamycin -small interfering RNA （mTOR siRNA） on proli- feration, apoptosis and extracellular matrix in premature rats lung fibroblasts exposed to hyperoxia

作　　者：杜江滨 李龙辉 王少华 刘晓雯 郑雪媚 邓建 Du Jiang-bin;Li Longhui;Wang Shaohua;Liu Xiaowen;Zheng Xuemei;Deng Jian(Department of Neonatology, Women and Children Health Institute Futian, Shenzhen 518045, Guangdong Province, China;Department of Neonatology, the First People＇s Hospital of Yueyaag, Yueyang 414000, Hunan Province, Chin)

机构地区：[1]深圳市福田区妇幼保健院新生儿科,518045 [2]岳阳市第一人民医院新生儿科,414000

出　　处：《中华实用儿科临床杂志》2018年第9期702-706,共5页Chinese Journal of Applied Clinical Pediatrics

基　　金：2014深圳市科技创新资金项目（45576799-3）;2015年深圳市卫计委博士类创新项目（201505022）

摘　　要：目的探讨雷帕霉素及哺乳动物雷帕霉素靶蛋白小干扰RNA（mammalian target of rapamycin－small interfering RNA，mTOR siRNA）对高体积分数氧（高氧）诱导的早产鼠肺成纤维细胞增殖、凋亡及Ⅰ型胶原（collagen Ⅰ，COLⅠ）、Ⅲ型胶原（collagen Ⅲ，COLⅢ）、纤维连接蛋白（fibronectin，FN）表达的影响。方法采用900 mL/L体积分数的氧气制作高氧损伤细胞模型，将早产鼠肺成纤维细胞分为空气对照组、高氧组、高氧＋雷帕霉素组和mTOR siRNA转染组，应用噻唑蓝（MTT）法评估细胞增殖，Annexin V－FITC和碘化丙啶（propidium lodide，PI）双染色法检测细胞凋亡；应用酶联免疫吸附试验（ELISA）法检测COLⅠ、COLⅢ和FN的水平，应用Western blot法检测凋亡相关基因Bcl－2和P53及结缔组织生长因子（connective tissue growth factor，CTGF）和转化生长因子β（transforming growth factor β，TGF－β）的蛋白表达情况。结果与空气对照组比，高氧组肺成纤维细胞增殖减少，凋亡增加，同时COLⅠ（28.30±0.53比17.43±0.37）、COLⅢ（27.86±1.02比17.43±0.37）和FN（32.87±0.42比21.57±0.47）含量及P53（0.810±0.119比0.160±0.018）、CTGF（0.590±0.017比0.220±0.007）和TGF－β（0.580±0.108比0.210±0.008）蛋白表达均增加，Bcl－2（0.150±0.004比0.600±0.130）蛋白表达减少，差异均有统计学意义（均P〈0.01）；雷帕霉素干预后与高氧组对比，肺成纤维细胞增殖增加，凋亡减少，COLⅠ（23.17±0.60比28.30±0.53）、COLⅢ（17.09±0.58比27.86±1.02）和FN（28.11±0.68比32.87±0.42）含量及P53（0.430±0.008比0.810±0.119）、CTGF（ 0.040±0.006比0.590±0.017）和TGF－β（0.380±0.008比0.580±0.108）蛋白表达均减少，Bcl－2（0.290±0.009比0.150±0.004）蛋白表达增加，差异均有统计学意义（均P〈0.01）；mTOR siRNA转染组较高氧＋雷帕霉素组比，肺成纤维细胞增殖增加，凋亡减少，COLⅠ（15ObjectiveTo investigate the effects of Rapamycin and mammalian target of rapamycin-small interfering RNA （mTOR siRNA） on the proliferation, apoptosis and collagen Ⅰ（COLⅠ）, collagen Ⅲ（COLⅢ） and fibronectin（FN） in premature rats lung fibroblasts exposed to hyperoxia.Methods900 mL/L volume fraction of oxygen was used to establish hyperoxia-damaged cell models, and the premature rats lung fibroblasts were divided into air control group, hyperoxia group, hyperoxia＋ rapamycin group and mammalian target of rapamycin-small interfering RNA transfection group.Cell proliferation was assessed by using 3-（4, 5-Dimethylthiazol-2-yl）-2, 5-diphenyltetrazolium bromide assay.Apoptosis were detected by Annexin V-FITC and propidium lodide （PI） double staining.The expressions of COLⅠ, COLⅢ and fibronectin was assessed by using enzyme linked immunosorbent assay and Bcl-2, P53 and pro-fibrotic factors of connective tissue growth factor（CTGF） and transforming growth factor β（TGF-β） by using Western blot.ResultsCompared with the air control group, the proliferation of lung fibroblasts decreased and the apoptosis increased in the hyperoxia group, while the contents of COLⅠ（28.30±0.53 vs. 17.43±0.37）, COLⅢ（27.86±1.02 vs. 17.43±0.37） and fibronectin（32.87±0.42 vs. 21.57±0.47）, P53（0.810±0.119 vs. 0.160±0.018）, TGF-β（0.580±0.108 vs. 0.210±0.008） and CTGF（0.590±0.017 vs. 0.220±0.007） were also increased but the expression of Bcl-2（0.150±0.004 vs. 0.600±0.130） protein was decreased, and the differences were all statistically significant （all P〈0.01）. Compared with the hyperoxia group, the proliferation of lung fibroblasts was increased in the hyperoxia＋ rapamycin group, but the apoptosis was decreased, the contents of COLⅠ（23.17±0.60 vs.28.30±0.53）, COLⅢ（17.09±0.58 vs.27.86±1.02） and fibronectin（28.11±0.68 vs.32.87±0.42）, P53（0.430±0.008 vs.0.810±0.119）, TGF-β（0.380±0.008 vs.0.580±0.108） and CTGF（0.040±0.

关 键 词：雷帕霉素靶蛋白小干扰RNA 雷帕霉素 肺成纤维细胞 细胞增殖 高体积分数氧肺损伤 

分 类 号：R722.6[医药卫生—儿科]