线粒体凋亡通路在严重烧伤大鼠肺纤维化中的作用

Effect of mitochondrial apoptosis on pulmonary fibrosis in rats with severe scald injury

目的探讨线粒体凋亡通路在严重烧伤大鼠肺纤维化中的作用。方法清洁级Wistar大鼠32只，用随机数字表法分为假伤组（A组）、烧伤组（B组）、烧伤12周康复组（C组）及烧伤12周康复＋二次烧伤组（D组）各8只。肺组织标本采集时间为：A组和B组于致伤后第4天；C组和D组则为接受为期12周的康复期后，再次分别接受第二次假伤和烫伤，并同样于伤后第4天搜集标本。Western印迹法检测肺组织B淋巴细胞瘤-2基因蛋白（Bcl-2）、Bcl-2相关X蛋白质（Bax）、剪切体含半胱氨酸的天冬氨酸蛋白水解酶3（Caspase-3）表达水平，末端脱氧核苷酸转移酶介导的dUTP缺口末端标记测定法（TUNEL）检测肺组织细胞凋亡水平，Masson三色染色和天狼星红苦味酸染色法用于检测肺组织胶原表达水平。结果Masson三色染色和天狼星红苦味酸染色病理均显示C组和D组肺组织纤维化明显。B、C、D组大鼠肺组织细胞凋亡率、剪切体Caspase-3和Bax／Bcl-2相对表达水平均显著高于A组，分别为（15．50±3．30）％、（7．88±3．10）％、（15．88±3．23）％比（2．10±1．07）％，（0．59±0．11）、（0．33±0．08）、（0．73±0．13）比（0．16±0．05）和（2．08±0．30）、（0．83±0．09）、（1．54±0．12）比（0．64±0．05）（均P〈0．05）。结论线粒体凋亡可能是促进烧伤后肺纤维化形成和发展的重要机制之一。

Objective To explore the role of mitochondrial apoptosis on pulmonary fibrosis in rats with severe scald injury. Methods According to the random digital table, a total number of 32 Wistar rats were divided into 4 groups： sham bum （group A）, bum group （group B）, 12-week post burn recovery group （group C） , and 12-week post bum recovery plus a second burn injury group （group D）. In group A and group B, lung tissues were harvested on post burn day 4. After received first bum injury 12 weeks, the group C and group D received separately a second sham burn injury and burn injury. Lung tissues were harvested on post burn day 4 after the second burn injury. All tissues were examined for cells apoptosis by Tenninal-deoxynucleoitidyl Transferase Mediated Nick End Labeling （TUNEL）. Pulmonary fibrosis was assessed by Masson tfiehrome staining and Sirius red staining. The protein expression levels of cleaved Caspase-3, Bax and Bcl-2 were assessed by Western blot. Restalts Both Masson trichrome staining and Sirius red staining showed obvious pulmonary fibrosis in group C and group D. The apoptosis rates of group B, C and D were significantly higher than that in group A （（15.50 ±3.30）%, （7.88 ±3.10）%, （ 15.88± 3.23 ） % vs （ 2. 10±1.07 ） % , all P 〈 0. 05 ）. Compared to group A, cleaved Caspase-3 levels were significantly higher in group B, C and D （（0.59 ±0.11）, （0.33±0.08）, （0.73 ±0.13） vs （0. 16 ±0. 05）, all P〈0. 05）. The ratio of Bax/Bel-2 in group B, C and D also increased significantly （（2.08±0.30）, （0.83±0.09）, （1.54±0.12） vs （0.64±0.05）, allP〈0. 05）. Conclusion Severe burn injury can induce pulmonary fibrosis and mitoehondrial apoptosis may play an important role in the development of pubnonary fibrosis.