肺纤维化大鼠上皮细胞凋亡及Bcl—xl和Bid动态变化

Expression of Bcl-xl/Bid and apoptosis in epithelial cells of pulmonary fibrosis rats

目的探讨肺纤维化大鼠中Bcl-2家族部分蛋白介导的线粒体途径引起上皮细胞凋亡的动态变化及相关关系。方法选取SD大鼠48只，随机分为4组：空白对照组（24只）、模型7d组（8只）、模型14d组（8只）、模型28d组（8只）。应用博莱霉素（5mg／kg）气管内注入建立实验性大鼠肺纤维化模型，空白对照组注入等量生理盐水，各组动物于第7、14、28天随机处死8只，取肺组织采用HE染色观察病理变化，TUNEL法检测肺组织细胞凋亡，RT—PCR和免疫组化法分别检测肺组织Bid、Bcl—xlmRNA以及蛋白的表达。结果实验性大鼠肺纤维化发病过程中，呈现典型的肺泡炎（7d）、肺泡炎与纤维化并存（14d）及稳定的肺纤维化（28d）等表现；同时间段模型组肺组织细胞凋亡指数、BidmRNA及蛋白表达均显著高于空白对照组（P〈0．01），Bcl—xl mRNA及蛋白表达显著低于空白对照组（P〈0．01）；模型组细胞凋亡与Bid蛋白及mRNA从第7天开始逐步升高，第28天达最高峰，BidmRNA与细胞凋亡指数呈正相关（r=0．8275，t=7．9375，P〈0．01）；模型组Bcl—xl蛋白与Bcl—xlmRNA表达在第7天开始下降，第28天达最低峰，Bcl—xlmRNA与细胞凋亡指数呈负相关（r=-0．6338，t=-4．489，P〈0．01）；Bcl—xl mRNA与BidmRNA无明显相关性（r=-0．3869，t=-2．2986，P〉0．05）。结论细胞凋亡在肺纤维形成中起重要作用，肺纤维化细胞凋亡可能与Bcl-2家族蛋白的表达失衡有关。

Objective To investigate the expressions of Bcl-2 family, which leads to apoptotic cell of the pulmonary fibrosis rats. Methods Fouty-eight Sprague-Dawley （SD） rats were randomly divided into four groups：control group, model group 7 d, 14 d,28 d. The model group were induced to produce pulmonary fibrosis with bleomycin endotracheally while the control group was given with normal saline instead in the same condition. Eight rats in each group were sacrificed on 7,14, and 28 days respectively after intratracheal instillation. Pathological changes in the lungs were evaluated by HE stain. The mRNA expressions of Bid and Bcl-xl were evaluated by RT-PCR quantitative analysis, and the immunohistochemical method was used to investigate the change of Bid and Bcl-xl protein. TUNEL was used to detect the change of cell apoptosis. Results The percentage o{ cell apoptosis in alveolar and bronachial epithelial cells of pulmonary fibrosis rats was higher than that in normal control group. The result of immunohistochemistry and RT-PCR suggested that the expressions of Bid mRNA and Bid protein in model groups were higher than those in normal control group. The expressions of Bcl-xl mRNA and Bcl-xl protein in model groups were lower than those in normal control group （ P 〈 0.01）. The expressions of Bid protein and mRNA also increased after 7 d by treated with bleomycin, increased on the 14th day and further increased on the 28th day. The expression of Bcl-xl protein and mRNA also decreased after 7 d by treated with bleomycin, reaches the trough in the 28th day. There was significant correlation between TUNEL results and the expression of Bid/Bcl-xl mRNA. Conclusions Higher expression of apoptosis in alveolar epithelial cells and bronchial cells of pulmonary fibrosis rats can contribute to fibrogenesis. The apoptosis index of lung cells may be related to the expression imbalance of Bcl-2 family protein.