金雀异黄素对肺泡Ⅱ型上皮细胞发生上皮―间质转化影响.docVIP

金雀异黄素对肺泡Ⅱ型上皮细胞发生上皮―间质转化影响 　　摘要：目的 观察金雀异黄素对转化生长因子-β1（TGF-β1）诱导人肺腺癌A549细胞发生上皮-间质转化（EMT）的影响，探讨其改善肺纤维化的作用机制。方法 CCK-8法检测不同浓度GEN和/或TGF-β1干预细胞72 h后的活性；RT-PCR检测上皮及间质标记物的转变以及在转变过程中发挥关键调节作用的转录因子；Western blot检测p38及JNK信号通路的变化。结果 TGF-β1连续刺激A549细胞72 h可诱导其转化为成纤维细胞表型，表现出更多间质标记物升高，内皮标记物降低；RT-PCR检测结果显示，金雀异黄素可呈浓度依赖性逆转这种现象；Western blot检测结果显示，金雀异黄素与TGF-β1协同刺激A549细胞24 h可显著抑制TGF-β1诱导p38及JNK信号通路的磷酸化水平。结论 金雀异黄素可抑制TGF-β1诱导A549细胞发生EMT而发挥抗纤维化的作用。 　　关键词：金雀异黄素；上皮-间质转化；A549；肺纤维化；转化生长因子-β1 　　DOI：10.3969/j.issn.1005-5304.2016.10.015 　　中图分类号：R285.5 文献标识码：A 文章编号：1005-5304（2016）10-0063-04 　　Abstract： Objective To investigate the effects of genistein on the epithelium-mesenchyme transition of alveolar type II cells； To discuss its mechanism of improving pulmonary fibrosis. Methods The activity of genistein with different concentrations and TGF-β1 intervention cells after 72 h were determined by CCK-8 method； epithelial and mesenchymal markers as well as the key regulatory factors in the transformation process were determined by RT-PCR； changes of p38 and JNK signaling pathways were determined by Western blot. Results The A549 cells were transformed into fibroblast phenotype stimulated by TGF-β1 for 72 h； mesenchymal markers increased and endothelial marker decreased. The results of RT-PCR indicated that genistein inhibited this phenomenon in a concentration-dependent manner. The results of Western blot showed that， genistein possibly inhibited the epithelial-mesenchymal transition through inhibiting the phosphorylation of p38 and JNK signaling pathways and its downstream transcription factors. Conclusion Genistein has anti-fibrosis effects through inhibiting A549 cells undergoing epithelial-mesenchymal transition. 　　Key words： genistein； epithelial-mesenchymal transition； A549； pulmonary fibrosis； TGF-β1 　　特发性肺纤维化（idiopathic pulmonary fibrosis，IPF）是一种以进行性肺纤维化为主要临床特点的疾病，其主要表现为肺间质成纤维细胞的大量聚集，破坏肺泡结构导致肺功能下降最终导致死亡。有研究表明，从IPF患者中分离出的成纤维细胞具有异质性，多种细胞如骨髓和肺泡上皮细胞在条件刺激下可转变为成纤维细胞[1-2]。其中肺泡上皮细胞在转化生长因子-β1（TGF-β