摘要
目的百草枯能够导致肺纤维化发生发展,但其发生机制尚不明确。文中探讨Wnt/β-catenin信号通路在百草枯致肺纤维化发生发展中的作用。方法 SD雄性大鼠48只,随机数字表法分为对照组和百草枯中毒组,每组24只。百草枯中毒组大鼠腹腔注射百草枯30 mg/kg建立肺纤维化模型,对照组腹腔注射等量等渗盐水。于百草枯中毒后14 d和28 d断颈处死各组大鼠,剪取肺组织,HE染色和马松染色检测肺纤维化程度,Western blot检测Wnt/β-catenin信号通路关键蛋白β-catenin的表达水平。培养人肺上皮细胞,实验分为3组,空白对照组:不作任何处理;百草枯组:给予50μmol/L的百草枯处理3d;抑制剂组:给予50μmol/L的百草枯处理,同时给予Wnt/β-catenin信号通路抑制剂DKK-1 20ng/m L处理3d。采用免疫荧光和Western blot检测Wnt/β-catenin信号通路关键蛋白β-catenin的表达水平,qRT-PCR检测肺上皮细胞标记E-Cadherin、Occludin和成纤维细胞标记蛋白α-SMA、Vimentin表达水平。结果大鼠百草枯中毒后,HE染色和马松染色结果显示,肺泡壁增厚,肺泡结构紊乱,胶原蛋白沉积,肺纤维化发生;Western blot检测结果表明,与对照组大鼠相比,百草枯中毒组大鼠中毒14 dβ-catenin的表达水平显著升高[(0.38±0.04)vs(0.67±0.06),P<0.01],中毒28 d时高达(1.05±0.08),升高更为明显(P<0.01)。与空白对照组相比,百草枯组E-Cadherin、Occludin表达水平显著下降(P<0.01),成纤维细胞标记α-SMA、Vimentin表达水平显著升高(P<0.01);而百草枯组相比,抑制剂组α-SMA和Vimentin表达水平显著下降,E-cadherin和Occludin的表达水平显著上升(P<0.01)。结论百草枯染毒导致大鼠肺纤维化发生,并导致肺组织Wnt/β-catenin信号通路激活,而阻断Wnt/β-catenin信号通路能够抑制百草枯诱导的上皮-间质发生,证实Wnt/β-catenin信号通路在百草枯致肺纤维化发生发展中起着重要的调控作用。
Objective Paraquat can lead to pulmonary fibrogenesis and its progression, but the underlying mechanisms are not yet unclear. This study aimed to explore the role of the Wnt/β- catenin signaling pathway in the occurrence and progression of paraquat-induced pulmonary fibrosis. Methods A total of 48 adult male Sprague-Dawley rats weighing 200-250 g were randomly divided into a paraquat （PQ） poisoning and a control group of equal number, the former injected intraperitoneally with paraquat at 30 mg/kg to induce pulmonary fibrosis while the latter with an equivalent volume of sterile saline. At 14 and 28 days after modeling, all the rats were sacrificed and their lung tissues collected for examination of the degrees of pulmonary fibrosis by HE and Masson staining and determination of the expression level of the key protein of the Wnt/β-catenin signaling pathway β-catenin by Western blot. Human pulmonary epithelial cells were cultured in vitro and divided into a blank control group, a PQ group, treated with 50 μmol/L of PQ for 3 days, and a DKK-1 group, treated with 50 μmol/L of PQ and 20 ng/ml of DKK-1 （ an antagonist of the Wnt/ β-catenin signaling pathway） for 3 days）. Then the expression of β-catenin was detected by immunofluorescence staining and Western blot and those of epithelial and fibroblasts markers E-Cadherin, Occludin, α-SMA and Vimentin were determined by qRT-PCR. Results HE and Masson staining revealed wall thickening and structural derangement of the pulmonary alveoli, collagen deposition, and pulmonary fibrogenesis. Compared with the controls, the rats in the PQ group showed markedly increased expression of β-eatenin at 14 days （0.38±0.04 vs 0.67±0.06, P〈0.01） , even more significantly at 28 days （ 1.05±0.08, P〈 0.01）. The levels of E-Cadherin and Occludin were remarkably down-regulated （P〈0.01） while those of α-SMA and Vimentin up-regulated in the animals of the PQ group as compared with the blank controls （P〈0.01）. On the contrary, the expressions of E
出处
《医学研究生学报》
CAS
北大核心
2017年第2期117-121,共5页
Journal of Medical Postgraduates
基金
国家自然科学基金(81401583)
全军后勤面上项目(CNJ14L002)
北京协和医学基金会睿E(睿意)急诊医学科研专项基金(R2015028
R2016002)
南京军区南京总医院科研基金(2014004)
