COPD大鼠原代肺动脉平滑肌细胞中MCP-1、TGF-β1和TLR4蛋白的检测

Detection of MCP-1,TGF-β1 and TLR4 protein in primary pulmonary artery smooth muscle cells of COPD rats

目的:探讨慢性阻塞性肺疾病(COPD)大鼠远端原代肺动脉平滑肌细胞(PASMCs)中单核细胞趋化蛋白-1(MCP-1)、转化生长因子-β1(TGF-β1)的分泌情况及其与Toll样受体4(TLR4)信号通路的相关性。方法:通过脂多糖(LPS)诱导建立大鼠COPD模型,分离培养、鉴定原代大鼠PASMCs,以LPS诱导PASMCs,然后分组(对照组、LPS组、TAK-242组,LPS+TAK-242组)培养细胞,用Western blot法检测各组PASMCs中TLR4的表达,ELISA法检测细胞培养上清液MCP-1、TGF-β1的浓度,并进行相关性分析。结果:LPS组较对照组PASMCs中TLR4的表达水平上调(P<0.05),细胞培养上清液中MCP-1、TGF-β1的含量升高(P<0.05);TAK-242组PASMCs中TLR4的表达水平下调(P<0.05),上清液中MCP-1、TGF-β1与TLR4无相关性。结论:LPS可诱导大鼠原代PASMCs合成分泌MCP-1、TGF-β1,同时上调TLR4表达水平。

Aim： To probe into the secretion of monocyte chemoattractant protein 1（ MCP-1）,transforming growth factor β1（ TGF-β1） in primary pulmonary artery smooth muscle cells（ PASMCs） of chronic obstructive pulmonary disease（ COPD） rats and the correlation with toll-like receptor 4（ TLR4） signaling pathways. Methods： The COPD rat model was established,and the PASMCs were separated,induced by LPS,and then divided into control group,LPS group,TAK-242 group,and LPS ＋ TAK-242 group. The expression of TLR4 in PASMCs of each group was detected by Western blot,and the concentrations of MCP-1 and TGF-β1 in cell culture fluid were determined by ELISA,and at last a correlation analysis was conducted. Results： In contrast with the control group,the expression of TLR4 in PASMCs of the LPS group was inecreased（ P〈0. 05） and the concentrations of MCP-1 and TGF-β1 in cell culture fluid were increased（ P〈0. 05）. The expression of TLR4 in PASMCs of the TAK-242 group was down-regulated（ P〈0. 05） and the concentrations of MCP-1 and TGF-β1 in cell culture fluid showed no correlation with the expression of TLR4. Conclusion： LPS can induce PASMCs of rats to synthesize and secrete MCP-1 and TGF-β1,and the TLR4 is up-expressed at the same time.