TLR4表达水平与COPD大鼠肺动脉平滑肌细胞合成分泌功能的关系

Relationship between expression level of TLR4 and secretion and synthesis function of pulmonary artery smooth muscle cells in COPD rats

目的探讨Toll样受体4(TLR4)表达水平与慢性阻塞性肺疾病(COPD)大鼠肺动脉平滑肌细胞(PASMCs)合成分泌功能的关系。方法建立大鼠COPD模型,HE染色判定模型建立,免疫组织化学染色观察对照组与COPD模型组肺组织动脉平滑肌层TLR4的表达。分离培养鉴定原代PASMCs,脂多糖(LPS)、TLR4的特异性抑制剂TAK-242干预细胞,实验分组:空白组、LPS组、TAK-242组、LPS+TAK-242组,Western blot检测各组PASMCs中TLR4的表达,ELISA法检测各组细胞培养上清液中Th1细胞因子干扰素(IFN)-γ、Th2细胞因子白介素(IL)-6及血小板衍化生长因子(PDGF)的浓度;并将TLR4的表达水平与上清液中炎性因子的分泌水平进行相关性分析。结果 COPD模型组大鼠肺组织动脉平滑肌层TLR4的表达明显高于正常大鼠;与空白组比较,LPS组PASMCs中TLR4的表达明显升高(P<0.05);LPS组PASMCs中合成分泌IFN-γ、IL-6、PDGF的水平较对照组明显升高,TAK-242阻断TLR4,PASMCs中TLR4的表达明显降低(P<0.05);细胞培养上清液中IFN-γ、IL-6、PDGF的表达水平较空白组明显降低(P<0.05);TLR4表达水平与细胞培养上清液IFN-γ、IL-6及PDGF的浓度呈显著正相关性(r=0.95、0.87、0.83,P<0.05)。结论TLR4可能参与调控PASMCs的合成分泌功能。

Objective To investigate the relationship between Toll-like receptor 4 （TLR4） expression level of rat pulmonary artery smooth muscle cells with chronic obstructive pulmonary disease （COPD） and its synthesis and se- cretion function. Methods The rat model of COPD was established. HE staining was used to determine whether the model was established successfully; Immunohistochemical staining was used to observe TLR4 expression in pul- monary artery smooth muscle layer of the control group and model group ; primarily cultured and identified pulmona- ry artery smooth muscle （ PASMCs）, lipopolysaccharide （ LPS）, TAK-242 the specificity inhibitors of TLR4 block TLR4, groups as follows： the control group, LPS stimulation group, TAK-242 group, LPS ＋ TAK-242 group, Western blot was used to detect the expression of TLR4 in PASMCs by LPS, Enzyme-linked immunosorbent assay （ELISA） analysis were used to detect the secretion of interferonr-γ（IFN）-γ, interleukin（IL）-6, platelet derivedgrowth factor （PDGF） of rat distal PASMCs in each group. Results Normal rats lung tissue express TLR4, how- ever TLR4 expression level in model group of COPD rats lung tissue obviously higher than normal group （ P 〈 0. 05）. The levels of IFN-γ, IL-6 and PDGF in the PPSMCs of LPS group were significantly higher than those in the control group TLR4 expression level was positively related to the concentration of IFN-γ, IL-6 and PDGF （r = 0. 91,0. 89,0. 83, P 〈 0. 05 ）. Conclusion TLR4 may participate in regulating the synthesis and secretion of PASMCs.