黄芪多糖通过调控TGF-β1/Smads信号通路对百草枯诱导的大鼠肺纤维化的影响

Effects of astragalus polysaccharide on paraquat-induced pulmonary fibrosis in rats by regulating TGF-β1/Smads signaling pathway

目的探索黄芪多糖通过调控TGF-β1/Smads信号通路对大鼠肺纤维化的影响。方法将大鼠分为空白对照(Control)组、黄芪多糖处理(APS)组、百草枯染毒(PQ)组以及PQ+APS组,百草枯诱导大鼠肺纤维化模型,检测大鼠肺湿重/干重比和羟脯氨酸含量,HE染色检测大鼠肺组织病理变化,Masson染色检测大鼠肺组织胶原沉积,ELISA试剂盒检测大鼠支气管肺泡灌洗液中肿瘤坏死因子(TNF-α)、白细胞介素1β(IL-1β)、IL-6的浓度,RT-PCR检测大鼠肺组织转化生长因子β1(TGF-β1)、胶原蛋白I(CollagenⅠ)和CollagenⅢmRNA表达,蛋白印迹检测大鼠肺组织上皮性钙黏附蛋白(E-cadherin)、平滑肌肌动蛋白α(α-SMA)、波形蛋白(Vimentin)、TGF-β1、Smad3、p-Smad3、Smad7蛋白表达。结果与Control组比较,PQ组肺湿重/干重比升高(t=12.922,P<0.001),羟脯氨酸含量升高(t=20.920,P<0.001),肺组织出现病理变化和胶原沉积,肺泡灌洗液中TNF-α(t=23.932,P<0.001)、IL-1β(t=34.826,P<0.001)、IL-6(t=17.985,P<0.001)浓度升高,TGF-β1(t=20.934,P<0.001)、Collagen I(t=26.853,P<0.001)、CollagenⅢ(t=18.493,P<0.001)mRNA表达升高,E-cadherin(t=25.456,P<0.001)蛋白表达降低,α-SMA(t=26.980,P<0.001)、Vimentin(t=23.862,P<0.001),TGF-β1(t=39.836,P<0.001)蛋白表达升高、p-Smad3/Smad3比率(t=19.606,P<0.001)升高,Smad7(t=30.904,P<0.001)蛋白表达降低;与PQ组比较,PQ+APS组肺湿重/干重比降低(t=9.174,P<0.001),羟脯氨酸含量降低(t=10.999,P<0.001),肺组织病理变化和胶原沉积减轻,肺泡灌洗液中TNF-α(t=8.654,P<0.001)、IL-1β(t=18.164,P<0.001)、IL-6(t=7.573,P<0.001)浓度降低,TGF-β1(t=8.879,P<0.001)、CollagenⅠ(t=12.687,P<0.001)、CollagenⅢ(t=11.333,P<0.001)mRNA表达降低,E-cadherin(t=14.255,P<0.001)蛋白表达升高,α-SMA(t=16.866,P<0.001)、Vimentin(t=18.439,P<0.001),TGF-β1(t=14.688,P<0.001)蛋白表达下调、p-Smad3/Smad3比率(t=11.384,P<0.001)下调,Smad7(t=13.131,P<0.001)蛋白表达升高。结论黄芪多糖可减轻百草枯诱导的大鼠肺纤维化,其作用机制与TGF-β1/Smads信号通路相关。

Objective To explore the effects of astragalus polysaccharide on paraquat-induced pulmonary fibrosis in rats by regulating TGF-β1/Smads signaling pathway.Methods Rats were divided into control group,APS,PQ and PQ+APS groups.Paraquat was used to establish the rat model of pulmonary fibrosis.Lung wet/dry weight ratio and hydroxyproline content were measured;the pathological changes were observed by HE staining.The fibrosis in lung tissues was observed by Masson staining,and the concentrations of tumor necrosis factorα(TNF-α),interleukin 1β(IL-1β),and IL-6 were measured by ELISA kits.RT-PCR was used to detect the gene expressionsof transforming growth factor-β1(TGF-β1),collagenⅠand collagenⅢ.Western blotting was used to detect the protein expressions of E-cadherin,α-smooth muscle actin(α-SMA),Vimentin,TGF-β1,Smad3,p-Smad3,and Smad7.Results Compared with the control group,in the PQ group the lung wet weight/dry weight ratio increased(t=12.922,P<0.001)and the hydroxyproline content increased(t=20.920,P<0.001).There were pathological changes and collagen deposition in lung tissues.TNF-α(t=23.932,P<0.001),IL-1β(t=34.826,P<0.001),and IL-6(t=17.985,P<0.001)in alveolar lavage fluid all increased in concentration.TGF-β1(t=20.934,P<0.001),collagenⅠ(t=26.853,P<0.001),and collagenⅢ(t=18.493,P<0.001)gene expressions increased.E-cadherin(t=25.456,P<0.001)protein expression decreased;α-SMA(t=26.980,P<0.001),Vimentin(t=23.862,P<0.001),and TGF-β1(t=39.836,P<0.001)protein expressions increased.p-Smad3/Smad3 ratio(t=19.606,P<0.001)increased,Smad7(t=30.904,P<0.001)protein expression decreased.Compared with PQ group,in PQ+APS group lung wet weight/dry weight ratio decreased(t=9.174,P<0.001),hydroxyl amino acid content(t=10.999,P<0.001)decreased,and lung tissue pathology and collagen deposition reduced.TNF-α(t=8.654,P<0.001),IL-1β(t=18.164,P<0.001),and IL-6(t=7.573,P<0.001)concentrations in alveolar lavage fluid decreased.TGF-β1(t=8.879,P<0.001),collagenⅠ(t=12.687,P<0.001)and collagenⅢ(t=11.333,P<0.001)gene expressions reduced.E-cadherin(t=14.255,P<0.001)protein expression increased;α-SMA(t=16.866,P<0.001),Vimentin(t=18.439,P<0.001),and TGF-β1(t=14.688,P<0.001)protein expressions as well as p-Smad3/Smad3 ratio(t=11.384,P<0.001)were down-regulated;Smad7(t=13.131,P<0.001)protein expression increased.Conclusion Astragalus polysaccharide can alleviate paraquat-induced pulmonary fibrosis in rats.Its mechanism may be related to inhibiting the TGF-β1/Smads signal pathway.