清肺口服液黄酮类成分改善RSV感染小鼠肺上皮屏障损伤的机制研究

Mechanism of Qingfei Oral Liquid Flavonoids in Improving Lung Epithelial Barrier Damage in RSV-Infected Mice

目的探讨清肺口服液黄酮类成分(Fla)对呼吸道合胞病毒(RSV)感染小鼠肺上皮屏障损伤的保护机制。方法将BALB/c小鼠随机分为正常组,模型组,Fla低、高剂量组(30、60 mg·kg^(-1)·d^(-1)),阳性药利巴韦林组(46 mg·kg^(-1)·d^(-1))。建立RSV感染的小鼠模型,持续给药4 d后,HE染色观察肺组织病理学变化;ELISA检测肺泡灌洗液(BALF)中白介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)的含量;以BALF蛋白含量和BALF/血清荧光比评估肺上皮通透性;Western blot检测肺组织紧密连接蛋白-1(Claudin-1)、闭合蛋白(Occludin)、闭锁小带蛋白-1(ZO-1)、p38丝裂原活化蛋白激酶(p38 MAPK)、p-p38 MAPK、Toll样受体4(TLR4)、髓样分化因子88(MyD88)的蛋白表达。结果与模型组相比,Fla高、低剂量组小鼠肺组织病理损伤减轻(P<0.05,P<0.01),BALF中IL-1β、TNF-α含量下降(P<0.05,P<0.01),BALF蛋白含量和BALF/血清荧光比降低(P<0.05,P<0.01),肺组织中Claudin-1、Occludin、ZO-1蛋白表达升高(P<0.05,P<0.01),p-p38 MAPK/p38 MAPK、TLR4、MyD88蛋白表达下降(P<0.05,P<0.01)。结论Fla可能通过抑制TLR4/MyD88/p38 MAPK通路,促进紧密连接蛋白表达,减轻肺部炎症,从而改善RSV导致的肺上皮屏障损伤。

OBJECTIVE To investigate the protective mechanism of flavonoids in Qingfei Oral Liquid(Fla)on pulmonary epithelial barrier damage in respiratory syncytial virus(RSV)-infected mice.METHODS BALB/c mice were randomly divided into normal group,model group,low-and high-dose groups of flavonoids in Fla(30,60 mg·kg^(-1)·d^(-1)),positive drug ribavirin group(46 mg·kg^(-1)·d^(-1)).RSV-infected mice model was established.After continuous administration for 4 d,the pathological changes of lung tissue were observed by HE staining;ELISA was used to detect the contents of interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α);Protein content in BALF and BALF/serum fluorescence ratio were used to evaluate lung epithelial permeability;Western blot was used to detect the protein expression of Claudin-1,Occludin,ZO-1,p38 mitogen-activated protein kinase(p38 MAPK),p-p38 MAPK,TLR4 and Myd88 in lung tissue.RESULTS Compared with the model group,the lung pathological damage in Fla group was alleviated(P<0.05,P<0.01),the contents of IL-1βand TNF-αin BALF decreased(P<0.05,P<0.01),the protein content in BALF and the ratio of BALF/serum fluorescence decreased(P<0.05,P<0.01),the levels of Claudin-1,Occludin and ZO-1 in the lung tissue increased(P<0.05,P<0.01),the protein expression of p-p38 MAPK/p38 MAPK,TLR4 and Myd88 decreased(P<0.05,P<0.01).CONCLUSION Fla might inhibit the TLR4/MyD88/p38 MAPK pathway,increase the tight junction protein,and reduce lung inflammation,thereby improving the lung epithelial barrier damage caused by RSV.