黄芩总黄酮改善哮喘模型小鼠气道炎症及对TLR3/TRIF通路表达的影响

Effects of Total Flavonoids of Scutellaria Baicalensis Georgi Improving Airway Inflammation and Its Effects on TLR3/TRIF Pathway Expression in Asthmatic Mice

目的探讨黄芩总黄酮对哮喘小鼠气道炎症的改善作用及对Toll样受体3(Toll-likereceptor3,TLR3)/β干扰素TIR结构域衔接蛋白(TIRdomain-containing adaptorinducing interferon-beta,TRIF)通路表达的影响。方法随机将50只4~6周龄雌性Balb/c小鼠分为正常对照组、模型组、阳性对照组(地塞米松,1mg/kg)、黄芩总黄酮低剂量组(25mg/kg)、黄芩总黄酮高剂量组(50mg/kg),每组各10只小鼠。除正常对照组外,其余各组构建支气管哮喘(以下简称为哮喘)小鼠模型。苏木精-伊红(hematoxylin-eosinstaining,HE)染色法观察小鼠肺组织病理改变,并根据病理图片分析小鼠气道重塑情况,计算支气管壁面积/内周长(W/Pi)、支气管平滑肌面积/内周长(S/Pi)和平滑肌细胞核数/内周长(N/Pi)值。采用酶联免疫吸附试验(enzyme linked immunosorben tassay,ELISA)法检测肺泡灌洗液(bronchoalveolar lavagefluid,BALF)中白细胞介素6(interleukin6,IL-6)和IL-8水平。实时荧光定量聚合酶链式反应(quantitative real-timepolymerase chain reaction,qPCR)检测小鼠肺组织中TLR3、TRIFmRNA表达水平。Westernbolt检测小鼠肺组织中TLR3、TRIF蛋白表达水平。结果病理结果显示,正常对照组的支气管腔和肺泡均保持良好的完整性;模型组的小鼠表现出黏膜水肿、炎性细胞浸润(黏膜下层和黏膜层)以及气道壁炎性细胞浸润数量增加,给予黄芩总黄酮后小鼠支气管腔和肺泡炎性细胞浸润减少,且黄芩总黄酮高剂量组炎性细胞数少于黄芩总黄酮低剂量组。与正常对照组相比,模型组W/Pi、S/Pi、N/Pi,IL-6、IL-8,TLR3、TRIFmRNA和蛋白水平,均显著增加(P均<0.05);与模型组相比,黄芩总黄酮低、高剂量组和阳性对照组W/Pi、S/Pi、N/Pi,IL-6、IL-8,TLR3、TRIFmRNA和蛋白水平,均显著降低(P均<0.05);黄芩总黄酮低剂量组W/Pi、S/Pi、N/Pi,IL-6、IL-8,TLR3、TRIFmRNA和蛋白水平,均显著高于黄芩总黄酮高剂量组(P均<0.05);黄芩总黄酮高剂量组与阳性对照组各指标比较差异均无统计学意义(P>0.05)。结论黄芩总黄酮可有效改善哮喘模型小鼠的气道炎症,其机制可能与抑制TLR3/TRIF信号通路,降低炎症反应,缓解气道重塑有关。

Objective To explore the effects of total flavonoids of Scutellaria baicalensis georgi on improving airway inflammation and Toll-like receptor 3(TLR3)/TIR domain-containing adaptor inducing interferon-beta(TRIF)pathway expression in asthmatic mice.Methods A total of female Balb/c mice aged 4 to 6 weeks were randomly divided into normal control group,model group,positive control group(dexamethasone,1 mg/kg),total flavonoids of Scutellaria baicalensis georgi low-dose group(25 mg/kg)and total flavonoids of Scutellaria baicalensis georgi high-dose group(50 mg/kg),with 10 rats in each.Except the normal control group,the asthmatic mouse model was constructed in the other groups.Hematoxylin-eosin staining(HE)were used to observe the pathological changes of lung tissues of mice,and the airway remodeling of mice was analyzed according to pathological pictures;bronchial wall area/inner perimeter(W/Pi),bronchial smooth muscle area/inner perimeter(S/Pi)and smooth muscle nucleus number/inner perimeter(N/Pi)were calculated.Interleukin 6(IL-6)and IL-8 levels in bronchoalveolar lavagefluid(BALF)were detected by enzyme linked immunosorbent assay(ELISA).The mRNA expression levels of TLR3 and TRIF in lung tissues of mice were detected by quantitative real-time polymerase chain reaction(qPCR).The expression levels of TLR3 and TRIF proteins in lung tissues of mice were detected by Western bolt.Results The pathological results showed that the bronchial lumen and alveoli in the normal control group maintained good integrity,while the mice in the model group showed increased mucosal edema,inflammatory cell infiltration(submucosa and mucosal layer)and the number of inflammatory cell infiltration in the airway wall,and decreased inflammatory cell infiltration in the bronchial lumen and alveoli after the administration of total flavonoids of Scutellaria baicalensis georgi,the number of inflammatory cells in the total flavonoids of Scutellaria baicalensis georgi high-dose group was lower than that in the low-dose group.Compared with normal control group,W/Pi,S/Pi,N/Pi,IL-6,IL-8,TLR3 and TRIF mRNA and protein levels in model groups significantly increased(all P<0.05);compared with model group,W/Pi,S/Pi,N/Pi,IL-6,IL-8,TLR3 and TRIF mRNA and protein levels in total flavonoids of Scutellaria baicalensis georgi low-dose and high-dose groups and positive control group significantly decreased(all P<0.05);W/Pi,S/Pi,N/Pi,IL-6,IL-8,TLR3,TRIF mRNA and protein levels in total flavonoids of Scutellaria baicalensis georgi low dose group were significantly higher than those in high dose group(all P<0.05);there was no significant difference in all indexes between total flavonoids of Scutellaria baicalensis georgi high dose group and positive control group(P>0.05).Conclusion Total flavonoids of Scutellaria baicalensis georgi can effectively improve airway inflammation in asthmatic mice,and its mechanism may be related to inhibiting TLR3/TRIF signaling pathway,reducing inflammatory response and alleviating airway removing.