小儿定喘颗粒改善Th17/Treg免疫失衡减轻RSV感染诱导哮喘小鼠气道炎症

Xiao'er Dingchuan Granule Improves Th17/Treg Immune Imbalance and Alleviates Air-way Inflammation Induced by RSV Infection in Asthmatic Mice

目的探讨小儿定喘颗粒对呼吸道合胞病毒(Respiratory syncytial virus,RSV)感染诱导哮喘小鼠肺组织炎症反应的影响及机制。方法将60只SPF级雄性Balb/c小鼠随机分为空白组,模型组,小儿定喘颗粒高、中、低剂量组,阳性药(地塞米松)对照组,每组10只。除空白组外,其它组采用呼吸道合胞病毒、复合鸡卵蛋白(Ovalbumin,OVA)雾化吸入激发哮喘,造模成功后连续给药2周。HE染色法观察小鼠肺组织病理学改变;Masson与过碘酸-雪夫染色(PAS)染色法观察气道胶原沉积和黏液情况;ELISA法检测肺组织、血清中IL-6、lL-10、IL-17和TGF-β细胞因子含量;Western Blot检测STAT3、p-STAT3、RORγt、Foxp3蛋白表达水平,qRT-PCR检测RORγt、Foxp3mRNA表达。结果HE染色显示模型组小鼠气道结构改变,炎性分泌物增多,Masson、PAS染色发现模型组中气道黏液、胶原沉积显著增高;与模型组相比,小儿定喘颗粒各剂量组及地塞米松组小鼠气道结构逐渐恢复,黏液储备与胶原沉积情况改善;与空白组相比,模型组肺组织、血清中IL-6、IL-17和TGF-β细胞因子含量明显升高,lL-10含量下降(P<0.01),小儿定喘颗粒各剂量组与地塞米松组IL-6、IL-17和TGF-β细胞因子表达下降,lL-10表达增高(P<0.01);与空白组组比较,模型组p-STAT3、RORγt显著升高,Foxp3蛋白表达显著降低(P<0.01);与空白组相比,模型组RORγt、Foxp3mRNA含量显著升高(P<0.01);与模型组比较,小儿定喘颗粒各剂量组及地塞米松组RORγt、Foxp3mRNA表达均显著降低(P<0.01)。结论小儿定喘颗粒可以改善Th17/Treg免疫失衡抑制,降低血清及肺组织中IL-6、lL-10、IL-17和TGF-β水平,减少炎细胞迁移、气道炎性浸润及胶原沉积情况,减轻RSV诱导的哮喘小鼠气道炎症反应。

Objective To explore the effect and mechanism of the optimal extraction process of Xiao'er Dingchuan Granules on re-spiratory syncytial virus(RSV)induced inflammatory response in lung tissue of asthmatic mice.Methods Sixty SPF grade male Balb/c mice were randomly divided into a normal group,a model group,a high-dose group of Xiao'er Dingchuan Granules,a medium dose group of Xiao'er Dingchuan Granules,a low-dose group of Xiao'er Dingchuan Granules,and a positive drug(dexa-methasone)control group,with 10 mice in each group.Except for the normal group,other groups were challenged with respirato-ry syncytial virus and ovalbumin(OVA)aerosol inhalation to induce asthma.After successful modeling,the medication was ad-ministered continuously for two weeks.HE staining method was used to observe the pathological changes of mouse lung tissue;Observing the deposition of collagen and mucus reserve in the airway using Masson and periodic acid Schiff staining(PAS);ELISA method for detecting IL-6,lL-10,IL-17,and TGF-βin lung tissue and serum.Cytokine content;Western Blot de-tection of STAT3,p-STAT3,RORγt,Foxp3 protein expression level,qRT-PCR detection of RORγt,Foxp3mRNA expression.Results The results showed that HE staining showed changes in the airway structure of the model group mice,with an increase in inflammatory secretions.Masson and PAS staining showed a significant increase in airway mucus and collagen deposition in the model group;Compared with the model group,the airway structure of mice in each dose group of Xiao'er Dingchuan Granule and dexamethasone group gradually recovered,and the mucus reserve and collagen deposition improved;Compared with the blank group,the model group showed IL-6,IL-17,and TGF-β1 levels in lung tissue and serum.The content of cytokines signifi-cantly increased,while the content of lL-10 decreased(P<0.01).The levels of IL-6,IL-17,and TGF-βin each dose group of Xiao'er Dingchuan Granule and dexamethasone groups were significantly increased.The expression of cytokines decreased and the expression of lL-10 increased(P<0.01);Compared with the control group,the model group showed p-STAT3 and RORγt significantly increased,while Foxp3 protein expression significantly decreased(P<0.01);Compared with the blank group,the model group has a RORγt.Foxp3mRNA expression significantly increased(P<0.01);Compared with the model group,the RORγt of each dose group of Xiao'er Dingchuan Granule and dexamethasone group.The expression of Foxp3mRNA was significantly reduced(P<0.01).Conclusion Xiao'er Dingchuan Granule improves Th17/Treg immune imbalance inhibition and reduces IL-6,lL-10,IL-17,and TGF-βin serum and lung tissue,it reduces the migration of inflammatory cells,air-way inflammatory infiltration,and collagen deposition and alleviates RSV induced airway inflammatory response in asthma mice.