襄五脂素对哮喘大鼠Th1/Th2失衡和GATA-3的影响

Effect of chicanine on Th1/Th2 imbalance and GATA-3 in asthmatic rats

目的探讨襄五脂素对哮喘动物模型的治疗作用及其机制。方法将大鼠随机分为5组,每组12只:对照组、模型组、低、中、高剂量组。除对照组之外,其他组大鼠均使用卵白蛋白(OVA)诱导构建哮喘大鼠模型。低、中和高剂量组大鼠分别灌胃10,20,40 mg/kg襄五脂素,对照组和模型组大鼠灌胃等体积的0.1%二甲基亚砜(DMSO)溶剂,共给药7 d。采用苏木素伊红(HE)和过碘酸-雪夫(PAS)染色观察大鼠肺组织形态和杯状细胞化生。ELISA法检测大鼠支气管肺泡灌洗液(BALF)中干扰素-γ(IFN-γ)、白细胞介素(IL)-12、IL-4、IL-5、IL-13的水平。流式细胞术检测脾脏Th1/Th2细胞亚群。qRT-PCR检测大鼠肺组织中E-cadherin、α-平滑肌肌动蛋白(α-SMA)、N-cadherin和vimentin的mRNA表达水平。Western blot检测大鼠肺组织中E-cadherin、α-SMA、N-cadherin、vimentin、p-p38(Thr180+Tyr182)、p38、GATA-3和Histone H3(细胞核内参蛋白)蛋白表达水平。结果与模型组比较,低、中和高剂量组大鼠的肺组织形态好转,炎性细胞、杯状细胞和黏液分泌减少;肺组织中E-cadherin的mRNA和蛋白相对表达量升高(P<0.05),α-SMA、N-cadherin和vimentin的mRNA和蛋白相对表达量降低(P<0.05),p-p38(Thr180+Tyr182)/p38比值和GATA-3/Histone H比值降低(P<0.05);BALF中IFN-γ和IL-12水平以及IFN-γ/IL-4比值均升高(P<0.05),IL-4、IL-5和IL-13水平降低(P<0.05);脾脏中CD4^(+)IFN-γ^(+)(Th1)细胞数量和Th1/Th2比值升高(P<0.05),CD4^(+)IL-4(Th2)细胞数量降低(P<0.05)。结论襄五脂素对哮喘大鼠具有显著治疗作用,可纠正哮喘大鼠Th1/Th2失衡并下调肺组织中p38介导的GATA-3磷酸化。

Objective To explore the therapeutic effect and its mechanism of chicanine on the animal model of asthma.Methods Rats were randomly divided into 5 groups:control group,model group,low,medium,and high dose groups,with 12 rats in each group.Except control group,all rats in the other groups were given ovalbumin(OVA)to induce the asthma model.Rats in low,medium,and high dose groups were given 10,20 and 40 mg/kg chicanine,respectively,and the rats in control group and model group were administered with equal volume of 0.1%dimethyl sulfoxide(DMSO)solvent by gavage for 7 d.The morphological changes of lung tissue and goblet cell metaplasia were observed by hematoxylin eosin(HE)and periodic acid-Schiff(PAS)staining.The levels of interferon-γ(IFN-γ),interleukin(IL)-12,IL-4,IL-5 and IL-13 in bronchoalveolar lavage fluid(BALF)were detected by ELISA.Th1/Th2 cell subsets in spleen were detected by flow cytometry.The relative mRNA expressions of E-cadherin,α-smooth muscle actin(α-SMA),N-cadherin and vimentin in lung tissue of rats were detected by qRT-PCR.The relative protein expressions of E-cadherin,α-SMA,N-cadherin,vimentin,p-p38(Thr180+Tyr182),p38,and GATA-3 in lung tissue of rats were detected by Western blot.Results Compared with model group,the morphology of lung tissue was improved in low,medium,and high dose groups,the secretion of inflammatory cells,goblet cells and mucus was decreased,the relative expressions of E-cadherin mRNA and protein in lung tissue were increased(P<0.05),the relative mRNA and protein expressions ofα-SMA,N-cadherin and vimentin were decreased(P<0.05),the ratio of p-p38(Thr180+Tyr182)/p38 ratio and GATA-3/Histone H3 ratio were decreased(P<0.05),the levels of IFN-γand IL-12 and the IFN-γ/IL-4 in BALF were increased(P<0.05),the levels of IL-4,IL-5 and IL-13 were decreased(P<0.05),the number of CD4^(+)IFN-γ^(+)(Th1)cells and Th1/Th2 ratio in spleen were increased(P<0.05),and the number of CD4^(+)IL-4(Th2)cells was decreased(P<0.05).Conclusion Chicanine has a significant therapeutic effect on asthmatic rats by correcting Th1/Th2 imbalance and downregulating p38-mediated GATA-3 phosphorylation in lung tissue of asthmatic rats.