大承气汤用于过敏性哮喘对MAPK信号通路和Th1/Th2型细胞因子影响的动物模型研究

Study on the effect of Dachengqi decoction on MAPK signaling pathway and Th1/Th2 cytokines in animal model of allergic asthma

目的研究大承气汤用于过敏性哮喘小鼠对丝裂原活化蛋白激酶(MAPK)信号通路和辅助性T细胞(Th)1/Th2型细胞因子影响。方法40只BALB/c小鼠通过随机数字表法分为空白对照组(A组)、模型组(B组)、地塞米松组(C组)和大承气汤组(D组),每组各10只。通过卵清白蛋白(OVA)致敏激发构建过敏性哮喘小鼠模型,分别在第0天、第14天致敏,第21~27天进行OVA雾化激发,C组及D组在雾化前1 h分别给予地塞米松及大承气汤干预,A组小鼠给予生理盐水处理。第28天实验结束,取肺组织及气管肺泡灌洗液(BALF),计算四组肺指数,检测四组BALF中细胞总数及各类炎症细胞(嗜酸性粒细胞、中性粒细胞、淋巴细胞、巨噬细胞)数目,检测MAPK信号通路蛋白[p38蛋白激酶(p38 MAPK)、细胞外调节蛋白激酶(ERK)、p-p38 MAPK、p-ERK1/2]表达水平,检测4组BALF炎症细胞因子[人干扰素-γ(INF-γ)、白细胞介素(IL)-4、IL-5、IL-17A]水平。结果肺指数水平:A组<D组<C组<B组(P<0.05)。BALF中总细胞数水平:A组<C组<D组<B组(P<0.05);嗜酸性粒细胞水平:A组<C组<D组<B组(P<0.05);中性粒细胞水平:A组<C组<D组<B组(P<0.05);淋巴细胞水平:A组<C组<D组<B组(P<0.05);巨噬细胞水平:A组>B组>D组>C组(P<0.05)。以β-actin为内参对照,p-p38/β-actin水平:A组=C组<B组(P<0.05),C组=D组<B组(P<0.05),A组<D组(P<0.05);p38/β-actin水平:A组=C组=D组=B组(P>0.05);p-ERK1/2/β-actin水平:A组<C组<D组<B组(P<0.05);ERK1/2/β-actin水平:A组=C组=D组=B组(P>0.05)。INF-γ水平:A组>C组>D组>B组(P<0.05);IL-4水平:A组<C组<D组<B组(P<0.05);IL-5水平:A组<C组<B组(P<0.05),A组<D组(P<0.05),C组=D组(P>0.05),B组=D组(P>0.05);IL-17A水平:A组<C组<D组<B组(P<0.05)。结论大承气汤可有效减轻过敏性哮喘小鼠炎症水平,其可能与抑制MAPK信号通路中磷酸化蛋白质表达及调节Th1/Th2平衡有关。

Objective To study the effects of Dachengqi decoction on the mitogen activated protein kinase(MAPK)signal pathway and helper T cell(Th)1/Th2 cytokines in mice with allergic asthma.Methods Forty BALB/c mice were randomly divided into a blank control group(group A),a model group(group B),a dexamethasone group(group C)and a Dachengqi decoction group(group D),with 10 mice in each group.A mouse model of allergic asthma was established by an ovalbumin(OVA)sensitization challenge.The sensitization was carried out on day 0 and 14th respectively,and the OVA aerosol challenge was carried out on days 21st-27th,and group C and D were treated with dexamethasone and Dachengqi decoction one hour before atomization,while group A was treated with normal saline.At the end of the experiment on the 28th day,the lung tissue and bronchoalveolar lavage fluid(BALF)were taken to calculate the lung index of the four groups,to detect the total number of cells and the number of various inflammatory cells(eosinophils,neutrophils,lymphocytes,macrophages)in the four groups of BALF,to measure the expression levels of MAPK signal pathway proteins[p38 protein kinase(p38 MAPK),extracellular regulated protein kinase(ERK),p-p38 MAPK,p-ERK1/2],and to detect the levels of BALF inflammatory cytokines[human interferon-γ(INF-γ),interleukin(IL)-4,IL-5,IL-17A]in the four groups.Results Lung index level:Group A<group D<group C<group B(P<0.05).The total number of cells in BALF:Group A<group C<group D<group B(P<0.05);Eosinophil level:Group A<group C<group D<group B(P<0.05);Neutrophil level:Group A<group C<group D<group B(P<0.05);Lymphocyte level:Group A<group C<group D<group B(P<0.05);Macrophage level:Group A>group B>group D>group C(P<0.05).Withβ-actin as the internal reference control,p-p38/β-actin level:Group A=group C<group B(P<0.05),group C=group D<group B(P<0.05),group A<group D(P<0.05);p38/β-actin level:Group A=group C=group D=group B(P>0.05);p-ERK1/2/β-actin level:Group A<group C<group D<group B(P<0.05);ERK1/2/β-actin level:Group A=group C=group D=group B(P>0.05).INF-γlevel:Group A>group C>group D>group B(P<0.05);IL-4 level:Group A<group C<group D<group B(P<0.05);IL-5 level:Group A<group C<group B(P<0.05),group A<group D(P<0.05),group C=group D(P>0.05),group B=group D(P>0.05);IL-17A level:Group A<group C<group D<group B(P<0.05).Conclusion Dachengqi decoction can effectively reduce the inflammatory level of allergic asthma mice,which may be related to inhibiting the expression of phosphorylated proteins in the MAPK signal pathway and regulating the balance of Th1/Th2.