疏风通络方调控哮喘小鼠Eotaxin/CCR3通路相关因子的实验研究

Experimental study of Shufeng Tongluo Decoction regulating Eotaxin/CCR3 pathway related factors in asthmatic mice

[目的]探索疏风通络方对哮喘小鼠肺组织嗜酸性粒细胞趋化因子(Eotaxin)、CC趋化因子受体3(CCR3)基因表达及含量影响。[方法]70只小鼠随机分为空白对照组、哮喘模型组、疏风通络方低剂量组、疏风通络方中剂量组、疏风通络方高剂量组、CCR3受体拮抗剂(SB328437)组、丝裂原活化蛋白激酶抑制剂(SB203580)组、丝裂原活化细胞外信号调节激酶抑制剂(PD98059)组、磷酸肌醇3-激酶抑制剂(LY294002)组、Ca蛋白拮抗剂(PTX)组,每组7只,除空白对照组外的9组小鼠均应用卵清蛋白致敏/激发的方法建立哮喘模型,哮喘模型组给予生理盐水灌胃,疏风通络方组分别给予低、中、高剂量的疏风通络方灌胃,其余各组给予生理盐水灌胃加腹腔注射相应拮抗剂0.2 mL,连续给药4 d后眼球放血处死小鼠,以聚合酶链式反应(RT-PCR)法测定肺组织Eotaxin-1(CCL11)mRNA、Eotaxin-2(CCL24)mRNA、CCR3 mRNA的表达,以免疫组化法测定肺组织CCR3含量,以酶联免疫吸附法(ELISA)测定肺组织CCL11、CCL24含量。[结果]1)与哮喘模型组比较,各中药组和SB328437组CCL11基因表达均有所降低。2)与哮喘模型组比较,疏风通络方低、中剂量组、SB328437组CCR3含量及基因表达均下降。3)各中药组和拮抗剂组分别与哮喘模型组比较,疏风通络方中剂量组CCL11与CCL24含量均下降,SB328437组CCL11含量明显下降,CCL24含量下降。[结论]疏风通络方具有降低哮喘模型小鼠肺组织中EOS趋化因子CCL11及其受体CCR3基因表达水平、减少哮喘模型小鼠肺组织Eotaxin及CCR3含量的作用,其抑制作用与SB328437类似。

[Objective]To explore the effect of Shufeng Tongluo Decoction on Eotaxin and CCR3 gene expression and content in lung tissue of asthmatic mice.[Methods]70 mice were randomly divided into control group,asthma model group,low dose group,medium dose group,high dose group,SB328437 group,SB203580 group,PD98059 group,LY294002 group and PTX group,with 7 mice in each group the mice in the Shufeng Tongluo Decoction group were given low,medium and high doses of Shufeng Tongluo Decoction by gavage,and the other groups were given normal saline by gavage and intraperitoneal injection of 0.2 mL of corresponding antagonist.After 4 days of continuous administration,the mice were killed by eyeball bloodletting.The expressions of Eotaxin-1(CCL11),Eotaxin-2(CCL24)and CCR3 mRNA in lung tissue were determined by RT-PCR and immunohistochemistry.The contents of CCR3 in lung tissue were determined by ELISA.The contents of CCL11 and CCL24 in lung tissue were determined by ELISA.[Results]1)Compared with asthma model group,the expression of CCL11 in TCM group and SB328437 group were decreased.2)Compared with asthma model group,CCR3 content and gene expression in low,medium dose and SB328437 groups of Shufeng Tongluo Decoction were decreased.3)Compared with the asthma model group,the content of CCL11 and CCL24 decreased in the middle dose Shufeng Tongluo Decoction group.The content of CCL11 and CCL24 decreased significantly in SB328437 group.[Conclusion]Shufeng Tongluo Decoction can reduce the expression of EOS chemokine CCL11 and its receptor CCR3 in the lung tissue of asthmatic mice,and reduce the content of eotaxin and CCR3 in the lung tissue of asthmatic mice,and its inhibitory effect is similar to that of SB328437.