PI3K信号通路对哮喘小鼠调节性T细胞与Th17失衡的调控作用  

Regulatory effect of PI3K signal pathway on Treg/Th17 imbalance in a murine asthma model

作　　者：聂颖[1] 张维溪[1] 崇蕾[1] 王婷[1] 李昌崇[1] 

机构地区：[1]温州医学院附属育英儿童医院呼吸科,浙江温州325027

出　　处：《中国病理生理杂志》2012年第2期332-337,共6页Chinese Journal of Pathophysiology

基　　金：浙江省自然科学基金资助项目(No.Y2090327);浙江省大学生科技创新活动计划项目(No.2010R413044);国家青年科学基金资助项目(No.81100015)

摘　　要：目的:探讨PI3K信号通路对哮喘小鼠T细胞中辅助性T细胞17(Th17)与CD4+CD 25+调节性T细胞(Treg)失衡的调控作用。方法:尼龙毛柱法分选对照组和哮喘组BALB/c小鼠脾脏T细胞,进而在细胞水平分组:对照组(A组)分为2个亚组:空白对照组(A1),PI3K抑制剂LY294002对照组(A2);哮喘组(B组)继续分为4个亚组:哮喘组(B1),5μmol/L PI3K抑制剂LY294002组(B2),10μmol/L PI3K抑制剂LY294002组(B3),20μmol/L PI3K抑制剂LY294002组(B4),共培养72 h,采用流式细胞仪检测CD4+CD 25+Treg的数量,酶联免疫吸附反应(ELISA)检测上清中白细胞介素(IL)-10和IL-17水平,逆转录-聚合酶链反应(RT-PCR)检测转录因子Foxp3和RORγt的mRNA水平。结果:(1)分选后获得的T细胞纯度为(74.12±3.08)%,细胞存活率为(92.82±3.21)%;(2)Treg的数量B1组较A1组显著降低(P<0.01);A2组显著高于A1组(P<0.05);B3和B4组较B1组显著增高(均P<0.01);(3)IL-17水平和RORγt mRNA表达B1组较A1组显著增高(P<0.01);A2组显著低于A1组(P<0.01,P<0.05);B2、B3、B4组均显著低于B1组(均P<0.01),呈剂量依赖性降低;而IL-10水平和Foxp3 mRNA表达B1组显著低于A1组(P<0.01);A2组显著高于A1组(P<0.01);B2、B3和B4组均显著高于B1组(P<0.01),且呈浓度依赖性增高;(4)RORγt/Foxp3 mRNA的比值B1组显著高于A1组(P<0.01);A2组显著低于A1组(P<0.01);B2、B3和B4组显著低于B1组(P<0.01),且呈浓度依赖性降低;RORγt/Foxp3 mRNA比值与IL-17水平呈正相关(r=0.89,P<0.01);RORγt/Foxp3 mRNA比值与IL-10呈负相关(r=-0.86,P<0.01)。结论:哮喘小鼠存在Th17/Treg失衡,PI3K信号通过调控Treg/Th17失衡而参与哮喘发病过程。AIM： To explore the regulatory effect of PI3K signal pathway on the regulatory T cells（Treg）/T helper 17 cells（Th17） imbalance in a murine asthma model.METHODS： BALB/c mice were used in the study.The mice were randomly divided into control group and asthma group.The mice in asthma group were intraperitoneally injected with the mixture of ovalbumin（OVA）/Al（OH）3 and then activated by exposure of the animals to OVA atomization.The T cells in the spleens from BALB/c mice were isolated by the method of nylon wool column,and the T cells were divided into different subgroups： the T cells from control group were divided into normal control group（A1） and PI3K inhibitor-treated group（A2）;the T cells from asthma group were divided into asthma group（B1）,5 μmol/L PI3K inhibitor-treated group（B2）,10 μmol/L PI3K inhibitor-treated group（B3） and 20 μmol/L PI3K inhibitor-treated group（B4）.The number of CD4＋CD25＋Treg was detected by flow cytometry.The levels of IL-10 and IL-17 were measured by ELISA.The transcription levels of Foxp3 and RORγt were determined by RT-PCR.RESULTS： The purity of the T cells was（74.12±3.08）% and the survival rate was（92.82±3.21）% after isolation.The number of CD4＋CD25＋ Treg in group B1 was significantly lower than that in group A1,group B3 and group B4,and that in group A2 was significantly higher than that in group A1.The levels of IL-17 and mRNA expression of RORγt in B1 group were much higher than those in A1 group,those in A2 group were much lower than those in A1 group,and those in B2,B3 and B4 groups were lower than those in B1 group in a dose-dependent manner.However,the levels of IL-10 and mRNA expression of Foxp3 in B1 group were significantly lower than those in group A1,those in A2 group were significantly higher than those in group A1,and those in B2,B3,and B4 groups were much higher than those in B1 group in a dose-dependent manner.The ratio of RORγt/Foxp3 mRNA in B1 group was higher than that in A1 group,that in A2 grou

关 键 词：哮喘 PI3K信号通路 辅助性T细胞17 调节性T细胞 白细胞介素17 白细胞介素10 

分 类 号：R363[医药卫生—病理学]