哮喘儿童外周血单个核细胞Tim-3 mRNA的表达及其与CD4^+CD25^+调节性T细胞的关系

Tim-3 mRNA Expression in Peripheral Blood Mononuclear Cells and Its Relationship with CD4^+CD25^+ Regulatory T Cells from Asthmatic Children

目的观察哮喘患儿外周血单个核细胞(PBMC)中Tim-3 mRNA的表达及其与CD4+CD25+调节性T细胞(Treg)的关系,探讨Tim-3在哮喘发生发展中的作用。方法收集哮喘门诊或住院患儿73例,其中哮喘缓解期38例(缓解组),轻至中度急性发作期35例(发作组)。利用RT-PCR检测哮喘患儿PBMC中Tim-3 mRNA的表达并做半定量分析,流式细胞术检测PBMC中的CD4+CD25+Treg的水平(CD4+CD25+Treg占CD4+T细胞的百分比)。利用酶联免疫吸附试验(ELISA)检测血浆中白细胞介素6(IL-6)、转化生长因子β(TGF-β)的水平,并分析Tim-3 mRNA与CD4+CD25+TregI、L-6水平的相关性。结果哮喘发作组PBMC中Tim-3 mRNA吸光度值为(0.86±0.17),与缓解组(0.39±0.11)和正常对照组(0.06±0.03)比较,差异具有统计学意义(均P<0.05),并且缓解组与正常对照组比较差异亦具有统计学意义(P<0.05)。哮喘发作组外周血CD4+CD25+Treg百分率为(8.35±1.67)%,与缓解组[(10.21±2.04)%]和正常对照组[(12.43±2.58)%]比较,差异均具有统计学意义(P<0.05),并且缓解组与正常对照组比较差异亦具有统计学意义(P<0.05);哮喘发作组血浆中IL-6水平为(78.35±14.59)pg/mL,与缓解组[(36.48±9.18)pg/mL]和正常对照组[(10.24±3.57)pg/mL]比较,差异均具有统计学意义(P<0.05),缓解组与正常对照组比较差异亦有统计学意义(P<0.05)。3组血浆中TGF-β水平没有明显差异;哮喘发作组和缓解组PBMC中Tim-3 mRNA的表达水平与CD4+CD25+Treg百分率均呈负相关(r=-0.81,-0.79,均P<0.05),与IL-6的水平呈正相关(r=0.87,0.83,均P<0.01)。结论哮喘患儿PBMC中Tim-3 mRNA表达增高参与哮喘的发生与发展,其机制可能与升高血浆IL-6,抑制CD4+CD25+Treg的生成有关。

Objective To detect the expression of T cell immunoglobulin mucin 3（Tim-3）mRNA in peripheral blood mononuclear cells（PBMC）isolated from asthmatic children and analyze its relationship with CD4＋CD25＋ regulatory T cells（Treg）,explore elementarily the role of Tim-3 in the occurrence and development of asthmatic inflammation.Methods Seventy-three asthmatic children were enrolled in this study,including 35 children with acute asthma exacerbation（acute attack group）and 38 with asthma remission（remittent group）.The Tim-3 mRNA expression was detected by using RT-PCR.The ratio of CD4＋CD25＋ Treg（CD4＋CD25＋ Treg/CD4＋ T）was measured by using flow cytometry.The levels of interleukin-6（IL-6）and transforming growth factor-β（TGF-β）were determined by using ELISA.The correlation among Tim-3 mRNA,CD4＋CD25＋ Treg and IL-6 level was analyzed.Results The expression of Tim-3 mRNA in children with acute asthma exacerbation was（0.86±0.17）,significantly higher than that in remission stage and normal control group（0.39±0.11 and 0.06±0.03,both P0.05）.The expression of Tim-3 mRNA in children with asthma remission was significantly higher than that in normal control group（P0.05）.The percentage of CD4＋CD25＋ Treg in children with acute asthma exacerbation was（8.35±1.67）%,significantly lower than that in remission stage and normal control group [（10.21±2.04）% and（12.43±2.58）%,both P0.05].The level of IL-6 in children with acute asthma exacerbation was（78.35±14.59）pg/mL,significantly higher than that in remission stage and normal control group（36.48±9.18 and 10.24±3.57 pg/mL,both P0.05）.The level of IL-6 in children with asthma remission was significantly higher than that in normal control group（P0.05）.There was no significant difference in level of TGF-β among three groups.The Tim-3 mRNA expression in asthma exacerbation and remission stage was correlated negatively with the level of CD4＋CD25＋ Treg（r=-0.81,-0.79,both P0.05）,and positively with the level of IL-6（r=0.87,0.83,both P0.01）.Conclusion The increased expression of Tim-3 mRNA in PBMC might take part in the occurrence and development of asthmatic inflammation,which may be related to the increased level of IL-6 and inhibition of CD4＋CD25＋ Treg generation.