【摘要】 目的 :研究与系统性红斑狼疮(systemic lupus erythematosus,SLE)患者调节性T细胞(regulatory T cell,Treg)功能密切相关的分子转化生长因子(transforming growth factor,TGF-β)的转录、表达、分泌水平的异常。方法:分离正常人和SLE患者的外周血单个核细胞(peripheral blood mononuclear cell,PBMC)和CD4+CD25+T细胞,以抗-CD3和抗-CD28刺激培养48 h,获取细胞,以三色流式细胞术检测CD4+CD25+LAP+T及CD8+CD25+LAP+T细胞,分析各亚群细胞比值;提取总m RNA,q RT-PCR测定TGF-β1的表达;收取上清用ELISA法检测总TGF-β1和游离的活化TGF-β1水平。结果 :活动性SLE患者CD4+T细胞中LAP+/CD4+T、CD25+LAP+/CD4+T、LAP+/CD4+CD25+比值较正常人显著升高,抗-CD3、抗-CD28抗体刺激后LAP+/CD4+T、CD25+LAP+/CD4+T、LAP+/CD4+CD25+比值进一步升高,与正常人比较差异有统计学意义(P<0.05);但均与疾病活动度无显著相关性;无论刺激与未刺激,SLE患者CD4+CD25+LAP+细胞中LAP染色的荧光强度亦高于相应正常组;而抗-CD3、抗-CD28抗体刺激后SLE患者PBMC及CD4+CD25+T细胞培养上清中总TGF-β1和游离的活化TGF-β1水平均显著低于正常人(P<0.05);q RT-PCR证实,受刺激后CD4+CD25+T细胞内TGF-β1的m RNA转录水平低于正常人。结论:尽管活动性SLE患者CD4+CD25+T细胞膜表面潜伏态TGF-β(LAP)的表达较正常人明显增高,但其TGF-β表达依然存在缺陷,主要表现为TGF-β1的转录、分泌水平降低,游离的活化TGF-β1产生障碍,从而可能削弱了Treg细胞的免疫抑制作用。新鲜分离的以及受刺激后的CD4+CD25+T细胞LAP表达增高可能反映了活动性SLE患者T细胞亚群的活化和CD4+CD25+LAP+Treg细胞的反应性扩增。更多还原

【Abstract】 Objective:To investigate the possible deficiencies in the transcription,secretion,and expression of regulatory T cells(Treg) associated TGF-β in patients with systemic lupus erythematosus(SLE). Methods:Peripheral blood mononuclear cells(PBMC)and CD4+CD25+T cells were isolated from patients with SLE and health controls. The cells were cultured and stimulated with antiCD3 and anti-CD28 for 48 hours. Three-color flow cytometry was performed to detect CD4+CD25+LAP+T and CD8+CD25+LAP+T cells;Q-RT-PCR was used to detect TGF-β m RNA level; ELISA was performed to detect the levels of total TGF-β1 and free active TGF-β1in the supernatants of the cell cultures. Results:The ratios of LAP+/CD4+T,CD25+LAP+/CD4+T and LAP+/CD4+CD25+in fresh PBMCs,or in cultured cells stimulated with anti-CD3 and anti-CD28,from active SLE patients were both significantly higher than those from health controls(P < 0.05),but there were no significantly correlations between the abnormally increased LAP expression and SLE disease activity index(SLEDAI). Whether stimulated or not,mean fluorescent intensity(MFI) of LAP in CD4+CD25+LAP+cells from patients with SLE was also higher than that from health controls(P < 0.05). With the stimulation,CD4+CD25+T cells from the patients secreted less TGF-β1,both in the forms of total TGF-β1 and free active TGF-β1(P < 0.05). Furthermore,CD4+CD25+T cells from active SLE patients displayed less TGF-β m RNA level than those from health controls did. Conclusion:In CD4+CD25+T cells from SLE,there are deficiencies in the transcription and secretion of TGF-β1,and in the production of free active TGF-β1,although the expression of LAP on the cell surface is increased,which may reflect the abnormal activation of T cells and a proliferative respond of CD4+CD25+LAP+Treg to auto-antigens. These deficiencies in TGF-β expression of CD4+CD25+T cells in active SLEpatients probably weaken the function of Treg cells.更多还原