系统性红斑狼疮患者外周血淋巴细胞CD28/CTLA-4分子表达的研究

CD28 and CTLA-4 expressions in peripheral blood T cells in patients with systemic lupus erythematosus

目的探讨系统性红斑狼疮(systemic lupus erythematosus,SLE)患者外周血淋巴细胞CD28/CTLA-4分子和CD28/CTLA-4mRNA的表达。方法研究对象为SLE患者49例(活跃期30例、缓解期19例)及对照组23例。外周血单个核细胞(peripheral blood mononuclear cell,PBMCs)经梯度密度离心法分离后分佛波醇乙酯(PMA)(10ng/ml)及伊屋诺霉素(500ng/ml)刺激组和不加刺激剂组两组培养。将PBMCs分别培养24、48、72及96小时。应用流式细胞计量术(flow cytometry,FCM)检测外周血淋巴细胞培养前后CD28及CTLA-4分子的表达。采用RT-PCR方法检测CD28mRNA和CTLA-4mRNA的表达。结果刺激前后SLE患者CD3+及CD8+T细胞上CD28分子表达量与对照组比较差异均无统计学意义(P>0.05)。刺激前活跃期SLE患者CD3+T细胞上CTLA-4分子表达量较对照组明显降低[(0.78±0.51)%vs(1.34±0.76)%,P<0.05]。刺激后72小时SLE患者CD3+T细胞及CD8+T细胞上CTLA-4分子表达量仍低于对照组,但差异无统计学意义(P>0.05)。刺激前后PBMCs中CD28mRNA及CTLA-4mRNA表达情况与刺激前后CD3+T细胞上CD28及CTLA-4分子变化情况相似。刺激前SLE患者CD3+T细胞上CTLA-4分子表达量与SLE活动指数(SLEDAI)呈显著的直线负相关关系(P<0.001)。结论 SLE患者T细胞CTLA-4分子存在表达及上调机制障碍,提示CD28分子及CTLA-4分子存在功能失衡,这种失衡可能通过一定的机制参与SLE的发病过程。

Objective To investigate the expressions of CD28/CTLA-4 molecules in peripheral blood T cells and their CD28/CTLA-4mRNA in patients with systemic lupus erythematosus（SLE）.Methods Forty-nine SLE patients were classified into two groups： active stage group（30 cases）and paracmasis stage group（19 cases）.Twenty-three normal individuals served as controls.After isolated by density gradient centrifugation,peripheral blood mononuclear cells（PBMCs）were cultured in two subgroups for 24h,48h,72h and 96h respectively.One was PMA（5 ng/ml）and ionomycin（500 ng/ml）group,the other was control group.Before and after culture,PBMCs were collected.The expressions of CD28 and CTLA-4 molecules in T cell were detected using flow cytometry（FCM）instruments after double staining by FITC or PE labeled monoclonal antibodies.The CD28/CTLA-4mRNA levels were assayed by reverse transcription-polymerase chain reaction（RT-PCR）.Results Either before or after stimulation by PMA and ionomycin,the expressions of CD28 molecules in CD3＋ and CD8＋ T cells in all SLE patients were not significantly different from those in controls（P0.05）.Before stimulation,the expression of CTLA-4 molecule in CD3＋T cells in active SLE patients（0.78±0.51）%was significantly lower than those in controls（1.34±0.76）%（P0.05）.The patterns of CD28/CTLA-4mRNA expressions in PBMCs were similar to those of CD28/CTLA-4 molecules in CD3＋ T cells.After stimulation,the expression of CTLA-4 molecules in T cells in SLE patients was up-regulated in some extent,but the amplitude of up-regulating was lower than that in normal controls at each time point.After stimulation,the expressions of CTLA-4 molecules in CD3＋ T cells in SLE patients were still lower than those in normal controls without statistical significance（P0.05）.The CTLA-4 molecule expression in CD3＋ T cells,before stimulation,was negatively correlated with SLEDAI（r=-0.641,P0.001）.Conclusions There is no defect in expression and up-regulation of CD28 molecule in T cell,while CTLA-4 molecule has a defective expression and up-regulation mechanism.This imbalance may induce functional disorders of costimulatory molecules and play an important role in the pathogenesis of SLE.