CD40信号介导同种反应中CD4^+T细胞生物学功能的研究

The Study on CD40 Signal that Induces Alloantigen Responsiveness of CD4^+T Cells by ex vivo Mixed Reaction

目的探讨CD40信号在血管内皮细胞(ECV)与人外周血CD4+T细胞体外共培养的同种反应中的生物学功能。方法采用免疫磁珠阴性选择法分离获得人外周血CD4+T细胞,将纯化的CD4+T细胞与高表达CD40分子的ECV体外共培养,同时应用功能型CD154单克隆抗体阻断CD40信号,通过检测不同时间点的共培养上清IL-2、IFN-γ水平及CD4+T细胞的增殖来评价CD40信号在同种反应中的生物学效应。结果CD4+T细胞与ECV共培养组上清中的IL-2和IFN-γ水平高于含有CD154单克隆抗体的阻断效应组,差异有统计学意义(P<0.05);此外,CD154单克隆抗体通过阻断CD40共信号,能有效地抑制CD4+T细胞活化,并使之对同种抗原的刺激,维持在较低的应答水平,在共培养的第6天差异最为显著(P<0.05)。结论CD40信号参与ECV介导的CD4+T细胞分泌IL-2和IFN-γ,并介导CD4+T细胞对同种抗原的免疫反应;CD154单克隆抗体等多种干预CD40信号的生物制剂可能在移植排斥的免疫负性调节中具有潜在的应用价值。

Objective To investigate the biological effects of CD40 signal in inducement of CD4^＋T ceils to alloantigen responsiveness. Methods Purified CD4^＋^ ceils and ECV were co-cultured with functional anti-CD154 mAb ex vivo. The cytokine levels of IL-2 and IFN-γ in the supernatant of cocultures were detected by ELISA. The proliferation of CD4^＋T cells stimulated by alloantigen was determined on a beta scintillation counter. Results On the basis of cytokine level and proliferation of CD4^＋ T ceils detections, it was evidenced that the blocking of CD40 signal with CD154 mAb inhibited the proliferation of CD4^＋T ceils co-cultured with ECV. Moreover the IL-2 and IFN-γ secretion of CD4^＋T ceils mainly depended on CD40 pathway. Conclusion The results demonstrate that the CD40-CD154 signal played an important role in the immuno-modulation of hypo-responsiveness to alloantigen stimulation. The means interfering with CD40 signal shows a potential application in basic and clinical transplantation rejection study.