雷帕霉素对同种移植耐受模型CD4^+CD25^+ T细胞活性的影响

Effect of Rapamycin on CD4^+CD25^+ regulatory T cells in allo-transplantation tolerance model

目的研究雷帕霉素(Rapa)对同种移植耐受个体CD4+CD25+T细胞体内负免疫调节作用的影响。方法建立同种皮肤移植模型,受体鼠术前预输注供体鼠脾细胞,术后给予环孢素A(CsA)进行耐受诱导。移植后第14天提取耐受诱导模型鼠的T细胞,经不同浓度Rapa和/或IL-2体外处理后,混合淋巴细胞反应(MLR)确定T细胞特异增殖水平;流式细胞术(FCM)检测CD4+CD25+T细胞比例变化;RT-PCR检测Foxp3mRNA表达情况;ELISA检测细胞培养不同时间后上清中IL-10的变化。然后将Rapa和/或IL-2处理的T细胞过继转移给同种移植后的BALB/c-SCID鼠,观察移植物存活状态。结果CsA加供体脾细胞预先注射可明显延长小鼠移植皮片的存活期(P<0.05);移植耐受状态的T细胞经Rapa和/或IL-2体外处理后CD4+CD25+T细胞比例升高、增殖水平明显降低、Foxp3表达量明显增加;过继转输给同种移植SCID鼠后,其移植皮片存活时间显著延长(P<0.05)。结论Rapa可体外扩增耐受诱导模型中CD4+CD25+T细胞,使CD4+CD25+T细胞相关的Foxp3和IL-10明显升高,过继免疫后,小鼠同种移植物存活时间明显延长,而低浓度IL-2可以协同Rapa的这一作用。

AIM： To study the effect of Rapamycin（Rapa） on CD4^＋CD25^＋ regulatory T cells in allo-transplantation tolerance model. METHODS： The model of skin allo-transplantation was established, The recipient BALB/c mice were injected with allogeneic donor spleen cells from 136 on the day before grafting and with cyclosporine（CsA） after transplantation. T cells were purified on the day 14 from the tolerant group and cultured with Rapa and/or IL-2 in different concentrations in vitro. Mixed lymphocyte reaction （MLR） was used to analyze the specific recall reaction of T cells. Percentages of CD4^＋CD25^＋ regulatory T cells were examined by flow cytometry （FCM）. The expression of Foxp3 mRNA was measured by RT-PCR, and the level of IL-10 in supernatant of T cells cultured in vitro was determined by ELISA. BALB/c-SCID mice underwent allo-transplantation were given adoptive transfer of T cells treated with Rapa and/or IL-2, then the survival conditions of the grafted skin were observed daily. RESULTS： CsA plus donor splenocyte injection can prolong allograft of BALB/c significantly （P〈0.05）. The number of CD4^＋CD25^＋ regulatory T cells and the expression of Foxp3 mRNA for T cell in the tolerant group treated with Rapa and/or IL-2 were obviously increased. Adoptive transfusion of T cells from tolerant mice treated with IL-2/Rapa obviously prolonged the allograft survival time in allografted-SCID mice. CONCLUSION： Rapa can increased the ratio of CD4^＋CD25^＋ regulatory T cells in vitro and prolonged the graft survival time obviously after adoptive immunity, and these effects are enhanced by low-dose of IL-2.