体外阻断CD_(40)-CD_(40)L共刺激途径减轻小鼠异基因骨髓移植中移植物抗宿主病的研究  

Study of reducing graft-versus-host disease by in vitro blockade of CD_(40) -CD_(40) ligand co-stimulatory pathway in allogeneic bone marrow transplantation mouse model

作　　者：黄绍良[1] 陈纯[1] 段连宁[2] 李浩威[2] 温冠媚[2] 李琳[3] 詹美意[4] 魏菁[5] 

机构地区：[1]中山大学附属第二医院儿科,广州510120 [2]中山大学中山医学院病理生理教研室 [3]广东省人民医院放疗科 [4]暨南大学医学院病理教研室 [5]中山大学附属第二医院医学研究中心

出　　处：《中华血液学杂志》2003年第6期290-294,共5页Chinese Journal of Hematology

摘　　要：目的　在异基因骨髓移植移植物抗宿主病 (GVHD)小鼠模型中 ,观察抗CD4 0 L单克隆抗体 (单抗 )体外预处理供鼠T细胞输注 ,观察其减轻GVHD的作用并探讨其作用机制。方法　供鼠(C5 7BL 6 H 2b)脾T细胞作为反应细胞 ,受鼠 (BALB cH 2d)脾细胞作为刺激细胞 ,分别加抗CD4 0 L单抗或不加单抗进行混合培养 ,培养第 5天的细胞作为经体外诱导后的脾T淋巴细胞 ,分别混合供鼠骨髓细胞后移植给接受 8.0Gy全身照射预处理的受鼠。比较受鼠GVHD发生和造血重建。在移植后不同时间点采用流式细胞仪检测受鼠脾细胞中T细胞表型的改变 ,用ELISA法检测外周血血清中Th1和Th2类细胞因子的水平。结果　移植后对照组受鼠均于 2 5d内死于GVHD ,实验组GVHD的发生率为2 0 % ,与对照组小鼠相比 ,存活率和存活时间明显增高和延长 (P <0 0 1) ;存活的实验组小鼠 (8只 )第4 0天时骨髓细胞中H 2Db 阳性细胞为 (93.5 4± 2 .32 ) %。实验组小鼠CD3+ CD4+ 、CD3+ CD8+ 、CD4+CD2 5+ 、CD4+ CD6 9+ 、CD8+ CD2 5+ 、CD4+ CD4 0 L+ 和CD8+ CD6 9+ T细胞比例明显低于对照组 (P <0 .0 5 ) ,CD8+ CD4 0 L+ 和CD4+ CD4 5RA+ T细胞比例在两组中变化差异无显著性 (P >0 .0 5 )。实验组受鼠血清中细胞因子水平明显低于对照组 (P <0 .0 5 )。结论　应?Objective To investigate the effect and its mechanism of reducing graft-versus-host disease (GVHD) by in vitro blockade of CD 40 -CD 40 L pathway in vitro, the donor T lymphocytes cultured in vitro with anti-CD 40 L mAb were transfused in bone marrow transplantation (BMT) GVHD mouse model. Methods C57BL/6 H-2b spleen T cells were isolated as responder cells, and BALB/c H-2d spleen cells as stimulator cells. They were cocultured with or without Anti-CD 40 L mAb as anti-CD 40 L mAb group and control group, respectively. At day 5, the mixed lymphocyte response(MLR)-culture cells mixed with bone marrow cells and transfused respectively into the TBI conditioned recipient mice. The mice were divided into two groups: group A, bone marrow cells(2×10 6) and spleen T lymphcytes(2×10 6) from MLR control group; group B, bone marrow cells(2×10 6) and spleen T lymphcytes(2×10 6) from MLR anti-CD 40 L mAb group. The GVHD incidence and hematopoietic reconstitution were observed. Peripheral blood sera and spleen cells of the recipients mice were harvested at scheduled time points for the measurement of cytokines and T cell immunophenotyping with flow cytometry. Results The incidence of GVHD in group A was 100%(10/10), and in group B was 20%(2/10). The percentage of H-2D b positive cells in group B (n=8) was (93.54±2.32)% at day 40 after transplantation. The levels of cytokines in serum from group B were significantly lower than those from group A (P<0.05). The expressions of CD 4 +, CD 8 +, CD 4 +CD 25+, CD 8 +CD 25+, CD 4 +CD 69+, CD 8 +CD 69+ and CD 4 +CD 40 L + were lower in group B than in group A (P<0.05). The expressions of CD 8 +CD 40 L + and CD 4 +CD 45RA+ were similar in the two groups(P>0.05). Conclusion Blockade of CD 40 -CD 40 L interaction in vitro could induce immune tolerance in vivo, reduce aGVHD in aGVHD mice model and form chimerism, which was mediated by inhibiting the Th1 and Th2 cytokines produc

关 键 词：体外阻断 CD40-CD40L 异基因 骨髓移植 移植物抗宿主病 

分 类 号：R392.1[医药卫生—免疫学]