抗TCRαβ单抗联合供体骨髓细胞输注诱导小鼠皮肤移植耐受的研究

Study on the Induction of Skin Transplantation Tolerancein Mice by Anti-TCRαβ mAb Combined with Donor Bone Marrow Cells Infusion

探讨抗TCRαβ单克隆抗体联合大剂量供体骨髓细胞输注方法对同种异基因小鼠皮肤移植耐受诱导的促进作用。第 0天 ,C5 7BL/ 6 (H 2 b,B6 )小鼠尾静脉注入 2× 10 8BALB/c (H 2 d,B/c )来源的骨髓细胞 ,同时腹腔注射抗TCRαβ单克隆抗体5 0 0 μg ;第 6天进行皮肤移植 ;在不同的时间对B6小鼠进行迟发型超敏反应 (DTH )、混合淋巴细胞反应 (MLR )等耐受状态进行检测 ,并进行MLR的IL 2逆转实验、过继转移实验和嵌合状态分析以初步探讨耐受形成机制。结果显示 ,经抗TCRαβ单克隆抗体联合大剂量供体骨髓细胞输注处理的B6小鼠的供体皮肤移植物平均存活 5 0 4d ,显著长于其他各组 (P <0 0 0 1)。相对于正常对照组小鼠 ,耐受B6小鼠在DTH和MLR中均表现出显著的低反应性 (P <0 0 0 1)。IL 2逆转实验结果表明克隆无能参与了移植耐受的形成。体内、外过继转移实验均显示耐受B6小鼠脾细胞中存在抑制细胞活性。嵌合体检查结果表明在耐受B6小鼠的胸腺和脾脏中均形成了混合嵌合体 ,在皮肤移植后第 15、 30和 70天时脾脏内供体来源嵌合体水平依次为 15 86 % ,10 5 7%和 1 77% ,胸腺中依次为 8 19% ,5 72 %和 1 87% ,嵌合体水平随时间而下降。这些表明 ,抗TCRαβ单抗联合大剂量供体骨髓细胞输注可以在异基因成年小?

To explore the role of anti-TCRαβ monoclonal antibody combined with high dose of donor bone marrow cells infusion in the induction of murine skin allografts tolerance. 2×10 8 bone marrow cells(BMC) Of BALB/c mice (H-2 d) were injected into recipient C57BL/6( H-2 b,B6) mice via the tail vein on day 0,meanwhile,an intraperitoneal injection of anti-TCRαβ monoclonal antibody(500 μg)was given.Skin grafting was performed on day 6 and DTH, MLR,IL-2 reverse assay of MLR, adoptive transfer assay and chimerism were performed at different time points to investigate the mechanism of tolerance. It was found that the mean survivaltime (MST) of BALB/c skin allografts in B6 recipients that were treated by anti-TCRαβ monoclonal antibody combined with high dose of donor bone marrow cells infusion was 50.4 days,which was significantly longer than that of other groups.DTH and MLR assay indicated that the tolerant mice displayed significant hyporesponsivness against donor antigen(P<0.001).The result of IL-2 reverse test showed that clone anergy was probably involved in the' formation of tolerance in the tolerant B6 mice.In vivo and in vitro adoptive transfer assay,suppressive activity in the spleens of tolerant B6 mice was observed.Chimerism exists in both the thymus and spleen of the tolerant B6 mice.The percentage of cells of BALB/c origin in the spleen of B6 mice in the tolerant mice was 15.86%,10.57%and 1.77% on day 15,30 and 70 after skin grafting,and in the thymus was 8.19%,5.72% and 1.87%,respectively.The chimerism level gradually declined with time.It concludes that treatment of anti-TCRαβ monoclonal antibody combined with high dose of donor bone marrow cells infusion can successfully induce a long-term tolerancein BALB/c mice to B6 skin graft and multiple mechanisms,including clone anergy,suppressor cells and chimerism were involved in the tolerance induction.