供体骨髓细胞输注促进小鼠皮肤移植耐受及其机理的研究

Improvement of transplantation tolerance of skin allograft by infusion of donor bone marrow cells and its mechanism

目的 探索“细胞 +环磷酰胺 (cyclophosphamide ,CP)”系统联合抗H 2 b 单克隆抗体、供体骨髓细胞输注诱导移植耐受的作用及其机制。方法 第 0天 ,经尾静脉给C5 7BL/ 6 (H 2 b,B6 )小鼠注入 10 8BALB/c(H 2 d,B/c)来源的脾细胞 ,第 2天 ,腹腔注射环磷酰胺 2 0 0mg/kg ,第 3、5天 ,分别经尾静脉注入抗H 2 b 单抗 ,剂量为 40 0 μg/ 0 .5ml,第 8天进行皮肤移植。皮肤移植后 1周 (第 15天 ) ,输入 2× 10 7供体BALB/c来源的骨髓细胞。观察皮肤移植物存活时间 ,并于第 30天对耐受B6小鼠作混合淋巴细胞反应 (mixedlymphocytereaction ,MLR) ,迟发型超敏反应 (delayedtypehypersensitivity ,DTH)等确定耐受的状态。并通过过继转移实验、嵌合体检查及脾细胞中细胞因子mRNA的表达情况 ,进一步探讨耐受形成的机制。结果 采用此诱导方案 ,B6小鼠对BALB/c小鼠的皮肤移植物长期存活。耐受可以被过继转移 ;耐受小鼠胸腺内的嵌合程度与耐受的维持密切相关 ;TH1型细胞因子在耐受小鼠中明显降低 ,而TH2型细胞因子明显升高。结论 “细胞 +CP”系统联合H 2 b 单克隆抗体、供体骨髓细胞输注能够诱导异基因小鼠皮肤移植耐受。耐受机制与胸腺嵌合体的存在密切相关。克隆无能 (aner gy)、抑制细胞和TH1/TH2偏移在耐受中也?

Objective To investigate the effect and mechanisms of transplantation tolerance induced by 'spleen cell(SC)+cyclophosphamide(CP)' protocol combined with anti-H2b monoclonal antibody and infusion of donor bone marrow cells. Methods C57BL/6(H-2b, B6) mice were given an intravenous infusion of 108 allogenic spleen cells from BALB/c (H-2d, B/c) on day 0, and intraperitoneal infusion of CP (200mg/kg) on day 2. On day 3 and 5, two doses of anti-H2b monoclonal antibody (400μg/0.5ml) were respectively given to B6 mice. Skin allograft was grafted on day 8. One week later (day 15), donor B/c bone marrow cells were intravenously infused to B6 mice. 30 days after induction of tolerance the MLR and DTH of tolerance B6 mice were examined to confirm the tolerance status. Chimeric analysis was used to explore tolerance mechanisms. Adoptive transfer assay and analysis of cytokine mRNA expression in tolerant mice thymus were performed to further explore tolerance mechanisms. Results Long-lasting skin allograft tolerance was achieved. The chimeric degrees of tolerant mice decreased with the time and were positively correlated with the tolerance status, indicating that the chimerism was substantially involved in the long-lasting skin allograft tolerance. The levels of IL-2 and IFN-γ mRNA expression in tolerant B6 mice were much lower than those in the control mice. The levels of IL-4 and IL-10 mRNA in tolerant mice were much higher. Conclusions Skin allograft tolerance of B6 mice induced by 'SC+CP' protocol combined with anti-H-2b monoclonal antibody can be significantly improved by the infusion of donor bone marrow cells. The chimeric degrees in the thymus of tolerant mice were positively correlated with the tolerance status. Clonal anergy, suppressor cells and deviation of T H1/T H2 cytokines were involved in the process of tolerance. [