Establishment of drug-induced tolerance for heart transplantation in large animals

最近，我们已经描述了一种药物（环磷酰胺（CP）加白消安（BU））诱导的小鼠皮肤移植耐受，cagularly克服完全H-2不匹配的障碍。利用这种方法，我们研究了这种方案是否可以延长心脏移植物的存活时间和抑制移植后心脏移植物血管病变（CAV）的发展。该方法的组成部分是在第0天静脉内给予1x 108个同种异体脾细胞，在第2天腹膜内给予200 mg/kg CP和30 mg/kg BU，在第3天在同一品系小鼠中静脉内注射去除T细胞的1x 107个同种异体骨髓细胞。在第28天进行心脏移植（HG）。流式细胞术检测外周血嵌合体，并通过心脏移植和皮肤移植的特异性接受来评估移植耐受性。通过FCM确定某些Vb家族的频率以评估供体反应性T细胞的缺失。H ...更多信息 在移植后的不同时间进行I al分析（Elastica货车Gieson）。采用RT-PCR方法检测移植心脏Th 1（IL-2、IFN-γ）和Th 2（IL-4、IL-10）细胞表达。在B10.D2（H-2d，IE+）→B10（H-2b，IE-）的完全MHC错配组合中，永久观察到稳定的多谱系混合嵌合体，受体B10小鼠外周中IE反应性Vb 11 + T细胞特异性减少。B10.D2移植物以供体特异性方式被永久接受，并且移植后CAV没有发展。移植耐受的诱导通过特异性接受供体品系的第二套皮肤移植来证实。供体B10.D2心脏移植后Th 1（IL-2、IFN-γ）和Th 2（IL-4、IL-10）细胞因子均无蓄积。这些结果表明，我们最近建立的药物诱导的耐受可以规律地诱导长g心脏移植耐受，而不发生CAV或移植物内Th 1或Th 2的mRNA积累。最近，我们描述了一种药物（环磷酰胺（CP）+白消安（BU））诱导的小鼠皮肤移植耐受，其可以完全克服H-2错配障碍。本研究旨在探讨该方案是否能延长移植心脏的存活时间和抑制移植后移植心脏血管病变（cardiac allograft vasculopathy，CAV）的发生。该方法的组成部分是在第0天静脉内给予1x 108个同种异体脾细胞，在第2天腹膜内给予200 mg/kg CP和30 mg/kg BU，在第3天在同一品系小鼠中静脉内注射去除T细胞的1x 107个同种异体骨髓细胞。在第28天进行心脏移植（HG）。流式细胞术检测外周血嵌合体，并通过心脏移植和皮肤移植的特异性接受来评估移植耐受性。通过FCM确定某些Vb家族的频率以评估供体反应性T细胞的缺失。在移植后的不同时间进行HI分析（弹性货车Gieson）。采用RT-PCR方法检测移植心脏Th 1（IL-2、IFN-γ）和Th 2（IL-4、IL-10）细胞表达。在B10.D2（H-2d，IE+）→B10（H-2b，IE-）的完全MHC错配组合中，永久观察到稳定的多谱系混合嵌合体，受体B10小鼠外周中IE反应性Vb 11 + T细胞特异性减少。B10.D2移植物以供体特异性方式被永久接受，并且移植后CAV没有发展。移植耐受的诱导通过特异性接受供体品系的第二套皮肤移植来证实。供体B10.D2心脏移植后Th 1（IL-2、IFN-γ）和Th 2（IL-4、IL-10）细胞因子均无蓄积。这些结果表明，我们最近建立的药物诱导的耐受可以规律地诱导长g心脏移植耐受，而不发生CAV或移植物内Th 1或Th 2的mRNA积累。少

Recently, we have described a drug (cyclophosphamide (CP) plus busulfan (BU))-induced skin allograft tolerance in mice that cagularly overcome fully H-2 mismatched barriers. Using this method, we have investigated whether or not this regimen can prolong survival of heart allografts and inhibit the development of post-transplant cardiac allograft vasculopathy (CAV).Method. The components of the method are intravenous administration of 1x108 allogeneic spleen cells on day 0, intraperitoneaction of 200 mg/kg CP and 30 mg/kg BU on day 2, and intravenous injection of T cell-depleted 1x107 allogeneic bone marrow cells in the same strain of mice on day 3. Heart grafting (HG) was performed on day 28. Chimerism in peripheral blood was followed by cytometric (FCM) analysis, and the transplantation tolerance was assessed by specific acceptance of heart grafting followed by set-skin grafting. The frequency of certain Vb families was determined by FCM to assess deletion of donor-reactive T cells. H … More i al analysis (elastica van Gieson) was performed at various timings after grafting. Th1 (IL-2, IFN-γ) and Th2 (IL-4, IL-10) cytopressions in the heart grafts were analyzed with RT-PCR.Results. In a fully MHC mismatched combination of B10.D2 (H-2d, IE+)→B10 (H-2b, IE-), stable multilineage mixed chimerism was observed permanently, and IE-reactive Vb11 + T cells were specifically reduced in the periphery in the recipient B10 mice. B10.D2 grafts were accepted permanently in a donor specific manner, and post-transplant CAV did not develop. Induction of the transplan tolerance was confirmed by specific acceptance of second set-skin grafting from donor strain. In the donor B10.D2 heart grafter Th1 (IL-2, IFN-γ) nor Th2 (IL-4, IL-10) cytokine was not accumulated.Conclusions. These results demonstrated that the drug-induced tolerance recently established by us can regularly induce a long-g heart allograft tolerance without development CAV or intragraft mRNA accumulation of Th1 or Th2.Recently, we have described a drug (cyclophosphamide (CP) plus busulfan (BU))-induced skin allograft tolerance in mice that can ularly overcome fully H-2 mismatched barriers. Using this method, we have investigated whether or not this regimen can prolong urvival of heart allografts and inhibit the development of post-transplant cardiac allograft vasculopathy (CAV).Methods. The components of the method are intravenous administration of 1x108 allogeneic spleen cells on day 0, intraperitoneaction of 200 mg/kg CP and 30 mg/kg BU on day 2, and intravenous injection of T cell-depleted 1x107 allogeneic bone marrow cells in the same strain of mice on day 3. Heart grafting (HG) was performed on day 28. Chimerism in peripheral blood was followed by cytometric (FCM) analysis, and the transplantation tolerance was assessed by specific acceptance of heart grafting followed by set-skin grafting. The frequency of certain Vb families was determined by FCM to assess deletion of donor-reactive T cells. Hi al analysis (elastica van Gieson) was performed at various timings after grafting. Th1 (IL-2, IFN-γ) and Th2 (IL-4, IL-10) cytopressions in the heart grafts were analyzed with RT-PCR.Results. In a fully MHC mismatched combination of B10.D2 (H-2d, IE+)→B10 (H-2b, IE-), stable multilineage mixed chimerism was observed permanently, and IE-reactive Vb11+ T cells were specifically reduced in the periphery in the recipient B10 mice. B10.D2 grafts were accepted permanently in a donor specific manner, and post-transplant CAV did not develop. Induction of the transplan tolerance was confirmed by specific acceptance of second set-skin grafting from donor strain. In the donor B10.D2 heart grafter Th1 (IL-2, IFN-γ) nor Th2 (IL-4, IL-10) cytokine was not accumulated.Conclusions. These results demonstrated that the drug-induced tolerance recently established by us can regularly induce a long-g heart allograft tolerance without development CAV or intragraft mRNA accumulation of Th1 or Th2. Less