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目的建立稳定的大鼠原位肝移植模型,并检测大鼠外周血中CD4+CD25+调节性T细胞(regulatory T cells,Treg)比例的变化。方法采用近交系成年雄性BN、LEW大鼠,改良“二袖套”法建立肝移植模型31例。术前随机分为3组:排斥组(LEW→BN,n=9),耐受组(BN→LEW,n=11),同基因组(BN→BN,n=11)。存活的受体分别于术后5 d、10 d、30 d取外周血,分离外周血中的单个核细胞,由流式细胞仪检测CD4+CD25+Treg比例。并于相应时间点每组随机各处死2只,取肝脏标本观察病理学变化。每组选取5例来观察生存期,存活>100 d时各处死2只,取肝脏标本。结果中位生存期:排斥组14 d,耐受组112 d,同基因组129 d。耐受组与同基因组术后未见明显排斥反应。术后7 d,排斥组肝脏病理Banff评分Ⅲ级。肝移植早期,排斥组和自发耐受组的外周血中CD4+CD25+Treg比例均增加,术后10 d耐受组CD4+CD25+Treg比例明显高于同基因组(P<0.01)。术后30 d,耐受组CD4+CD25+Treg比例仍维持在较高水平。结论 BN与LEW大鼠组合可建立稳定可靠的急性排斥模型与自发耐受模型。CD4+CD25+Treg可能参与自发免疫耐受的形成。
Objective To establish a stable orthotopic liver transplantation model in rats and detect the ratio of CD4 + CD25 + regulatory T cells (Tregs) in peripheral blood of rats. Methods Inbred male adult BN and LEW rats were used to establish 31 liver transplantation models. The patients were randomly divided into three groups: rejection group (LEW → BN, n = 9), tolerated group (BN → LEW, n = 11) and isobaric group (BN → BN, n = 11). Peripheral blood was collected from the surviving recipients at 5 d, 10 d and 30 d after operation, respectively. Peripheral blood mononuclear cells (PBMCs) were isolated and the ratio of CD4 + CD25 + Tregs was detected by flow cytometry. At the corresponding time point, each group randomly divided into two deaths, take the liver specimens observed pathological changes. Five patients in each group were selected to observe the survival time, survival of> 100 d each killed two, take the liver specimens. Results The median survival time was 14 d in the rejection group, 112 d in the tolerated group and 129 d in the same genomic group. There was no significant rejection in the tolerated and syngeneic groups. At 7 days after operation, the pathological grade of the liver in the rejection group was grade Ⅲ. The proportion of CD4 + CD25 + Treg in peripheral blood of rejection group and spontaneous tolerance group increased at the early stage of liver transplantation. The proportion of CD4 + CD25 + Treg in the tolerated group was significantly higher than that in the same genotype at 10 days after transplantation (P <0.01). After 30 days, the proportion of CD4 + CD25 + Treg in resistant group remained at a high level. Conclusion BN combined with LEW rats can establish a stable and reliable acute rejection model and spontaneous tolerance model. CD4 + CD25 + Treg may be involved in the formation of spontaneous immune tolerance.