大鼠原位肝移植急性排斥反应模型的建立及技术改进

EastabHshment and surgical renovations of acute rejection model in rat orthotopic liver transplantation

目的建立大鼠原位肝脏移植的急性排斥模型并观察其排斥反应的特点。方法选择近交系雄性DA大鼠为供体，Lewis大鼠为受体，改良“二袖套”法建立肝移植模型48例，肝动脉重建采用“袖管式微血管缝合法”（ microvascular sleeve anatomosis） 。受体随机分为两组，A组（非干预组，n=24）：术后不用免疫抑制剂；B组（FK506处理组，n=24）：术后每日用FK506每日0．2mg／kg体重灌胃。分别于术后3、5、7d随机解剖受体大鼠6只，取材肝组织对移植肝进行病理学观察，同时静脉采血测定血清谷丙转氨酶（ALT）、白蛋白（Alb）和总胆红素（TBIL）浓度；每组各留6只观察其生存时间和死亡原因。结果本方法建立模型的手术时间（包括供体手术）和重建动脉的时间明显缩短，手术成功率和肝动脉的通畅率均为100％。DA→Lewis组合的模型术后的血清生化指标、肝脏病理学改变、生存情况均符合肝移植急性排斥的特点，排斥反应于术后3～5d出现，并逐渐增强，术后7d迅速达到高峰并维持；该模型可以为抗排斥药物FK506预防而达到长期存活。A、B两组中位生存时间分别为14、73d；A、B两组的累积生存率曲线差异有统计学意义（Log—Rank取值为11．78，P〈0．01）。结论DA到Lewis大鼠品系组合可以建立稳定可靠的肝移植急性排斥模型；该模型具有研究周期短、病理改变典型、可重复性好等优点。

Objective To establish acute rejection （AR） model in rat orthotopic liver transplantation （OLT） and observe the characteristics of AR of liver grafts from inbred rats. Methods Forty-eight OLT models were established by modified two-cuff technique involving performing a simple sleeve anastomosis for graft arterialization,with inbred male DA rats as the donors and inbred male Lewis rats as the recipients. The recipients were randomly divided into 2 groups：non-intervention group （n = 24） without immunosuppression and FKS06 treatment group （n =24） lavaged with 0.2 mg/ （kg · d） FKS06.6. Recipients were sacrificed respectively on the day 3,5,7 after OLT to momitor the changes of serum biochemistry including alanine transaminase （ALT）, total bilirubin （TBIL） and albumin （ALB） and grafts histology. Six recipients were kept in each group to observe the survival time. Results The total operative time and in particular the time needed to conduct the arterial anastomosis was significantly reduced with the use of the two-cuff and arterial sleeve anastomosis techiniques. Both rate of operative success and hepatic artery patency were 100%. The typical characteristics of AR from DA-to-Lewis rats after OLT including the features of the serum biochemistry such as ALT,ALB and TBIL,histological manifestations and survival time were similar to those of AR of liver grafts that could be reversed by FKS06. AR occurred 3-5 days after transplantation, became increasingly severe over time,peaked on postoperative day 7 and maintained thereafter. The mean survival time of two groups was 14 and 73 days respectively, and there was significant difference between groups A and B in cumulative survival rate. Conclusion AR model of liver transplantation between DA and Lewis rat strains is stable. This model has many merits of short term research cycle, typical pathological characteristics and reproducible features.