大鼠小肠移植早期应用趋化因子受体拮抗剂抑制移植物急性排斥反应

Early adoption a chemokine receptor antagonist Met-RANTES suppressing acute rejection of small bowel allograft in rats

目的探讨趋化因子受体拮抗剂(MetRANTES)在小肠移植早期应用对排斥反应的免疫抑制作用及其与他克莫司的协同效应。方法SD大鼠为供者,Wistar大鼠为受者,建立异基因节段性异位小肠移植模型。移植后将大鼠分为4组,每组24只。第1组为对照组,移植前后不作任何处理;第2组为MetRANTES组,小肠移植后(0～7d)腹腔注射MetRANTES200μg/d;第3组为小剂量他克莫司(FK506)组,小肠移植后(0～7d)腹腔注射FK5060.5mg·kg-1·d-1;第4组为MetRANTES联合FK506组,2种药物的应用方法与第2、3组相同。观察移植大鼠的一般状况和存活时间以及免疫细胞浸润情况,并于移植术后3、5、7d分别取各组大鼠移植肠标本(n=6)进行组织病理学检查,采用免疫荧光染色和激光扫描共聚焦显微镜技术对移植肠RANTES和CD4+、CD8+、CD25+T淋巴细胞的表达进行连续定量测定。结果第1～4组大鼠平均存活时间分别为(7.37±1.19)d、(22.32±8.60)d、(23.64±6.58)d和(30.55±4.18)d,第2、3、4组大鼠与第1组相比,存活时间明显延长(P<0.01);第4组大鼠存活时间更长,与第2、3组比较,差异均有统计学意义(P<0.01)。第1组全部死于急性排斥反应及感染,组织病理学检查显示移植后第3、5、7d分别符合轻、中、重度排斥反应。第2、3、4组病理学检查无明显排斥反应征象。第1组大鼠的移植肠RANTES表达在术后各时段均显著高于其他3组(P<0.01),其动态变化与急性排斥反应的进程呈正相关;第2组和第4组大鼠移植肠RANTES、CD4+、CD8+和CD25+T细胞的表达均明显低于第1组(P<0.01)。结论MetRANTES能明显抑制小肠移植急性排斥反应,有效保护移植肠功能,显著延长移植物的存活时间,并可增强小剂量他克莫司的免疫抑制作用。

Objective To explore the immunosuppression effect of early adopting Met-RANTES after small bowel transplantation on allograft rejection and its synergic effect with tacrolimus. Methods In mature male SD rats as donors and Wistar rats as recipients, the model of heterotopic segmental small bowel transplantation was set up. Rats were divided into 4 groups after transplantation, and each group had 24 rats： group 1 , alIogeneic small bowel transplant untreated group as control group; group 2 , allogeneic small bowel transplant treated with Met-RANTES （200 μg/day, i. p, 0～7 days）; group 3 , allogeneic small bowel transplant treated with low dose of tacrolimus （FK506, 0. 5 mg·kg^-1 ·day^-1 , i. p, 0～7 days）; group 4 , allogeneic small bowel transplant treated with Met- RANTES and FK506 in the same way with group 2 and 3. After transplantation clinical course, survival time and immuocyte infiltration were observed. Grafts （n = 6） of each group were sampled respectively at day 3, 5, and 7 after the transplantation. All samples were examined pathologically. By using immunofluorescent staining and laser scanning confocal microscope techniques, successive quantitative measurement was conducted to detect the expression of graft RANTES and CD4^＋ , CD8^＋ , CD25^＋ T lymphocytes. Results Average survival time in groups 1, 2, 3 and 4 was （7. 37 ± 1.19） days, （22. 32 ± 8. 60） days, （23. 64. 32 ± 6. 58） days, and （30. 55 ± 4. 18） days. Compared with that in the group 1, the average survival time in groups 2, 3 and 4 was obviously prolonged （P〈0.01）. The average survival time in group 4 was much longer than that in groups 2 and 3 （P〈0. 01 ）. Rats in group 1 died of acute rejection and infection. The histological examination revealed that in the first group mild, moderate and severe rejection occurred on the post-operative day 3, 5, and 7. While in groups 2, 3 and 4, no obvious rejection occurred. The RANTES expression in group 1 in each postoperative period was significantly higher than that of the other three groups （P〈0. 01 ）. There was a positive correlation between the dynamic change of the RANTES expression and the process of acute rejection. The CD4^＋ , CD8^＋ , CD25^＋ T lymphocytse and the RANTES expression of the grafts in groups 2 and 4 were significantly lower than in group 1 （P〈0. 01 ）. Conclusions Met-RANTES may obviously suppress acute allograft rejection in small bowel transplantation, effectively protect the function of grafts, and significantly prolong the survival time of the recipients. In addition, Met-RANTES may enhance the imrnunosuppressive function of FK506 with low dose.