摘要目的 观察细胞毒性T淋巴细胞相关抗原4融合蛋白(CTLA4Ig)与西罗莫司(SRL)联用阻断共刺激通路对异种胰岛移植物存活的影响.方法 取C57BL/6小鼠,腹腔注射链佐星,制成糖尿病模型.采用随机单位组设计分组法将糖尿病小鼠分为7组,各组均于小鼠左肾包膜下移植SD大鼠胰岛300胰岛当量.CTLA4Ig组分别于移植当天及移植后第2、4、6天腹腔注射CTLA4Ig 0.5 mg/d;SRL组分别于移植当天及移植后第1、2天给予SRL灌胃,0.2 mg·kg-1·d-1,其后隔天用药1次,共用2周;MRI组分别于移植当天及移植后第2、4天腹腔注射仓鼠抗小鼠CD154单克隆抗体(MR1)0.5 mg/d;CTLA4Ig和SRL联用组(SRL联用组)、CTLA4Ig和MRl联用组(MR1联用组)以及CTLA4Ig、MR1和SRL联用组(三药联用组)各药物的剂量与用法同上述各组;对照组仅行胰岛移植,不予以药物.观察至移植后200 d,通过监测受者血糖水平来判断排斥反应的发生情况.记录各组移植物的存活(即无排斥反应)时间.发生排斥反应者,或未发生排斥反应、移植物存活时间＞200 d者,取移植胰岛,行HE染色及免疫荧光染色,进行组织学观察.结果 对照组移植物存活时间中位数为17 d,该组最终均发生排斥反应.SRL组、MRl组和CTLA4Ig组移植物存活时间中位数分别为34 d、98 d和77 d.均明显长于对照组(P＜0.05),三组中分别有90%(9/10)、62.5%(5/8)和83.3%(5/6)的小鼠发生排斥反应.SRL联用组移植物存活时间中位数为130 d,明显长于上述4组(P＜0.01),有50%(3/6)的小鼠发生排斥反应.MR1联用组以及三药联用组移植物存活时间中位数均＞200 d,分别有42.9%(3/7)和25%(2/8)的小鼠发生排斥反应.组织学检查结果显示,对照组发生排斥反应时,其移植胰岛破坏严重,可见大量CD4+和CD8+淋巴细胞及巨噬细胞浸润,并可见IgG、IgM和补体C3沉积.其它组发生排斥反应者的组织学改变与对照组相似.SRL联用组存活200 d的小鼠,其移植胰岛组织中未见或仅有少量炎症细胞浸润,胰岛素和胰高血糖素染色阳性,未见IgG、IgM和补体C3沉积.结论 短期联合使用CTLA4Ig和SRL能显著延长小鼠体内大鼠来源的胰岛的存活时间.

abstractsObjective To study the effect of CTLA4Ig and rapamycin therapy on islet xenotransplantation. Methods Streptozotocin-induced diabetic C57BL/6 mice were transplanted (Tx) under the kidney capsule with rat islets and randomly divided into following groups: group 1, control group, i.e. islet Tx without any therapy; group 2, rapamycin group (0. 2 mg/kg by oral garage at day 0, 1, 2 and every other day to 14); group 3, anti-CD154 mAb (MRI) group (0. 5 mg i. p on day 0, 2 and 4); group 4, CTLA4Ig group (0. 5 nag i. p on day 0, 2, 4 and 6); group 5 (combination therapy with CTLA4Ig and rapamycin); group 6 (combination therapy with CTLA4Ig and MR1);group 7 (combination therapy with CTLA4Ig, MR1 and rapamycin). Mice were followed for islet function by glycemia and histology was performed at time of rejection and 200 days after Tx. Results Rapamycin, MR1, CTLA4Ig therapy alone significantly prolonged rat xenograft survival compared to control group [median graft survival (MGS) 34 days, 98 days and 77 days vs. 17 days, respectively,P＜0. 05), but rejection still occurred. Combination therapy of CTLA4Ig + rapamycin allowed significantly prolonged graft survival compared with control group (MGS 130 days) (P＜0. 01 ). Combination therapy of CTLA4Ig + MR1 and CTLA4Ig + MR1 + rapamycin allowed indefinite graft survival (MGS ＞200 days and ＞200 days respectively). Histological analysis showed severe xenografts destruction at rejection. At day of 200 post-Tx, CTLA4Ig + rapamycin treated mouse grafts showed positive staining for insulin and glucagon, and no cellular infiltration within the grafts. Conclusion Long-term survival of concordant islet xenografts was achieved by combination therapy of CTLA4Ig and rapamycin.