RNA干扰阻断B7/CD28共刺激通路在小鼠心脏移植中的抗排斥作用  

The anti-rejection effect of blocking BT/CD28 co-stimulatory pathway by RNA interference in mice heart transplantation

作　　者：王炜[1] 刘彤[1] 朱理玮[1] 王鹏志[1] 

机构地区：[1]天津医科大学总医院普通外科,300052

出　　处：《中华器官移植杂志》2006年第11期648-651,共4页Chinese Journal of Organ Transplantation

基　　金：基金项目:此项目受天津市重中之重项目资助(05YFGDSF02600)

摘　　要：目的探讨RNA干扰(RNAi)技术阻断B7/CD28共刺激通路对小鼠异体心脏移植排斥反应的影响及其机制。方法经体外转录合成针对CD80 mRNA和CD86 mRNA序列特异性小片段干扰RNA(siRNA),转染供者骨髓来源的树突状细胞(DC),半定量逆转录聚合酶链反应、流式细胞仪检测DC转染CD80siRNA、CD86siRNA前后CD80 mRNA和CD86 mRNA的表达水平以及细胞表面CD80及CD86的表达情况。在小鼠异位心脏移植前7d．经静脉给受者输注经siRNA干扰后的DC(干扰DC组),同时设立同种异体对照组、环孢素A(CsA)治疗组(术后皮下注射CsA 5 mg/d)、同系移植对照组和未干扰DC组(移植前输注未转染DC),观察各组移植心脏的存活时间,对移植物的排斥反应进行病理分级,并测定移植物组织中白细胞介素2(IL-2)、γ干扰素(IFN-γ)及IL-10的mRNA表达水平。结果siRNA转染DC后,其CD80 mRNA及CD86 mRNA的表达受到明显抑制,CD80、CD86的阳性率分别由84%和67%下降至35%和30%。与同种异体对照组和未干扰DC组比较,干扰DC组移植心脏存活时间明显延长(P＜0．01),组织排斥反应病理分级显著降低(P <0．01),移植心脏组织中IL-2 mRNA和IFN-γmRNA的表达水平明显降低(P＜0．01),而IL-10 mRNA的表达水平明显升高(P＜0．01)。结论利用RNAi敲减供者骨髓来源的DC表面B7分子的表达,以阻断B7/CD28共刺激通路,具有抑制小鼠心脏移植排斥反应的作用,其机理可能是通过诱导T淋巴细胞无能并使T辅助细胞分化向T_H2型方向偏移。Objective To explore the influence of blocking B7/CD28 co-stimulatory pathway by RNA interference on the rejection response in mice heterogeneous heart transplantation and its mechanism. Methods siRNA of which sequence specified to CD80 and CD86mRNA was synthesized in vitro respectively and transfected into donor derived myeloid dendritic cells （DCs）. The expression levels of CD80 and CD86 mRNA and surface antigen CD80, CD86 were assayed by semi-quantitative reverse transcription polymerase chain reaction and flow cytometry before and after CD80 siRNA and CD86 siRNA transfection. Seven days prior to heterogeneous heart transplantation in mice, DCs modified by siRNA were transfused into recipients intraveneously （DC interference group）. At the same time, group of allograft transplantation, cyclosporine A （CsA）-treated （CsA injected subcutaneously postoperatively, 5 mg·kg^-1·d^-1） group, group of isograft transplantation, and non-interference DC group （transfusion of non-interfered DCs pre-transplanting） were assigned. The graft survivals were individually recorded and the graft rejection grading was pathologically evaluated. Interleukin 2 （IL- 2）, interferon γ （IFN-γ）, and IL-10 mRNA expression levels in grafts tissue were determined. Results After siRNA transfected into DC, the expression levels of CD80 and CD86 mRNA were downregulated significantly and the the antigen CD80^＋ and CD86^＋ reduced from 84 %, 67% to 35% and 30 % respectively. As compared with groups of allograft and non-interference DC, survival of the grafts was significantly longer in DC interference group （P〈0.01）, pathological grade of rejection significantly lower （P〈0.01）, IL-2 and IFN-γ mRNA expression levels lower, and IL-10 mRNA expression levels higher in grafts tissue （P〈0. 01 ）. Conclusion Knocking down the molecule B7 expression level in donor-derived myeloid DCs through RNAi, which could block B7/CD28 co-stimulatory pathway, could exhibit inhibitive effect on rejection response in

关 键 词：RNA干扰 抗原 CD80 心脏移植 免疫抑制法 树突细胞 

分 类 号：R654.2[医药卫生—外科学]