B7特异性RNA干扰对皮肤移植的影响.docVIP

B7特异性RNA干扰对皮肤移植的影响 袁成良1 邹自英2 魏世刚1 李永忠3 刘定海1 刘利洪1 1德阳市人民医院检验科，四川德阳，618000 2成都军区总医院中心实验室， 四川成都 610083 3德阳市人民医院烧伤科 四川德阳，618000 [摘要] 目的：探讨RNA干扰技术阻断B7/CD28共刺激信号对小鼠异体皮肤移植排斥反应的影响。方法：设计并合成B7特异性siRNA，在脂质体的介导下转染树突状细胞，转染后72h收集细胞，用蛋白印迹法检测B7-1、B7-2蛋白的表达。在小鼠异体皮肤移植前7d，经静脉将转染B7特异性siRNA的DC输入小鼠体内（DC转染组），同时设立同种异体移植对照组、同系移植对照组、环孢素A（CsA）治疗组（术后每日皮下注射CsA 5mg/kg）和未转染DC组（移植前输注未转染DC），观察各组移植皮瓣的存活时间和排斥反应情况，并用MTT法检测混合淋巴细胞增殖反应，术后1月观察迟发性超敏反应情况。结果：Western Blot结果显示转染72h后，siRNA1对B7-1蛋白表达的抑制率为(77.2±6.26)%；siRNA2对B7-2蛋白表达的抑制率为(73.3±5.42)%。与同种异体移植对照组和未转染DC组比较，DC转染组移植皮瓣成活时间明显延长（P0.01），组织排斥反应程度较轻，混合淋巴细胞反应和DTH反应显著降低（P0.01）。结论：B7特异性siRNA可明显抑制树突细胞B7基因的表达，以阻断B7/CD28共刺激通路，抑制小鼠皮肤移植排斥反应，为进一步研究siRNA诱导免疫耐受提供了新方法。 [关键词] RNA干扰，树突状细胞，B7，皮肤移植，免疫抑制 Effect of RNA interference for B7 on the rejection response in skin transplantation YUAN Cheng-Liang1 WEI Shi-Gang1 LI Young-Zhong2 LIU Ding-Hai1 LIU Li-Hong1 ZOU Zi-Ying2 1Department of Laboratory,People’s Hospital of Deyang City, Deyang 618000,Sichuan Province,China 2 Laboratory Center , Chengdu Military Command General Hospital, Chengdu,610083, Sichuan Province China; 3Department of Burns,People’s Hospital of Deyang City, Deyang 618000,Sichuan Province,China [Abstract] Objective：To investigate the effect of blocking B7/CD28 co-stimulatory pathway by RNA interference on the rejection response in mice herogeneous skin transplantation. Methods：Two different siRNA (siRNA1-targeted against B7-1, siRNA2-targeted against B7-2) were designed and synthysized and transfected into DC with liposome. At 72h after transfection ,the expression of B7 gene were assayed by Western blot. Seven days prior to heterogeneous skin transplantation in mice ,DCs modified by siRNA were transfused into recipients intraveneously(DC transfection group). At the same time ,group of allograft transplantation, group of isograft transplantation,cyclosporine A (CsA-treated) (CsA injected subcutaneously postoperatively,5 mg/kg/d)group, and non-transfection DC group (transfusion of n