腺病毒载体介导的共刺激分子融合蛋白对实验性自身免疫性心肌炎的作用  

Effects of adenovirus-mediated gene transfer of ICOSIg and CTLA4Ig fusion protein on experimental autoimmune myocarditis

作　　者：刘巍[1] 高成[2] 周保国[3] 王秀荣[4] 李冬梅[4] 李悦[1] 李为民[1] 

机构地区：[1]哈尔滨医科大学第一临床医学院心内科,150001 [2]哈尔滨医科大学第一临床医学院神经外科,150001 [3]哈尔滨医科大学第一临床医学院普外科,150001 [4]中国农业科学院哈尔滨兽医研究所

出　　处：《中华微生物学和免疫学杂志》2006年第5期452-457,共6页Chinese Journal of Microbiology and Immunology

摘　　要：目的探讨腺病毒载体介导的共刺激分子融合蛋白CTLA4Ig与ICOSIg联合治疗对实验性自身免疫性心肌炎(EAM)的作用。方法猪心肌肌球蛋白免疫Lewis大鼠制成EAM模型。分别构建CTLA-4胞外域、ICOS胞外域与人IgGFc段融合的腺病毒表达载体,常规方法生产表达上述融合蛋白的腺病毒用于治疗,免疫第14天注射后观察至第28天。第28天超声心动图检测心脏功能,苏木素-伊红(HE)染色观察心肌炎症程度,Westernblot检测心肌CTLA-4、ICOS、ICOSL及B7-1、B7-2蛋白表达水平,ELISA检测血浆IL-2、IL-4和IFN-γ水平。结果CTLA4Ig、ICOSIg单独或联合治疗均使大鼠心功能指标、心肌炎症程度明显改善。Westernblot显示联合治疗组CTLA-4、ICOSL及ICOS、B7-1蛋白表达下调,而B7-2表达差异无统计学意义。细胞因子平衡向TH2方向偏离。结论CTLA4Ig及ICOSIg联合阻断共刺激分子通路减轻EAM自身免疫性心肌损伤,改善大鼠心脏功能。其机制可能通过下调心肌组织CTLA-4、ICOS、ICOSL及B7-1蛋白表达。Objective To explore the effects of adenovirus vector-mediated gene transfer of ICOSIg and CTLA4Ig fusion protein on experimental autoimmune myocardifis（EAM）. Methods Expression vector containing CTLA4Ig（pAdeno-CTLA4Ig） and ICOSIg（pAdeno-ICOSIg） was constructed by fusion of human CTLA-4 or ICOS and IgG Fc segment. After the adenovirus vector was digested by Pac Ⅰ enzyme, it was transfected into HEK 293 cells. Adenovirus expresses CTLA4Ig or ICOSIg was produced. EGFP constructed into adenovirus vector was used as controls. EAM was induced in Lewis rats by injection of porcine cardiac myosin. All the immunized Lewis rats were divided into 4 groups. Group A（ n = 15） received adenovirus containing CTLA4Ig and ICOSIg from day 14 to day 28; group B（ n = 15）, group C （ n = 15） and group D（ n = 15） received adenovirus containing CTLA4Ig, ICOSIg and EGFP respectively. Group E（ n = 10） was normal controls that did not receive immunization. On day 28, all the rats in 5 groups were killed after echocardiography examination. Histopathological examination was used to observe inflammation in the myocardium. Western blot was used to detect CTLA-4, ICOS, ICOSL, B7-1 and B7-2 protein expression .Results Alliance application of CTLA4Ig and ICOSIg exerts therapeutic effects on EAM. On day 28, cardiac function and myocardial inflammation improved significantly compared to group D. Exparesion of CTLA-4, ICOS and ICOSL, B7-2 was significantly decreased in group A, B and C compared with group D. Blockade of costimulatory pathway with alliance therapy of CTLA4Ig and ICOSIg alleviated autoimmune damage in EAM and improved cardiac function. Down-regulation of costimulatory molecules and anti-inflammation may be involved in the mechanisms.

关 键 词：实验性自身免疫性心肌炎 ICOS CTLA-4 基因治疗 融合蛋白 

分 类 号：R542.2[医药卫生—心血管疾病]