酶联免疫斑点技术在心脏移植术后早期诊断急性排斥反应中的作用  

The role of ELISPOT in early diagnosis of acute rejection

作　　者：朱鹏[1] 陈义发[1] 陈孝平[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院肝脏外科中心, 武汉430030

出　　处：《中华器官移植杂志》2008年第4期197-200,共4页Chinese Journal of Organ Transplantation

基　　金：国家自然科学基金（30571754）;留学回国人员科研启动基金（教外司留[2005]383）

摘　　要：目的探讨酶联免疫斑点技术（ELISPOT）在心脏移植术后早期诊断急性排斥反应中的作用及在供者评估方面的应用价值。方法采用小鼠腹部心脏移植模型，将实验小鼠分为3组，每组25对。（1）移植排斥组：供者为C57BL／6小鼠，受者为BALB／C小鼠，移植后无特殊处理；（2）实验处理组：供者为（257BL／6小鼠，受者为BALB／C小鼠，受者在移植术前1天经尾静脉输注特异性供者脾细胞，术前1d及术后1周按每克体重应用环孢素A5μg；（3）同系移植组：供、受者均为BALB／C小鼠，移植术后无特殊处理。术后观察移植心存活时间及病理改变；应用ELISPOT技术检测受者脾细胞悬液中供者γ干扰素（IFN-γ）活性分泌细胞频数。结果移植排斥组和实验处理组移植心平均存活时间分别为（7．8±0．77）d和（14．80±1．01）d，同系移植组存活时间均超过28d，各组之间的差异有统计学意义（P〈0．05）。移植排斥组与实验处理组和同系移植组相比较，移植心的心肌细胞变性坏死严重，并有大量炎性细胞浸润。移植术后第4天，移植排斥组、实验处理组和同系移植组受者脾细胞悬液中供者特异性IFN-γ活性分泌细胞频数分别为（288±16）个／2×10^5、（32±10）个／2×10^5和（6±2）个／2×10^5；第7天为（416±19）个／2×10^5、（44±8）个／2×10^5和（7±2）个／2×10^5；其与相应的移植心存活时间存在明显负相关，而与术后第7天病理分级存在明显正相关。结论应用ELLS-POT技术检测受者术后早期供者特异性IFN-7活性分泌细胞频数可作为急性排斥反应的一个早期诊断指标。此技术具有较高的灵敏性与特异性，可用于术前供者配型。Objective To study the method for early diagnosis of acute rejection and the role of ELISPOT in donor evaluation. Methods All ventral heterotopic cardiac transplantation models were divided into three groups （each group having 25 recipients） as follows： rejection group （C57BL/6 donor heart to BALB/c recipient, no specific treatment after transplantation）, treated group （C57BL/6 donor heart to BALB/c recipient, donor specific splenocytes transfusion before transplantation plus 5 μg/g cyclosporin A per day applied continuously from 1 day before operation to 7 day posttransplantation）, isograft group （BALB/c donor heart to BALB/c recipient, no specific treatment after transplantation）. Mean survival time （MST） and histopathologic changes were observed. By using ELISPOT assay, IFN-γ-secreting splenocytes were detected and counted. Results MST of heart allografts in rejection and treated groups was （7. 8±0. 77） and （14. 80±1.01） days respectively. The survival time of grafts in isograft group was all more than 28 days. There was significant difference among three groups. The number of infiltrating cells in rejection group was much more than the other groups as well as the extent of histopathologic changes. The number of donor-specific IFN-γ-secreting splenocytes in three groups on the posttransplantation day 4 and 7 was （288 ± 16）, （32 ± 10）, （6 ± 2）/2 ×10^5 and （416 ± 19）, （44 ± 8） ,（7± 2）/2 × 10^5 , respectively, which had negative correlation with MST and positive correlation with histopathologic changes. Conclusions The detection of donor-specific IFN-γ-secreting splenocytes with ELISPOT assay might be a useful indicator for early diagnosis of acute rejection. This assay had high sensitivity and specificity enough for pretransplant donor evaluation.

关 键 词：免疫酶技术 心脏移植 Γ干扰素 小鼠 

分 类 号：R686[医药卫生—骨科学]