输注分子嵌合小鼠前体T细胞抑制异源小鼠T细胞增殖反应的研究  

Research on molecular chimeric mice pre-T cells infusion to reduce response to allogeneic T cells stimulation

作　　者：陆礼[1] 宋东旭[1] 李卫东[1] 付蔚华[1] 

机构地区：[1]天津医科大学总医院普通外科,300052

出　　处：《国际生物医学工程杂志》2014年第1期36-38,52,I0006,共5页International Journal of Biomedical Engineering

基　　金：国家自然科学基金资助项目（81041118）

摘　　要：目的 研究分子嵌合前体T细胞（pre-T细胞）对异基因T淋巴细胞增殖能力的影响,旨在为研究同种异体器官移植诱导免疫耐受提供新策略.方法 RT-PCR获取C57BL/6小鼠MHC-Ⅰ等位基因H-2Kb和H-2Db基因,并与pIRES质粒连接,构建重组质粒并转染体外培养BALB/c小鼠pre-T细胞构建分子嵌合细胞,并设立未转染及转染空质粒对照组.各组细胞回输BALB/c小鼠,7d后提取各组小鼠脾脏T细胞,将其与C57BL/6小鼠T淋巴细胞混合培养,观察刺激指数（SI）.结果 构建质粒测序表明,插入序列的结果包含GenBank检索的C57BL/6小鼠H-2Kb和H-2Db序列,流式细胞仪检测质粒pIRES-H-2Kb和pIRES-H-2Db转染后,pre-T细胞表面H-2Kb及H-2Db蛋白的表达增高,与未转染组及转染空质粒组相比,其差异均具有统计学意义（P＜0.05）.单向混合淋巴细胞培养结果显示：2质粒共注射pre-T细胞组的SI值（0.764±0.074）较空质粒组（0.983±0.081）和未转染组（0.994±0.142）明显下降,其差异有统计学意义（P＜0.05）.结论 分子嵌合细胞可降低受体T细胞对异基因T淋巴细胞刺激的增殖反应,有望应用于体内诱导免疫耐受.Objective To research the effect of molecular chimeric mice pre-T cells on proliferation ability of allogeneic mouse T cells.Methods The MHC-Ⅰ gene （H-2Kband H-2Db gene） were extracted and amplified by RT-PCR,the identified pre-T cells were transfected by the constructed eukaryotic expression vector of C57BL/6mouse MHC-I （pIRES-H-2Db and pIRES-H-2Kb）,non-transfected group and sham pIRES-transfected control group were set.The molecular chimeric cells were transfused back to BALB/c mouse.After 7 days,T lymphocyte cells of each group were extracted,the ability of molecular chimeric cells inducing spleen T lymphocyte response to allogeneic T cells was observed through mixed lymphocyte culture （MLC）.Results Sequencing of the plasmid we have constructed showed that insertion sequence contained C57BL/6 mice H-2Kb and H-2Db series which could be retrieved from GenBank.The result of flow cytometry analysis indicated that H-2Kb and H-2Db protein had an increased expression in pre-T cells,the difference with other two groups was statistically significant （P＜0.05）.The result of MLC demonstrated that the stimulation index （SI） of T lymphocyte in co-injection transfected pIRES-H-2Kb and pIRES-H-2Db pre-T cells group （0.764±0.074） were significantly decreased compared with non-transfected group（0.983±0.081）and sham pIRES-transfected group（0.994±0.142） （P＜0.05）.Conclusions The molecular chimeric pre-T cells infusion can reduce spleen T lymphocyte response to allogeneic T cells and it may induce immune tolerance in vivo.

关 键 词：MHC-Ⅰ基因 前体T细胞 分子嵌合 刺激指数 

分 类 号：R392.12[医药卫生—免疫学]