MHC-Ⅱ类分子与hTSH受体共表达系统的建立  

Establishment a Cell line Co-expressing MHC ClassⅡMolecules and hTSHR

作　　者：田竹芳[1] 雒文田[2] 吴晓燕[2] 朱本章[2] 

机构地区：[1]西安市中心医院,陕西西安710003 [2]西安交通大学第一医院,陕西西安7100061

出　　处：《实用医技杂志》2004年第10A期1949-1952,共4页Journal of Practical Medical Techniques

摘　　要：目的 :应用电击基因导入法 ,建立 MHC- 类分子与 h TSHR共表达体系 (h Ml2 ) ,为 GD发病机制的研究奠定基础。方法 :采用电击基因导入法将质粒 p CRTM3- h TSHR转染于 M12细胞 (H- 2 d) ,应用 G4 18筛选出阳性细胞 ,经有限稀释获得单克隆细胞株。用逆转录聚合酶链反应 (RT- PCR)法检测单克隆细胞内 h TSHR胞外段 m RNA的表达 ;用甲亢病人血清粗提 Ig G及 b TSH刺激后测定上清中 c AMP值以检测单克隆细胞株表达的 h TSHR是否具有天然 h TSHR功能。结果 :应用有限稀释法获得 14株单克隆细胞。RT- PCR结果显示 :3号、5号、6号、8号、11号细胞内有高水平的 h TSHR胞外段 m RNA的转录。应用甲亢病人血清粗提 Ig G及 b TSH刺激后 ,6号、8号细胞上清中c AMP值明显升高 ,与对照组之间存在显著差别 (P<0 .0 5 )。结论 :应用电击基因导入法成功地建立了 MHC- 类分子与 h TSH受体共表达体系。Objective Using electroporation to establish a stable cell line co-expressing MHC classⅡmolecules and hTSHR for study the pathogenesis of Graves’disease(GD).Methods M12 cells were transfected withp CRTM 3-hTSHR plasmids using electroporation.Cells were selected for neomycin resistance using G418,and positive cells were cloned by limiting dilution.Stable transfectants were selected by RT-PCR and their ability to increase cAMP levels in the presence of bTSH and crude IgG of GD.Results PCR product sproved that hTSHR mRNA were transcribed in No.3、5、6、8、11 cell lines.Cell lines were stimulated by crude IgG of GD and bTSH,cAMP incr eased significantly in culture supernatants of No.6 and No.8 cells lines,it suggest that the hTSHR expressing On the No.6 and No.8 cell lines have the ability of native hTSHR.Conclusion We successfully established the cell lines that co-express MHC class Ⅱ molecules and hTSHR.

关 键 词：单克隆 TSH受体 MHC 共表达 胞外段 细胞内 病人 基因导入 稀释 应用 

分 类 号：R581[医药卫生—内分泌] R392[医药卫生—内科学]