TRPV1通道异源表达体系的建立和功能的初步研究  

Construction and preliminary functional study of heterologous expression system of TRPV1 channel

作　　者：李文兰[1] 王德凤[1] 周海玉[2] 詹红丹[2] 戴逸飞[2] 周炜炜[2] 戴丽[2] 隋峰[2] 霍海如[2] 

机构地区：[1]哈尔滨商业大学生命科学与环境科学研究中心,黑龙江哈尔滨150076 [2]中国中医科学院中药研究所,北京100070

出　　处：《中国药理学通报》2016年第3期439-441,共3页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金面上项目(No 81274112;81373986;81473372);国家自然科学基金青年项目(No 81403125;81403322);北京市自然科学基金(No 7152106)

摘　　要：目的构建能够表达TRPV1通道用于实验研究的HEK293T细胞表达体系.方法采用脂质体转染方式将TRPV1质粒转入人胚胎肾HEK293T细胞,建立TRPV1型通道瞬时异源表达体系,并对转染后的TRPV1表达体系进行了鉴定,同时对该通道功能特性进行了研究.结果经荧光显微镜观察,转染率可达40% ～50%;WesternBlot研究发现转染后的HEK293T细胞在与TRPV1通道蛋白相应的位置上具有明显的条带,提示TRPV1通道蛋白在转染后的细胞中有异源性表达;共聚焦显微成像显示,转染后的HEK293T细胞在受到TRPV1通道激动剂刺激后,胞内荧光强度明显变强,提示TRPV1通道介导钙离子功能正常.结论研究结果表明,基于HEK293T细胞的TRPV1通道瞬时转染的异源表达体系成功构建.Aim The TRPV1 plasmid was transiently transfected into human embryonic kidney HEK 293 T cells to establish the heterologous expression system of TRPV1-channel.Methods The transfection efficiency was confirmed under fluorescence microscope and the TRPV1 protein expression was identified by using Western blot,and the functional characteristics of the channel were studied by using the method of confocal microscopy.Results The transfection rate could reach 40%~50%;the transfected cells were found to have a clear band at the corresponding position that TRPV1 should be,which indicated that TRPV1 channel protein was expressed in the transfected cells.The confocal microscopy imaging result showed that the transfected HEK 293 T cells were activated by TRPV1 channel agonist.Conclusion Transient transfection of HEK 293 T cells with TRPV1 channel is successfully constructed and the heterologous TRPV1 channel is verified to have normal calcium-mediating function.

关 键 词：瞬时转染 TRPV1通道 HEK293T细胞 异源性表达 背根神经节 基因转染 

分 类 号：R322.61[医药卫生—人体解剖和组织胚胎学] R341[医药卫生—基础医学]