人肠系膜血管平滑肌细胞BK_(Ca)通道在HEK293细胞上的表达方法研究  

Methodological study on expression of BK_(Ca) of human mesentery artery smooth muscle cells in HEK 293

作　　者：程俊[1] 曾晓荣[1] 谭晓秋[1] 李鹏云[1] 文静[1] 毛亮[1] 杨艳[1] 

机构地区：[1]泸州医学院心血管医学研究所,医学电生理教育部重点实验室,四川泸州646000

出　　处：《泸州医学院学报》2014年第6期549-552,共4页Journal of Luzhou Medical College

基　　金：国家自然科学基金资助项目(No:81173661);泸州市-泸州医学院联合专项(2013LZLY-J49);创新团队(No.KYTD201005)

摘　　要：目的:本研究旨在HEK 293细胞系上成功表达人肠系膜血管平滑肌细胞大电导钙激活钾通道(large-conductance calcium-activated potassium channels,BKCa),建立一种稳定高效表达人肠系膜血管平滑肌细胞BKCa的HEK 293细胞系方法,进一步揭示在工具细胞表达上的BKCa的基本特性,为以后的诸多实验提供参考价值。方法:应用基因工程技术构建人肠系膜血管平滑肌细胞BKCa通道基因表达载体,用脂质体转染法转染HEK 293细胞,建立了稳定高效表达BKCa通道的HEK 293细胞系的方法。结果:在HEK 293细胞系上表达的BKCa单通道电流的电导值为229.56±4.62 p S,具有电压依赖性,钙离子敏感性等特性;在HEK 293细胞系表达的BKCa全细胞宏观电流具有明显的电压依赖性和外向整流特性。结论:成功建立了一种在HEK293细胞系上表达的BKCa通道的实验方法,并成功记录到BKCa通道的单通道电流和全细胞宏观电流,且与天然细胞上BKCa通道电流特性基本一致。Objective： To successfully express large conductance calcium-activated potassium channels（BKCa）of human mesenteric artery smooth muscle cells in HEK 293 cell line, and to reveal the characteristics of BKCa expression on HEK 293 cells. Methods： BKCachannels gene expression vector of human mesenteric vascular smooth muscle cell was constructed by gene engineering technology, and lipofectamine was used to transfect HEK293 cells to establish a stable and efficient method of expression of BKCachannel in HEK 293 cell line. Results：The conductance value of BKCasingle channel current expressed on HEK 293 cells was 229.56 ± 4.62 pS, with voltage dependence, and Ca^2＋sensitivity; whole cell macroscopic currents of BKCaexpressed in HEK 293 cell line was voltage dependent and had outward rectifying. Conclusion： An experimental method of expressing BKCa channel in HEK 293 cell line is successfully established, and the single channel current of BKCachannel and whole cell macroscopic currents are recorded, which it is consistent with the current characteristics of BKCa channel in natural cells.

关 键 词：大电导钙激活钾通道 HEK 293 膜片钳 

分 类 号：R33-33[医药卫生—人体生理学]