超极化激活环核苷酸门控通道在人肺组织中的表达及功能  

Expression and function of hyperpolarization-activated cyclic nucleotid-gated cation channel in human lung

作　　者：丁炎[1] 崔湧[2] 昌建鈞 候亚鹏 周祉妤 聂宏光[1] Ding Yan;Cui Yong;Chang Jianjun;Hou Yapeng;Zhou Zhiyu;Nie(Hongguang Institute of Metabolic Disease Research and Drug Development, China Medical University, Shenyang 110122, China;Department of Anesthesiology, the First Hospital of China Medical University, Shenyang 110001, China)

机构地区：[1]中国医科大学代谢病分子机制与药物研究所,沈阳110122 [2]中国医科大学附属第一医院麻醉科,沈阳110001

出　　处：《中国医药》2018年第2期203-206,共4页China Medicine

基　　金：国家自然科学基金(81270098、81670010);辽宁省高等学校基本科研项目(LQNK201745)~~

摘　　要：目的探讨超极化激活环核苷酸门控(HCN)通道在人肺组织标本及人肺上皮细胞系中的表达情况,并进行相关功能学研究。方法收集临床肺癌手术患者癌旁正常肺组织标本,体外培养人肺上皮H441。采用聚合酶链反应和蛋白质免疫印迹技术检测人肺组织标本和人肺上皮细胞中HCN通道的表达水平。应用尤斯灌流室实验装置测定H441单层细胞的短路电流,检测HCN通道的相关功能。结果聚合酶链反应结果显示,HCN 4和HCN 1通道扩增产物经琼脂糖凝胶电泳出现116 bp和91 bp的目的条带,HCN 4和HCN 1亚型在人肺组织和H441细胞系中均有mRNA水平的表达,而HCN 2和HCN 3亚型未见表达。用10%聚丙烯酰胺凝胶电泳分离后,在99 000和130 000分别出现HCN 4和HCN 1的目的条带,HCN 4和HCN 1亚型在人肺组织和H441细胞系中均有表达,HCN 2和HCN 3亚型未见表达。尤斯灌流室系统测定短路电流的变化,3、10、30、100、300μmol/L的ZD7288能够剂量依赖性抑制人H441单层细胞的短路电流,根据Boltzmann方程拟合后的半数抑制率为85.8μmol/L。结论 HCN 4/1通道亚型在人肺组织中具有组织和功能学表达,为临床寻求合适药物治疗肺脏相关疾病提供了理论基础。Objective To explore the expression and function of hyperpolarization-activated cyclic nucleotid-gated cation channel（HCN） in human lung tissue and lung epithelial cells. Methods Tumor-adjacent normal lung tissues were separated from lung cancer patients. Expressions of HCN in human lung tissue and vitro cultured human lung epithelial cell line H441 were tested by polymerase chain reaction and western blotting. Short circuit currents in H441 cell monolayers were measured by Ussing chamber apparatus to determine the function of HCN. Results Polymerase chain reaction showed that HCN 4 and HCN 1 expressed as 116 bp and 91 bp in agarose gel electrophoresis; HCN 4 and HCN 1 mRNA significantly expressed in human lung tissue and H441 cells; HCN 2 and HCN 3 mRNA were not detected. Western blotting showed HCN 4 and HCN 1 expression in human lung tissue and H441 cells; HCN 2 and HCN 3 were not detected. In Ussing chamber test, 3, 10, 30, 100, 300 μmol/L ZD7288 could inhibit the short circuit currents in H441 cell monolayers in a dose dependent manner, the half inhibition rate was 85.8 μmol/L derived from the Boltzmann equation. Conclusion HCN 4/1 channel isoforms have functional expression in human lung.

关 键 词：超极化激活环核苷酸门控通道 肺组织 短路电流 

分 类 号：R322.35[医药卫生—人体解剖和组织胚胎学]