摘要
目的:探讨Cl C-3氯通道是否为IK1钾通道的调节靶点,重点研究鼻咽癌细胞IK1钾通道对Cl C-3氯通道功能及蛋白表达的影响。方法:采用siRNA转染技术抑制低分化鼻咽癌上皮细胞(CNE-2Z)IK1基因的表达;real-time PCR技术检测Cl C-3 mRNA的表达;Western blot检测Cl C-3的蛋白表达;细胞免疫荧光结合激光共聚焦显微镜技术检测Cl C-3和IK1蛋白在细胞内分布;全细胞膜片钳记录细胞氯电流。结果:IK1 siRNA可以成功转染CNE-2Z细胞,有效抑制鼻咽癌细胞IK1钾离子通道的表达;用IK1 siRNA抑制鼻咽癌细胞IK1钾离子通道的表达后,Cl C-3的mRNA表达上调而Cl C-3蛋白却表达减少:在低分化鼻咽癌上皮细胞,低渗刺激可激活氯通道,产生一个较大的氯电流,在成功转染IK1 siRNA的细胞,此氯电流明显减弱。结论:敲低IK1钾离子通道可抑制Cl C-3氯离子通道的表达和功能。
AIM： To investigate whether the Cl C-3 chloride channel is an acting target of the IK1 potassium channel,and to study the action of IK1 potassium channel on the functional activities and expression of Cl C-3 chloride channels. METHODS： IK1 gene was silenced by IK1 siRNA in poorly-differentiated nasopharyngeal carcinoma cells（ CNE-2Z）. Real-time PCR and Western blot were used to detect the expression of Cl C-3 at mRNA and protein levels. The distribution of Cl C-3 protein in the cells was observed under confocal immunofluorescence microscope. The chloride current was recorded by the patch-clamp technique. RESULTS： IK1 siRNA was successfully transfected into the CNE-2Z cells and knocked down the expression of IK1 potassium. The mRNA expression of Cl C-3 was increased by the IK1 siRNA. IK1 siRNA inhibited the expression of Cl C-3 protein. A chloride current was activated by hypotonic challenges,and the hypotonicity-induced current was reduced in the cells which successfully transfected with IK1 siRNA. CONCLUSION： The knockdown of IK1 potassium channels inhibits the expression and function of Cl C-3 chloride channel.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2015年第12期2113-2119,共7页
Chinese Journal of Pathophysiology
基金
国家自然科学基金资助项目(No.81272223)
关键词
CL
C-3氯通道
IK1钾通道
鼻咽癌
膜片钳技术
ClC-3 chloride channel
IK1 potassium channel
Nasopharyngeal carcinoma
Patch-clamp techniques
