caveolin-1对心脏herg钾通道功能调控作用的研究

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1、·1236·医学研究生学报2013年12月第26卷第12期JMedPostgra，Vo1．26，No．12，December，2013论著(基础研究)Caveolin一1对心脏HERG钾通道功能调控作用的研究马青艳，余宏，林吉进[摘要】目的既往研究发现caveolin一1参与了多种心脏离子通道的调控作用，但对心脏HERG钾通道是否具有调控作用，则未见报道。研究caveolin一1对心脏HERG钾通道功能的调控作用。方法①基因转染：应用Lipofectamine2000将pcDNA3．0-caveolin一1质粒和p

2、cDNA3．0-herg转染HEK293细胞，通过G418筛选获得稳定表达细胞株；②脂筏破坏：通过在培养液中加入10mmol／L的甲基．B．环糊精，去除HEK293／HERG细胞膜上的脂筏；③Caveolin．1表达抑制：应用siRNA转染HEK293／HERG细胞抑制caveolin一1表达；@Caveolin一1过度表达：将pcDNA3．0-herg转染HEK293／caveolin一1细胞株，获得caveolin一1过度表达的HEK293／HERG细胞；⑤应用膜片钳技术记录经不同处理后的HERG通道电流。结果

3、①细胞膜脂筏破坏与caveolin一1表达抑制均能显著增大HERG钾通道电流幅度和显著加速尾电流去激活速率；②Caveolin一1过度表达则显著减小了HERG通道电流的振幅并显著缓慢了尾电流的去激活速率。结论Caveolin．1对心脏HERG钾通道的功能具有负性调控作用。【关键词】HERG钾通道；Caveolin一1；RNA干扰；脂筏；膜片钳技术[中图分类号】R54[文献标志码】A[文章编号]1008—8199(2013)12．1236-05RegulatoryeffectofCaveolin-1onthecard

4、iacHERGpotassiumchannelMAQing—yan，YUHong，LINJi-jin(DepartmentofCardiology，GuangdongProvincialCardiovascularInstitute，GuangdongGeneralHospital，Guang—dongAcademyofMedicalSciences，Guangzhou510080，Guangdong，China)[Abstract]ObjectiYePreviousstudiesshowthatcaveolin一

5、1isinvolvedintheregulationofmultipleionchannels．However．noreporthasbeenseenonitsregulatoryeffectonthecardiacHERGchannelfunction．Thisstudywastoinvestigatetheregulatoryfunc．tionofcaveolin一1onthecardiacHERGpotassiumchanne1．MethodsHEK293cellsweretransfectedwithpcD

6、NA3．0一caveolin一1orpcDNA3．0-hergbyLipofectamine2000，andG418wasusedtoscreenstablyexpressedcelllines．，I1}lelipidraftsinthece11membraneweredisruptedbyincubationwith10mmol／Lmethyl—B．cyclodextrin．Theexpressionofcaveolin．1wasknockeddownbytransfectedcaveolin一1一specifi

7、csiRNAandcaveolin一1WasoverexpressedbytransfectionwithpcDNA3．0一caveolin．1．n1eHERGchannelcurrentswerere—cordedbythepatch—clamptechnique．ResultsAsignificantincreaseintheHERGcurrentamplitudeandafasterdeactivationofthetai1currentwereobservedintheHEK293／HERGcellswit

8、hlipidraftsdisruptionandinthecaveolin．1一inhibitedHEK293／HERGcells．TheHERGcurrentamplitudewassignificantlyreducedandthetailcurentdeactivatedmoreslowlyintheHEK293／HERGce11swiththeove