Caveolin-1对心脏HERG钾通道功能调控作用的研究

Regulatory effect of Caveolin-1 on the cardiac HERG potassium channel

目的既往研究发现caveolin-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-1对心脏HERG钾通道功能的调控作用。方法①基因转染:应用Lipofectamine 2000将pcDNA3.0-caveolin-1质粒和pcDNA3.0-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-β-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-1表达;④Caveolin-1过度表达:将pcDNA3.0-herg转染HEK293/caveolin-1细胞株,获得caveolin-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①细胞膜脂筏破坏与caveolin-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-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。

Objective Previous studies show that caveolin-1 is involved in the regnlation of multiple ion channels. However, no report has been seen on its regulatory effect on the cardiac HERG channel function. This study was to investigate the regulatory func- tion of caveolin-1 on the cardiac HERG potassium channel. Methods HEK293 cells were transfected with pcDNA3.0-caveolin-1 or pcDNA3.0-berg by Lipofectamine 2000, and G418 was used to screen stably expressed cell lines. The lipid rafts in the cell membrane were disrupted by incubation with 10 mmol/L methyl-13-eyelodextrin. The expression of eaveolin-1 was knocked down by transfected caveolin-l-specific siRNA and eavcolin-1 was overexpressed by transfection with pcDNA3.0-caveolin-1. The HERG channel currents were recorded by the patch-clamp technique. Results A significant increase in the HERG current amplitude and a faster deactivation of the tail current were observed in the HEK293/HERG cells with lipid rafts disruption and in the caveolin-l-inhibited HEK293/HERG cells. The HERG current amplitude was significantly reduced and the tail current deactivated more slowly in the HEK293/HERG cells with the overexpression of caveolin-1. Conclusion Caveolin-1 negatively regulates the function of the cardiac HERG channel.