Stabilizing effects of eicosapentaenoic acid on Kv1.5 channel protein expressed in mammalian cells
Stabilizing effects of eicosapentaenoic acid on Kv1.5 channel protein expressed in mammalian cells
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DOI：
10.1016/j.ejphar.2008.12.016
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发表时间：
2009-02-14
影响因子：
5
通讯作者：
Hisatome, Ichiro
中科院分区：
文献类型：
--
作者：
Koshida, Shunya;Kurata, Yasutaka;Hisatome, Ichiro
研究了二十碳五烯酸（EPA）和二十二碳六烯酸（DHA）对Kv1.5通道蛋白稳定性的影响。采用免疫印迹、免疫沉淀、免疫荧光和膜片钳技术检测转染的非洲绿猴肾成纤维细胞和大鼠心房中Kv1.5 （Kv1.5- flag）的表达和功能。EPA和DHA都以剂量依赖性的方式立即阻断Kv1.5通道电流，并伴有磷酸化的降低。低浓度EPA （0.3 ~ 10 μ M）慢性处理（12 h）可增加Kv1.5- flag蛋白水平和Kv1.5通道电流，但其门控动力学未发生变化，半衰期延长；较高浓度EPA和DHA （30 ~ 100 μ M）均降低了Kv1.5-FLAG的表达。高浓度EPA还降低了Kv1.5 mRNA和突触相关蛋白97的表达。低浓度EPA使内质网、高尔基体和细胞膜中Kv1.5的表达增加。用蛋白转运抑制剂brefeldin A或秋水仙碱和Kv1.5通道阻滞剂4-氨基吡啶预处理可消除epa诱导的Kv1.5通道表达和电流的增加。口服EPA （30 mg/kg）可提高大鼠心房内源性Kv1.5水平。这些结果表明，长期使用低浓度的EPA可以稳定内质网和高尔基体中的Kv1.5通道蛋白，从而增强细胞膜上的Kv1.5通道电流。(C) 2008 Elsevier B.V.版权所有
We investigated the effects of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on the stability of Kv1.5 channel protein. The expression and function of Kv1.5 (Kv1.5-FLAG) in transfected African green monkey kidney fibroblast cells as well as rat atrium were estimated by immunoblotting, immunoprecipitation, immunofluorescence and patch-clamp techniques. Both EPA and DHA immediately blocked Kv1.5 channel current in a dose-dependent manner, accompanied by reduction of their phosphorylation. Chronic treatment (for 12 h) with EPA at lower concentrations (0.3-10 mu M) increased the level of Kv1.5-FLAG protein as well as Kv1.5 channel current without changes in its gating kinetics, prolonging its half-life; in contrast, both EPA and DHA at higher concentrations (30-100 mu M) decreased the expression of Kv1.5-FLAG. EPA at the higher concentrations also decreased mRNA of Kv1.5 and synapse-associated protein 97 expression. EPA at the lower concentrations increased Kv1.5 expression in the endoplasmic reticulum, Golgi apparatus and cell membrane. EPA-induced increase of Kv1.5 channel expression and current was abolished by pretreatment with the protein transport inhibitor brefeldin A or colchicines, and by the Kv1.5 channel blocker 4-aminopyridine. Oral administration of EPA (30 mg/kg) increased the level of endogenous Kv1.5 in rat atria. These results indicate that chronic treatment with EPA at lower concentrations stabilizes Kv1.5 channel protein in the endoplasmic reticulum and Golgi apparatus thereby enhancing the Kv1.5 channel current on the cell membrane. (C) 2008 Elsevier B.V. All rights reserved.