Asano M, Nakajima T, Iwasawa K, Asakura Y, Morita T, Nakamura F, Tomaru T, Wang Y, Goto A, Toyo-oka T, Soma M, Suzuki S, Okuda Y
Second Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Japan.
Eur J Pharmacol. 1999 Aug 27;379(2-3):199-209. doi: 10.1016/s0014-2999(99)00476-8.
The purpose of this study was to clarify how eicosapentaenoic acid (EPA), an omega-3 polyunsaturated fatty acid, modulates the vascular action of vasopressin in rat aortic smooth muscle cell lines. The effects of EPA on Ca2+ mobilization and DNA synthesis elicited by vasopressin were investigated and compared to those of Ca2+ channel blocking agents, by means of Ca2+ measurements and the incorporation of [3H]thymidine. Patch-clamp techniques were also employed. Vasopressin (100 nM) elicited an initial peak of intracellular Ca2+ ([Ca2+]i), followed by a sustained phase due to Ca2+ entry. Nifedipine or nicardipine (1 microM), a potent L-type Ca2+ channel blocker, partly inhibited the sustained phase, but La3+ completely abolished it. EPA (10 microM) also inhibited it even in the presence of nicardipine. Under voltage-clamp conditions with CsCl-internal solution, depolarizing pulses positive to -30 mV from a holding potential of -40 mV elicited a slow inward current. The inward current was blocked by La3+, nicardipine, and nifedipine (1 microM), suggesting that the inward current mainly consisted of the voltage-dependent L-type Ca2+ channel (ICa.L). EPA (1-30 microM) also inhibited ICa.L in a concentration-dependent manner. The inhibitory effect of EPA was observed at concentrations higher than 1 microM, and its half-maximal inhibitory concentration (IC50) was 7.6 microM. Vasopressin induced a long-lasting inward current at a holding potential of -40 mV. The vasopressin-induced current was considered as a non-selective cation current (Icat) with a reversal potential of approximately +0 mV. Both nifedipine and nicardipine (10 microM) failed to inhibit it significantly, but La3+ completely abolished Icat. EPA also inhibited vasopressin-induced Icat in a concentration-dependent manner; its IC50 value was 5.9 microM. Vasopressin (100 nM) stimulated [3H]thymidine incorporation. Exclusion of extracellular Ca2+ with EGTA or La3+ markedly inhibited it. EPA (3-30 microM) also inhibited the incorporation induced by vasopressin, while nifedipine and nicardipine (1 microM) only partly inhibited it. These results suggested that EPA, unlike nifedipine and nicardipine, inhibited vasopressin-induced Ca2+-entry and proliferation in rat vascular smooth muscle cells, where the inhibitory effects of EPA on Icat as well as ICa.L might be involved. Thus, EPA would exert hypotensive and antiatherosclerotic effects.
本研究的目的是阐明ω-3多不饱和脂肪酸二十碳五烯酸（EPA）如何调节大鼠主动脉平滑肌细胞系中血管加压素的血管作用。通过钙离子测量和[3H]胸苷掺入，研究了EPA对血管加压素引发的钙离子动员和DNA合成的影响，并与钙离子通道阻滞剂的作用进行了比较。还采用了膜片钳技术。血管加压素（100 nM）引发细胞内钙离子（[Ca2+]i）的初始峰值，随后由于钙离子内流进入持续阶段。强效L型钙离子通道阻滞剂硝苯地平或尼卡地平（1 μM）部分抑制了持续阶段，但La3+完全消除了该阶段。即使在存在尼卡地平的情况下，EPA（10 μM）也能抑制该阶段。在使用氯化铯内部溶液的电压钳条件下，从-40 mV的保持电位去极化至-30 mV以上的脉冲引发缓慢内向电流。该内向电流被La3+、尼卡地平和硝苯地平（1 μM）阻断，表明该内向电流主要由电压依赖性L型钙离子通道（ICa.L）组成。EPA（1 - 30 μM）也以浓度依赖性方式抑制ICa.L。EPA在高于1 μM的浓度下观察到抑制作用，其半数最大抑制浓度（IC50）为7.6 μM。血管加压素在-40 mV的保持电位下诱导出持久的内向电流。血管加压素诱导的电流被认为是一种非选择性阳离子电流（Icat），其反转电位约为 +0 mV。硝苯地平和尼卡地平（10 μM）均未能显著抑制该电流，但La3+完全消除了Icat。EPA也以浓度依赖性方式抑制血管加压素诱导的Icat；其IC50值为5.9 μM。血管加压素（100 nM）刺激[3H]胸苷掺入。用EGTA或La3+排除细胞外钙离子可显著抑制该过程。EPA（3 - 30 μM）也抑制血管加压素诱导的掺入，而硝苯地平和尼卡地平（1 μM）仅部分抑制。这些结果表明，与硝苯地平和尼卡地平不同，EPA抑制大鼠血管平滑肌细胞中血管加压素诱导的钙离子内流和增殖，其中EPA对Icat以及ICa.L的抑制作用可能起了作用。因此，EPA可能具有降压和抗动脉粥样硬化作用。
