A novel isothiourea derivative selectively inhibits the reverse mode of Na+/Ca2+ exchange in cells expressing NCX1
A novel isothiourea derivative selectively inhibits the reverse mode of Na+/Ca2+ exchange in cells expressing NCX1
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DOI：
10.1074/jbc.271.37.22391
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发表时间：
1996-09-13
影响因子：
4.8
通讯作者：
Shigekawa, M
中科院分区：
文献类型：
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作者：
Iwamoto, T;Watano, T;Shigekawa, M
新合成的No.7943 (2-[2-[4-(4-硝基苯甲氧基)苯基]乙基]异硫脲甲磺酸盐)是Na+/Ca2+交换器(NCX1)的选择性抑制剂，它剂量依赖性地抑制心肌细胞、平滑肌细胞和平滑肌细胞中Na-i(+)依赖性Ca-45(2+)摄取和Na-i(+)依赖性[Ca2+](i)增加。 NCX1转染的成纤维细胞（IC50=1.2-2.4μM）。无需预先与试剂孵育即可观察到抑制作用，并通过用缓冲液洗涤细胞 1 分钟完全逆转。有趣的是，No.7943 在抑制 Na-o(+) 依赖性 Ca-45(2+) 外流和 Na-o(+) 诱导的 [Ca2+](i) 下降（IC50 = >30 mu M）方面的效力要弱得多，表明它选择性地阻断完整细胞中 Na+/Ca2+ 交换的反向模式。在主要由内向外囊泡组成的心脏肌膜制剂中，该药物抑制 Na-i(+) 依赖性 Ca-45(2+) 摄取和 Na-o(+) 依赖性 Ca-45(2+) 流出，效力相似但略低（IC50 = 5.4-13 mu M）。细胞和肌膜囊泡中的抑制对于 Ca2+ 和 Na+ 来说都是非竞争性的。这些结果表明，No.7943 主要作用于完整细胞中转运位点以外的外部交换位点，尽管它能够从质膜两侧抑制交换剂，但高达 10 μM 的 No.7943 不会影响许多其他离子转运蛋白或几个心脏动作电位参数。这些浓度的该药物也不影响心肌细胞的舒张期 [Ca2+](i) 或自发搏动。此外，No.7943 显着抑制 Ca2+ 悖论条件下心肌细胞内的 Ca2+ 超载。因此，No.7943 不仅可用作研究 Na+/Ca2+ 交换器的转运机制和生理作用的工具，而且还具有作为病理条件下通过 Na+/Ca2+ 交换器介导的过量 Ca2+ 内流的选择性阻断剂的治疗潜力。
No.7943 (2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea methanesulfonate), a selective inhibitor of the Na+/Ca2+ exchanger (NCX1), has been newly synthesized, It dose-dependently inhibited Na-i(+)-dependent Ca-45(2+) uptake and Na-i(+)-dependent [Ca2+](i) increase in cardiomyocytes, smooth muscle cells, and NCX1-transfected fibroblasts (IC50 = 1.2-2.4 mu M). Inhibition was observed without prior incubation with the agent and was completely reversed by washing cells with buffer for 1 min. Interestingly, No.7943 was much less potent in inhibiting Na-o(+)-dependent Ca-45(2+) efflux and Na-o(+)-induced [Ca2+](i) decline (IC50 = >30 mu M), indicating that it selectively blocks the reverse mode of Na+/Ca2+ exchange in intact cells. In cardiac sarcolemmal preparations consisting mostly of inside-out vesicles, the agent inhibited Na-i(+)-dependent Ca-45(2+) uptake and Na-o(+)-dependent Ca-45(2+) efflux with similar, but slightly lower, potencies (IC50 = 5.4-13 mu M). Inhibition was noncompetitive with respect to Ca2+ and Na+ in both cells and sarcolemmal vesicles. These results suggest that No.7943 primarily acts on external exchanger site(s) other than the transport sites in intact cells, although it is able to inhibit the exchanger from both sides of the plasma membrane, No.7943 at up to 10 mu M does not affect many other ion transporters nor several cardiac action potential parameters. This agent at these concentrations also did not influence either diastolic [Ca2+](i) or spontaneous beating in cardiomyocytes. Furthermore, No.7943 markedly inhibited Ca2+ overloading into cardiomyocytes under the Ca2+ paradox conditions. Thus, No.7943 is not only useful as a tool with which to study the transport mechanism and physiological role of the Na+/Ca2+ exchanger but also has therapeutic potential as a selective blocker of excessive Ca2+ influx mediated via the Na+/Ca2+ exchanger under pathological conditions.