Department of Environmental and Preventive Medicine, Hyogo College of Medicine, Mukogawa-cho, Nishinomiya, Hyogo, Japan.
J Pharm Pharmacol. 2012 Jun;64(6):855-61. doi: 10.1111/j.2042-7158.2012.01485.x. Epub 2012 Mar 12.
The aim of this study was to determine whether diacylglycerol kinase (DGK) is involved in transplasmalemmal Ca²⁺ influx of platelets.
Effects of R59949, an inhibitor of diacylglycerol kinase, on intracellular Ca²⁺ concentration (Ca²⁺ ) and mRNA expression of DGK isozymes were investigated using washed human platelet suspensions.
Thrombin-induced increase in Ca²⁺ was significantly inhibited by pretreatment of platelets with R59949, while thapsigargin-induced increase in Ca²⁺ was comparable in platelets with and without R59949 pretreatment. Thapsigargin-induced increase in Ca²⁺ was markedly attenuated in the presence of SKF-96365. In the presence of SKF-96365, thrombin-induced increase in Ca²⁺ was significantly attenuated, and additional treatment with R59949 caused a further decrease in Ca²⁺ . Pretreatment of platelets with 1-butanol significantly attenuated thrombin-induced increase in Ca²⁺ , while thrombin-induced increase in Ca²⁺ was augmented in the presence of propranolol. mRNA expression of DGK-α and DGK-γ, which are known to be inhibited by R59949, in platelets was confirmed by RT-PCR analysis.
R59949 inhibited a store-depletion-insensitive component of transplasmalemmal Ca²⁺ entry induced by thrombin, while store-operated Ca²⁺ entry was not affected by R59949. The results of this study suggest that phosphatidic acid is involved in thrombin-induced Ca²⁺ influx of platelets.
本研究旨在探讨二酰基甘油激酶（DGK）是否参与血小板质膜 Ca²⁺内流。
采用人血小板悬液，观察二酰基甘油激酶抑制剂 R59949 对细胞内 Ca²⁺浓度([Ca²⁺]i)和 DGK 同工酶 mRNA 表达的影响。
R59949 预处理可显著抑制凝血酶诱导的[Ca²⁺]i 增加，而 R59949 预处理对血小板内 Ca²⁺浓度（[Ca²⁺]i）无影响。SKF-96365 存在时，钙诱导剂诱导的[Ca²⁺]i 增加明显减弱。在 SKF-96365 存在的情况下，凝血酶诱导的[Ca²⁺]i 增加明显减弱，而 R59949 的进一步处理导致[Ca²⁺]i 的进一步降低。血小板 1-丁醇预处理可显著减弱凝血酶诱导的[Ca²⁺]i 增加，而普萘洛尔存在时，凝血酶诱导的[Ca²⁺]i 增加增强。通过 RT-PCR 分析证实了 R59949 抑制的 DGK-α 和 DGK-γ的 mRNA 表达，这两种同工酶已知被 R59949 抑制。
R59949 抑制了凝血酶诱导的质膜 Ca²⁺内流的一种不依赖于储存耗竭的成分，而储存操作的 Ca²⁺内流不受 R59949 影响。本研究结果表明，磷酸脂酸参与了凝血酶诱导的血小板 Ca²⁺内流。
