Derian C K, Santulli R J, Tomko K A, Haertlein B J, Andrade-Gordon P
Drug Discovery Research, The R. W. Johnson Pharmaceutical Research Institute Spring House, PA 19477, USA.
Thromb Res. 1995 Jun 15;78(6):505-19. doi: 10.1016/0049-3848(95)00084-5.
The thrombin receptor on human platelets is activated by thrombin to stimulate platelet aggregation through the tethered ligand SFLLRN. This study examined the effects of thrombin and SFLLRN on aggregation and calcium mobilization ([Ca2+]i) in rat, guinea pig, rabbit, dog, monkey, and human platelets, and the role of protein kinases in regulating these functions. Thrombin induced platelet aggregation and [Ca2+]i in all species studied; however, only guinea pig, monkey and human platelets were responsive to SFLLRN. Similar species specific effects were obtained with [Ca2+]i studies. The kinetic profile for [Ca2+]i differed among species, suggesting that regulatory mechanisms for calcium differed between agonists and among species. Staurosporine, a non-selective inhibitor of protein kinases, inhibited platelet aggregation induced by thrombin or SFLLRN in all species. Staurosporine inhibited thrombin-induced [Ca2+]i in guinea pigs, had no effect in rat, and increased [Ca2+]i in all other species. Staurosporine inhibited SFLLRN-induced [Ca2+]i in guinea pig, yet had no effect in monkey or human. Tyrphostin 23, a specific inhibitor of tyrosine protein kinases, inhibited thrombin-induced aggregation of rabbit, monkey, dog and human platelets. SFLLRN-induced aggregation was also inhibited by tyrphostin 23. Tyrphostin 23 inhibited [Ca2+]i induced by either thrombin or SFLLRN in all species. Based on the differential response to agonist stimulation, we propose that thrombin can activate platelets via SFLLRN-dependent and independent mechanisms, which could involve yet unrecognized subtypes of the thrombin receptor or distinct cellular activating mechanisms. Furthermore, differential regulation of calcium mobilization and aggregation was observed in those platelets responding to either thrombin or SFLLRN.
人血小板上的凝血酶受体被凝血酶激活，通过拴系配体SFLLRN刺激血小板聚集。本研究检测了凝血酶和SFLLRN对大鼠、豚鼠、兔、狗、猴和人血小板聚集及钙动员（[Ca2+]i）的影响，以及蛋白激酶在调节这些功能中的作用。凝血酶在所有研究的物种中均诱导血小板聚集和[Ca2+]i升高；然而，只有豚鼠、猴和人血小板对SFLLRN有反应。[Ca2+]i研究也获得了类似的物种特异性效应。不同物种间[Ca2+]i的动力学曲线不同，提示激动剂之间以及不同物种间钙的调节机制存在差异。蛋白激酶的非选择性抑制剂星形孢菌素在所有物种中均抑制凝血酶或SFLLRN诱导的血小板聚集。星形孢菌素抑制豚鼠凝血酶诱导的[Ca2+]i升高，对大鼠无影响，而在所有其他物种中则使[Ca2+]i升高。星形孢菌素抑制豚鼠SFLLRN诱导的[Ca2+]i升高，但对猴或人无影响。酪氨酸蛋白激酶的特异性抑制剂 tyrphostin 23抑制兔、猴、狗和人血小板凝血酶诱导的聚集。tyrphostin 23也抑制SFLLRN诱导的聚集。tyrphostin 23在所有物种中均抑制凝血酶或SFLLRN诱导的[Ca2+]i升高。基于对激动剂刺激的不同反应，我们提出凝血酶可通过依赖SFLLRN和不依赖SFLLRN的机制激活血小板，这可能涉及尚未被认识的凝血酶受体亚型或不同的细胞激活机制。此外，在对凝血酶或SFLLRN有反应的血小板中观察到钙动员和聚集的差异调节。
