Apoptosis signal-regulating kinase 1 (ASK1) is a member of mitogen-activated protein kinase kinase kinase (MAP3K) family, which recently has been implicated in the regulation of p38 MAPK/PLA2/thromboxane (TxA2) generation, as well as P2Y12 signalling in murine platelets. ASK1 has therefore been proposed as a potential target for anti-thrombotic therapy. At present it is unknown whether ASK1 also contributes to TxA2 formation and platelet function in human. In this study we therefore examined the role of ASK1 using the ASK1 inhibitor selonsertib (GS-4997). We established that ASK1 is responsible for p38 phosphorylation and TxA2 formation in murine platelets, with both GS4997 and p38 inhibitors reducing TxA2 formation. Similar to murine platelets, activation of human platelets resulted in the rapid and transient phosphorylation of ASK1 and the MAP2Ks MMK3/4/6. In contrast, phosphorylation of p38 and its substrate; MAPKAP-kinase2 (MAPKAPK2) was much more sustained. In keeping with these findings, inhibition of ASK1 blocked early, but not later p38/MAPKAPK2 phosphorylation. The latter was dependent on non-canonical autophosphorylation as it was blocked by the p38 inhibitor; SB203580 and the SYK inhibitor; R406. Furthermore, ASK1 and p38 inhibitors had no effect on PLA2 phosphorylation, TxA2 formation and platelet aggregation, demonstrating that this pathway is redundant in human platelets. Together, these results demonstrate that ASK1 contributes to TxA2 formation in murine, but not human platelets and highlight the importance of confirming findings from genetic murine models in humans.

中文翻译：

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ASK1 介导的 p38MAPK 激活在人血小板功能中的冗余作用。


细胞凋亡信号调节激酶 1 (ASK1) 是丝裂原激活蛋白激酶激酶激酶 (MAP3K) 家族的成员，最近被发现参与 p38 MAPK/PLA2/血栓烷 (TxA2) 生成以及 P2Y12 信号传导的调节在小鼠血小板中。因此，ASK1 被提议作为抗血栓治疗的潜在靶点。目前尚不清楚ASK1是否也有助于人类TxA2的形成和血小板功能。因此，在本研究中，我们使用 ASK1 抑制剂 selonsertib (GS-4997) 检查了 ASK1 的作用。我们确定 ASK1 负责小鼠血小板中 p38 磷酸化和 TxA2 形成，GS4997 和 p38 抑制剂均可减少 TxA2 形成。与鼠血小板类似，人血小板的激活导致 ASK1 和 MAP2Ks MMK3/4/6 快速且短暂的磷酸化。相反，p38及其底物的磷酸化； MAPKAP-激酶2 (MAPKAPK2) 的持续性要好得多。与这些发现一致，抑制 ASK1 会阻断早期而非晚期的 p38/MAPKAPK2 磷酸化。后者依赖于非典型自磷酸化，因为它被 p38 抑制剂阻断； SB203580和SYK抑制剂； R406。此外，ASK1 和 p38 抑制剂对 PLA2 磷酸化、TxA2 形成和血小板聚集没有影响，表明该途径在人类血小板中是多余的。总之，这些结果表明 ASK1 有助于小鼠 TxA2 的形成，但不促进人类血小板的形成，并强调了在人类遗传小鼠模型中确认研究结果的重要性。