Dual regulation of cyclic AMP formation by thrombin in HEL cells, a leukaemic cell line with megakaryocytic properties.
Dual regulation of cyclic AMP formation by thrombin in HEL cells, a leukaemic cell line with megakaryocytic properties.
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HEL 细胞（一种具有巨核细胞特性的白血病细胞系）中凝血酶对环 AMP 形成的双重调节。
DOI：
10.1042/bj2810073
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发表时间：
1992
期刊：
影响因子：
--
通讯作者：
Woolkalis,MJ
中科院分区：
文献类型：
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作者：
Brass,LF;Woolkalis,MJ
凝血酶被认为是通过切割细胞表面受体来刺激应答细胞，所述细胞表面受体经由G蛋白与细胞内第二信使产生酶偶联。为了更好地理解这一过程，我们研究了巨核细胞系HEL细胞中凝血酶对腺苷酸环化酶的调节，并将其与血小板进行了比较。一个显着的差异被发现。在HEL-cell膜制剂，凝血酶抑制环腺苷酸（cAMP）的形成百日咳毒素敏感的机制与血小板中观察到的。与此相反，当加入到完整的HEL细胞，凝血酶激活腺苷酸环化酶，并引起cAMP形成的增加协同所产生的毛喉素和前列腺素I2。这种增加，这是没有看到与血小板，伴随着增加cAMP代谢的磷酸二酯酶。与其他对凝血酶的反应一样，cAMP形成的增加需要具有蛋白水解活性的凝血酶，并受到同源脱敏的影响。通过添加来自凝血酶受体的N-末端的多肽可以引起等效的反应，该多肽已经显示出激活受体。然而，凝血酶的作用不能通过添加佛波酯和Ca 2+离子载体A23187来再现，也不能用花生四烯酸代谢抑制剂来防止。预孵育的细胞与肾上腺素，抑制GS介导的腺苷酸环化酶的激活，或百日咳毒素，抑制磷脂酶C的激活，凝血酶诱导的cAMP的形成没有影响。这些结果表明，凝血酶可以通过两种不同的机制来调节cAMP的形成。首先，凝血酶可以以GI依赖性方式抑制腺苷酸环化酶。这种效应在HEL-cell膜制剂中占主导地位，就像在血小板中一样，但当凝血酶加入到完整的HEL细胞中时检测不到。相反，在完整的HEL细胞中，凝血酶激活腺苷酸环化酶。虽然明显受体介导的，这种反应似乎不涉及Gi，Gs，蛋白激酶C，类花生酸形成或细胞溶质Ca 2+浓度的变化。
Thrombin is thought to stimulate responsive cells by cleaving cell-surface receptors coupled to intracellular second-messenger-generating enzymes via G-proteins. In order to understand this process better, we have examined the regulation of adenylate cyclase by thrombin in the megakaryoblastic HEL cell line and compared it with platelets. A notable difference was found. In HEL-cell membrane preparations, thrombin inhibited cyclic AMP (cAMP) formation by a pertussis-toxin-sensitive mechanism comparable with that observed in platelets. In contrast, when added to intact HEL cells, thrombin activated adenylate cyclase and caused an increase in cAMP formation synergistic with that produced by forskolin and prostaglandin I2. This increase, which was not seen with platelets, was accompanied by an increase in cAMP metabolism by phosphodiesterase. Like other responses to thrombin, the increase in cAMP formation required proteolytically active thrombin and was subject to homologous desensitization. An equivalent response could be evoked by the addition of a polypeptide, derived from the N-terminus of the thrombin receptor, that has been shown to activate the receptor. The effects of thrombin could not, however, be reproduced by the addition of phorbol ester and the Ca2+ ionophore, A23187, nor be prevented with inhibitors of arachidonate metabolism. Preincubation of the cells with adrenaline, which inhibited Gs-mediated activation of adenylate cyclase, or pertussis toxin, which inhibited phospholipase C activation, had no effect on thrombin-induced cAMP formation. These results suggest that thrombin can regulate cAMP formation by two different mechanisms. First, thrombin can inhibit adenylate cyclase in a Gi-dependent manner. This effect predominates in HEL-cell membrane preparations, as it does in platelets, but is not detectable when thrombin is added to intact HEL cells. Instead, in intact HEL cells thrombin activates adenylate cyclase. Although clearly receptor-mediated, this response does not appear to involve Gi, Gs, protein kinase C, eicosanoid formation or changes in the cytosolic Ca2+ concentration.