Full activation of mouse platelets requires ADP secretion regulated by SERCA3 ATPase-dependent calcium stores
Full activation of mouse platelets requires ADP secretion regulated by SERCA3 ATPase-dependent calcium stores
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DOI：
10.1182/blood-2015-10-678383
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发表时间：
2016-08-25
期刊：
影响因子：
20.3
通讯作者：
Rosa, Jean-Philippe
中科院分区：
文献类型：
--
作者：
Elaib, Ziane;Adam, Frederic;Rosa, Jean-Philippe
肌质内质网钙（Ca 2+）腺苷三磷酸酶（ATP酶）3（SERCA 3）在血小板生理学中的作用仍然知之甚少。在这里，我们表明SERCA 3敲除（SERCA 3（-/-））小鼠表现出延长的尾部出血时间和再出血。在体内氯化铁诱导的血栓形成模型中，动脉和小静脉中的血栓形成延迟。体外实验证实血小板粘附和血栓在胶原蛋白上生长不良。通过腺苷三磷酸双磷酸酶去除5 '-二磷酸腺苷（ADP）可将对照血小板的粘附和血栓生长降低至SERCA 3（-/-）血小板的水平。低胶原或低凝血酶浓度诱导的SERCA 3（-/-）血小板聚集、致密颗粒分泌和Ca 2+动员均弱于对照组。因此，SERCA 3（-/-）血小板在凝血酶刺激后表现出总储存Ca 2+和Ca 2+储存再摄取的部分缺陷。重要的是，ADP，而不是5-羟色胺，挽救了SERCA 3（-/-）血小板的聚集，分泌和Ca 2+动员，表明特异性。电子显微镜检查、米帕林染色和总5-羟色胺含量显示致密颗粒正常，排除了致密颗粒缺陷。ADP诱导正常的血小板聚集，排除ADP激活途径的缺陷。与SERCA 2b抑制剂毒胡萝卜素相反，SERCA 3特异性抑制剂2，5-二-（叔丁基）-1，4-苯并氢醌减少了对照血小板的Ca 2+动员和分泌。这证实了催化活性SERCA 3在ADP分泌中的特定作用。因此，SERCA 3依赖性Ca 2+储存似乎在SERCA 3（-/-）血小板中耗尽。最后，α（IIb）β（3）整联蛋白阻断不影响SERCA 3依赖性分泌，因此证明不依赖于α（IIb）β（3）接合。总之，这些结果表明，SERCA 3依赖性Ca 2+储存控制着一条特异性ADP分泌途径，这条途径是由低浓度激动剂诱导的血小板完全分泌所必需的，与α（IIb）β无关（3）。
The role of the sarco-endoplasmic reticulum calcium (Ca2+) adenosine triphosphatase (ATPase) 3 (SERCA3) in platelet physiology remains poorly understood. Here, we show that SERCA3 knockout (SERCA3(-/-)) mice exhibit prolonged tail bleeding time and rebleeding. Thrombus formation was delayed both in arteries and venules in an in vivo ferric chloride-induced thrombosis model. Defective platelet adhesion and thrombus growth over collagen was confirmed in vitro. Adenosine 5'-diphosphate (ADP) removal by apyrase diminished adhesion and thrombus growth of control platelets to the level of SERCA3(-/-) platelets. Aggregation, dense granule secretion, and Ca2+ mobilization of SERCA3(-/-) platelets induced by low collagen or low thrombin concentration were weaker than controls. Accordingly, SERCA3(-/-) platelets exhibited a partial defect in total stored Ca2+ and in Ca2+ store reuptake following thrombin stimulation. Importantly ADP, but not serotonin, rescued aggregation, secretion, and Ca2+ mobilization in SERCA3(-/-) platelets, suggesting specificity. Dense granules appeared normal upon electron microscopy, mepacrine staining, and total serotonin content, ruling out a dense granule defect. ADP induced normal platelet aggregation, excluding a defect in ADP activation pathways. The SERCA3-specific inhibitor 2,5-di-(tert-butyl)-1,4-benzohydroquinone diminished both Ca2+ mobilization and secretion of control platelets, as opposed to the SERCA2b inhibitor thapsigargin. This confirmed the specific role of catalytically active SERCA3 in ADP secretion. Accordingly, SERCA3-dependent Ca2+ stores appeared depleted in SERCA3(-/-) platelets. Finally, alpha(IIb)beta(3) integrin blockade did not affect SERCA3-dependent secretion, therefore proving independent of alpha(IIb)beta(3) engagement. Altogether, these results show that SERCA3-dependent Ca2+ stores control a specific ADP secretion pathway required for full platelet secretion induced by agonists at low concentration and independent of alpha(IIb)beta(3).