Innovative Therapies in Hemostasis, INSERM 1140, Université de Paris, Paris, France.
Clinical Biochemistry Department, Necker Hospital, Assistance Publique-Hôpitaux de Paris (AP-HP), Paris, France.
Blood Adv. 2021 Mar 23;5(6):1706-1718. doi: 10.1182/bloodadvances.2020002847.
Blood platelets are essential for controlling hemostasis. They are released by megakaryocytes (MKs) located in the bone marrow, upon extension of cytoplasmic protrusions into the lumen of bone marrow sinusoids. Their number increases in postpulmonary capillaries, suggesting a role for oxygen gradient in thrombopoiesis (ie, platelet biogenesis). In this study, we show that initiation of thrombopoiesis from human mature MKs was enhanced under hyperoxia or during pro-oxidant treatments, whereas antioxidants dampened it. Quenching mitochondrial reactive oxygen species (mtROS) with MitoTEMPO decreased thrombopoiesis, whereas genetically enhancing mtROS by deacetylation-null sirtuin-3 expression increased it. Blocking cytosolic ROS production by NOX inhibitors had no impact. Classification according to the cell roundness index delineated 3 stages of thrombopoiesis in mature MKs. Early-stage round MKs exhibited the highest index, which correlated with low mtROS levels, a mitochondrial tubular network, and the mitochondrial recruitment of the fission activator Drp1. Intermediate MKs at the onset of thrombopoiesis showed high mtROS levels and small, well-delineated mitochondria. Terminal MKs showed the lowest roundness index and long proplatelet extensions. Inhibiting Drp1-dependent mitochondrial fission of mature MKs by Mdivi-1 favored a tubular mitochondrial network and lowered both mtROS levels and intermediate MKs proportion, whereas enhancing Drp1 activity genetically had opposite effects. Reciprocally, quenching mtROS limited mitochondrial fission in round MKs. These data demonstrate a functional coupling between ROS and mitochondrial fission in MKs, which is crucial for the onset of thrombopoiesis. They provide new molecular cues that control initiation of platelet biogenesis and may help elucidate some unexplained thrombocytopenia.
血小板对于控制止血至关重要。它们由骨髓中的巨核细胞（MKs）释放，细胞质突起延伸到骨髓窦腔时释放。它们在肺毛细血管后数量增加，表明氧梯度在血小板生成（即血小板发生）中起作用。在这项研究中，我们表明，在高氧或在促氧化剂处理下，人类成熟 MKs 的血小板生成起始增强，而抗氧化剂则减弱了它。用 MitoTEMPO 淬灭线粒体活性氧（mtROS）会减少血小板生成，而通过去乙酰化无效 Sirtuin-3 表达来增强 mtROS 则会增加血小板生成。用 NOX 抑制剂阻断细胞质 ROS 产生没有影响。根据细胞圆形度指数的分类，将成熟 MKs 的血小板生成划分为 3 个阶段。早期圆形 MKs 表现出最高的指数，与低 mtROS 水平、线粒体管状网络和分裂激活因子 Drp1 的线粒体募集相关。处于血小板生成起始阶段的中期 MKs 表现出高 mtROS 水平和小而清晰的线粒体。晚期 MKs 表现出最低的圆形度指数和长的伸展出的原血小板。用 Mdivi-1 抑制成熟 MKs 中依赖 Drp1 的线粒体分裂有利于管状线粒体网络的形成，并降低 mtROS 水平和中间 MKs 的比例，而通过遗传增强 Drp1 活性则有相反的效果。相反，淬灭 mtROS 限制了圆形 MKs 中的线粒体分裂。这些数据表明 ROS 和线粒体分裂在 MKs 中存在功能偶联，这对于血小板生成的起始至关重要。它们提供了控制血小板发生起始的新分子线索，可能有助于阐明一些未解释的血小板减少症。
