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Ferrata Storti Foundation
Haematologica 2021 Volume 106(5):1368-1380
Hematopoiesis
Cytoskeletal-based mechanisms differently regulate in vivo and in vitro proplatelet formation
Alicia Bornert,1 Julie Boscher,1 Fabien Pertuy,1 Anita Eckly,1 David Stegner,2 Catherine Strassel,1 Christian Gachet,1 François Lanza1 and Catherine Léon1
1Université de Strasbourg, INSERM, EFS Grand-Est, BPPS UMR-S 1255, Strasbourg, France and 2Institute of Experimental Biomedicine, University Hospital Würzburg & Rudolf Virchow Center for Experimental Biomedicine, University of Würzburg, Würzburg, Germany
ABSTRACT
Platelets are produced by bone marrow megakaryocytes through cytoplasmic protrusions, named native proplatelets (nPPT), into blood vessels. Proplatelets also refer to protrusions observed in megakaryocyte culture (cultured proplatelets [cPPT]) which are morpho- logically different. Contrary to cPPT, the mechanisms of nPPT formation are poorly understood. We show here in living mice that nPPT elongation is in equilibrium between protrusion and retraction forces mediated by myosin-IIA. We also found, using wild-type and b1-tubulin-deficient mice, that microtubule behavior differs between cPPT and nPPT, being absolutely required in vitro, while less critical in vivo. Remarkably, micro- tubule depolymerization in myosin-deficient mice did not affect nPPT elongation. We then calculated that blood Stokes’ forces may be sufficient to promote nPPT extension, independently of myosin and microtubules. Together, we propose a new mechanism for nPPT extension that might explain contradictions between severely affected cPPT production and moderate platelet count defects in some patients and animal models.
Introduction
Blood platelets are key elements of hemostasis for the prevention of bleeding and are produced by a unique mechanism. They arise from megakaryocytes (MK), spe- cialized cells in the bone marrow.1,2 Upon differentiation from hematopoietic pro- genitors, MKs undergo endomitosis, leading to a giant cell, whose cytoplasm is filled by a highly developed intracellular membrane network called the demarcation membrane system (DMS).3 At a mature stage, MK lie adjacent to the bone marrow sinusoid vessels and initiate cytoplasmic protrusion through the vessel wall. These protrusions further elongate inside the blood circulation, attached to their mother cells. These extensions are named proplatelets (PPT) and are fueled by the DMS that acts as a membrane reservoir to allow PPT growth.4 Once inside the blood stream, PPT are released into the circulation as large fragments that have been proposed to further remodel in downstream organs to release bona fide platelets, small anucle- ated MK fragments having a discoid shape.2,5,6
The first PPT term was proposed following in situ scanning electron microscopy (SEM) observation of “long intrasinusoidal "proplatelet" processes which originate from the cell body of extravascularly located megakaryocytes”.7 Later on, in vivo observations by time lapse imaging in living animal confirmed the morphology of PPT as elongated protrusions in wild-type (WT) mice under physiological condi- tions.7-14 The same denomination was also given to the cytoplasmic MK extensions in culture or in bone marrow explants.8,15-19 Yet, the morphology of PPT observed in vitro strongly differs from that observed in situ/in vivo. Early in vitro observations of marrow explants8,15,16,20 and later of progenitor-differentiated MK in culture17,19,21 sim- ilarly recorded PPT presenting branched thin shafts (1-4 mm in diameter) leading to an entanglement of PPT surrounding the MK body.17-19 These morphological differ- ences between these two types of MK extensions raise the possibility that the mechanisms at stake could as well differ between in vitro cultured PPT (hereafter
Correspondence:
CATHERINE LÉON
catherine.leon@efs.sante.fr
Received: September 23, 2019. Accepted: April 14, 2020. Pre-published: April 23, 2020.
https://doi.org/10.3324/haematol.2019.239111
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