Abstract Number: OC 74.3
Meeting: ISTH 2021 Congress
Background: To produce platelets, megakaryocytes (MKs) must cross the sinusoidal barrier of the bone marrow (BM). We have previously shown that MKs use podosomes to create transendothelial pores, enabling their transmigration. Although integrins are presumed dispensable for thrombopoiesis since patients with Glanzmann’s thrombasthenia have normal platelet counts, they are essential in the formation of podosomes in several cells.
Aims: We evaluated the role of ß1 et ß3 integrins, the two forms expressed by MKs, in the final stages of megakaryopoiesis, i.e. podosome formation and transmigration.
Methods: We generated double knockout mice lacking ß1 and ß3 integrins (Itgb1-/-b3-/-) in the MK/platelet lineage. Using complementary imaging approaches, podosome formation was investigated in vitro and in situ, and transmigration was examined in situ and in vivo.
Results: We observed no difference in podosome formation in vitro (density, number of MKs displaying podosomes) between Itgb1-/-b3-/- and control MKs immobilized on fibronectin, matrigel or endothelial cells. In situ, Itgb1-/-b3-/- MKs extended normal invasive podosomes leading to the formation of transendothelial pores, suggesting that the integrins are dispensable to MK transendothelial penetration. However, electron microscopy revealed the presence of entire Itgb1-/- b3-/- MKs inside the vascular lumen of sinusoid. This was confirmed by immunofluorescence on thick fixed BM sections (23.6±3.1% vs 3.8±1.4%) and by dynamic intravital 2-photon microscopy. Accordingly, large polypoid MK nucleus were found in the pulmonary microvessel of these mice. Transmigration of entire Itgb1-/-b3-/- MKs is unlikely to result from an increased size of endothelial pores, indicating other mechanisms. We investigated integrin-mediated adhesion and found that Itgb1-/-b3-/- MKs plated on fibronectin displayed a round shape compared to spread control MKs, suggesting weaker adhesive properties of these cells.
Conclusions: ß1 and ß3 integrins limit the transmigration of entire MKs, likely by modulating their anchoring to their BM microenvironment and/or the deformability of their polylobulated nucleus.
To cite this abstract in AMA style:Scandola C, Rinckel J-, Proamer F, Hoffmann D, Bornert A, Léon C, Laquerrière P, Lanza F, de la Salle H, Gachet C, Eckly A. ß1 and ß3 Integrins Limit the Passage of Entire Megakaryocytes across the Sinusoids of the Bone Marrow [abstract]. Res Pract Thromb Haemost. 2021; 5 (Suppl 2). https://abstracts.isth.org/abstract/s1-and-s3-integrins-limit-the-passage-of-entire-megakaryocytes-across-the-sinusoids-of-the-bone-marrow/. Accessed November 29, 2021.
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