Abstract Number: PB1618
Meeting: ISTH 2020 Congress
Theme: Platelets and Megakaryocytes » Megakaryocytes and Thrombopoiesis
Background: Thrombocytopenia, or low circulating platelet count, is often caused by a defect in megakaryocyte (MK) development. MKs reside primarily in the bone marrow, where they develop from undifferentiated multipotent cells into highly specialized, polyploidy cells that undergo the process of proplatelet formation (PPF). It is currently not clear if and how Rap1, a member of the Ras small GTPase superfamily, contributes to platelet generation.
Aims: We hypothesized that the two Rap1 isoforms, Rap1a and Rap1b, are critical in MK development and/or PPF.
Methods: We studied MK development in primary murine MKs isolated from conditional knockout mice lacking Rap1a (Rap1amKO), Rap1b (Rap1bmKO), or both isoforms (Rap1a,bmKO) in MKs and platelets. Flow cytometry, immunohistochemistry and 2-photon/confocal/electron microscopy imaging were used to analyze MKs in vivo, ex vivo, in situ, and in vitro.
Results: Rap1amKO mice exhibited a normal peripheral platelet count (PPC) but significantly increased MK numbers in the bone marrow and spleen. Compared to controls, MKs from Rap1amKO mice were smaller and less differentiated (lower ploidy). Rap1bmKO mice exhibited a mild macrothrombocytopenia but normal MK numbers and ploidy. However, PPF in Rap1bmKO MKs was impaired, both in vivo and ex vivo (few but very large proplatelets). Rap1a,bmKO mice exhibited a marked macrothrombocytopenia, impaired MK differentiation and a dramatic defect in PPF. Rap1bmKO and Rap1a,bmKO MKs exhibited defects in demarcation membrane system (DMS) biogenesis and the association of the DMS with F-actin in proplatelet extensions. Consistent with the abnormal DMS, LIM kinase and cofilin signaling was altered in Rap1a,bmKO mice, and LIM kinase inhibitor treatment normalized the platelet count in the mutant mice.
Conclusions: These results provide the first definitive proof for a critical role of Rap1 GTPases in MK biology. Future studies will have to show whether some inherited or acquired thrombocytopenias are caused, at least in part, by altered Rap1 signaling in MKs.
To cite this abstract in AMA style:
Ghalloussi D, Harousseau G, Eckly A, Kenny JP, Kirkpatrick C, O'Shaughnessy EC, Hahn K, Gachet C, Stefanini L, Eto K, Bergmeier W. Rap1a/b Isoforms Differentially Regulate Megakaryocyte Biology [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/rap1a-b-isoforms-differentially-regulate-megakaryocyte-biology/. Accessed March 21, 2024.« Back to ISTH 2020 Congress
ISTH Congress Abstracts - https://abstracts.isth.org/abstract/rap1a-b-isoforms-differentially-regulate-megakaryocyte-biology/