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Thromb Haemost 2017; 117(12): 2322-2333
DOI: 10.1160/TH17-06-0433
Cellular Haemostasis and Platelets
Schattauer GmbH Stuttgart

Developmental Stage–Specific Manifestations of Absent TPO/c-MPL Signalling in Newborn Mice

Viola Lorenz   *
,
Haley Ramsey   *
,
Zhi-Jian Liu
,
Joseph Italiano Jr.
,
Karin Hoffmeister
,
Sihem Bihorel
,
Donald Mager
,
Zhongbo Hu
,
William B. Slayton
,
Benjamin T. Kile
,
Martha Sola-Visner   *
,
Francisca Ferrer-Marin   *
Further Information

Publication History

23 June 2017

08 September 2017

Publication Date:
06 December 2017 (online)

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Abstract

Congenital amegakaryocytic thrombocytopaenia (CAMT) is a disorder caused by c-MPL mutations that impair thrombopoietin (TPO) signalling, resulting in a near absence of megakaryocytes (MKs). While this phenotype is consistent in adults, neonates with CAMT can present with severe thrombocytopaenia despite normal MK numbers. To investigate this, we characterized MKs and platelets in newborn c-MPL –/– mice. Liver MKs in c-MPL –/– neonates were reduced in number and size compared with wild-type (WT) age-matched MKs, and exhibited ultrastructural abnormalities not found in adult c-MPL –/– MKs. Platelet counts were lower in c-MPL –/– compared with WT mice at birth and did not increase over the first 2 weeks of life. In vivo biotinylation revealed a significant reduction in the platelet half-life of c-MPL –/– newborn mice (P2) compared with age-matched WT pups, which was not associated with ultrastructural abnormalities. Genetic deletion of the pro-apoptotic Bak did not rescue the severely reduced platelet half-life of c-MPL –/– newborn mice, suggesting that it was due to factors other than platelets entering apoptosis early. Indeed, adult GFP+ (green fluorescent protein transgenic) platelets transfused into thrombocytopenic c-MPL –/– P2 pups also had a shortened lifespan, indicating the importance of cell-extrinsic factors. In addition, neonatal platelets from WT and c-MPL –/– mice exhibited reduced P-selectin surface expression following stimulation compared with adult platelets of either genotype, and platelets from c-MPL –/– neonates exhibited reduced glycoprotein IIb/IIIa (GPIIb/IIIa) activation in response to thrombin compared with age-matched WT platelets. Taken together, our findings indicate that c-MPL deficiency is associated with abnormal maturation of neonatal MKs and developmental stage-specific defects in platelet function.

Keywords

animal models - bone marrow - megakaryocytes - platelet physiology - thrombocytopaenia

Authors' Contributions

V.L., H.R, Z-J.L., S. B., D.M., Z.H. and F.F-M. designed and performed experiments, collected and analysed data, and wrote the manuscript; J.I. Jr. performed and interpreted the electron microscopy studies; K.H., W.B.S. and B.T.K. assisted with experimental design, data interpretation and preparation of the manuscript; M.S-V. and F.F-M. supervised and designed experiments, interpreted data and wrote the manuscript.


* Viola Lorenz, Haley Ramsey, Martha Sola-Visner and Francisca Ferrer-Marin contributed equally to this work.


Supplementary Material

 

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