Studies of alpha-granule proteins in cultured human megakaryocytes.
alpha-Granule protein storage is important for producing platelets with normal haemostatic function. The low to undetectable levels of several megakaryocyte-synthesized alpha-granule proteins in normal plasma suggest megakaryocytes are important to sequester these proteins in vivo. alpha-Granule protein storage in vitro has been studied using other cell types, with differences observed in how some proteins are processed compared to platelets. Human megakaryocytes, cultured from cord blood CD34(+) cells and grown in serum-free media containing thrombopoietin, were investigated to determine if they could be used as a model for studying normal alpha-granule protein processing and storage. ELISA indicated that cultured megakaryocytes contained the alpha-granule proteins multimerin, von Willebrand factor, thrombospondin-1, beta-thromboglobulin and platelet factor 4, but no detectable fibrinogen and factor V. A significant proportion of the alpha-granule protein in megakaryocyte cultures was contained within the cells (averages: 41-71 %), consistent with storage. Detailed analyses of multimerin and von Willebrand factor confirmed that alpha-granule proteins were processed to mature forms and were predominantly located in the alpha-granules of cultured megakaryocytes.Thrombopoietin-stimulated cultured megakaryocytes provide a useful model for studying alpha-granule protein processing and storage.
Department of Pathology, McMaster University, Hamilton, Ontario, Canada.
Dragoslava Kika
DK
Veljkovic
Elisabeth M
EM
Cramer
Gulie
G
Alimardani
Serge
S
Fichelson
Jean-Marc
JM
Massé
Catherine P M
CP
Hayward
D001798:Blood Proteins / Q000032:analysis / Q000378:metabolism; D002478:Cells, Cultured; D003594:Cytoplasmic Granules / Q000737:chemistry*; D006801:Humans; D008533:Megakaryocytes / Q000737:chemistry* / Q000166:cytology / Q000648:ultrastructure; D010978:Platelet Factor 4 / Q000032:analysis / Q000378:metabolism; D011506:Proteins / Q000032:analysis / Q000378:metabolism*; D013926:Thrombopoietin / Q000494:pharmacology; D019700:Thrombospondin 1 / Q000032:analysis / Q000378:metabolism; D001620:beta-Thromboglobulin / Q000032:analysis / Q000378:metabolism; D014841:von Willebrand Factor / Q000032:analysis / Q000378:metabolism
D001798:Blood Proteins; D011506:Proteins; D019700:Thrombospondin 1; D001620:beta-Thromboglobulin; C069282:multimerin; D014841:von Willebrand Factor; D010978:Platelet Factor 4; D013926:Thrombopoietin