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Biology of human megakaryocyte factor V
AM Gewirtz, M Keefer, K Doshi, AE Annamalai, HC Chiu and RW Colman
To learn more about human megakaryocyte coagulation cofactor V (FV), we
studied the expression of this protein in normal bone marrow megakaryocytes
and in megakaryocytes cloned from their colony-forming unit in FV-depleted
plasma clot cultures. Mouse monoclonal antibodies directed against either
the light chain or an activation peptide of human FV and a rabbit
polyclonal, monospecific FV antiserum were used as probes for these
experiments in conjunction with a variety of immunochemical detection
techniques. All morphologically recognizable megakaryocytes were shown to
contain FV. The origin of this protein appeared to be both from FV bound to
the cell as well as from endogenous FV in the majority of cells examined.
The existence of a population of small bone marrow mononuclear cells that
simultaneously expressed platelet glycoproteins and FV was also noted. Such
cells represented approximately 70% of all small cells positive for
platelet glycoproteins. In contrast, only about 40% of megakaryocyte
colonies cloned in FV-deficient medium contained cells with
immunochemically detectable FV. FV expression was most clearly demonstrated
in large cells in the colonies, whereas smaller, presumably less mature
cells labeled weakly or not at all. Synthesis of FV by human megakaryocytes
was documented using elutriation-enriched cells incubated in 35S-
methionine-containing medium. Megakaryocyte lysates and medium conditioned
by these cells were subjected to immunoaffinity column purification. Column
eluates analyzed by sodium dodecyl sulfate- polyacrylamide gel
electrophoresis and autoradiography revealed radioactive bands comigrating
with the heavy and light chains of thrombin-activated FV. These studies
suggest that human megakaryocytes both bind and synthesize FV. Expression
of these traits appears to be related to cell maturation, with binding
ability appearing earlier than the ability to synthesize this protein.
Finally, although the ability to bind FV appears to be universal among
megakaryocytes, our culture data suggest that synthesis may be a
restricted, or constitutively expressed property of these cells.
Volume 67,
Issue 6,
pp. 1639-1648,
06/01/1986
Copyright © 1986 by The American Society of Hematology
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