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Synthesis of coagulation factor V by cultured aortic endothelium
TJ Cerveny, DN Fass and KG Mann
Bovine aortic endothelium has been examined with respect to the synthesis
of coagulation factor V. After cultured cells reached confluency, samples
of supernatant culture media and solubilized cells were analyzed for factor
V in a two-stage bioassay and in a double- antibody radioimmunoassay. In
addition, preconfluent cells were pulsed for 4 days with 35S-methionine in
methionine-free media. After the 4- day pulse, supernatant media were
chromatographed on a factor V monoclonal antibody-Sepharose resin to
isolate 35S-labeled factor V. The isolated material and 125I-factor V
standards were analyzed by electrophoresis and autoradiography. The
bioassay indicated an increase, with time, of unactivated factor V in the
culture supernatant, whereas solubilized cells were negative for factor V.
The radioimmunoassay indicated an increase, with time, of factor V antigen
in the culture supernatants, and the solubilized cells yielded a constant
level of antigen per cell. Autoradiograms of electrophoretograms of
immunoadsorbed 35S-culture supernatant with 125I- factor V/Va standards
revealed labeled proteins with electrophoretic mobilities compatible with
125I-factor V/Va standards. The data obtained from three different
sources-bioassay, radioimmunoassay, and 35S-methionine incorporation-all
indicate that factor V is synthesized by cultured bovine aortic
endothelium.
Volume 63,
Issue 6,
pp. 1467-1474,
06/01/1984
Copyright © 1984 by The American Society of Hematology
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