Biosynthesis of coagulation Factor V by a human hepatocellular carcinoma cell line

D. B. Wilson, H. H. Salem, J. S. Mruk, I. Maruyama, P. W. Majerus

Research output: Contribution to journalArticle

51 Scopus citations

Abstract

A human hepatocellular carcinoma line, HepG2, was found to secrete coagulation Factor V. Factor V activity in HepG2 culture fluid increased nearly linearly during a 20-h time course (5 ng Factor Va/h per 106 cells). Thrombin treatment increased Factor V activity in HepG2 culture medium 6- to 9-fold, indicating that the medium accumulates a mixture of Factors V and Va. To demonstrate de novo synthesis of Factor V, HepG2 cells were incubated in culture medium containing [35S]methionine. Labeled Factor V was immunoprecipitated from the medium and was shown to co-migrate with purified plasma Factor V upon sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorography. When medium was treated with thrombin before immunoprecipitation and fluorography, the 330,000-M(r) [35S]methionine-labeled Factor V was converted to Factor Va. Factor Va coagulant activities from HepG2 cells and human plasma were inhibited in parallel by anti-Factor V antibody, indicating that HepG2 and plasma Factor Va have the same intrinsic activity. If normal hepatocytes synthesize Factor V at the same rate as HepG2 cells, then hepatocyte secretion can account for the total Factor V present in plasma. The production of Factor V by cultured human umbilical vein endothelial cells was also examined. Spent culture medium from endothelial cells contained only Factor Va and the amount was <1% of the activity found in medium from HepG2 cells under comparable conditions. The amount of Factor V activity in endothelial cell culture fluid did not change with time in culture.

Original languageEnglish
Pages (from-to)654-658
Number of pages5
JournalJournal of Clinical Investigation
Volume73
Issue number3
DOIs
StatePublished - Jan 1 1984

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