Regulation of complement factor H synthesis in U-937 cells by phorbol myristate acetate, lipopolysaccharide, and IL-1.
The capacity of the human monocyte cell line U-937 to synthesize complement factor H was examined. The kinetics of secretion of factor H into cell culture supernatant were followed by a sensitive solid phase RIA capable of measuring 0.1 ng of protein. Daily secretion of factor H was low and almost linear and was approximately 3 ng of factor H per 10(6) cells. Factor H synthesis was inhibited by cycloheximide but returned to the levels seen in untreated cultures after removal of the inhibitor. LPS and IL-1 both effected a time- and dose-dependent enhancement of factor H synthesis. Induction of U-937 cells with PMA to differentiate into macrophage-like cells also resulted in increased factor H synthesis. RIA of cell lysates, immunofluorescence microscopy, as well as FACS analysis all revealed that factor H Ag was also associated with the U-937 cell membrane. The population of U-937 cells bearing membrane-associated factor H was decreased from 77 to 43% after stimulation for 48 h with LPS (1 microgram/ml). [35S]Methionine metabolic labeling and SDS-PAGE analysis of factor H immunoprecipitates from unstimulated and stimulated culture supernatants and cell lysates demonstrated a major polypeptide, m.w. 150,000, and a minor component, m.w. 42,000. Western blot analysis of factor H in fresh normal plasma also detected both 150,000 and 42,000 m.w. factor H proteins. This is in agreement with the recent demonstration of a 4.4- and 1.8-kb mRNA for factor H in human liver. These data demonstrate that U-937 cells synthesize factor H that is structurally and antigenically similar to factor H in normal plasma. The exact nature of the membrane-bound factor H and its functions and mechanism of attachment to the cell membrane remain to be elucidated.
Minta
J O
JO
Department of Pathology, University of Toronto, Ontario, Canada.
D002460:Cell Line; D003180:Complement C3b Inactivator Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D017242:Complement Factor H; D005455:Fluorescent Antibody Technique; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008565:Membrane Proteins / Q000032:analysis; D008715:Methionine; D009000:Monocytes / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
C493495:CFH protein, human; D003180:Complement C3b Inactivator Proteins; D007375:Interleukin-1; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D017242:Complement Factor H; D008715:Methionine; D013755:Tetradecanoylphorbol Acetate