Biosynthesis of the third component of complement by the human monocyte-like cell line, U-937

Abstract

The human monocyte-like cell line, U-937, was shown to synthesize and secrete C3 by hemolytic assays, radioimmunoassays and metabolic labeling experiments. The daily synthesis of antigenic C3 by unstimulated U-937 cells was low (about 3 ng/10 6 cells/24hr) over a 3 day period. Induction of the cells to differentiate into macrophage-like cells with phorbol myristate acetate (PMA), resulted in 5-fold augmentation of C3 synthesis and secretion into the culture medium. Using a plaque assay for enumerating C3 production by single cells, approx. 5% of unstimulated U-937 cells were found to secrete hemolytically active C3. The proportion of C3-plaque forming cells was increased about 6-fold in PMA stimulated cells. The synthesis of C3 by U-937 cells was reversibly inhibited by cycloheximide. Data from SDS-PAGE analyses showed that U-937 cells synthesized C3 as a precursor polypeptide chain and was capable of processing this pro-molecule into the secreted two chain form. C3 antigen immunoprecipitated from stimulated U-937 cell lysates showed an increased amount of low mol. wt material as compared to C3 antigen immunoprecipitated from the lysates of unstimulated cells. This may be attributable to increased intracellular proteolytic activity in the PMA stimulated cells. The studies show that the U-937 cell line provides a useful model for studies on the synthesis and processing of complement proteins and the physiological regulation of complement production.