Immunologic and Physical Characterization of Human Chorionic Gonadotropin and Its Subunits in Cultures of Human Malignant Trophoblast

Abstract

Culture fluids and cell homogenates from cultures of BeWo human malignant trophoblast cells were examined for the presence of chorionic gonadotropin (hCG) and its subunits by gel filtration on a calibrated Sephadex G-200 column. Culture fluids, cell homogenates and gel filtration eluates were radioimmunoassayed in homologous hCG, hCG.alpha. and hCG.beta. assays. The gel filtration pattern from the culture fluid consisted of 3 discrete antigenic peaks, including hCG and hCG.beta. of normal size and hCG.alpha. of apparently larger than normal size. There was approximately twice as much hCG as hCG.beta. or hCG.alpha. in the culture fluid. The gel filtration profile from the cell homogenate consisted of a broad peak of hCG.beta. but only small amounts of hCG and hCG.alpha.. The major hCG.beta. and hCG.alpha. peaks were of apparently smaller molecular weight than highly purified hCG.beta. and hCG.alpha. standards, respectively. The dose-response behavior of trophoblast cell homogenate in the homologous radioimmunoassays for hCG, hCG.alpha. and hCG.beta. provided additional evidence that hCG.beta. was the predominant species in the cells. Incubation of cells with 1 mM dibutyryl cyclic AMP plus 1 mM theophylline (dbT) stimulated the secretion of hCG and hCG.beta. of normal size, and of the large form of hCG.alpha.. The gel filtration profile of stimulated cell homogenates consisted predominantly of a small molecular weight form of hCG.beta.. The bulk of the hCG in the cell is present in the form of its subunits, and incubation with dbT stimulates the production of both subunits in addition to hCG.