Suppression by beta-mercaptoethanol of the intracellular hormonal dynamics of human chorionic gonadotropin-beta subunit (hCG-beta) in BeWo choriocarcinoma cells

Abstract

The effects of beta-mercaptoethanol (ME) on the steady-state level of mRNA of the human chorionic gonadotropin-beta subunit (hCG-beta) and the intracellular hormonal dynamics of the product protein were examined in BeWo cells, a choriocarcinoma cell line, using Northern blot analysis and a radioimmunoassay (RIA) specific for hCG-beta. ME reduced both medium and intracellular contents of hCG-beta in a dose-dependent manner, with its minimum effective dose being 0.01 per cent. The highest dose used (0.1 per cent) caused more than 90 per cent inhibition with both parameters, without affecting the cell number and the cell viability as verified by trypan blue exclusion. Significant reductions in both the medium and intracellular contents began to occur 6 h after the onset of incubation with ME. The ME-induced suppressions were reversible. Northern blot analysis showed that ME had no effects on the steady-state level of hCG-beta mRNA. When medium and cell lysates collected from ME-free incubations were incubated with 0.03 per cent ME, there were significant reductions of immunoreactive hCG-beta with both the medium and cell lysates. The magnitude of reduction, however, was much greater with the latter (75 per cent) than with the former (25 per cent). In contrast, the hCG-beta immunoreactivity of the RIA reference preparation was unaffected by incubation with ME. These results suggested that the major target(s) of ME action were the intracellularly located hCG-beta molecule, presumably its intramolecular disulphide bonds. It must also be pointed out that the hCG-beta molecule synthesized and secreted by BeWo cells have some structural deviation from the reference standard molecule of normal trophoblastic origin to explain the differential susceptibility to ME.