In this study, human trophoblast cells were isolated from term placentas by trypsin-DNase digestion and Percoll gradient centrifugation. After the cells were cultured in vitro, bioactive inhibin and immunoreactive inhibin were measured in the culture medium and cellular lysate by an ovine pituitary cell culture system and an immunoenzymatic assay. The trophoblast cells were capable of producing inhibin, as indicated by the observation that enhancement of inhibin content and secretion was dependent upon the cell number and time in culture. High levels of inhibin were observed in the culture medium and the cellular lysate with cell numbers of over 1 x 106 cells/well, whereas the inhibin levels decreased until they were nearly undetectable when the cell number was less than 0.25 x 106 cells/well. In the culture medium, a sharp increase of inhibin levels was observed after 2 days of culture. Bio-inhibin and immunoinhibin concentrations in the culture medium increased tenfold and fourfold, respectively, from day 2 to day 4. The peak level of bio-inhibin (3.5 U/mL) occurred on day 4 and that of immuno-inhibin (3.55 U/mL) on day 6. In contrast, the maximal level of bio-inhibin (4.45 U/mL) and immunoinhibin (4.25 U/mL) in the cellular lysate was observed on day 2. The patterns of the changes in bio-inhibin and immuno-inhibin were quite similar either in the culture medium (r=0.87, P <.05) or in the cellular lysate (r r=0.92, P < .05). With immunoperoxidase staining using rabbit antiserum to inhibin a-subunit, inhibin immunoreactivity was localized in the cytoplasm rather than in the nuclei or on the cell membrane of trophoblasts. Our data indicate that human trophoblast cells can produce and secrete bioactive and immunoreactive inhibins in vitro.
(C) 1992 The American College of Obstetricians and Gynecologists