Inhibin and activin production and subunit expression in human placental cells cultured in vitro

Abstract

Inhibins and activins are dimeric proteins, with each subunit being one of three related protein subunits (α, βA or βB). The mRNA levels of these subunits were studied quantitatively during in-vitro differentiation of human cytotrophoblast cells into syncytium, using Northern blot analysis and semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis. The corresponding protein concentrations were determined by specific enzyme-linked immunosorbent assays for inhibin A, B, pro αC and activin A in cellular protein extracts and culture medium (n = 5). Immunofluorescence studies showed syncytium formation after 48 h. The α subunit was present before plating and increased at 48 h (P < 0.001) while the βA subunit was weak before plating and increased at 24 h. The βB subunit was not detected. With respect to corresponding protein synthesis, inhibin A (α + βA) had risen after 48 h in cellular protein extract and after 72 h in culture medium, while activin A (βA + βB) was detected after 24 h, with no significant variations in culture medium. There was a good correlation between inhibin A and α subunit expression (r = 0.736, P < 0.001), as well as between activin A and βA subunit expression (r = 0.755, P < 0.001). This study showed that mRNA expression parallels protein synthesis of inhibin and activin in trophoblast cells. Inhibin A synthesis appears to be dependent on α subunit mRNA expression, rather than on the βA subunit which controls activin A synthesis. This study has also shown that isolated cytotrophoblast cells do not produce dimeric inhibin. However, during the transformation of cytotrophoblast cells into syncytium, βA subunit mRNA expression may be an indicator of cell aggregation, while α subunit mRNA expression may be an indicator of cell fusion.