The human villous cytotrophoblast: interactions with extracellular matrix proteins, endocrine function, and cytoplasmic differentiation in the absence of syncytium formation.

Human syncytiotrophoblasts are derived from villous cytotrophoblasts by cell fusion. Coincident with this morphologic transformation, trophoblasts acquire specific endocrine functions, including elaboration of chorionic gonadotropin (hCG). We wondered if syncytia formation was a prerequisite for biochemical differentiation or simply was one part of the differentiation program. By growing purified human cytotrophoblasts under serum-free conditions and manipulating the culture surface, we were able to disassociate morphologic from biochemical differentiation. We have shown previously (Endocrinology 1986, 118:1567) that human cytotrophoblasts grown in the presence of fetal calf serum flatten out, aggregate, and form functional syncytiotrophoblasts in vitro over 24-96 hr. Here we demonstrate that when grown in the absence of serum, the cells do not undergo these morphologic changes, but remain as individual spherical cells. If the culture surface was precoated with fibronectin or a variety of collagens, but not albumin, the cells regained their ability to flatten, aggregate, and form syncytia. Attachment to and syncytia formation on fibronectin was blocked by the addition of the R-G-D-S-containing peptide, Gly-Arg-Gly-Asp-Ser-Pro. Attachment to and syncytia formation on type I collagen was blocked by anti-human fibronectin F(ab')2 fragments, while association with type IV collagen was not affected by this antibody, suggesting that fibronectin mediates trophoblast association with type I collagen, but not type IV. Although syncytia formation did not occur when cytotrophoblasts were cultured under serum-free conditions in the absence of ECM proteins, biochemical differentiation was not affected. These cells secreted hCG at the same rate under serum-free conditions whether they were plated on plastic only--which prevented syncytia formation--or fibronectin, laminin or, type IV collagen--which allowed syncytia formation to occur. Furthermore, cytoplasmic differentiation in the absence of syncytia formation was confirmed by performing transmission electron microscopy on cytotrophoblasts grown under serum-free conditions in the presence of 8-bromo-cAMP. We conclude that syncytia formation is not a prerequisite for biochemical differentiation, but simply part of the trophoblast differentiation program.