An investigation of TGF-b receptors and the actions of TGF-b1 and TGF-b2 in placental trophoblast cells /

Transforming Growth Factor Beta (TGF-$ beta$) is highly pleiotropic, involved in controlling cellular proliferation, differentiation extracellular matrix (ECM) remodelling. TGF-$ beta$ is a potential mediator of placental trophoblast functions, including differentiation, hormone production, endometrial invasion and immunosuppression. Trophoblast cells are the epithelial-like covering of the chorionic villae at the fetal-maternal interface. In this study equilibrium binding assays and functional assays were used to investigate the binding characteristics and the actions of the TGF-$ beta$1 and TGF-$ beta$2 isoforms on an established human choriocarcinoma trophoblastic cell line, BeWo. Equilibrium saturation experiments indicated that the BeWo cells exhibited similar average affinities and total number of binding sites for TGF-$ beta$1 and TGF-$ beta sb2$. The K$ sb{ rm D}$ values obtained from Scatchard analyses were approximately 65 pM for $ sp{125}$I-TGF-$ beta$1 and 40 pM for $ sp{125}$I-TGF-$ beta$2, with 70,000 and 85,000 sites per cell respectively. Competitive equilibrium experiments indicated that TGF-$ beta$1 and TGF-$ beta$2 were equipotent (apparent half maximal inhibition (IC$ sb{50}$) $ sp sim70$ pM) and that all binding sites were capable of recognizing both isoforms. Two frequently used functional assays to determine dose responsiveness to TGF-$ beta$ isoforms were investigated with BeWo cells. A $ sp3$H-thymidine incorporation assay was used to measure TGF-$ beta$ effects and this indicated that neither TGF-$ beta$1 nor TGF-$ beta$2 had any significant effect on BeWo cells. The synthesis of an ECM protein, fibronectin, was monitored using both gelatin-Sepharose affinity chromatography and immunoprecipitation with anti-human fibronectin polyclonal antibodies. However, neither TGF-$ beta$1 nor TGF-$ beta$2 had any effect on fibronectin synthesis in BeWo cells as measured by these two methods. The significance of the results obtained from the functio