Characterization and regulation of epidermal growth factor receptors in human placental cell cultures

We have confirmed that cultured human placental cells rapidly release hCG. Preincubation with epidermal growth factor (EGF) for 24 h significantly increased the amount of hCG released and also increased human placental lactogen release by these cells. To better understand the mechanisms of action of EGF on the feto-placental unit, we studied EGF receptor binding and regulation by examining the characteristics and specificity of EGF receptors in human placental syncytiotrophoblast cultures. Maximal [125I]EGF binding occurred at pH 7.5 and 4 C, and exhibited a high degree of specificity. In the presence or absence of Bacitracin at 4 C, specific binding values were similar, and labeled EGF was physically intact, as assessed by trichloroacetic acid precipitation or rebinding to human placental membranes. The percent specific binding was proportional to cell and ligand concentrations and was significantly increased in term (52.9 +/- 1.2%; n = 11) compared to early gestation placental cells (22.7 +/- 3.4%; n = 7; P less than 0.001). Both term and midterm EGF displacement curves generated curvilinear Scatchard plots, suggesting receptor heterogeneity. Pretreatment of cells with EGF resulted in a dose and time-dependent decrease in specific binding, which was maximal (80%) at 200 ng/ml EGF. This loss of binding was due to decreases in the number of both high and low affinity receptor sites, with no significant change in the apparent affinity. The induction of EGF receptor loss by EGF was a specific effect on the EGF receptor. Preincubation of these same cells with insulin caused a decrease in the number of insulin receptors, while the number of EGF receptors remained unaltered. Conversely, preincubation with EGF, in a dose that down-regulated EGF receptors, did not alter insulin receptor number or affinity. Down-regulation of EGF receptors was reversible, with 50% recovery by 16 h. However, cycloheximide (10 micrograms/ml) blocked EGF-induced down-regulation and receptor recovery. The presence of EGF receptors in human placental cells and the ontogenic changes found suggest that EGF may be involved in the regulation of fetal growth and development. These studies indicate the feasibility of using human placental cells in culture as a model system to probe hormone-cell interaction in the fetoplacental unit.