Epidermal Growth Factor and Phorbol Ester Actions on Human Osteosarcoma Cells CHARACTERIZATION OF RESPONSIVE AND NONRESPONSIVE

Epidermal growth factor (EGF) and the tumor-promoting diterpene phorbol 12-myristate 13-acetate (PMA) have been shown previously to increase the production of prostaglandin E2 (PGE2) in bone, a biochemical action which leads to resorption of extracellular matrix and calcium release. In this report, we have identified specific, high affinity receptors for EGF and phorbol esters on 4 lines of PGE2-producing human osteosarcoma cells: G-292, TX-4, MG-63, and SaOS-2. EFG receptors in all 4 lines had similar affinities and binding capacities and exhibited homologous down modulation after incubation with EGF. In addition, each cell line showed specific high affinity binding of [3H]phorbol 12,13-dibutyrate. The lines differed, however, in their biological responses to EGF and PMA. G292 and TX-4 cells responded to EGF and PMA with a 3to &fold increase in the production of PGEn. There was good agreement between the Kd values for binding and EDso values for enhanced prostaglandin production. In these cells, incubation with both EGF and PMA gave a synergistic PGEz response. MG-63 cells increased the synthesis of PGEz in response to PMA but not in response to EGF, while SaOS-2 cells did not give this biological response to either agent. Endogenous phospholipase activity, measured by [3H]arachidonate release from prelabeled cells, was tightly correlated with the PGEa response. However, responsiveness to PMA or EGF was differentiated by dependence on external calcium. PMA increased PGEz production in the absence or presence of calcium. The prostaglandin response to EGF, however, required the presence of external calcium. EGF also increased by 2-fold the uptake of “‘Ca2’ into G-292 cells; PMA did not. Thus, stimulation of PGEz production by EGF occurs via a mechanism dependent on external calcium, possibly involving calcium influx, while the action of PMA is independent of extracellular calcium. The failure of certain osteosarcoma cell lines to respond to EGF may be linked to the absence of EGF-induced calcium uptake.