ENDOTHELIN RAPIDLY STIMULATES TYROSINE PHOSPHORYLATION IN OSTEOBLAST-LIKE CELLS

The mitogenic activity of endothelin (ET) was studied in osteoblast-li ke cells, MC3T3-E1. [H-3] Thymidine incorporation induced by ET was ma rkedly lower than that of platelet-derived growth factor (PDGF). ET sy nergistically stimulated H-3 thymidine incorporation induced by PDGF w ith an apparent ED50 value of 2.5 nM. Treatment of MC3T3-E1 cells with ET and subsequent immunoblotting of the cell extracts with antiphosph otyrosine antibodies followed by labeling with [I-125] protein A resul ted in the identification of several phosphotyrosine-containing protei ns. The intensity of these labeled phosphoproteins significantly incre ased when the cells were treated with a combination of ET and PDGF. Ge nistein, an inhibitor of tyrosine kinases, blocked [H-3] thymidine inc orporation as well as protein tyrosine phosphorylation stimulated by e ither ET, PDGF or the combination of ET and PDGF. These findings sugge st that tyrosine phosphorylation could play a role in the comitogenic activity of ET in osteoblast-like cells.