ATP activates DNA synthesis by acting on P2X receptors in human osteoblast-like MG-63 cells

In human osteoblast-like MG-63 cells, extracellular ATP increased [H-3] thy midine incorporation and cell proliferation and synergistically enhanced pl atelet-derived growth factor- or insulin-like growth factor I-induced [H-3] thymidine incorporation. ATP-induced [H-3] thymidine incorporation was mim icked by the nonhydrolyzable ATP analogs adenosine 5'-O-(3-thiotriphosphate ) and adenosine 5'-adenylylimidodiphosphate and was inhibited by the P2 pur inoceptor antagonist suramin, suggesting involvement of P2 purinoceptors. T he P2Y receptor agonist UTP and UDP and a P2Y receptor antagonist reactive blue 2 did not affect [H-3] thymidine incorporation, whereas the P2X recept or antagonist pyridoxal phosphate-6-azophenyl-2',4-disulfonic acid inhibite d ATP-induced [H-3] thymidine incorporation, suggesting that ATP-induced DN A synthesis was mediated by P2X receptors. RT-PCR analysis revealed that MG -63 cells expressed P2X(4), P2X(5), P2X(6), and P2X(7), but not P2X(1), P2X (2), and P2X(3), receptors. In fura 2-loaded cells, not only ATP, but also UTP, increased intracellular Ca2+ concentration, and inhibitors for several Ca2+-activated protein kinases had no effect on ATP-induced DNA synthesis, suggesting that an increase in intracellular Ca2+ concentration is not ind ispensable for ATP-induced DNA synthesis. ATP increased mitogen-activated p rotein kinase activity in a Ca2+-independent manner and synergistically enh anced platelet-derived growth factor- or insulin-like growth factor I-induc ed kinase activity. Furthermore, the mitogen-activated protein kinase kinas e inhibitor PD-98059 totally abolished ATP-induced DNA synthesis. We conclu de that ATP increases DNA synthesis and enhances the proliferative effects of growth factors through P2X receptors by activating a mitogen-activated p rotein kinase pathway.