ERYTHROPOIESIS INVITRO - ROLE OF CALCIUM

The in vitro plasma clot technique was employed to examine the role of Ca during the interaction of erythropoietin and mouse erythroid progenitor cells. Erythropoietin-induced erythroid colony formation was increased 24% by the carboxylic ionophore A23187 (10 nM), whereas a 35% increase was produced by the carboxylic ionophore Ro 2-2985/1 (1 nM). EGTA [ethylene glycol bis (.beta.-aminoethyl ether) tetraacetic acid] (3 mM) inhibited erythropoietin-induced erythroid colony formation. Inhibition of erythroid colony formation by EGTA could be reversed by Ca2+, but not by Mn2+, Mg2+, Zn2+ or Fe2+. At least 30 min exposure of marrow cells to erythropoietin in vitro was required for production of erythroid colonies. EGTA substantially inhibited erythropoietin-induced erythroid colony formation even when the marrow cells were exposed to the hormone for up to 2 h before addition of the chelator. Marrow cells incubated first in Ca-free medium with erythropoietin and then cultured in the presence of Ca but not erythropoietin, failed to form erythroid colonies although colony formation occurred when erythropoietin was provided. Ca is apparently required for extracellular and intracellular events during the interaction of erythropoietin with its target cells.