Effects of cyclic nucleotides on hemoglobin synthesis in fetal calf liver cells in culture

Abstract

Fetal calf liver cells were cultured in a serum-free medium in the presence of dibutyryl cAMP (dbcAMP), dibutyryl cGMP (dbcGMP), or cGMP. After a 20-h incubation with the nucleotides the synthesis of the fetal hemoglobins F0 and F1 was measured by the incorporation of [3H]leucine. The three cyclic nucleotides stimulated hemoglobin synthesis at a concentration of 1 μM. dbcGMP (10 nM) inhibited the synthesis of F0 and F1 but at a concentration of 1 μM significantly stimulated the synthesis of both hemoglobins. dbcAMP (10 μM) preferentially stimulated the synthesis of the transient hemoglobin F1 whereas cCMP (1 μM)stimulated specifically the synthesis of the definitive hemoglobin F0. Prostaglandin E1 (100 nM) stimulated the synthesis of F1 in a manner similar to that observed for dbcAMP. AMP, GMP, CMP, and sodium butyrate did not stimulate hemoglobin synthesis at the concentrations indicated above. The analysis of the α- and γ-globin chains by high pressure liquid chromatography indicated that the synthesis of both chains are stimulated by the three cyclic nucleotides. These changes in hemoglobin synthesis taking place at different concentrations of cyclic nucleotides may be of importance in the maturation of erythroid cells, because in these cells differentiation is normally coupled with cell proliferation. Furthermore, some of these cyclic nucleotides may be second messengers of erythropoietic hormones. We have found that the intracellular levels of cGMP were increased 15 min after addition of step III sheep plasma erythropoietin to the cultures, whereas the amounts of cAMP remained unchanged. It is then possible that erythropoietin itself or a factor present in the erythropoietin preparation may act by a cGMP-dependent mechanism.