Binding of erythropoietin to CFU-E derived from fetal mouse liver cells
Fukamachi, H.; Saito, T.; Tojo, A.; Kitamura, T.; Urabe, A.; Takaku, F.
Experimental Hematology 15(8): 833-837
1987
ISSN/ISBN: 0301-472X PMID: 3305053 Document Number: 292912
The binding of recombinant erythropoietin (EPO) to fetal mouse liver cells (FMLC) was investigated using a radioiodinated derivative which retained full biological activity. FMLC were fractionated using a preformed Percoll density gradient. Using the fractionated FMLC, the ability to form CFU-E colonies in a semisolid culture was examined, and the binding of [125I]EPO was measured. The highest specific binding of [125I]EPO was observed in a fraction with a density between 1.062 and 1.076 g/ml. The same fraction showed the highest ability to form CFU-E-derived colonies. After suspension culture of FMLC with EPO for 2 days, differentiated erythroid cells with higher density markedly increased. The specific binding of [125I]EPO to these cells almost disappeared with differentiation. Scatchard analysis with cells of the CFU-E-enriched fraction showed a nonlinear curve, suggesting the existence of two classes of binding sites. One binding site was high-affinity (Kd1 = 0.41 nM), and the other low-affinity (Kd2 = 3.13 nM). These results suggest that the expression of EPO receptors on the erythroid cells is highest in CFU-E.