Erythroid progenitors in polycythemia vera: demonstration of their hypersensitivity to erythropoietin using serum free cultures

Abstract

In order to demonstrate whether “spontaneous” erythroid colonies observed in vitro in polycythemia vera (PV) using standard colony assays were independent from erythropoietin (epo) or exquisitely sensitive to the hormone, we used methyl cellulose serum-free cultures, in which serum was completely replaced by iron-saturated transferrin, α-thioglycerol albumin, and low density lipoproteins. Connaught Step III epo was used. In 6 cases of PV, no epo-independent colony (CFU-E or BFU-E derived) was observed in serum-free conditions, while spontaneous colonies were present after plating the same PV bone marrow in culture with serum. The epo dose-response curves showed a tenfold increase in the sensitivity to epo of PV erythroid progenitors compared to normal controls. In PV, the first CFU-E and BFU-E colonies were observed after addition of 0.001–0.01 lU/ml of epo, while in controls they appeared at an epo concentration between 0.01 and 0.1 lU/ml. Numbers of spontaneous colonies in cultures with serum compared with the epo dose-response curve in the same patient in serum-free cultures are much higher than expected from the small amount of epo present in the serum. These results confirm that PV erythroid progenitors able to differentiate spontaneously in standard culture conditions are in fact dependent of epo and hypersensitive to the hormone. They show that these abnormal progenitors do not represent a homogeneous population, but exhibit different degrees in their hypersensitivity to epo. Since the small amount of epo present in the normal serum cannot explain the growth of spontaneous colonies by itself, a hypersensitivity to other serum factors cannot be excluded.