We have developed a novel serum-free (SF) medium for the growth of normal human adult circulating burst-forming units erythroid (BFU-E). This medium, which utilizes a defatted albumin devoid of contaminants having erythropoietic activity, allowed us to reliably investigate the growth factor requirements of circulating BFU-E. In the presence of a defined BPA composed of recombinant human interleukin-3 (rHu IL-3) and hemin, optimal growth of BFU-E required the addition of erythropoietin (rHu Epo), insulin-like growth factor-I (rHu IGF-I) and retinyl acetate or all-trans-retinoic acid. The retinoids synergised with either Epo or IGF-I to increase 3 to 4-fold the number of burst-component colonies (BCC). IGF-I could entirely replace Epo for erythroid burst-formation, but required a 100-fold higher concentration than the latter. The effect of IGF-I was shown to occur in the presence of an anti-Epo antibody which inhibited the effect of Epo.  These findings show that IGF-I has an Epo-like activity and strongly indicate that an Epo-independent pathway exists for normal human erythropoiesis in vitro.  Removal of adherent cells prevented  burst-formation; this was partially restored by the addition of stem cell factor (rHu SCF).

The sensitivities of circulating BFU-E from patients with Polycythemia vera (PV) to Epo, IGF-I and IL-3 were examined in the SF medium.  We found that the Epo and IL-3 sensitivities of PV and normal BFU-E were identical; in contrast, the sensitivity of BFU-E to IGF-I in PV was >100-fold greater than normal. Thus, the erythrocytosis characteristic of PV cannot be explained by hypersensitivity to either Epo or IL-3.  Instead, the apparent Epo-independence of erythroid progenitors in PV can be explained by their pronounced hypersensitivity to IGF-I, given that we show that IGF-I can replace Epo in SF medium. Moreover, burst-formation in PV was unaffected by adherent cell removal. This suggests that the PV phenotype is due to a combination of an IGF-I-hypersensitivity with a relative independence from the influence of adherent cells.