Serum-free culture of enriched hematopoietic progenitors reflects physiologic levels of fetal hemoglobin biosynthesis

Y. Fujimori, M. Ogawa, S. C. Clark, G. J. Dover

Research output: Contribution to journalArticlepeer-review

18 Scopus citations


Adult erythroid progenitors produce significantly higher fetal hemoglobin (HbF) levels in cultures containing fetal calf serum (FCS) and erythropoietin (Ep) than in vivo. The precise mechanisms for this increased HbF production in culture have not been elucidated. We examined HbF biosynthesis by enriched human progenitors in serum-free (SF) culture. We measued globin chain biosynthesis by combination of isoelectric focusing and autoradiography and examined percent nucleated erythrocytes containing HbF (%FNRBC) using microscopic immunodiffusion. CD34 (My10)-positive marrow cells from a normal subject yielded an almost negligible amount of γ-globin in SF culture stimulated by 100 U/mL interleukin-3 (IL-3) and 2 U/mL Ep, while corresponding FCS culture revealed significant γ-globin biosynthesis. The %FNRBC of the erythrooid bursts in SF cultures derived from nine normal adults (2.0% ± 0.9% F cells) was 3.0% ± 3.4%, while in FCS culture, it was 25% ± 12% (mean ± SD). Dosages of IL-3 between 10 and 10,000 U/mL did not increase %FNRBC in FCS or SF conditions. Mean Hb contents of nucleated erythrocytes (NRBC) assayed by microdensitometry of pericellular immunoprecipate were similar in FCS and SF cultures. The number of erythroid bursts per 2 x 103 CD34-positive marrow cells was 48 ± 20 in FCS and 36 ± 12 in SF cultures. In two experiments, progenitors grown for 7 days under SF conditions were isolated and recultured in either SF or FCS conditions for 7 days, and the resulting erythroid bursts were analyzed for FNRBC. The bursts that had been returned to FCS cultured yielded values of %FNRBC intermediate between those obtained from progenitors grown entirely in SF or FCS cultures, indicating that serum effect is not solely due to growth selection for certain subpopulations of erythroid burst-forming units. This experiment also demonstrated that the factors present in serum responsible for HbF augmentation act at both early and late stages during erythroid burst development. SF culture of peripheral blood progenitors of one subject with heterocellular hereditary persistence of fetal hemoglobin (HPFH) yielded elevated levels of FNRBC (19% ± 5%) that accurately reflected the F cell (18%) of the circulating blood. Similarly, FNRBC in cultures of progenitors from one umbilical cord blood sample (86% F reticulocytes) was 87 ± 3% FNRBC. The SF culture for enriched human progenitors, which nearly reflects the physiologic HbF programs of the donor, should facilitate studies of the exact mechanisms of postnatal reactivation of HbF production.

Original languageEnglish (US)
Pages (from-to)1718-1722
Number of pages5
Issue number8
StatePublished - 1990
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology


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