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Y Fujimori, M Ogawa, SC Clark and GJ Dover
Department of Medicine, Medical University of South Carolina, Charleston.
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
measured 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
gamma-globin in SF culture stimulated by 100 U/mL interleukin-3 (IL-3) and
2 U/mL Ep, while corresponding FCS culture revealed significant
gamma-globin biosynthesis. The %FNRBC of the erythroid 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 of SF
conditions. Mean Hb contents of nucleated erythrocytes (NRBC) assayed by
microdensitometry of pericellular immunoprecipitate were similar in FCS and
SF cultures. The number of erythroid bursts per 2 x 10(3) 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 cultures 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.
This article has been cited by other articles:
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| Copyright © 1990 by American Society of Hematology Online ISSN: 1528-0020 | |||||||||
