1: Exp Hematol. 2003 Sep;31(9):760-9. 

Erratum in:
    Exp Hematol. 2003 Nov;31(11):1139-40.

In vitro expansion of human cord blood CD36+ erythroid progenitors: temporal
changes in gene and protein expression.

Scicchitano MS, McFarland DC, Tierney LA, Narayanan PK, Schwartz LW.

GlaxoSmithKline, Safety Assessment Department, King of Prussia, Pa. 19406, USA.
Marshall.2.Scicchitano@gsk.com

OBJECTIVE: Erythropoiesis involves proliferation and differentiation of
committed erythroid progenitors to mature red blood cells. The objective of this
study was to characterize growth characteristics of human CD36+ erythroid
progenitors and to profile temporal expression of lineage-specific transcription
factors, structural proteins, and growth factor receptors involved in
erythropoiesis. MATERIALS AND METHODS: Erythropoietin-induced differentiation of
human cord blood CD36+ erythroid progenitors was profiled for GATA-1, GATA-2,
NFE2, EKLF, SCL, PU.1, Id1, Evi-1, c-myb, Hox2.2, c-kit, EpoR, glycophorin A
(GPA), CD71, beta- and gamma-globin, and protein 4.2 gene and/or protein
expression and DNA content analysis on days 4, 7, and 15 of culture. RESULTS:
Real-time RT-PCR analysis revealed upregulation of GATA-1, Id1, glycophorin A,
and protein 4.2 mRNA expression on day 7 when compared to day 4 and decreased
expression on day 15. EKLF, GATA-2, Hox2.2, c-myb, Evi-1, c-kit, and PU.1 mRNA
expression decreased on days 7 and 15. NFE2, CD71, SCL, and EPO-R mRNA
expression remained similar on days 4 and 7 but decreased on day 15. Expression
of globin genes beta- and gamma-globin increased on both day 7 and day 15
compared to day 4. Values from flow cytometric quantitation of glycophorin A,
transferrin receptor (CD71), and hemoglobin A proteins correlated with gene
expression results. DNA analysis demonstrated that most cells lacked DNA content
by day 15, a finding consistent with enucleation and terminal erythroid
differentiation. CONCLUSION: These data indicate that in vitro liquid cultures
of committed CD36+ erythroid progenitor cells retain, in part, many features of
erythropoiesis at the cellular and molecular level and may provide a useful
model for assessment of disease-related or drug-induced erythropoietic
abnormalities.

PMID: 12962721 [PubMed - indexed for MEDLINE]


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2: J Cell Physiol. 1999 Sep;180(3):390-401. 

Increased expression of the distal, but not of the proximal, Gata1 transcripts
during differentiation of primary erythroid cells.

Vannucchi AM, Linari S, Lin CS, Koury MJ, Bondurant MC, Migliaccio AR.

Division of Hematology, University of Florence and Azienda Ospedale Careggi,
Italy.

Gata1 is expressed from either one of two alternative promoters, the erythroid
(proximal to the AUG) and the testis (distal to the AUG) promoter, both used by
hemopoietic cells. To clarify the role of the distal and proximal Gata1
transcripts in erythroid differentiation, we determined by specific reverse
transcriptase-polymerase chain reactions their relative levels of expression
during the differentiation of erythroid precursors purified from the spleen of
mice treated with phenylhydrazine (PHZ) or infected with the anemia-inducing
strain of the Friend virus (FVA cells). PHZ cells are erythroid precursors that
progress in vivo to erythroblasts in 3 days. Both PHZ and FVA cells
synchronously proliferate and differentiate in vitro in the presence of
erythropoietin (EPO). The levels of total and of distal, but not of proximal,
Gata1 transcripts increased by five- to eightfold during in vivo and in vitro
differentiation of FVA and PHZ cells. The increase in expression was temporally
associated with an increase in the expression of Eklf, Scl, and Nfe2, three
genes required for erythroid differentiation, and preceded by 24 h the
repression of Gata2 and Myb expression. The day 1 PHZ cells that survived 18 h
in the absence of EPO do not express globin genes and express detectable levels
of distal but not of proximal Gata1 transcripts. These cells activate the
expression of the globin genes within 2 h when exposed to EPO. Therefore, during
erythroid differentiation of primary cells, increased expression of distal Gata1
transcripts underlies the increase in the expression of total Gata1 associated
with the establishment of the erythroid differentiation program.

PMID: 10430179 [PubMed - indexed for MEDLINE]


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