Erythroblasts from Friend virus infected‐ and phenylhydrazine‐treated mice accurately model erythroid differentiation

Abstract

Summary. The dynamics of gene expression during terminal erythroid differentiation have been examined in three murine models; the erythroleukaemia cell line HCD-57 and splenic erythroblasts isolated from mice treated with either the anaemia-inducing strain of Friend virus (FVA cells) or the haemolytic agent phenylhydrazine (PHZ cells). In response to erythropoietin (EPO) and haemin, HCD-57 cells proliferated and synthesized haemoglobin, but failed to complete terminal differentiation as indicated by lack of change in both gene expression and morphological appearance. In contrast, EPO-induced terminal differentiation in FVA and PHZ cells in vitro was accompanied by increases in haemoglobin positivity, morphological maturation and a shared pattern of gene expression. EPO receptor (EPO-R) mRNA levels peaked before globin gene expression which was maximal at 24 h. Peak GATA-1 and EKLF mRNA levels also preceded the globin gene peak, but the highest NF-E2 levels coincided with maximal globin levels, suggesting a role for NF-E2 in the maintenance, rather than the initiation of globin gene expression. Peak expression of d-aminolaevulinic acid synthase (ALAS) coincided with peak globin expression. FVA and PHZ cells represent more effective models than the HCD-57 cell line for the investigation of erythroid gene expression during EPO-regulated terminal erythropoiesis.

A common epitope is shared by activated signal transducer and activator of transcription-5 (STAT5) and the phosphorylated erythropoietin receptor: implications for the docking model of STAT activation

“…57 Phenylhydrazine administration generates a population of splenic erythroblasts that accurately represents erythroid differentiation as monitored by the expression of EPO-R, GATA-1, EKLF, NF-E2, ␦-aminolevulinic acid synthase, ␣-globin, and ␤-major globin. 58 In this study, we have shown that phenylhydrazineprimed erythroblasts are EPO-responsive and the EPO-R is immunoprecipitated by the activation-specific STAT5 antibody. -6), Ba/F3 EPO-R Y1Y2Y4Y8 (lanes 7-10), Ba/F3 Y1Y2Y4 (lanes [11][12][13][14], Ba/F3 Y1Y2 (lanes [15][16][17][18], and Ba/F3 EPO-R F8 (lanes 19-21) cells stimulated in the presence or absence of IL-3 (I) or EPO (E).…”

A novel role for STAT1 in regulating murine erythropoiesis: deletion of STAT1 results in overall reduction of erythroid progenitors and alters their distribution

“…38 Because approximately 90% of the phenylhydrazine-primed splenocytes are EPO-responsive erythroblasts that are capable of undergoing normal erythroid differentiation in the presence of EPO, this is a physiologically relevant model of erythroid development. 39 This is the first report to examine stress erythropoiesis signaling in primary erythroblasts from genetargeted animals. Because the response to phenylhydrazine was identical in wild-type and STAT1-deficient mice, we were able to directly compare signaling in splenocytes derived from each animal.…”

Differential regulation of SOCS genes in normal and transformed erythroid cells

“…High levels of BFU-E are found in the spleen immediately after phenylhydrazine treatment (Sasaki et al, 2000;Vannucchi et al, 2000), and the maturation of these progenitors has been shown to model accurately erythroid terminal differentiation (Hodges et al, 1999). Splenocytes enriched for BFU-E were obtained 1 day postphenylhydrazine treatment and cultured in the presence of Epo for 48 h. Figure 1a (Vannucchi et al, 2000).…”