Real-time monitoring of stress erythropoiesis in vivo using Gata1 and β-globin LCR luciferase transgenic mice

Abstract

Erythroid progenitors have the potential to proliferate rapidly in response to environmental stimuli. This process is referred to as stress erythropoiesis, with erythropoietin (EPO) playing central roles in its promotion. In this study, we wanted to elucidate the molecular mechanisms governing the regulation of stress erythropoiesis and the maintenance of red-cell homeostasis. This was achieved by our development of a noninvasive real-time monitoring system for erythropoiesis using transgenic mouse lines expressing luciferase under the control of the mouse Gata1 hematopoietic regulatory domain (G1-HRD-luc) or human ␤-globin locus control region (Hbb-LCR-luc). Optical bioluminescence images revealed that the luciferase was specifically expressed in spleen and bone marrow and was induced rapidly in response to anemia and hypoxia stimuli. The G1-HRD-luc activity tracked the emergence and disappearance of proerythroblast-stage progenitors, whereas the Hbb-LCR-luc activity tracked erythroblasts and later stage erythroid cells. Increased plasma EPO concentration preceded an increase in G1-HRD-luc, supporting our contention that EPO acts as the key upstream signal in stress erythropoiesis. Hence, we conclude that G1-HRD-luc and Hbb-LCR-luc reporters are differentially activated during stress erythropoiesis and that the transgenic mouse lines used serve as an important means for understanding the homeostatic regulation of erythropoiesis. IntroductionThe lifespan of circulating erythroid cells is limited, rendering continuous erythropoiesis a necessity throughout the life of the animal. Erythroid progenitors have the potential to proliferate rapidly in response to anemia and hypoxia stimuli, a process referred to as stress erythropoiesis and mediated by erythropoietin (EPO). 1,2 EPO is a glycoprotein hormone, 3 and its expression is induced by various erythropoietic stimuli. During erythropoiesis, hematopoietic stem cells (HSCs) first give rise to common myeloid progenitors (CMPs) with the potential to differentiate toward erythroid, megakaryocytic, and granulocytic/ monocytic cell lineages. 4 CMP progenitors differentiate into bipotential megakaryocytic and erythroid progenitors (MEPs). 4 The earliest erythroid-committed progenitors are those corresponding to erythroid burst-forming units (BFU-Es), which further differentiate through erythroid colony-forming units (CFU-Es) and proerythroblasts into erythroblasts. These erythroid progenitors, which account for a rare population of hematopoietic cells under steady state conditions, have the potential to proliferate rapidly in response to acute erythropoietic stimuli. [5][6][7] In particular, the number of CFU-Es increases markedly during stress erythropoiesis. The receptor for erythropoietin (EpoR), which is expressed abundantly in CFU-E stage progenitors, 8 plays a crucial role in promoting the erythropoietic response. 9,10 Erythropoietic stimuli result in tissue hypoxia and induce EPO gene expression through the activation of hypoxia-inducible transcription factors. EP...

A mouse model of adult-onset anaemia due to erythropoietin deficiency

“…3c). We also analysed spleen, the erythropoietic organ more responsible than bone marrow for stress erythropoiesis in rodents 29,30 . In control mice, phlebotomy-induced anaemia enlarged the spleen for compensatory stress erythropoiesis, but the size and weight of the spleen remained largely normal in ISAM regardless of severe anaemia ( Fig.…”

The dynamics of acute malaria infections. I. Effect of the parasite's red blood cell preference

“…These could be determined by transferring labelled RBCs of different ages into singly infected mice and determining their loss following infection. An indirect approach would be to compare competition between malaria strains in untreated mice, and mice treated with erythropoietin (the hormone that stimulates erythropoiesis, the production of RBCs) just prior to infection (Suzuki et al 2006). Specifically, we predict that such a treatment would result in an increase in the relative density of strains that can infect younger RBCs.…”

Differential Contribution of the Gata1 Gene Hematopoietic Enhancer to Erythroid Differentiation

“…The 5Ј homologous regions of the G1BAC-GFP and G1BAC-Luc targeting vectors contain a fragment that includes part of the first intron up to exon II (SacI-NcoI) (Fig. 1), a GFP or Luc gene, and a poly(A) signal, which were derived from the G1HRD-GFP and G1HRD-Luc constructs, respectively (37,38). The 3Ј homologous region of the G1BAC-GFP targeting vector contains an EcoRI-EcoRI fragment spanning from exon III to exon VI.…”

“…In vivo bioluminescence imaging was performed using an IVIS imaging system (Xenogen, Alameda, CA) as previously described (37). For stimulation of erythropoiesis, phenylhydrazine (60 mg/kg) was injected intraperitoneally for two consecutive days.…”