%0 Journal Article %~ Pubmed %A Mizuochi, Chiyo %A Fraser, Stuart T %A Biasch, Katia %A Horio, Yuka %A Kikushige, Yoshikane %A Tani, Kenzaburo %A Akashi, Koichi %A Tavian, Manuela %A Sugiyama, Daisuke %T Intra-Aortic Clusters Undergo Endothelial to Hematopoietic Phenotypic Transition during Early Embryogenesis. %B %D 2012 %V 7 %N 4 %P e35763 %@ 1932-6203 %X Intra-aortic clusters (IACs) attach to floor of large arteries and are considered to have recently acquired hematopoietic stem cell (HSC)-potential in vertebrate early mid-gestation embryos. The formation and function of IACs is poorly understood. To address this issue, IACs were characterized by immunohistochemistry and flow cytometry in mouse embryos. Immunohistochemical analysis revealed that IACs simultaneously express the surface antigens CD31, CD34 and c-Kit. As embryos developed from 9.5 to 10.5 dpc, IACs up-regulate the hematopoietic markers CD41 and CD45 while down-regulating the endothelial surface antigen VE-cadherin/CD144, suggesting that IACs lose endothelial phenotype after 9.5 dpc. Analysis of the hematopoietic potential of IACs revealed a significant change in macrophage CFC activity from 9.5 to 10.5 dpc. To further characterize IACs, we isolated IACs based on CD45 expression. Correspondingly, the expression of hematopoietic transcription factors in the CD45(neg) fraction of IACs was significantly up-regulated. These results suggest that the transition from endothelial to hematopoietic phenotype of IACs occurs after 9.5 dpc. %Z FOR Codes: 60103 110202 %0 Journal Article %~ Pubmed %A Funnell, Alister P W %A Norton, Laura J %A Mak, Ka Sin %A Burdach, Jon %A Artuz, Crisbel M %A Twine, Natalie A %A Wilkins, Marc R %A Power, Carl A %A Hung, Tzong-Tyng %A Perdomo, Jos?? %A Koh, Philip %A Bell-Anderson, Kim S %A Orkin, Stuart H %A Fraser, Stuart T %A Perkins, Andrew C %A Pearson, Richard C M %A Crossley, Merlin %T The CACCC-binding protein KLF3/BKLF represses a subset of KLF1/EKLF target genes and is required for proper erythroid maturation in vivo. %B Molecular and Cellular Biology %D 2012 %V %N %P %@ 1098-5549 %X The CACCC-box binding protein Erythroid Kr??ppel-like Factor (EKLF/KLF1) is a master regulator that directs the expression of many important erythroid genes. We have previously shown that EKLF drives transcription of a second Klf, Basic Kr??ppel-like Factor/Klf3. We have now tested the in vivo role of KLF3 in erythroid cells by examining Klf3 knockout mice. KLF3 deficient adults exhibit a mild compensated anemia, including enlarged spleens, increased red pulp, a higher percentage of erythroid progenitors, together with elevated reticulocytes and abnormal erythrocytes in the peripheral blood. Impaired erythroid maturation is also observed in the fetal liver. We have found that KLF3 levels rise as erythroid cells mature to become TER119(+). Consistent with this, microarray analysis of both TER119(-) and TER119(+) erythroid populations revealed that KLF3 is most critical at the later stages of erythroid maturation and is indeed primarily a transcriptional repressor. Notably, many of the genes repressed by KLF3 are also known to be activated by EKLF. However, the majority of these are not currently recognized as erythroid-specific genes. These results reveal the molecular and physiological function of KLF3, defining it as a feedback repressor that counters the activity of EKLF at selected target genes to achieve normal erythropoiesis. %Z FOR Codes: 60103 110202 %0 Journal Article %~ Pubmed %A Baron, Margaret H %A Isern, Joan %A Fraser, Stuart T %T The embryonic origins of erythropoiesis in mammals. %B Blood %D 2012 %V 119 %N 21 %P 4828-37 %@ 1528-0020 %X Erythroid (red blood) cells are the first cell type to be specified in the postimplantation mammalian embryo and serve highly specialized, essential functions throughout gestation and postnatal life. The existence of 2 developmentally and morphologically distinct erythroid lineages, primitive (embryonic) and definitive (adult), was described for the mammalian embryo more than a century ago. Cells of the primitive erythroid lineage support the transition from rapidly growing embryo to fetus, whereas definitive erythrocytes function during the transition from fetal life to birth and continue to be crucial for a variety of normal physiologic processes. Over the past few years, it has become apparent that the ontogeny and maturation of these lineages are more complex than previously appreciated. In this review, we highlight some common and distinguishing features of the red blood cell lineages and summarize advances in our understanding of how these cells develop and differentiate throughout mammalian ontogeny. %Z FOR Codes: 60103 110202 %0 Journal Article %~ Pubmed %A Isern, Joan %A He, Zhiyong %A Fraser, Stuart T %A Nowotschin, Sonja %A Ferrer-Vaquer, Anna %A Moore, Rebecca %A Hadjantonakis, Anna-Katerina %A Schulz, Vincent %A Tuck, David %A Gallagher, Patrick G %A Baron, Margaret H %T Single-lineage transcriptome analysis reveals key regulatory pathways in primitive erythroid progenitors in the mouse embryo. %B Blood %D 2011 %V 117 %N 18 %P 4924-34 %@ 1528-0020 %X Primitive erythroid (EryP) progenitors are the first cell type specified from the mesoderm late in gastrulation. We used a transgenic reporter to image and purify the earliest blood progenitors and their descendants from developing mouse embryos. EryP progenitors exhibited remarkable proliferative capacity in the yolk sac immediately before the onset of circulation, when these cells comprise nearly half of all cells of the embryo. Global expression profiles generated at 24-hour intervals from embryonic day 7.5 through 2.5 revealed 2 abrupt changes in transcript diversity that coincided with the entry of EryPs into the circulation and with their late maturation and enucleation, respectively. These changes were paralleled by the expression of critical regulatory factors. Experiments designed to test predictions from these data demonstrated that the Wnt-signaling pathway is active in EryP progenitors, which display an aerobic glycolytic profile and the numbers of which are regulated by transforming growth factor-??1 and hypoxia. This is the first transcriptome assembled for a single hematopoietic lineage of the embryo over the course of its differentiation. %Z FOR Codes: 110202 %0 Journal Article %~ Pubmed %A Sugiyama, Daisuke %A Inoue-Yokoo, Tomoko %A Fraser, Stuart T %A Kulkeaw, Kasem %A Mizuochi, Chiyo %A Horio, Yuka %T Embryonic regulation of the mouse hematopoietic niche. %B TheScientificWorldJournal [electronic resource] %D 2011 %V 11 %N %P 1770-80 %@ 1537-744X %X Hematopoietic stem cells (HSCs) can differentiate into several types of hematopoietic cells (HCs) (such as erythrocytes, megakaryocytes, lymphocytes, neutrophils, or macrophages) and also undergo self-renewal to sustain hematopoiesis throughout an organism's lifetime. HSCs are currently used clinically as transplantation therapy in regenerative medicine and are typically obtained from healthy donors or cord blood. However, problems remain in HSC transplantation, such as shortage of cells, donor risks, rejection, and graft-versus-host disease (GVHD). Thus, increased understanding of HSC regulation should enable us to improve HSC therapy and develop novel regenerative medicine techniques. HSC regulation is governed by two types of activity: intrinsic regulation, programmed primarily by cell autonomous gene expression, and extrinsic factors, which originate from so-called "niche cells" surrounding HSCs. Here, we focus on the latter and discuss HSC regulation with special emphasis on the role played by niche cells. %Z FOR Codes: 60403 110202 60103 %0 Journal Article %~ Pubmed %A Fraser, Stuart T %A Midwinter, Robyn G %A Berger, Birgit S %A Stocker, Roland %T Heme Oxygenase-1: A Critical Link between Iron Metabolism, Erythropoiesis, and Development. %B %D 2011 %V 2011 %N %P 473709 %@ 1687-9112 %X The first mature cells to arise in the developing mammalian embryo belong to the erythroid lineage. This highlights the immediacy of the need for red blood cells during embryogenesis and for survival. Linked with this pressure is the necessity of the embryo to obtain and transport iron, synthesize hemoglobin, and then dispose of the potentially toxic heme via the stress-induced protein heme oxygenase-1 (HO-1, encoded by Hmox1 in the mouse). Null mutation of Hmox1 results in significant embryonic mortality as well as anemia and defective iron recycling. Here, we discuss the interrelated nature of this critical enzyme with iron trafficking, erythroid cell function, and embryonic survival. %Z FOR Codes: 110202 %0 Journal Article %~ Pubmed %A Isern, Joan %A Fraser, Stuart T %A He, Zhiyong %A Baron, Margaret H %T Developmental niches for embryonic erythroid cells. %B Blood cells, molecules & diseases %D 2010 %V 44 %N 4 %P 207-8 %@ 1096-0961 %X Primitive erythroid cells (EryP) are the first differentiated cell type to be specified during mammalian embryogenesis. EryP arise from a pool of lineage-restricted progenitors in the yolk sac (YS) and then enter the newly formed embryonic circulation to mature in a stepwise, synchronous fashion. Numbering in the millions in the mid-gestation mouse embryo, EryP are the dominant circulating blood cell prior to the rapid generation of adult-type definitive erythroid (EryD) cells in the fetal liver. The identification of maturational events in this lineage presented a significant challenge, as EryD begin to outnumber EryP in the bloodstream from approximately E14.5 onwards. We used human epsilon-globin gene regulatory elements to drive lineage-specific expression of a histone-H2B::EGFP fusion protein, allowing us to label the chromatin of EryP during their development and to track and quantify EryP nuclei following their expulsion from the cell. Using this transgenic fluorescent reporter mouse line, we have monitored primitive erythropoiesis in three distinct niches: the YS, where EryP progenitors arise; the circulation, where EryP continue to divide and mature; and the fetal liver, where EryP complete the terminal stages of their differentiation. %Z FOR Codes: 111401 60103 %0 Journal Article %~ Pubmed %A Fraser, Stuart T %A Isern, Joan %A Baron, Margaret H %T Use of transgenic fluorescent reporter mouse lines to monitor hematopoietic and erythroid development during embryogenesis. %B Methods in enzymology %D 2010 %V 476 %N %P 403-27 %@ 1557-7988 %X The use of fluorescent reporter proteins such as GFP, RFP, and their variants to tag and track cells within the embryo has revolutionized developmental biology. Expression of these proteins within restricted populations has been achieved through the use of lineage-specific regulatory elements. This approach has proven especially powerful in the hematopoietic system, where it has been possible to monitor the generation, expansion, maturation, and migration of primitive erythroid cells, macrophages, and megakaryocytes during embryogenesis at unprecedented resolution. Such analyses have provided novel insights into the development of these lineages. In this chapter, we discuss the design considerations and methodologies involved in the production and analysis of transgenic mouse lines in which fluorescent reporters are expressed in the hematopoietic system of the mouse embryo. %Z FOR Codes: 60103 110202 %0 Journal Article %~ Pubmed %A Isern, Joan %A Fraser, Stuart T %A He, Zhiyong %A Zhang, Hailan %A Baron, Margaret H %T Dose-dependent regulation of primitive erythroid maturation and identity by the transcription factor Eklf. %B Blood %D 2010 %V 116 %N 19 %P 3972-80 %@ 1528-0020 %X The primitive erythroid (EryP) lineage is the first to differentiate during mammalian embryogenesis. Eklf/Klf1 is a transcriptional regulator that is essential for definitive erythropoiesis in the fetal liver. Dissection of the role(s) of Eklf within the EryP compartment has been confounded by the simultaneous presence of EryP and fetal liver-derived definitive erythroid (EryD) cells in the blood. To address this problem, we have distinguished EryP from their definitive counterparts by crossing Eklf(+/-) mutant and ??-globin::histone H2B-GFP transgenic mice. Eklf-deficient EryP exhibit membrane ruffling and a failure to acquire the typical discoidal erythroid shape but they can enucleate. Flow cytometric analyses of H2B-GFP(+) EryP revealed that Eklf heterozygosity results in the loss of Ter119 surface expression on EryP but not on EryD. Null mutation of Eklf resulted in abnormal expression of a range of surface proteins by EryP. In particular, several megakaryocyte markers were ectopically expressed by maturing Eklf-null EryP. Unexpectedly, the platelet tetraspanin CD9 was detected on nucleated wild-type EryP but not on mature EryD and thus provides a useful marker for purifying circulating EryP. We conclude that Eklf gene dosage is crucial for regulating the surface phenotype and molecular identity of maturing primitive erythroid cells. %Z FOR Codes: 60103 60403 110202