Lineage regulators direct BMP and Wnt pathways to cell-specific programs during differentiation and regeneration.

BMP and Wnt signaling pathways control essential cellular responses through activation of the transcription factors SMAD (BMP) and TCF (Wnt). Here, we show that regeneration of hematopoietic lineages following acute injury depends on the activation of each of these signaling pathways to induce expression of key blood genes. Both SMAD1 and TCF7L2 co-occupy sites with master regulators adjacent to hematopoietic genes. In addition, both SMAD1 and TCF7L2 follow the binding of the predominant lineage regulator during differentiation from multipotent hematopoietic progenitor cells to erythroid cells. Furthermore, induction of the myeloid lineage regulator C/EBPα in erythroid cells shifts binding of SMAD1 to sites newly occupied by C/EBPα, whereas expression of the erythroid regulator GATA1 directs SMAD1 loss on nonerythroid targets. We conclude that the regenerative response mediated by BMP and Wnt signaling pathways is coupled with the lineage master regulators to control the gene programs defining cellular identity.

Acute Myeloid Leukemia Causes T Cell Exhaustion and Depletion in a Humanized Graft-versus-Leukemia Model.

Allogeneic hematopoietic stem cell transplantation (alloSCT) is, in many clinical settings, the only curative treatment for acute myeloid leukemia (AML). The clinical benefit of alloSCT greatly relies on the graft-versus-leukemia (GVL) effect. However, AML relapse remains the top cause of posttransplant death; this highlights the urgent need to enhance GVL. Studies of human GVL have been hindered by the lack of optimal clinically relevant models. In this article, we report, the successful establishment of a novel (to our knowledge) humanized GVL model system by transplanting clinically paired donor PBMCs and patient AML into MHC class I/II knockout NSG mice. We observed significantly reduced leukemia growth in humanized mice compared with mice that received AML alone, demonstrating a functional GVL effect. Using this model system, we studied human GVL responses against human AML cells in vivo and discovered that AML induced T cell depletion, likely because of increased T cell apoptosis. In addition, AML caused T cell exhaustion manifested by upregulation of inhibitory receptors, increased expression of exhaustion-related transcription factors, and decreased T cell function. Importantly, combined blockade of human T cell-inhibitory pathways effectively reduced leukemia burden and reinvigorated CD8 T cell function in this model system. These data, generated in a highly clinically relevant humanized GVL model, not only demonstrate AML-induced inhibition of alloreactive T cells but also identify promising therapeutic strategies targeting T cell depletion and exhaustion for overcoming GVL failure and treating AML relapse after alloSCT.

Interleukin-4 treatment reduces leukemia burden in acute myeloid leukemia.

Interleukin-4 (IL-4) is a signature cytokine pivotal in Type 2 helper T cell (Th2) immune response, particularly in allergy and hypersensitivity. Interestingly, IL-4 increases endogenous levels of prostaglandin D2 (PGD2 ) and its metabolites, Δ12 -prostaglandin J2 (Δ12 -PGJ2 ) and 15-deoxy-Δ12,14 -prostaglandin J2 (15d-PGJ2 ), collectively called cyclopentenone PGs (CyPGs). However, the therapeutic role of IL-4 in hematologic malignancies remains unclear. Here, we employed a murine model of acute myeloid leukemia (AML), where human MLL-AF9 fusion oncoprotein was expressed in hematopoietic progenitor cells, to test the effect of IL-4 treatment in vivo. Daily intraperitoneal treatment with IL-4 at 60 µg/kg/d significantly alleviated the severity of AML, as seen by decreased leukemia-initiating cells (LICs). The effect of IL-4 was mediated, in part, by the enhanced expression of hematopoietic- PGD2  synthase (H-PGDS) to effect endogenous production of CyPGs, through autocrine and paracrine signaling mechanisms. Similar results were seen with patient-derived AML cells cultured ex vivo with IL-4. Use of GW9662, a peroxisome proliferator-activated receptor gamma (PPARγ) antagonist, suggested endogenous CyPGs-PPARγ axis mediated p53-dependent apoptosis of LICs by IL-4. Taken together, our results reveal a beneficial role of IL-4 treatment in AML suggesting a potential therapeutic regimen worthy of clinical trials in patients with AML.

Epo receptor signaling in macrophages alters the splenic niche to promote erythroid differentiation.

Anemic stress induces stress erythropoiesis, which rapidly generates new erythrocytes to restore tissue oxygenation. Stress erythropoiesis is best understood in mice where it is extramedullary and occurs primarily in the spleen. However, both human and mouse stress erythropoiesis use signals and progenitor cells that are distinct from steady-state erythropoiesis. Immature stress erythroid progenitors (SEPs) are derived from short-term hematopoietic stem cells. Although the SEPs are capable of self-renewal, they are erythroid restricted. Inflammation and anemic stress induce the rapid proliferation of SEPs, but they do not differentiate until serum erythropoietin (Epo) levels increase. Here we show that rather than directly regulating SEPs, Epo promotes this transition from proliferation to differentiation by acting on macrophages in the splenic niche. During the proliferative stage, macrophages produce canonical Wnt ligands that promote proliferation and inhibit differentiation. Epo/Stat5-dependent signaling induces the production of bioactive lipid mediators in macrophages. Increased production of prostaglandin J2 (PGJ2) activates peroxisome proliferator-activated receptor γ (PPARγ)-dependent repression of Wnt expression, whereas increased production of prostaglandin E2 (PGE2) promotes the differentiation of SEPs.

Mechanisms of erythrocyte development and regeneration: implications for regenerative medicine and beyond.

Hemoglobin-expressing erythrocytes (red blood cells) act as fundamental metabolic regulators by providing oxygen to cells and tissues throughout the body. Whereas the vital requirement for oxygen to support metabolically active cells and tissues is well established, almost nothing is known regarding how erythrocyte development and function impact regeneration. Furthermore, many questions remain unanswered relating to how insults to hematopoietic stem/progenitor cells and erythrocytes can trigger a massive regenerative process termed 'stress erythropoiesis' to produce billions of erythrocytes. Here, we review the cellular and molecular mechanisms governing erythrocyte development and regeneration, and discuss the potential links between these events and other regenerative processes.

Monocyte-derived macrophages expand the murine stress erythropoietic niche during the recovery from anemia.

Anemic stress induces a physiological response that includes the rapid production of new erythrocytes. This process is referred to as stress erythropoiesis. It is best understood in the mouse where it is extramedullary and utilizes signals and progenitor cells that are distinct from bone marrow steady-state erythropoiesis. The development of stress erythroid progenitors occurs in close association with the splenic stress erythropoiesis niche. In particular, macrophages in the niche are required for proper stress erythropoiesis. Here we show that the expansion of the niche occurs in concert with the proliferation and differentiation of stress erythroid progenitors. Using lineage tracing analysis in 2 models of anemic stress, we show that the expansion of the splenic niche is due to the recruitment of monocytes into the spleen, which develop into macrophages that form erythroblastic islands. The influx in monocytes into the spleen depends in part on Ccr2-dependent signaling mediated by Ccl2 and other ligands expressed by spleen resident red pulp macrophages. Overall, these data demonstrate the dynamic nature of the spleen niche, which rapidly expands in concert with the stress erythroid progenitors to coordinate the production of new erythrocytes in response to anemic stress.

Selenoproteins regulate stress erythroid progenitors and spleen microenvironment during stress erythropoiesis.

Micronutrient selenium (Se) plays a key role in redox regulation through its incorporation into selenoproteins as the 21st amino acid selenocysteine (Sec). Because Se deficiency appears to be a cofactor in the anemia associated with chronic inflammatory diseases, we reasoned that selenoproteins may contribute to erythropoietic recovery from anemia, referred to as stress erythropoiesis. Here, we report that loss of selenoproteins through Se deficiency or by mutation of the Sec tRNA (tRNA[Sec]) gene (Trsp) severely impairs stress erythropoiesis at 2 stages. Early stress erythroid progenitors failed to expand and properly differentiate into burst-forming unit-erythroid cells , whereas late-stage erythroid progenitors exhibited a maturation defect that affected the transition of proerythroblasts to basophilic erythroblasts. These defects were, in part, a result of the loss of selenoprotein W (SelenoW), whose expression was reduced at both transcript and protein levels in Se-deficient erythroblasts. Mutation of SelenoW in the bone marrow cells significantly decreased the expansion of stress burst-forming unit-erythroid cell colonies, which recapitulated the phenotypes induced by Se deficiency or mutation of Trsp Similarly, mutation of SelenoW in murine erythroblast (G1E) cell line led to defects in terminal differentiation. In addition to the erythroid defects, the spleens of Se-deficient mice contained fewer red pulp macrophages and exhibited impaired development of erythroblastic island macrophages, which make up the niche supporting erythroblast development. Taken together, these data reveal a critical role of selenoproteins in the expansion and development of stress erythroid progenitors, as well as the erythroid niche during acute anemia recovery.

Crth2 receptor signaling down-regulates lipopolysaccharide-induced NF-κB activation in murine macrophages via changes in intracellular calcium.

Prostaglandin D2 and its cyclopentenone metabolites [cyclopentenone prostaglandins (CyPGs)], Δ12prostaglandin J2 and 15-deoxy-Δ12,14-prostaglandin J2, act through 2 GPCRs, d-type prostanoid 1 and the chemoattractant receptor homologous molecule expressed on type 2 T-helper cells (Crth2). In addition to its role in allergy and asthma, the role of Crth2 in the resolution of inflammation, to mediate the proresolving functions of endogenous CyPGs, is not well understood. We investigated the regulation of LPS or zymosan-induced inflammatory response by signals from the Crth2 receptor in macrophages that lack Crth2 expression [knockout (KO)]. Increased expression of proinflammatory genes, including Tnf-α, was observed in Crth2 KO cells. Targeting the endogenous biosynthetic pathway of CyPGs with indomethacin or HQL79, which inhibit cyclooxygenases or hematopoietic prostaglandin D synthase, respectively, or use of Crth2 antagonists recapitulated the proinflammatory phenotype as in Crth2 KO cells. Ligand-dependent activation of Crth2 by 13,14-dihydro-15-keto-prostaglandin D2 increased Ca2+ influx through store-operated Ca2+ entry (SOCE) accompanied by the up-regulation of stromal interaction molecule 1 and calcium release-activated calcium modulator 1 expression, suggesting that the proresolution effects of CyPG-dependent activation of SOCE could be mediated by Crth2 during inflammation. Interestingly, Crth2 signaling down-regulated the Ca2+-regulated heat stable protein 1 that stabilizes Tnf-α mRNA via the increased expression of microRNA 155 to dampen inflammatory responses triggered through the TNF-α-NF-κB axis. In summary, these studies present a novel regulatory role for Crth2 during inflammatory response in macrophages.-Diwakar, B. T., Yoast, R., Nettleford, S., Qian, F., Lee, T.-J., Berry, S., Huffnagle, I., Rossi, R. M., Trebak, M., Paulson, R. F., Prabhu, K. S. Crth2 receptor signaling down-regulates lipopolysaccharide-induced NF-κB activation in murine macrophages via changes in intracellular calcium.

Development of SKI-349, a dual-targeted inhibitor of sphingosine kinase and microtubule polymerization.

Our sphingosine kinase inhibitor (SKI) optimization studies originated with the optimization of the SKI-I chemotype by replacement of the substituted benzyl rings with substituted phenyl rings giving rise to the discovery of SKI-178. We have recently reported that SKI-178 is a dual-targeted inhibitor of both sphingosine kinase isoforms (SphK1/2) and a microtubule disrupting agent (MDA). In mechanism-of-action studies, we have shown that these two separate actions synergize to induce cancer cell death in acute myeloid leukemia (AML) cell and animal models. Owning to the effectiveness of SKI-178, we sought to further refine the chemotype while maintaining "on-target" SKI and MDA activities. Herein, we modified the "linker region" between the substituted phenyl rings of SKI-178 through a structure guided approach. These studies have yielded the discovery of an SKI-178 congener, SKI-349, with log-fold enhancements in both SphK inhibition and cytotoxic potency. Importantly, SKI-349 also demonstrates log-fold improvements in therapeutic efficacy in a retro-viral transduction model of MLL-AF9 AML as compared to previous studies with SKI-178. Together, our results strengthen the hypothesis that simultaneous targeting of the sphingosine kinases (SphK1/2) and the induction of mitotic spindle assembly checkpoint arrest, via microtubule disruption, might be an effective therapeutic strategy for hematological malignancies including AML.

Activation of PPARγ by endogenous prostaglandin J2 mediates the antileukemic effect of selenium in murine leukemia.

Supplementation with nontoxic doses of micronutrient selenium has been shown to alleviate chronic myelogenous leukemia (CML) via the elimination of leukemia stem cells (LSCs) in mice. This treatment provides a new and novel method for eliminating the LSCs that are otherwise not targeted by existing therapies. The antileukemic effect of selenium was dependent on the production of endogenous cyclopentenone prostaglandins (CyPGs), Δ-12 prostaglandin J2 (Δ12-PGJ2), and 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2). Here, we show that these endogenous CyPGs, produced by mice maintained on selenium-supplemented diets, alleviate the symptoms of CML through their ability to activate the nuclear hormone receptor, peroxisome proliferator activated receptor γ (PPARγ). GW9662, a potent PPARγ antagonist, blocked the antileukemic effect of selenium supplementation by significantly reducing CyPGs. This effect was mediated by an increase in 15-prostaglandin dehydrogenase (15-Pgdh) activity, which oxidizes and inactivates Δ12-PGJ2 and 15d-PGJ2 In contrast, treatment with the PPARγ agonist pioglitazone mimicked selenium supplementation. This treatment led to decreased 15-Pgdh activity and increased CyPG levels, which inhibited CML progression. Selenium-dependent activation of PPARγ mediated by endogenous CyPGs decreased Stat5 expression leading to the downregulation of Cited2, a master regulator of LSC quiescence. These studies suggest a potential role for selenium supplementation as an adjuvant therapy in CML.

In vitro culture of stress erythroid progenitors identifies distinct progenitor populations and analogous human progenitors.

Tissue hypoxia induces a systemic response designed to increase oxygen delivery to tissues. One component of this response is increased erythropoiesis. Steady-state erythropoiesis is primarily homeostatic, producing new erythrocytes to replace old erythrocytes removed from circulation by the spleen. In response to anemia, the situation is different. New erythrocytes must be rapidly made to increase hemoglobin levels. At these times, stress erythropoiesis predominates. Stress erythropoiesis is best characterized in the mouse, where it is extramedullary and utilizes progenitors and signals that are distinct from steady-state erythropoiesis. In this report, we use an in vitro culture system that recapitulates the in vivo development of stress erythroid progenitors. We identify cell-surface markers that delineate a series of stress erythroid progenitors with increasing maturity. In addition, we use this in vitro culture system to expand human stress erythroid progenitor cells that express analogous cell-surface markers. Consistent with previous suggestions that human stress erythropoiesis is similar to fetal erythropoiesis, we demonstrate that human stress erythroid progenitors express fetal hemoglobin upon differentiation. These data demonstrate that similar to murine bone marrow, human bone marrow contains cells that can generate BMP4-dependent stress erythroid burst-forming units when cultured under stress erythropoiesis conditions.

Δ12-prostaglandin J3, an omega-3 fatty acid-derived metabolite, selectively ablates leukemia stem cells in mice.

Targeting cancer stem cells is of paramount importance in successfully preventing cancer relapse. Recently, in silico screening of public gene-expression datasets identified cyclooxygenase-derived cyclopentenone prostaglandins (CyPGs) as likely agents to target malignant stem cells. We show here that Δ(12)-PGJ(3), a novel and naturally produced CyPG from the dietary fish-oil ω-3 polyunsaturated fatty acid eicosapentaenoic acid (EPA; 20:5) alleviates the development of leukemia in 2 well-studied murine models of leukemia. IP administration of Δ(12)-PGJ(3) to mice infected with Friend erythroleukemia virus or those expressing the chronic myelogenous leukemia oncoprotein BCR-ABL in the hematopoietic stem cell pool completely restored normal hematologic parameters, splenic histology, and enhanced survival. More importantly, Δ(12)-PGJ(3) selectively targeted leukemia stem cells (LSCs) for apoptosis in the spleen and BM. This treatment completely eradicated LSCs in vivo, as demonstrated by the inability of donor cells from treated mice to cause leukemia in secondary transplantations. Given the potency of ω-3 polyunsaturated fatty acid-derived CyPGs and the well-known refractoriness of LSCs to currently used clinical agents, Δ(12)-PGJ(3) may represent a new chemotherapeutic for leukemia that targets LSCs.

Self-renewal of leukemia stem cells in Friend virus-induced erythroleukemia requires proviral insertional activation of Spi1 and hedgehog signaling but not mutation of p53.

Friend virus induces erythroleukemia through a characteristic two-stage progression. The prevailing model proposes that during the initial, polyclonal stage of disease most of the infected cells terminally differentiate, resulting in acute erythrocytosis. In the late stage of disease, a clonal leukemia develops through the acquisition of new mutations--proviral insertional activation of Spi1/Pu.1 and mutation of p53. Previous work from our laboratory demonstrated that Friend virus activates the bone morphogenic protein 4 (BMP4)-dependent stress erythropoiesis pathway, which leads to the rapid expansion of stress erythroid progenitors, which are the targets for Friend virus in the spleen. We recently showed that stress erythroid progenitors have intrinsic self-renewal ability and therefore could function as leukemia stem cells (LSCs) when infected with Friend virus. Here, we show that the two stages of Friend virus-induced disease are caused by infection of distinct stress progenitor populations in the spleen. The development of leukemia relies on the ability of the virus to hijack the intrinsic self-renewal capability of stress erythroid progenitors leading to the generation of LSCs. Two signals are required for the self-renewal of Friend virus LSCs proviral insertional activation of Spi1/Pu.1 and Hedgehog-dependent signaling. Surprisingly, mutation of p53 is not observed in LSCs. These data establish a new model for Friend virus-induced erythroleukemia and demonstrate the utility of Friend virus as a model system to study LSC self-renewal.

Intra-Peritoneal Transplantation for Generating Acute Myeloid Leukemia in Mice.

There is an unmet need for novel therapies to treat acute myeloid leukemia (AML) and the associated relapse that involves persistent leukemia stem cells (LSCs). An experimental AML rodent model to test therapies based on successfully transplanting these cells via retro-orbital injections in recipient mice is fraught with challenges. The aim of this study was to develop an easy, reliable, and consistent method to generate a robust murine model of AML using an intra-peritoneal route. In the present protocol, bone marrow cells were transduced with a retrovirus expressing human MLL-AF9 fusion oncoprotein. The efficiency of lineage negative (Lin-) and Lin-Sca-1+c-Kit+ (LSK) populations as donor LSCs in the development of primary AML was tested, and intra-peritoneal injection was adopted as a new method to generate AML. Comparison between intra-peritoneal and retro-orbital injections was done in serial transplantations to compare and contrast the two methods. Both Lin- and LSK cells transduced with human MLL-AF9 virus engrafted well in the bone marrow and spleen of recipients, leading to a full-blown AML. The intra-peritoneal injection of donor cells established AML in recipients upon serial transplantation, and the infiltration of AML cells was detected in the blood, bone marrow, spleen, and liver of recipients by flow cytometry, qPCR, and histological analyses. Thus, intra-peritoneal injection is an efficient method of AML induction using serial transplantation of donor leukemic cells.

Nitric oxide regulates metabolism in murine stress erythroid progenitors to promote recovery during inflammatory anemia.

Inflammation skews bone marrow hematopoiesis increasing the production of myeloid effector cells at the expense of steady-state erythropoiesis. A compensatory stress erythropoiesis response is induced to maintain homeostasis until inflammation is resolved. In contrast to steady-state erythroid progenitors, stress erythroid progenitors (SEPs) utilize signals induced by inflammatory stimuli. However, the mechanistic basis for this is not clear. Here we reveal a nitric oxide (NO)-dependent regulatory network underlying two stages of stress erythropoiesis, namely proliferation, and the transition to differentiation. In the proliferative stage, immature SEPs and cells in the niche increased expression of inducible nitric oxide synthase ( Nos2 or iNOS ) to generate NO. Increased NO rewires SEP metabolism to increase anabolic pathways, which drive the biosynthesis of nucleotides, amino acids and other intermediates needed for cell division. This NO-dependent metabolism promotes cell proliferation while also inhibiting erythroid differentiation leading to the amplification of a large population of non-committed progenitors. The transition of these progenitors to differentiation is mediated by the activation of nuclear factor erythroid 2-related factor 2 (Nfe2l2 or Nrf2). Nrf2 acts as an anti-inflammatory regulator that decreases NO production, which removes the NO-dependent erythroid inhibition and allows for differentiation. These data provide a paradigm for how alterations in metabolism allow inflammatory signals to amplify immature progenitors prior to differentiation. Key points: Nitric-oxide (NO) dependent signaling favors an anabolic metabolism that promotes proliferation and inhibits differentiation.Activation of Nfe2l2 (Nrf2) decreases NO production allowing erythroid differentiation.

Activation of GPR44 decreases severity of myeloid leukemia via specific targeting of leukemia initiating stem cells.

Relapse of acute myeloid leukemia (AML) remains a significant concern due to persistent leukemia-initiating stem cells (LICs) that are typically not targeted by most existing therapies. Using a murine AML model, human AML cell lines, and patient samples, we show that AML LICs are sensitive to endogenous and exogenous cyclopentenone prostaglandin-J (CyPG), Δ12-PGJ2, and 15d-PGJ2, which are increased upon dietary selenium supplementation via the cyclooxygenase-hematopoietic PGD synthase pathway. CyPGs are endogenous ligands for peroxisome proliferator-activated receptor gamma and GPR44 (CRTH2; PTGDR2). Deletion of GPR44 in a mouse model of AML exacerbated the disease suggesting that GPR44 activation mediates selenium-mediated apoptosis of LICs. Transcriptomic analysis of GPR44-/- LICs indicated that GPR44 activation by CyPGs suppressed KRAS-mediated MAPK and PI3K/AKT/mTOR signaling pathways, to enhance apoptosis. Our studies show the role of GPR44, providing mechanistic underpinnings of the chemopreventive and chemotherapeutic properties of selenium and CyPGs in AML.

Selenoprotein-dependent up-regulation of hematopoietic prostaglandin D2 synthase in macrophages is mediated through the activation of peroxisome proliferator-activated receptor (PPAR) gamma.

The plasticity of macrophages is evident from their dual role in inflammation and resolution of inflammation that are accompanied by changes in the transcriptome and metabolome. Along these lines, we have previously demonstrated that the micronutrient selenium increases macrophage production of arachidonic acid (AA)-derived anti-inflammatory 15-deoxy-Δ(12,14)-prostaglandin J(2) (15d-PGJ(2)) and decreases the proinflammatory PGE(2). Here, we hypothesized that selenium modulated the metabolism of AA by a differential regulation of various prostaglandin (PG) synthases favoring the production of PGD(2) metabolites, Δ(12)-PGJ(2) and 15d-PGJ(2). A dose-dependent increase in the expression of hematopoietic-PGD(2) synthase (H-PGDS) by selenium and a corresponding increase in Δ(12)-PGJ(2) and 15d-PGJ(2) in RAW264.7 macrophages and primary bone marrow-derived macrophages was observed. Studies with organic non-bioavailable forms of selenium and the genetic manipulation of cellular selenium incorporation machinery indicated that selenoproteins were necessary for H-PGDS expression and 15d-PGJ(2) production. Treatment of selenium-deficient macrophages with rosiglitazone, a peroxisome proliferator-activated receptor γ ligand, up-regulated H-PGDS. Furthermore, electrophoretic mobility shift assays indicated the presence of an active peroxisome proliferator-activated receptor-response element in murine Hpgds promoter suggesting a positive feedback mechanism of H-PGDS expression. Alternatively, the expression of nuclear factor-κB-dependent thromboxane synthase and microsomal PGE(2) synthase was down-regulated by selenium. Using a Friend virus infection model of murine leukemia, the onset of leukemia was observed only in selenium-deficient and indomethacin-treated selenium-supplemented mice but not in the selenium-supplemented group or those treated with 15d-PGJ(2). These results suggest the importance of selenium in the shunting of AA metabolism toward the production of PGD(2) metabolites, which may have clinical implications.

Stress erythropoiesis: new signals and new stress progenitor cells.

PURPOSE OF REVIEW: Acute anemic stress induces a physiological response that includes the rapid development of new erythrocytes. This process is referred to as stress erythropoiesis, which is distinct from steady state erythropoiesis. Much of what we know about stress erythropoiesis comes from the analysis of murine models. In this review, we will discuss our current understanding of the mechanisms that regulate stress erythropoiesis in mice and discuss outstanding questions in the field. RECENT FINDINGS: Stress erythropoiesis occurs in the murine spleen, fetal liver and adult liver. The signals that regulate this process are Hedgehog, bone morphogenetic protein 4 (BMP4), stem cell factor and hypoxia. Recent findings show that stress erythropoiesis utilizes a population of erythroid-restricted self-renewing stress progenitors. Although the BMP4-dependent stress erythropoiesis pathway was first characterized during the recovery from acute anemia, analysis of a mouse model of chronic anemia demonstrated that activation of the BMP4-dependent stress erythropoiesis pathway provides compensatory erythropoiesis in response to chronic anemia as well. SUMMARY: The BMP4-dependent stress erythropoiesis pathway plays a key role in the recovery from acute anemia and new data show that this pathway compensates for ineffective steady state erythropoiesis in a murine model of chronic anemia. The identification of a self-renewing population of stress erythroid progenitors in mice suggests that therapeutic manipulation of this pathway may be useful for the treatment of human anemia. However, the development of new therapies will await the characterization of an analogous pathway in humans.

Metabolic regulation of stress erythropoiesis, outstanding questions, and possible paradigms.

Steady state erythropoiesis produces new erythrocytes at a constant rate to replace the senescent cells that are removed by macrophages in the liver and spleen. However, infection and tissue damage disrupt the production of erythrocytes by steady state erythropoiesis. During these times, stress erythropoiesis is induced to compensate for the loss of erythroid output. The strategy of stress erythropoiesis is different than steady state erythropoiesis. Stress erythropoiesis generates a wave of new erythrocytes to maintain homeostasis until steady state conditions are resumed. Stress erythropoiesis relies on the rapid proliferation of immature progenitor cells that do not differentiate until the increase in serum Erythropoietin (Epo) promotes the transition to committed progenitors that enables their synchronous differentiation. Emerging evidence has revealed a central role for cell metabolism in regulating the proliferation and differentiation of stress erythroid progenitors. During the initial expansion stage, the immature progenitors are supported by extensive metabolic changes which are designed to direct the use of glucose and glutamine to increase the biosynthesis of macromolecules necessary for cell growth and division. At the same time, these metabolic changes act to suppress the expression of genes involved in erythroid differentiation. In the subsequent transition stage, changes in niche signals alter progenitor metabolism which in turn removes the inhibition of erythroid differentiation generating a bolus of new erythrocytes to alleviate anemia. This review summarizes what is known about the metabolic regulation of stress erythropoiesis and discusses potential mechanisms for metabolic regulation of proliferation and differentiation.