c-Myc–mediated control of cell fate in megakaryocyte-erythrocyte progenitors

Abstract It has been found that c-Myc protein plays a critical role in controlling self-renewal versus differentiation in hematopoietic stem cells. We report that c-Myc also controls the fate of megakaryocyte-erythrocyte progenitors through regulating the differentiation of erythroid and megakaryocytic progenitors. In addition to the significant reduction of granulocytes/macrophages and B and T lymphocytes because of the reduction of their corresponding progenitors, we found significantly increased numbers of megakaryocytic progenitors and mature megakaryocytes in bone marrow and spleens of c-Myc-knockout (c-Myc−/−) mice. Differentiation of erythrocytes was blocked at the erythroid progenitor stage. This increased megakaryocytopoiesis is a cell-intrinsic defect of c-Myc-mutant hematopoietic stem cells, as shown by transplantation studies. Furthermore, we found that c-Myc is required for polyploidy formation but not for cytoplasmic maturation of megakaryocytes. Megakaryocytes from c-Myc−/− mice are significantly smaller in size and lower in ploidy than those of control mice; however, because of the dramatic increase in megakaryocyte number, although fewer platelets are produced by each megakaryocyte, a greater than 3-fold increase in platelet number was consistently observed in c-Myc−/− mice. Thus, c-Myc−/− mice develop a syndrome of severe thrombocytosis-anemia-leukopenia because of significant increases in megakaryocytopoiesis and concomitant blockage of erythrocyte differentiation and reductions in myelolymphopoiesis.

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Growth Factor Independence 1 b (Gfi1b) as a New Regulator of Hematopoietic Stem Cell Fate

Blood ◽

10.1182/blood.v116.21.837.837 ◽

2010 ◽

Vol 116 (21) ◽

pp. 837-837

Author(s):

Cyrus Khandanpour ◽

Lothar Vassen ◽

Marie-Claude Gaudreau ◽

Christian Kosan ◽

Tarik Moroy

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Cell Fate ◽

Conflicts Of Interest ◽

Fold Increase ◽

Surface Proteins ◽

Expression Array ◽

Hematopoietic Stem

Abstract Abstract 837 Donor matched transplantation of bone marrow or hematopoietic stem cells (HSCs) are widely used to treat hematological malignancies, but are associated with high mortality. Methods for expansion of HSC numbers and their mobilization into the bloodstream of a donor could significantly improve therapy. We show here that the zinc finger transcriptional repressor Gfi1b is highly expressed in hematopoietic stem cells (defined as CD 150+, CD 48-, Lin-, Sca1+ and c-kit+) cells and is down-regulated more than 10 fold upon differentiation into multipotential progenitors (defined as CD 150+ or CD150-, CD 48+, Lin-, Sca1+ and c-kit+). Constitutive germline deletion of Gfi1b is lethal at midgestation due to impaired development of erythrocytes and megakaryocytes. We have therefore developed a conditional knock-out of Gfi1b to study its role specifically in the adult hematopoietic system. Deletion of Gfi1b leads to a 30-fold increase of HSC numbers in bone marrow and around a100 fold increase in spleen and peripheral blood. This was due to a higher rate of HSCs undergoing cell cycling. Concomitantly, the number of quiescent HSCs was reduced 5–6 times. We then performed an gene expression array of wt and Gfi1b deficient HSCs and observed that loss of Gfi1b leads to an altered RNA expression of integrins and adhesion molecules, for instance CXCR4, VCAM-1 and Tenascin C, which usually retain HSCs in a dormant state in the endosteal niche. These changes were also confirmed on protein level. Finally, we could observe a higher levels of Reactive Oxygen Species (ROS) in the Gfi1b deficient HSCs compared to wt HSCs. We verified whether elevated level of ROS are causative for the expansion of HSCs and noticed that application of N-Acetyl-Cystein, which counteracts the effects of ROS, limits significantly the expansion of HSCs, underscoring the important role of ROS in the expansion of Gfi1b deficient HSCs. Despite markedly increased proliferation, Gfi1b-/- HSCs can reconstitute lymphoid and myeloid lineages to the same extent as wt HSCs when transplanted in competition with wt HSCs. Furthermore, Gfi1b deficient HSCs also feature an expansion after transplantation and expand 5–10 fold more than wt HSC when transplanted initially in equal numbers with wt HSCs. It is possible that lower expression of CXCR4, VCAM-1 and other surface proteins leads to release and egression of Gfi1b deficient HSCs from the hypoxic endosteal stem cell niche and exposes the HSCs to more oxygen which in turn increases ROS levels. Elevated ROS could promote entry of Gfi1b-/- HSCs into cell cycle. In conclusion Gfi1b regulates HSC dormancy, pool size and potentially also the egress and mobilization of HSCs and might offer a new therapeutic approach to improve human HSC transplantation. Disclosures: No relevant conflicts of interest to declare.

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Endothelial stroma programs hematopoietic stem cells to differentiate into regulatory dendritic cells through IL-10

Blood ◽

10.1182/blood-2006-01-007187 ◽

2006 ◽

Vol 108 (4) ◽

pp. 1189-1197 ◽

Cited By ~ 88

Author(s):

Hua Tang ◽

Zhenhong Guo ◽

Minghui Zhang ◽

Jianli Wang ◽

Guoyou Chen ◽

...

Keyword(s):

Stem Cells ◽

Dendritic Cells ◽

T Cell ◽

Hematopoietic Stem Cells ◽

Critical Role ◽

Regulatory Function ◽

Hematopoietic Stem ◽

Regulatory Dendritic Cells

Abstract Regulatory dendritic cells (DCs) have been reported recently, but their origin is poorly understood. Our previous study demonstrated that splenic stroma can drive mature DCs to proliferate and differentiate into regulatory DCs, and their natural counterpart with similar regulatory function in normal spleens has been identified. Considering that the spleen microenvironment supports hematopoiesis and that hematopoietic stem cells (HSCs) are found in spleens of adult mice, we wondered whether splenic microenvironment could differentiate HSCs into regulatory DCs. In this report, we demonstrate that endothelial splenic stroma induce HSCs to differentiate into a distinct regulatory DC subset with high expression of CD11b but low expression of Ia. CD11bhiIalo DCs secreting high levels of TGF-β, IL-10, and NO can suppress T-cell proliferation both in vitro and in vivo. Furthermore, CD11bhiIalo DCs have the ability to potently suppress allo-DTH in vivo, indicating their preventive or therapeutic perspectives for some immunologic disorders. The inhibitory function of CD11bhiIalo DCs is mediated through NO but not through induction of regulatory T (Treg) cells or T-cell anergy. IL-10, which is secreted by endothelial splenic stroma, plays a critical role in the differentiation of the regulatory CD11bhiIalo DCs from HSCs. These results suggest that splenic microenvironment may physiologically induce regulatory DC differentiation in situ.

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Scl isoforms act downstream of etsrp to specify angioblasts and definitive hematopoietic stem cells

Blood ◽

10.1182/blood-2009-09-244640 ◽

2010 ◽

Vol 115 (26) ◽

pp. 5338-5346 ◽

Cited By ~ 42

Author(s):

Xi Ren ◽

Gustavo A. Gomez ◽

Bo Zhang ◽

Shuo Lin

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Critical Role ◽

Vascular Endothelial ◽

Hematopoietic Stem ◽

Vegf Signaling ◽

Endothelial Growth Factor ◽

Endothelial Development ◽

Definitive Hematopoiesis ◽

Runx1 Expression

Abstract Recent lineage studies suggest that hematopoietic stem cells (HSCs) may be derived from endothelial cells. However, the genetic hierarchy governing the emergence of HSCs remains elusive. We report here that zebrafish ets1-related protein (etsrp), which is essential for vascular endothelial development, also plays a critical role in the initiation of definitive hematopoiesis by controlling the expression of 2 stem cell leukemia (scl) isoforms (scl-α and scl-β) in angioblasts. In etsrp morphants, which are deficient in endothelial and HSC development, scl-α alone partially rescues angioblast specification, arterial-venous differentiation, and the expression of HSC markers, runx1 and c-myb, whereas scl-β requires angioblast rescue by fli1a to restore runx1 expression. Interestingly, when vascular endothelial growth factor (Vegf) signaling is inhibited, HSC marker expression can still be restored by scl-α in etsrp morphants, whereas the rescue of arterial ephrinb2a expression is blocked. Furthermore, both scl isoforms partially rescue runx1 but not ephrinb2a expression in embryos deficient in Vegf signaling. Our data suggest that downstream of etsrp, scl-α and fli1a specify the angioblasts, whereas scl-β further initiates HSC specification from this angioblast population, and that Vegf signaling acts upstream of scl-β during definitive hematopoiesis.

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The Making of Hematopoiesis: Developmental Ancestry and Environmental Nurture

International Journal of Molecular Sciences ◽

10.3390/ijms19072122 ◽

2018 ◽

Vol 19 (7) ◽

pp. 2122 ◽

Cited By ~ 7

Author(s):

Geoffrey Brown ◽

Rhodri Ceredig ◽

Panagiotis Tsapogas

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Progenitor Cells ◽

Cell Fate ◽

Hematopoietic Stem ◽

Fate Determination ◽

Intermediate Progenitors ◽

Lineage Tree ◽

Stem And Progenitor Cells ◽

The Many

Evidence from studies of the behaviour of stem and progenitor cells and of the influence of cytokines on their fate determination, has recently led to a revised view of the process by which hematopoietic stem cells and their progeny give rise to the many different types of blood and immune cells. The new scenario abandons the classical view of a rigidly demarcated lineage tree and replaces it with a much more continuum-like view of the spectrum of fate options open to hematopoietic stem cells and their progeny. This is in contrast to previous lineage diagrams, which envisaged stem cells progressing stepwise through a series of fairly-precisely described intermediate progenitors in order to close down alternative developmental options. Instead, stem and progenitor cells retain some capacity to step sideways and adopt alternative, closely related, fates, even after they have “made a lineage choice.” The stem and progenitor cells are more inherently versatile than previously thought and perhaps sensitive to lineage guidance by environmental cues. Here we examine the evidence that supports these views and reconsider the meaning of cell lineages in the context of a continuum model of stem cell fate determination and environmental modulation.

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Rapid and sustained hematopoietic recovery in lethally irradiated mice transplanted with purified Thy-1.1lo Lin-Sca-1+ hematopoietic stem cells

Blood ◽

10.1182/blood.v83.12.3758.3758 ◽

1994 ◽

Vol 83 (12) ◽

pp. 3758-3779 ◽

Cited By ~ 123

Author(s):

N Uchida ◽

HL Aguila ◽

WH Fleming ◽

L Jerabek ◽

IL Weissman

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Blood Cells ◽

Critical Role ◽

White Blood Cells ◽

Cell Types ◽

Dose Dependence ◽

Hematopoietic Stem ◽

Hematopoietic Recovery

Abstract Hematopoietic stem cells (HSCs) are believed to play a critical role in the sustained repopulation of all blood cells after bone marrow transplantation (BMT). However, understanding the role of HSCs versus other hematopoietic cells in the quantitative reconstitution of various blood cell types has awaited methods to isolate HSCs. A candidate population of mouse HSCs, Thy-1.1lo Lin-Sca-1+ cells, was isolated several years ago and, recently, this population has been shown to be the only population of BM cells that contains HSCs in C57BL/Ka-Thy-1.1 mice. As few as 100 of these cells can radioprotect 95% to 100% of irradiated mice, resulting long-term multilineage reconstitution. In this study, we examined the reconstitution potential of irradiated mice transplanted with purified Thy-1.1lo Lin-Sca-1+ BM cells. Donor-derived peripheral blood (PB) white blood cells were detected as early as day 9 or 10 when 100 to 1,000 Thy-1.1lo Lin-Sca-1+ cells were used, with minor dose-dependent differences. The reappearance of platelets by day 14 and thereafter was also seen at all HSC doses (100 to 1,000 cells), with a slight dose-dependence. All studied HSC doses also allowed RBC levels to recover, although at the 100 cell dose a delay in hematocrit recovery was observed at day 14. When irradiated mice were transplanted with 500 Thy-1.1lo Lin-Sca-1+ cells compared with 1 x 10(6) BM cells (the equivalent amount of cells that contain 500 Thy-1.1lo Lin-Sca-1+ cells as well as progenitor and mature cells), very little difference in the kinetics of recovery of PB, white blood cells, platelets, and hematocrit was observed. Surprisingly, even when 200 Thy1.1lo Lin-Sca- 1+ cells were mixed with 4 x 10(5) Sca-1- BM cells in a competitive repopulation assay, most of the early (days 11 and 14) PB myeloid cells were derived from the HSC genotype, indicating the superiority of the Thy-1.1lo Lin-Sca-1+ cells over Sca-1- cells even in the early phases of myeloid reconstitution. Within the Thy-1.1lo Lin-Sca-1+ population, the Rhodamine 123 (Rh123)hi subset dominates in PB myeloid reconstitution at 10 to 14 days, only to be overtaken by the Rh123lo subset at 3 weeks and thereafter. These findings indicate that HSCs can account for the early phase of hematopoietic recovery, as well as sustained hematopoiesis, and raise questions about the role of non-HSC BM populations in the setting of BMT.

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Human hematopoietic stem cells stimulated to proliferate in vitro lose engraftment potential during their S/G2/M transit and do not reenter G0

Blood ◽

10.1182/blood.v96.13.4185 ◽

2000 ◽

Vol 96 (13) ◽

pp. 4185-4193 ◽

Cited By ~ 181

Author(s):

Hanno Glimm ◽

IL-Hoan Oh ◽

Connie J. Eaves

Keyword(s):

Cell Cycle ◽

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Transforming Growth Factor ◽

Ex Vivo ◽

Cell Activity ◽

Human Cord Blood ◽

Hematopoietic Stem ◽

A Cell

Abstract An understanding of mechanisms regulating hematopoietic stem cell engraftment is of pivotal importance to the clinical use of cultured and genetically modified transplants. Human cord blood (CB) cells with lymphomyeloid repopulating activity in NOD/SCID mice were recently shown to undergo multiple self-renewal divisions within 6 days in serum-free cultures containing Flt3-ligand, Steel factor, interleukin 3 (IL-3), IL-6, and granulocyte colony-stimulating factor. The present study shows that, on the fifth day, the transplantable stem cell activity is restricted to the G1fraction, even though both colony-forming cells (CFCs) and long-term culture-initiating cells (LTC-ICs) in the same cultures are approximately equally distributed between G0/G1and S/G2/M. Interestingly, the G0 cells defined by their low levels of Hoechst 33342 and Pyronin Y staining, and reduced Ki67 and cyclin D expression (representing 21% of the cultured CB population) include some mature erythroid CFCs but very few primitive CFCs, LTC-ICs, or repopulating cells. Although these findings suggest a cell cycle–associated change in in vivo stem cell homing, the cultured G0/G1 and S/G2/M CD34+ CB cells exhibited no differences in levels of expression of VLA-4, VLA-5, or CXCR-4. Moreover, further incubation of these cells for 1 day in the presence of a concentration of transforming growth factor β1 that increased the G0/G1 fraction did not enhance detection of repopulating cells. The demonstration of a cell cycle–associated mechanism that selectively silences the transplantability of proliferating human hematopoietic stem cells poses both challenges and opportunities for the future improvement of ex vivo–manipulated grafts.

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Chromosomal Instability and Telomere Shortening in Long-Term Culture of Hematopoietic Stem Cells: Insights from a Cell Culture Model of RPS14 Haploinsufficiency

Cytogenetic and Genome Research ◽

10.1159/000356096 ◽

2014 ◽

Vol 142 (1) ◽

pp. 14-20 ◽

Cited By ~ 4

Author(s):

K. Thomay ◽

A. Schienke ◽

B. Vajen ◽

U. Modlich ◽

A. Schambach ◽

...

Keyword(s):

Stem Cells ◽

Cell Culture ◽

Hematopoietic Stem Cells ◽

Chromosomal Instability ◽

Cell Culture Model ◽

Telomere Shortening ◽

Hematopoietic Stem ◽

Culture Model ◽

A Cell

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Keap1-Nrf2 system regulates cell fate determination of hematopoietic stem cells

Genes to Cells ◽

10.1111/gtc.12126 ◽

2014 ◽

Vol 19 (3) ◽

pp. 239-253 ◽

Cited By ~ 27

Author(s):

Shohei Murakami ◽

Ritsuko Shimizu ◽

Paul-Henri Romeo ◽

Masayuki Yamamoto ◽

Hozumi Motohashi

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Cell Fate ◽

Cell Fate Determination ◽

Hematopoietic Stem ◽

Fate Determination

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Bone Morphogenetic Proteins Regulate the Developmental Program of Human Hematopoietic Stem Cells

Journal of Experimental Medicine ◽

10.1084/jem.189.7.1139 ◽

1999 ◽

Vol 189 (7) ◽

pp. 1139-1148 ◽

Cited By ~ 270

Author(s):

Mickie Bhatia ◽

Dominique Bonnet ◽

Dongmei Wu ◽

Barbara Murdoch ◽

Jeff Wrana ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Bone Morphogenetic Proteins ◽

Transforming Growth Factor ◽

Critical Role ◽

Type I ◽

Hematopoietic Tissue ◽

Hematopoietic Stem ◽

Proliferation And Differentiation

The identification of molecules that regulate human hematopoietic stem cells has focused mainly on cytokines, of which very few are known to act directly on stem cells. Recent studies in lower organisms and the mouse have suggested that bone morphogenetic proteins (BMPs) may play a critical role in the specification of hematopoietic tissue from the mesodermal germ layer. Here we report that BMPs regulate the proliferation and differentiation of highly purified primitive human hematopoietic cells from adult and neonatal sources. Populations of rare CD34+CD38−Lin− stem cells were isolated from human hematopoietic tissue and were found to express the BMP type I receptors activin-like kinase (ALK)-3 and ALK-6, and their downstream transducers SMAD-1, -4, and -5. Treatment of isolated stem cell populations with soluble BMP-2, -4, and -7 induced dose-dependent changes in proliferation, clonogenicity, cell surface phenotype, and multilineage repopulation capacity after transplantation in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. Similar to transforming growth factor β, treatment of purified cells with BMP-2 or -7 at high concentrations inhibited proliferation yet maintained the primitive CD34+CD38− phenotype and repopulation capacity. In contrast, low concentrations of BMP-4 induced proliferation and differentiation of CD34+ CD38−Lin− cells, whereas at higher concentrations BMP-4 extended the length of time that repopulation capacity could be maintained in ex vivo culture, indicating a direct effect on stem cell survival. The discovery that BMPs are capable of regulating repopulating cells provides a new pathway for controlling human stem cell development and a powerful model system for studying the biological mechanism of BMP action using primary human cells.

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