RESUMEN
According to current models of hematopoiesis, lymphoid-primed multi-potent progenitors (LMPPs) (Lin(-)Sca-1(+)c-Kit(+)CD34(+)Flt3(hi)) and common myeloid progenitors (CMPs) (Lin(-)Sca-1(+)c-Kit(+)CD34(+)CD41(hi)) establish an early branch point for separate lineage-commitment pathways from hematopoietic stem cells, with the notable exception that both pathways are proposed to generate all myeloid innate immune cell types through the same myeloid-restricted pre-granulocyte-macrophage progenitor (pre-GM) (Lin(-)Sca-1(-)c-Kit(+)CD41(-)FcγRII/III(-)CD150(-)CD105(-)). By single-cell transcriptome profiling of pre-GMs, we identified distinct myeloid differentiation pathways: a pathway expressing the gene encoding the transcription factor GATA-1 generated mast cells, eosinophils, megakaryocytes and erythroid cells, and a pathway lacking expression of that gene generated monocytes, neutrophils and lymphocytes. These results identify an early hematopoietic-lineage bifurcation that separates the myeloid lineages before their segregation from other hematopoietic-lineage potential.
Asunto(s)
Diferenciación Celular , Linaje de la Célula , Linfocitos/fisiología , Células Mieloides/fisiología , Células Progenitoras Mieloides/fisiología , Animales , Antígenos CD/metabolismo , Células Cultivadas , Biología Computacional , Factor de Transcripción GATA1/genética , Factor de Transcripción GATA1/metabolismo , Hematopoyesis , Inmunidad Innata , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Análisis de Secuencia de ARN , Análisis de la Célula Individual , Análisis de Matrices Tisulares , Tirosina Quinasa 3 Similar a fms/genética , Tirosina Quinasa 3 Similar a fms/metabolismoRESUMEN
The stepwise commitment from hematopoietic stem cells in the bone marrow to T lymphocyte-restricted progenitors in the thymus represents a paradigm for understanding the requirement for distinct extrinsic cues during different stages of lineage restriction from multipotent to lineage-restricted progenitors. However, the commitment stage at which progenitors migrate from the bone marrow to the thymus remains unclear. Here we provide functional and molecular evidence at the single-cell level that the earliest progenitors in the neonatal thymus had combined granulocyte-monocyte, T lymphocyte and B lymphocyte lineage potential but not megakaryocyte-erythroid lineage potential. These potentials were identical to those of candidate thymus-seeding progenitors in the bone marrow, which were closely related at the molecular level. Our findings establish the distinct lineage-restriction stage at which the T cell lineage-commitment process transits from the bone marrow to the remote thymus.
Asunto(s)
Linfocitos B/citología , Linaje de la Célula/inmunología , Células Progenitoras Linfoides/citología , Células Mieloides/citología , Células Precursoras de Linfocitos B/citología , Linfocitos T/citología , Animales , Separación Celular , Citometría de Flujo , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/inmunología , Células Progenitoras Linfoides/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena en Tiempo Real de la Polimerasa , Timo/citologíaRESUMEN
Although an essential role for canonical Notch signaling in generation of hematopoietic stem cells in the embryo and in thymic T-cell development is well established, its role in adult bone marrow (BM) myelopoiesis remains unclear. Some studies, analyzing myeloid progenitors in adult mice with inhibited Notch signaling, implicated distinct roles of canonical Notch signaling in regulation of progenitors for the megakaryocyte, erythroid, and granulocyte-macrophage cell lineages. However, these studies might also have targeted other pathways. Therefore, we specifically deleted, in adult BM, the transcription factor recombination signal-binding protein J κ (Rbpj), through which canonical signaling from all Notch receptors converges. Notably, detailed progenitor staging established that canonical Notch signaling is fully dispensable for all investigated stages of megakaryocyte, erythroid, and myeloid progenitors in steady state unperturbed hematopoiesis, after competitive BM transplantation, and in stress-induced erythropoiesis. Moreover, expression of key regulators of these hematopoietic lineages and Notch target genes were unaffected by Rbpj deficiency in BM progenitor cells.
Asunto(s)
Médula Ósea/metabolismo , Eritropoyesis , Mielopoyesis , Receptores Notch/metabolismo , Transducción de Señal , Estrés Fisiológico , Animales , Proteína de Unión a la Señal Recombinante J de las Inmunoglobulinas/genética , Proteína de Unión a la Señal Recombinante J de las Inmunoglobulinas/metabolismo , Ratones , Ratones Transgénicos , Receptores Notch/genéticaRESUMEN
Despite their known transforming properties, the effects of leukemogenic FLT3-ITD mutations on hematopoietic stem and multipotent progenitor cells and on hematopoietic differentiation are not well understood. We report a mouse model harboring an ITD in the murine Flt3 locus that develops myeloproliferative disease resembling CMML and further identified FLT3-ITD mutations in a subset of human CMML. These findings correlated with an increase in number, cell cycling, and survival of multipotent stem and progenitor cells in an ITD dose-dependent manner in animals that exhibited alterations within their myeloid progenitor compartments and a block in normal B cell development. This model provides insights into the consequences of constitutive signaling by an oncogenic tyrosine kinase on hematopoietic progenitor quiescence, function, and cell fate.
Asunto(s)
Proliferación Celular , Células Madre Hematopoyéticas/metabolismo , Leucemia Mielomonocítica Crónica/metabolismo , Células Madre Multipotentes/metabolismo , Mutación , Trastornos Mieloproliferativos/metabolismo , Tirosina Quinasa 3 Similar a fms/metabolismo , Animales , Diferenciación Celular , Supervivencia Celular , Células Cultivadas , Exones , Regulación Neoplásica de la Expresión Génica , Genotipo , Células Madre Hematopoyéticas/patología , Humanos , Estimación de Kaplan-Meier , Leucemia Experimental/metabolismo , Leucemia Experimental/patología , Leucemia Mielomonocítica Crónica/genética , Leucemia Mielomonocítica Crónica/mortalidad , Leucemia Mielomonocítica Crónica/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Células Madre Multipotentes/patología , Trastornos Mieloproliferativos/genética , Trastornos Mieloproliferativos/patología , Fenotipo , Transducción de Señal , Tirosina Quinasa 3 Similar a fms/genéticaRESUMEN
MicroRNAs (miRs) are involved in many aspects of normal and malignant hematopoiesis, including hematopoietic stem cell (HSC) self-renewal, proliferation, and terminal differentiation. However, a role for miRs in the generation of the earliest stages of lineage committed progenitors from HSCs has not been identified. Using Dicer inactivation, we show that the miR complex is not only essential for HSC maintenance but is specifically required for their erythroid programming and subsequent generation of committed erythroid progenitors. In bipotent pre-MegEs, loss of Dicer up-regulated transcription factors preferentially expressed in megakaryocyte progenitors (Gata2 and Zfpm1) and decreased expression of the erythroid-specific Klf1 transcription factor. These results show a specific requirement for Dicer in acquisition of erythroid lineage programming and potential in HSCs and their subsequent erythroid lineage differentiation, and in particular indicate a role for the miR complex in achieving proper balance of lineage-specific transcriptional regulators necessary for HSC multilineage potential to be maintained.
Asunto(s)
Linaje de la Célula , ARN Helicasas DEAD-box/fisiología , Células Eritroides/citología , Células Eritroides/metabolismo , Células Madre Hematopoyéticas/citología , Células Progenitoras de Megacariocitos/citología , Ribonucleasa III/fisiología , Animales , Biomarcadores/metabolismo , Western Blotting , Diferenciación Celular , ARN Helicasas DEAD-box/antagonistas & inhibidores , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Células Madre Hematopoyéticas/metabolismo , Integrasas/metabolismo , Células Progenitoras de Megacariocitos/metabolismo , Ratones , Ratones Noqueados , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ribonucleasa III/antagonistas & inhibidores , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
There is much interest in the mechanisms that regulate adult tissue homeostasis and their relationship to processes governing foetal development. Mice deleted for the Wilms' tumour gene, Wt1, lack kidneys, gonads, and spleen and die at mid-gestation due to defective coronary vasculature. Wt1 is vital for maintaining the mesenchymal-epithelial balance in these tissues and is required for the epithelial-to-mesenchyme transition (EMT) that generates coronary vascular progenitors. Although Wt1 is only expressed in rare cell populations in adults including glomerular podocytes, 1% of bone marrow cells, and mesothelium, we hypothesised that this might be important for homeostasis of adult tissues; hence, we deleted the gene ubiquitously in young and adult mice. Within just a few days, the mice suffered glomerulosclerosis, atrophy of the exocrine pancreas and spleen, severe reduction in bone and fat, and failure of erythropoiesis. FACS and culture experiments showed that Wt1 has an intrinsic role in both haematopoietic and mesenchymal stem cell lineages and suggest that defects within these contribute to the phenotypes we observe. We propose that glomerulosclerosis arises in part through down regulation of nephrin, a known Wt1 target gene. Protein profiling in mutant serum showed that there was no systemic inflammatory or nutritional response in the mutant mice. However, there was a dramatic reduction in circulating IGF-1 levels, which is likely to contribute to the bone and fat phenotypes. The reduction of IGF-1 did not result from a decrease in circulating GH, and there is no apparent pathology of the pituitary and adrenal glands. These findings 1) suggest that Wt1 is a major regulator of the homeostasis of some adult tissues, through both local and systemic actions; 2) highlight the differences between foetal and adult tissue regulation; 3) point to the importance of adult mesenchyme in tissue turnover.
Asunto(s)
Glomerulonefritis/genética , Homeostasis/genética , Insuficiencia Multiorgánica/genética , Proteínas WT1/fisiología , Animales , Atrofia/genética , Atrofia/patología , Linaje de la Célula/genética , Transición Epitelial-Mesenquimal/genética , Femenino , Eliminación de Gen , Regulación de la Expresión Génica , Glomerulonefritis/patología , Gónadas/embriología , Gónadas/metabolismo , Gónadas/patología , Hematopoyesis/genética , Factor I del Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/metabolismo , Glomérulos Renales/embriología , Glomérulos Renales/metabolismo , Glomérulos Renales/patología , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Células Madre Mesenquimatosas/metabolismo , Ratones , Ratones Transgénicos , Insuficiencia Multiorgánica/patología , Páncreas Exocrino/embriología , Páncreas Exocrino/metabolismo , Páncreas Exocrino/patología , Podocitos/metabolismo , Podocitos/patología , Bazo/embriología , Bazo/metabolismo , Bazo/patología , Tamoxifeno/farmacología , Proteínas WT1/genéticaRESUMEN
BACKGROUND: The in vivo clinical significance of malignant stem cells remains unclear. METHODS: Patients who have the 5q deletion (del[5q]) myelodysplastic syndrome (interstitial deletions involving the long arm of chromosome 5) have complete clinical and cytogenetic remissions in response to lenalidomide treatment, but they often have relapse. To determine whether the persistence of rare but distinct malignant stem cells accounts for such relapses, we examined bone marrow specimens obtained from seven patients with the del(5q) myelodysplastic syndrome who became transfusion-independent while receiving lenalidomide treatment and entered cytogenetic remission. RESULTS: Virtually all CD34+, CD38+ progenitor cells and stem cells that were positive for CD34 and CD90, with undetectable or low CD38 (CD38−/low), had the 5q deletion before treatment. Although lenalidomide efficiently reduced these progenitors in patients in complete remission, a larger fraction of the minor, quiescent, CD34+,CD38-/low, CD90+ del(5q) stem cells as well as functionally defined del(5q) stem cells remained distinctly resistant to lenalidomide. Over time, lenalidomide resistance developed in most of the patients in partial and complete remission, with recurrence or expansion of the del(5q) clone and clinical and cytogenetic progression. CONCLUSIONS: In these patients with the del(5q) myelodysplastic syndrome, we identified rare and phenotypically distinct del(5q) myelodysplastic syndrome stem cells that were also selectively resistant to therapeutic targeting at the time of complete clinical and cytogenetic remission. (Funded by the EuroCancerStemCell Consortium and others.)
Asunto(s)
Antineoplásicos/farmacología , Deleción Cromosómica , Cromosomas Humanos Par 5/genética , Resistencia a Medicamentos/genética , Síndromes Mielodisplásicos/patología , Células Madre Neoplásicas/efectos de los fármacos , Talidomida/análogos & derivados , ADP-Ribosil Ciclasa 1/análisis , Anciano , Anciano de 80 o más Años , Antígenos CD34/análisis , Antineoplásicos/uso terapéutico , Femenino , Expresión Génica , Humanos , Lenalidomida , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/tratamiento farmacológico , Síndromes Mielodisplásicos/genética , Células Madre Neoplásicas/inmunología , Fenotipo , Inducción de Remisión , Talidomida/farmacología , Talidomida/uso terapéutico , Antígenos Thy-1/análisisRESUMEN
Hoxb4 overexpression promotes dramatic expansion of bone marrow (BM) hematopoietic stem cells (HSCs) without leukemic transformation and induces development of definitive HSCs from early embryonic yolk sac and differentiating embryonic stem cells. Knockout studies of Hoxb4 showed little effect on hematopoiesis, but interpretation of these results is obscured by the lack of direct evidence that Hoxb4 is expressed in HSCs and possible compensatory effects of other (Hox) genes. To evaluate accurately the pattern of Hoxb4 expression and to gain a better understanding of the physiologic role of Hoxb4 in the hemato-poietic system, we generated a knock-in Hoxb4-yellow fluorescent protein (YFP) reporter mouse model. We show that BM Lin(-)Sca1(+)c-Kit(+) cells express Hoxb4-YFP and demonstrate functionally in the long-term repopulation assay that definitive HSCs express Hoxb4. Similarly, aorta-gonad-mesonephrous-derived CD45(+)CD144(+) cells, enriched for HSCs, express Hoxb4. Furthermore, yolk sac and placental HSC populations express Hoxb4. Unexpectedly, Hoxb4 expression in the fetal liver HSCs is lower than in the BM, reaching negligible levels in some HSCs, suggesting an insignificant role of Hoxb4 in expansion of fetal liver HSCs. Hoxb4 expression therefore would not appear to correlate with the cycling status of fetal liver HSCs, although highly proliferative HSCs from young BM show strong Hoxb4 expression.
Asunto(s)
Proteínas Bacterianas/genética , Rastreo Celular/métodos , Genes Reporteros , Hematopoyesis/genética , Células Madre Hematopoyéticas/fisiología , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/fisiología , Proteínas Luminiscentes/genética , Factores de Transcripción/genética , Factores de Transcripción/fisiología , Animales , Proteínas Bacterianas/metabolismo , Embrión de Mamíferos , Femenino , Genes Reporteros/fisiología , Hematopoyesis/fisiología , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/metabolismo , Proteínas de Homeodominio/metabolismo , Proteínas Luminiscentes/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Modelos Animales , Modelos Biológicos , Embarazo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Factores de Transcripción/metabolismoRESUMEN
Lymphoid-primed multipotent progenitors with down-regulated megakaryocyte-erythroid (MkE) potential are restricted to cells with high levels of cell-surface FLT3 expression, whereas HSCs and MkE progenitors lack detectable cell-surface FLT3. These findings are compatible with FLT3 cell-surface expression not being detectable in the fully multipotent stem/progenitor cell compartment in mice. If so, this process could be distinct from human hematopoiesis, in which FLT3 already is expressed in multipotent stem/progenitor cells. The expression pattern of Flt3 (mRNA) and FLT3 (protein) in multipotent progenitors is of considerable relevance for mouse models in which prognostically important Flt3 mutations are expressed under control of the endogenous mouse Flt3 promoter. Herein, we demonstrate that mouse Flt3 expression initiates in fully multipotent progenitors because in addition to lymphoid and granulocyte-monocyte progenitors, FLT3(-) Mk- and E-restricted downstream progenitors are also highly labeled when Flt3-Cre fate mapping is applied.
Asunto(s)
Perfilación de la Expresión Génica , Células Madre Hematopoyéticas/metabolismo , Células Madre Multipotentes/metabolismo , Tirosina Quinasa 3 Similar a fms/genética , Animales , Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Trasplante de Médula Ósea , Linaje de la Célula/genética , Membrana Celular/metabolismo , Células Precursoras Eritroides/citología , Células Precursoras Eritroides/metabolismo , Citometría de Flujo , Células Precursoras de Granulocitos/citología , Células Precursoras de Granulocitos/metabolismo , Células Madre Hematopoyéticas/citología , Humanos , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Células Progenitoras de Megacariocitos/citología , Células Progenitoras de Megacariocitos/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Monocitos/citología , Monocitos/metabolismo , Células Madre Multipotentes/citología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tirosina Quinasa 3 Similar a fms/metabolismoRESUMEN
GATA3 has been identified as a master regulator of T helper cells, as well as being important for early thymic progenitors and T-cell commitment. However, Gata3 expression initiates already at the hematopoietic stem cell (HSC) level, implicating a potential role also in the regulation of HSCs. Herein we used a conditional Gata3 knockout strategy in which Gata3 expression was completely deleted from the earliest stage of embryonic hematopoietic development after emergence of HSCs from hemogenic endothelium. Through a detailed analysis of HSCs at the phenotypic and functional level, we demonstrate that steady-state levels of HSCs are normal in Gata3(fl/fl)Vav-Cre(tg/+) mice. Moreover, through long-term primary and secondary transplantation experiments, we also unequivocally demonstrate that Gata3 has a redundant role in post-transplantation HSC self-renewal.
Asunto(s)
Proliferación Celular , Factor de Transcripción GATA3/fisiología , Células Madre Hematopoyéticas/fisiología , Animales , Células Cultivadas , Factor de Transcripción GATA3/genética , Factor de Transcripción GATA3/metabolismo , Hematopoyesis/fisiología , Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transducción de Señal/genética , Transducción de Señal/fisiologíaRESUMEN
Acquisition of homozygous activating growth factor receptor mutations might accelerate cancer progression through a simple gene-dosage effect. Internal tandem duplications (ITDs) of FLT3 occur in approximately 25% cases of acute myeloid leukemia and induce ligand-independent constitutive signaling. Homozygous FLT3-ITDs confer an adverse prognosis and are frequently detected at relapse. Using a mouse knockin model of Flt3-internal tandem duplication (Flt3-ITD)-induced myeloproliferation, we herein demonstrate that the enhanced myeloid phenotype and expansion of granulocyte-monocyte and primitive Lin(-)Sca1(+)c-Kit(+) progenitors in Flt3-ITD homozygous mice can in part be mediated through the loss of the second wild-type allele. Further, whereas autocrine FLT3 ligand production has been implicated in FLT3-ITD myeloid malignancies and resistance to FLT3 inhibitors, we demonstrate here that the mouse Flt3(ITD/ITD) myeloid phenotype is FLT3 ligand-independent.
Asunto(s)
Dosificación de Gen/fisiología , Duplicación de Gen/fisiología , Pérdida de Heterocigocidad/fisiología , Proteínas de la Membrana/genética , Trastornos Mieloproliferativos/genética , Tirosina Quinasa 3 Similar a fms/fisiología , Alelos , Animales , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/patología , Células de la Médula Ósea/fisiología , Proliferación Celular , Células Cultivadas , Técnicas de Sustitución del Gen , Pérdida de Heterocigocidad/genética , Masculino , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Fenotipo , Secuencias Repetidas en Tándem/genética , Secuencias Repetidas en Tándem/fisiología , Tirosina Quinasa 3 Similar a fms/metabolismoRESUMEN
Mice deficient in c-fms-like tyrosine kinase 3 (FLT3) signaling have reductions in early multipotent and lymphoid progenitors, whereas no evident myeloid phenotype has been reported. However, activating mutations of Flt3 are among the most common genetic events in acute myeloid leukemia and mice harboring internal tandem duplications within Flt3 (Flt3-ITD) develop myeloproliferative disease, with characteristic expansion of granulocyte-monocyte (GM) progenitors (GMP), possibly compatible with FLT3-ITD promoting a myeloid fate of multipotent progenitors. Alternatively, FLT3 might be expressed at the earliest stages of GM development. Herein, we investigated the expression, function, and role of FLT3 in recently identified early GMPs. Flt3-cre fate-mapping established that most progenitors and mature progeny of the GM lineage are derived from Flt3-expressing progenitors. A higher expression of FLT3 was found in preGMP compared with GMP, and preGMPs were more responsive to stimulation with FLT3 ligand (FL). Whereas preGMPs and GMPs were reduced in Fl(-/-) mice, megakaryocyte-erythroid progenitors were unaffected and lacked FLT3 expression. Notably, mice deficient in both thrombopoietin (THPO) and FL had a more pronounced GMP phenotype than Thpo(-/-) mice, establishing a role of FL in THPO-dependent and -independent regulation of GMPs, of likely significance for myeloid malignancies with Flt3-ITD mutations.
Asunto(s)
Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/fisiología , Leucemia Mieloide Aguda/fisiopatología , Tirosina Quinasa 3 Similar a fms/genética , Tirosina Quinasa 3 Similar a fms/metabolismo , Animales , Linaje de la Célula/fisiología , Expresión Génica/fisiología , Granulocitos/citología , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patología , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/citología , Células Mieloides/citología , Transducción de Señal/fisiología , Trombopoyetina/genéticaRESUMEN
Originally cloned from hematopoietic stem cell (HSC) populations and its ligand being extensively used to promote ex vivo HSC expansion, the FMS-like tyrosine kinase 3 (FLT3; also called FLK2) receptor and its ligand (FL) were expected to emerge as an important physiologic regulator of HSC maintenance and expansion. However, the role of FLT3 receptor and ligand in HSC regulation remains unclear and disputed. Herein, using Fl-deficient mice, we establish for the first time that HSC expansion in fetal liver and after transplantation is FL independent. Because previous findings in Flk2(-/-) mice were compatible with an important role of FLT3 receptor in HSC regulation and because alternative ligands might potentially interact directly or indirectly with FLT3 receptor, we here also characterized HSCs in Flk2(-/-) mice. Advanced phenotypic as well as functional evaluation of Flk2(-/-) HSCs showed that the FLT3 receptor is dispensable for HSC steady-state maintenance and expansion after transplantation. Taken together, these studies show that the FLT3 receptor and ligand are not critical regulators of mouse HSCs, neither in steady state nor during fetal or posttransplantation expansion.
Asunto(s)
Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/fisiología , Proteínas de la Membrana/metabolismo , Tirosina Quinasa 3 Similar a fms/metabolismo , Animales , División Celular/fisiología , Citometría de Flujo , Supervivencia de Injerto/fisiología , Hematopoyesis/fisiología , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Fenotipo , Tirosina Quinasa 3 Similar a fms/genéticaRESUMEN
Extensive studies of mice deficient in one or several cytokine receptors have failed to support an indispensable role of cytokines in development of multiple blood cell lineages. Whereas B1 B cells and Igs are sustained at normal levels throughout life of mice deficient in IL-7, IL-7Ralpha, common cytokine receptor gamma chain, or flt3 ligand (FL), we report here that adult mice double deficient in IL-7Ralpha and FL completely lack visible LNs, conventional IgM+ B cells, IgA+ plasma cells, and B1 cells, and consequently produce no Igs. All stages of committed B cell progenitors are undetectable in FL-/- x IL-7Ralpha-/- BM that also lacks expression of the B cell commitment factor Pax5 and its direct target genes. Furthermore, in contrast to IL-7Ralpha-/- mice, FL-/- x IL-7Ralpha-/- mice also lack mature B cells and detectable committed B cell progenitors during fetal development. Thus, signaling through the cytokine tyrosine kinase receptor flt3 and IL-7Ralpha are indispensable for fetal and adult B cell development.
Asunto(s)
Linfocitos B/fisiología , Diferenciación Celular/fisiología , Proteínas de la Membrana/metabolismo , Receptores de Interleucina-7/metabolismo , Animales , Linfocitos B/citología , Proteínas de la Membrana/genética , Proteínas de la Membrana/inmunología , Ratones , Ratones Noqueados , Ganglios Linfáticos Agregados/metabolismo , Receptores de Interleucina-7/genética , Receptores de Interleucina-7/inmunología , Transducción de Señal/inmunología , Transducción de Señal/fisiologíaRESUMEN
Studies from our and other laboratories have over the last 2 years implicated the existence of multipotent progenitors (MPPs) with combined granulocyte-macrophage, B cell, and T cell potential, but little or no megakaryocyte-erythroid (MkE) potential in the adult bone marrow Lineage(-)SCA-1(+)KIT(+) (LSK) compartment of multipotent stem and progenitor cells. The evidence for the existence of LSKCD34(+)FLT3(hi) lymphoid-primed MPPs (LMPPs) implicates that a strict separation into common myeloid and lymphoid pathways might not be the first lineage commitment step of hematopoietic stem cells (HSCs). Together with the evidence for existence of common myeloid and common lymphoid progenitors (CMPs and CLPs, respectively), the identification of LMPPs also suggests that at least the granulocyte-macrophage lineage can be generated through alternative pathways. However, the existence of LMPPs has recently been questioned, as there is evidence that at least a fraction of LSKCD34(+)FLT3(hi) cells sustains MkE potential. Thus, in more recent studies we have in more detail compared the molecular signature of adult LMPPs to populations of LSK cells enriched for cells with pluripotent HSC activity. Notably, we have found at the global as well as single-cell level that LMPPs when compared with pluripotent HSCs downregulate the transcriptional priming of genes typically expressed in cells of the MkE lineage, while upregulating early lymphoid genes. Although other studies have suggested that the earliest HSC commitment steps might differ in fetal and adult hematopoiesis, we have also obtained evidence suggesting that the LMPP is defined already during fetal development.
Asunto(s)
Linfocitos/citología , Células Madre/citología , Animales , Antígenos CD34/biosíntesis , Linfocitos B/citología , Linaje de la Célula , Granulocitos/metabolismo , Hematopoyesis , Células Madre Hematopoyéticas/citología , Humanos , Macrófagos/metabolismo , Proteínas de la Membrana/biosíntesis , Ratones , Modelos Biológicos , Linfocitos T/citologíaRESUMEN
Hematopoietic stem cells (HSC)(1) supply organisms with life-long output of mature blood cells. To do so, the HSC pool size has to be maintained by HSC self-renewing divisions. PRDM3 and PRDM16 have been documented to regulate HSC self-renewal, maintenance and function. We found Prdm11 to have similar expression patterns in the hematopoietic stem and progenitor cell (HSPC) compartments as Prdm3 and Prdm16. Therefore, we undertook experiments to test if PRDM11 regulates HSC self-renewal, maintenance and function by investigating the Prdm11(-/-) mice. Our data shows that phenotypic HSPCs are intact in bone marrow (BM) of one-year-old Prdm11(-/-) mice. In addition, Prdm11(-/-) mice were able to fully regenerate the hematopoietic system upon BM transplantation (BMT) into lethally irradiated mice with a mild drop in lymphoid output only. Taken together, this suggests that PRDM11, in contrast to PRDM3 and PRDM16, is not directly involved in regulation of HSPCs in mice.
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Proteínas Portadoras/metabolismo , Células Madre Hematopoyéticas/citología , Proteínas Represoras/metabolismo , Células Madre/citología , Animales , Biomarcadores/metabolismo , Células de la Médula Ósea/citología , Trasplante de Médula Ósea , Proteínas Portadoras/genética , Células Madre Hematopoyéticas/metabolismo , Megacariocitos/citología , Ratones , Ratones Noqueados , Recuento de Plaquetas , Proteínas Represoras/genética , Células Madre/metabolismo , Factores de Transcripción , Irradiación Corporal TotalRESUMEN
Whether signals mediated via growth factor receptors (GFRs) might influence lineage fate in multipotent progenitors (MPPs) is unclear. We explored this issue in a mouse knockin model of gain-of-function Flt3-ITD mutation because FLT3-ITDs are paradoxically restricted to acute myeloid leukemia even though Flt3 primarily promotes lymphoid development during normal hematopoiesis. When expressed in MPPs, Flt3-ITD collaborated with Runx1 mutation to induce high-penetrance aggressive leukemias that were exclusively of the myeloid phenotype. Flt3-ITDs preferentially expanded MPPs with reduced lymphoid and increased myeloid transcriptional priming while compromising early B and T lymphopoiesis. Flt3-ITD-induced myeloid lineage bias involved upregulation of the transcription factor Pu.1, which is a direct target gene of Stat3, an aberrantly activated target of Flt3-ITDs, further establishing how lineage bias can be inflicted on MPPs through aberrant GFR signaling. Collectively, these findings provide new insights into how oncogenic mutations might subvert the normal process of lineage commitment and dictate the phenotype of resulting malignancies.
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Leucemia Mieloide Aguda/patología , Células Madre Multipotentes/citología , Células Mieloides/citología , Tirosina Quinasa 3 Similar a fms/fisiología , Animales , Diferenciación Celular/fisiología , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Transformación Celular Neoplásica/patología , Modelos Animales de Enfermedad , Citometría de Flujo/métodos , Expresión Génica , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Ratones , Análisis por Micromatrices , Células Madre Multipotentes/inmunología , Células Madre Multipotentes/metabolismo , Células Madre Multipotentes/patología , Células Mieloides/inmunología , Células Mieloides/metabolismo , Células Mieloides/patología , Transducción de Señal , Tirosina Quinasa 3 Similar a fms/genética , Tirosina Quinasa 3 Similar a fms/metabolismoRESUMEN
The prevailing model for adult hematopoiesis postulates that the first lineage commitment step results in a strict separation of common myeloid and common lymphoid pathways. However, the recent identification of granulocyte/monocyte (GM)-lymphoid restricted lymphoid-primed multipotent progenitors (LMPPs) and primitive common myeloid progenitors (CMPs) within the "HSC" compartment provide compelling support for establishment of independent GM-megakaryocyte/erythroid (GM-MkE) and GM-lymphoid commitment pathways as decisive early lineage fate decisions. These changes in lineage potentials are corroborated by corresponding changes in multilineage transcriptional priming, as LMPPs down-regulate MkE priming but become GM-lymphoid transcriptionally primed, whereas CMPs are GM-MkE primed. These distinct biological and molecular relationships are established already in the fetal liver.
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Diferenciación Celular/fisiología , Linaje de la Célula/fisiología , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/fisiología , Modelos Biológicos , Células Madre Adultas/clasificación , Animales , Células Madre Fetales/clasificación , Células Madre Hematopoyéticas/clasificación , Humanos , Transducción de SeñalRESUMEN
Evidence for a novel route of adult hematopoietic stem-cell lineage commitment through Lin-Sca-1+Kit+Flt3hi (LSKFlt3hi) lymphoid-primed multipotent progenitors (LMPPs) with granulocyte/monocyte (GM) and lymphoid but little or no megakaryocyte/erythroid (MkE) potential was recently challenged, as LSKFlt3hi cells were reported to possess MkE potential. Herein, residual (1%-2%) MkE potential segregated almost entirely with LSKFlt3hi cells expressing the thrombopoietin receptor (Mpl), whereas LSKFlt3hiMpl- LMPPs lacked significant MkE potential in vitro and in vivo, but sustained combined GM and lymphoid potentials, and coexpressed GM and lymphoid but not MkE transcriptional lineage programs. Gradually increased transcriptional lymphoid priming in single LMPPs from Rag1GFP mice was shown to occur in the presence of maintained GM lineage priming, but gradually reduced GM lineage potential. These functional and molecular findings reinforce the existence of GM/lymphoid-restricted progenitors with dramatically down-regulated probability for committing toward MkE fates, and support that lineage restriction occurs through gradual rather than abrupt changes in specific lineage potentials.
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Linfocitos/metabolismo , Células Madre Multipotentes/metabolismo , Células Mieloides/metabolismo , Receptores de Trombopoyetina/metabolismo , Animales , Antígenos Ly/genética , Antígenos Ly/metabolismo , Regulación hacia Abajo/genética , Proteínas de Homeodominio/metabolismo , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Transgénicos , Proteínas Proto-Oncogénicas c-kit/genética , Proteínas Proto-Oncogénicas c-kit/metabolismo , Receptores de Trombopoyetina/genética , Tirosina Quinasa 3 Similar a fms/genética , Tirosina Quinasa 3 Similar a fms/metabolismoRESUMEN
PURPOSE OF REVIEW: This review addresses recently reported evidence for alternative cellular pathways for haematopoietic stem cell lineage commitment. RECENT FINDINGS: Using various approaches, several laboratories suggested the existence of adult as well as foetal multipotent progenitor cells with combined B cell, T cell and granulocyte/macrophage potential, but little or no megakaryocyte/erythroid potential. Compared with haematopoietic stem cells, these multipotent progenitor cells exhibited downregulated transcriptional expression of genes of the megakaryocyte/erythroid lineages and upregulated expression of lymphoid lineage genes. The existence of these lineage-restricted multipotent progenitor cells suggests that the first lineage commitment step of haematopoietic stem cells does not result in strict separation into myelopoiesis and lymphopoiesis, and that there might be alternative pathways for commitment toward different lineage fates. These findings have been questioned by other studies, however. To resolve this controversy and establish the complete road map for haematopoietic lineage commitment, improved tools and more stringent standards for how to identify and characterize lineage fate options of distinct stem and progenitor cells are needed. SUMMARY: Current and future progress in establishing the complete cellular roadmap for haematopoietic lineage commitment will permit identification and characterization of key regulators of lineage fate decisions in haematopoietic stem cells.