Down syndrome (DS), with trisomy of chromosome 21 (HSA21), is the commonest human aneuploidy. Pre... more Down syndrome (DS), with trisomy of chromosome 21 (HSA21), is the commonest human aneuploidy. Pre-leukemic myeloproliferative changes in DS foetal livers precede the acquisition of GATA1 mutations, transient myeloproliferative disorder (DS-TMD) and acute megakaryocytic leukemia (DS-AMKL). Trisomy of the Erg gene is required for myeloproliferation in the Ts(17 16 )65Dn DS mouse model. We demonstrate here that genetic changes specifically attributable to trisomy of Erg lead to lineage priming of primitive and early multipotential progenitor cells in Ts(17 16 )65Dn mice, excess megakaryocyte-erythroid progenitors, and malignant myeloproliferation. Gene expression changes dependent on trisomy of Erg in Ts (17 16 )65Dn multilineage progenitor cells were correlated with those associated with trisomy of HSA21 in human DS hematopoietic stem and primitive progenitor cells. These data suggest a role for ERG as a regulator of hematopoietic lineage potential, and that trisomy of ERG in the context of DS foetal liver hemopoiesis drives the pre-leukemic changes that predispose to subsequent DS-TMD and DS-AMKL.
The formation of megakaryocyte colonies in agar cultures of murine bone marrow or spleen cells ca... more The formation of megakaryocyte colonies in agar cultures of murine bone marrow or spleen cells can be stimulated by the addition of interleukin-3 (IL-3), erythropoietin (EPO), thrombopoietin (TPO), or IL-6. However, greater numbers of colonies developed if combinations of two or more of these stimuli were used, particularly combinations including stem cell factor, with maximal numbers of colonies developing with the combination of stem cell factor plus IL-3 plus EPO. The data indicate that most committed progenitor cells in the megakaryocyte lineage were unusual in that they required stimulation by two or more hematopoietic growth factors. In tests using a range of growth factors, G-CSF was exceptional in that it consistently specifically inhibited megakaryocyte colony formation stimulated by EPO, TPO, or IL-6 but not that stimulated by IL-3. The mechanisms involved in this inhibitory action of G-CSF are unknown, but the inhibitory action could be of relevance for the dose-dependent lowering of platelet levels observed in some subjects injected with G-CSF.
There are only three known stimuli for eosinophil formation-GM-CSF, interleukin-5 (IL-5), and IL-... more There are only three known stimuli for eosinophil formation-GM-CSF, interleukin-5 (IL-5), and IL-3. Because mice with inactivation of the gene encoding the common β receptor chain for GM-CSF, IL-5, and IL-3 (βc -/mice) cannot respond to GM-CSF or IL-5 and do not produce IL-3, they should lack eosinophils. However, they produce reduced numbers of eosinophils, indicating the existence of at least one additional stimulatory factor. Use of βc -/mouse marrow cells failed to detect a novel eosinophil-stimulating factor in cell line-or organ-conditioned media, but stem cell factor (SCF) was found to have a previously unrecognized ability to stimulate the formation of eosinophil colonies or mixed neutrophil-eosinophil colonies. This action of SCF was also observable with marrow cells from other mouse strains and was enhanced by the addition of G-CSF but no other factor tested. Recloning of SCF-stimulated blast colonies showed that progenitors forming pure eosinophil or mixed neutrophil-eosinophil colonies can have a common ancestor but many appear to arise independently from different preprogenitor cells. The observed activity of SCF in marrow cultures was relatively weak, but its action may be stronger in vivo, and SCF needs to be added to GM-CSF, IL-5, and IL-3 in the list of cytokines able to stimulate eosinophil formation.
, SOCS-1, SOCS-2, SOCS-3, SOCS-5, SOCS-6, or SOCS-7 indicated that the inhibitory SOCS proteins, ... more , SOCS-1, SOCS-2, SOCS-3, SOCS-5, SOCS-6, or SOCS-7 indicated that the inhibitory SOCS proteins, with the possible exception of SOCS-3, were not involved in the G-CSF-initiated suppression of megakaryocyte colony formation. Stem Cells 2005;23:55-62
Proceedings of the National Academy of Sciences, 2013
Previous studies have shown that mouse bone marrow cells can produce mast cells when stimulated i... more Previous studies have shown that mouse bone marrow cells can produce mast cells when stimulated in vitro by stem cell factor (SCF) and interleukin-3 (IL-3). Experiments to define the marrow cells able to generate mast cells showed that the most active subpopulations were the Kit + Sca1progenitor cell fraction and the more ancestral Kit + Sca1 + blast colony-forming cell fraction. In clonal cultures, up to 64% of blast colony-forming cells were able to generate mast cells when stimulated by SCF and IL-3, and, of these, the most active were those in the CD34 -Flt3Rlong-term repopulating cell fraction. Basophils, identified by the monoclonal antibody mMCP-8 to mouse mast cell serine protease-8, were also produced by 50% of blast colony-forming cells with a strong concordance in the production of both cell types by individual blast colony-forming cells. Enriched populations of marrow-derived basophils were shown to generate variable numbers of mast cells after a further incubation with SCF and IL-3. The data extend the repertoire of lineage-committed cells able to be produced by multipotential hematopoietic blast colony-forming cells and show that basophils and mast cells can have common ancestral cells and that basophils can probably generate mast cells at least under defined in vitro conditions.
Proceedings of the National Academy of Sciences, 2009
units, spleen (CFU-S) supporting the possibility that the 2 cells may be identical. The colony po... more units, spleen (CFU-S) supporting the possibility that the 2 cells may be identical. The colony populations generated by BL-CFC-S were similar in their phenotype and proliferative capacity to progenitor cells in whole bone marrow but the progeny of BL-CFC-F were skewed with an abnormally high proportion of Kit ؊ Flt3R ؉ cells whose clonogenic cells tended to generate only macrophage progeny. Both blast colony populations had a high percentage of GR1 ؉ and Mac1 ؉ cells but BL-CFC-F colonies also contained a significant population of B220 ؉ and IL-7R ؉ cells relevant to the superior ability of BL-CFC-F colony cells to generate B lymphocytes and the known dependency of this process on Flt3 ligand and IL-7. The commitment events and phenotypic changes during the generation of differing progenitor cells in blast colonies can now be clonally analyzed in a convenient in vitro culture system. blast colonies ͉ lineage commitment ͉ hematopoietic stem cells
Murine Ba/F3 cells were transfected with cDNA for the ␣-chain of the murine interleukin-5 (IL-5) ... more Murine Ba/F3 cells were transfected with cDNA for the ␣-chain of the murine interleukin-5 (IL-5) receptor and cloned lines of these cells were able to proliferate in response to as little as 2.5 pg/ml of IL-5. The bioassay was demonstrated to be specific for IL-5 and was able to measure IL-5 produced in culture by organs from adult C57BL/6 and BALB/c mice. The highest levels of IL-5 were produced by lung tissue but thymus and bladder consistently produced IL-5 and more variable production was observed by the heart, spleen, muscle, bone shaft, uterus and testes. Bone marrow cells produced no detectable IL-5. Observed levels of production of IL-5 were similar when using organs from mice lacking high-affinity receptors for IL-5 and from nu/nu, RAG-1−/− and NOD/SCID mice lacking T lymphocytes. In inflammatory peritoneal exudates induced by the injection of casein plus bacteria, levels of induced IL-5 were higher if the mice lacked high-affinity receptors for IL-5. The data indicate that T lymphocytes are not the dominant cellular source of IL-5 in organ-conditioned media and that local IL-5 production can occur with a wide range of normal murine organs. Leukemia (2001) 15, 1248-1255.
Polycomb Repressive Complex 2 (PRC2) is a chromatin modifier that regulates stem cells in embryon... more Polycomb Repressive Complex 2 (PRC2) is a chromatin modifier that regulates stem cells in embryonic and adult tissues. Loss of function studies of PRC2 components has been complicated by early embryonic dependence on PRC2 activity and the partial functional redundancy of Ezh1 and Ezh2, which encode the enzymatic component of PRC2. Here, we investigated the role of PRC2 in hematopoiesis by conditional deletion of Suz12, a core component of PRC2. Complete loss of Suz12 resulted in failure of hematopoiesis, both in the embryo and the adult, with a loss of maintenance of hematopoietic stem cells (HSCs). In contrast, partial loss of PRC2 enhanced HSC self-renewal. Deletion in individual blood cell lineages revealed that, while Suz12 was required for lymphoid development, it was dispensable for the development of granulocytic, monocytic and megakaryocytic cells. Collectively, these data reveal the multi-faceted role of PRC2 in hematopoiesis, with divergent dose-dependent effects in HSC an...
Down syndrome (DS), with trisomy of chromosome 21 (HSA21), is the commonest human aneuploidy. Pre... more Down syndrome (DS), with trisomy of chromosome 21 (HSA21), is the commonest human aneuploidy. Pre-leukemic myeloproliferative changes in DS foetal livers precede the acquisition of GATA1 mutations, transient myeloproliferative disorder (DS-TMD) and acute megakaryocytic leukemia (DS-AMKL). Trisomy of the Erg gene is required for myeloproliferation in the Ts(17 16 )65Dn DS mouse model. We demonstrate here that genetic changes specifically attributable to trisomy of Erg lead to lineage priming of primitive and early multipotential progenitor cells in Ts(17 16 )65Dn mice, excess megakaryocyte-erythroid progenitors, and malignant myeloproliferation. Gene expression changes dependent on trisomy of Erg in Ts (17 16 )65Dn multilineage progenitor cells were correlated with those associated with trisomy of HSA21 in human DS hematopoietic stem and primitive progenitor cells. These data suggest a role for ERG as a regulator of hematopoietic lineage potential, and that trisomy of ERG in the context of DS foetal liver hemopoiesis drives the pre-leukemic changes that predispose to subsequent DS-TMD and DS-AMKL.
Marrow cells from mice lacking high-affinity receptors for granulocyte-macrophage colony-stimulat... more Marrow cells from mice lacking high-affinity receptors for granulocyte-macrophage colony-stimulating factor (GM-CSF; betac-/- mice) were shown to bind and internalize much less GM-CSF than cells from normal (betac+/+) mice. betac-/- mice were used to determine the effect of negligible receptor-mediated clearance on detectible GM-CSF responses to the intravenous injection of endotoxin or the intraperitoneal injection of casein plus microorganisms. Unlike the minor serum GM-CSF responses to endotoxin seen in betac+/+ mice, serum GM-CSF levels rose 30-fold to 9 ng/mL in betac-/- mice even though loss of GM-CSF in the urine was greater than in betac+/+ mice. Organs from betac-/- and betac+/+ mice had a similar capacity to produce GM-CSF in vitro, as did peritoneal cells from both types of mice when challenged in vitro by casein. However, when casein was injected intraperitoneally, betac-/- mice developed higher and more sustained levels of GM-CSF than did betac+/+ mice. The data indicat...
Upon ligand binding, RIPK1 is recruited to tumor necrosis factor receptor superfamily (TNFRSF) an... more Upon ligand binding, RIPK1 is recruited to tumor necrosis factor receptor superfamily (TNFRSF) and Toll-like receptor (TLR) complexes promoting prosurvival and inflammatory signaling. RIPK1 also directly regulates caspase-8-mediated apoptosis or, if caspase-8 activity is blocked, RIPK3-MLKL-dependent necroptosis. We show that C57BL/6 Ripk1(-/-) mice die at birth of systemic inflammation that was not transferable by the hematopoietic compartment. However, Ripk1(-/-) progenitors failed to engraft lethally irradiated hosts properly. Blocking TNF reversed this defect in emergency hematopoiesis but, surprisingly, Tnfr1 deficiency did not prevent inflammation in Ripk1(-/-) neonates. Deletion of Ripk3 or Mlkl, but not Casp8, prevented extracellular release of the necroptotic DAMP, IL-33, and reduced Myd88-dependent inflammation. Reduced inflammation in the Ripk1(-/-)Ripk3(-/-), Ripk1(-/-)Mlkl(-/-), and Ripk1(-/-)Myd88(-/-) mice prevented neonatal lethality, but only Ripk1(-/-)Ripk3(-/-)Cas...
Proceedings of the National Academy of Sciences, 2003
Injection of neonatal bone marrow cells from mice lacking the gene encoding suppressor of cytokin... more Injection of neonatal bone marrow cells from mice lacking the gene encoding suppressor of cytokine signaling 1 (SOCS1) into irradiated syngeneic 129͞Sv or C57BL͞6 mice led to a decreased survival, more rapidly occurring in 129͞Sv than in C57BL͞6 mice. Moribund mice did not exhibit the acute or chronic diseases developed by Socs1 ؊/؊ mice but developed a pathology characteristic of graft-versus-host disease with typical chronic inflammatory lesions in the liver, skin, lungs, and gut. The results indicate that cells derived from the Socs1 ؊/؊ bone marrow are autoaggressive but did not identify the cell types involved. Failure of the engrafted Socs1 ؊/؊ marrow cells to reproduce the tissue damage typical of Socs1 ؊/؊ disease indicates that loss of SOCS1 from target tissues may also be required for the development of the Socs1 ؊/؊ diseases, such as fatty degeneration of the liver, polymyositis, or corneal inflammation.
Proceedings of the National Academy of Sciences, 2002
Mice with inactivation of the gene encoding the suppressor of cytokine signaling-1 (SOCS-1) die i... more Mice with inactivation of the gene encoding the suppressor of cytokine signaling-1 (SOCS-1) die in neonatal life with an IFN-␥dependent inflammatory disease dominated by fatty degeneration and necrosis of the liver. To establish the long-term pathological consequences of loss of SOCS-1 in mice, where initial survival was made possible by also deleting the IFN-␥ gene, a comparison was made of the lifespan of groups of SOCS-1 ؊/؊ IFN-␥ ؊/؊ , SOCS-1 ؉/؉ IFN-␥ ؊/؊ and SOCS-1 ؉/؉ IFN-␥ ؉/؉ mice. Mice lacking the genes for both SOCS-1 and IFN-␥ exhibited an accelerated death rate compared with control groups. Disease states developing selectively in SOCS-1 ؊/؊ IFN-␥ ؊/؊ mice were polycystic kidneys, pneumonia, chronic skin ulcers, and chronic granulomas in the gut and various other organs. Mice of all three groups developed cataracts, but disease development was accelerated in the groups lacking IFN-␥. SOCS-1 ؊/؊ IFN-␥ ؊/؊ mice exhibited a slightly increased predisposition to the development of T lymphoid leukemia, either spontaneous or radiation-induced. The development of polycystic kidneys may be caused by a developmental defect in renal-tubule organization noted in neonatal SOCS-1 ؊/؊ mice. The chronic infections and granulomas of SOCS-1 ؊/؊ IFN-␥ ؊/؊ mice may be based on autoaggression of SOCS-1 ؊/؊ T lymphoid and related cells or a functional deficiency of these cells when lacking SOCS-1.
Proceedings of the National Academy of Sciences, 2006
Genetically primed adult C57BL mice were deleted of exon 5 of the gene encoding the transcription... more Genetically primed adult C57BL mice were deleted of exon 5 of the gene encoding the transcription factor PU.1 by IFN activation of Cre recombinase. After a 13-week delay, conditionally deleted (PU.1 ؊/؊ ) mice began dying of myeloid leukemia, and 95% of the mice surviving from early postinduction death developed transplantable myeloid leukemia whose cells were deleted of PU.1 and uniformly Gr-1 positive. The leukemic cells formed autonomous colonies in semisolid culture with varying clonal efficiency, but colony formation was enhanced by IL-3 and sometimes by granulocyte-macrophage colony-stimulating factor. Nine of 13 tumors analyzed had developed a capacity for autocrine IL-3 or granulocyte-macrophage colony-stimulating factor production, and there was evidence of rearrangement of the IL-3 gene. Acquisition of autocrine growth-factor production and autonomous growth appeared to be major events in the transformation of conditionally deleted PU.1 ؊/؊ cells to fully developed myeloid leukemic populations.
Proceedings of the National Academy of Sciences, 2007
Murine granulocytic cells, in becoming leukemic, need to acquire enhanced self-generation and a c... more Murine granulocytic cells, in becoming leukemic, need to acquire enhanced self-generation and a capacity for autocrine growth stimulation. Mice transplanted with bone marrow cells transduced with the Mixl1 homeobox gene develop a very high frequency of myeloid leukemia derived from the transduced cells. Preleukemic mice contained a high frequency of transduced clonogenic granulocytic cells. They exhibited an abnormally high capacity for self-replication and could generate immortalized granulocytic cell lines that remained absolutely dependent on either GM-CSF or IL-3 and were not leukemic. Organs from mice repopulated by marrow cells transduced either with Mixl1 or the control murine stem cell virus vector exhibited a capacity to produce IL-3 in vitro, activity being highest with the lungs, marrow, bladder, and thymus.
Proceedings of the National Academy of Sciences, 2007
In agar cultures of marrow cells from adult female BALB/c chimeric GATA-1 Plt13/؉ mice, a high fr... more In agar cultures of marrow cells from adult female BALB/c chimeric GATA-1 Plt13/؉ mice, a high frequency of unusual dispersed colonies was noted. Analysis showed that these were colonies of mast cells and that mast cell colony-forming cells (progenitors) could be detected in clonal cultures of adult marrow, neonatal marrow, or fetal liver if the combined stimulus of stem cell factor and interleukin-3 was used. Mast cell progenitors were in active cell cycle and showed an extensive capacity for self-generation. Mast cell colonies both from control GATA-1 ؉/؉ mice and GATA-1 Plt13/؉ mice could generate growth factor-dependent cloned cell lines that grew for >18 months. Surprisingly, the majority of the excessive numbers of mast cell progenitors in chimeric GATA-1 Plt13/؉ mice were transcribing the inactive Plt13 allele of GATA-1, suggesting that GATA-1 normally acts to restrict the emergence of committed mast cell progenitors. In sharp contrast, all eosinophil progenitors in these mice were transcribing the normal GATA-1 allele. No excess tissue mast cells were observed in GATA-1 Plt13/؉ mice, suggesting that the excess mast cell progenitors in these mice might be generating mast cells with a defective in vivo proliferative or tissue homing capacity. mast cell colonies ͉ mast cell lines ͉ eosinophil progenitors Author contributions: D.M., I.M., and W.S.A. designed research; D.M., I.M., S.M., and L.D.R. performed research; D.M., I.M., S.M., L.D.R., and W.S.A. analyzed data; and D.M. wrote the paper.
Down syndrome (DS), with trisomy of chromosome 21 (HSA21), is the commonest human aneuploidy. Pre... more Down syndrome (DS), with trisomy of chromosome 21 (HSA21), is the commonest human aneuploidy. Pre-leukemic myeloproliferative changes in DS foetal livers precede the acquisition of GATA1 mutations, transient myeloproliferative disorder (DS-TMD) and acute megakaryocytic leukemia (DS-AMKL). Trisomy of the Erg gene is required for myeloproliferation in the Ts(17 16 )65Dn DS mouse model. We demonstrate here that genetic changes specifically attributable to trisomy of Erg lead to lineage priming of primitive and early multipotential progenitor cells in Ts(17 16 )65Dn mice, excess megakaryocyte-erythroid progenitors, and malignant myeloproliferation. Gene expression changes dependent on trisomy of Erg in Ts (17 16 )65Dn multilineage progenitor cells were correlated with those associated with trisomy of HSA21 in human DS hematopoietic stem and primitive progenitor cells. These data suggest a role for ERG as a regulator of hematopoietic lineage potential, and that trisomy of ERG in the context of DS foetal liver hemopoiesis drives the pre-leukemic changes that predispose to subsequent DS-TMD and DS-AMKL.
The formation of megakaryocyte colonies in agar cultures of murine bone marrow or spleen cells ca... more The formation of megakaryocyte colonies in agar cultures of murine bone marrow or spleen cells can be stimulated by the addition of interleukin-3 (IL-3), erythropoietin (EPO), thrombopoietin (TPO), or IL-6. However, greater numbers of colonies developed if combinations of two or more of these stimuli were used, particularly combinations including stem cell factor, with maximal numbers of colonies developing with the combination of stem cell factor plus IL-3 plus EPO. The data indicate that most committed progenitor cells in the megakaryocyte lineage were unusual in that they required stimulation by two or more hematopoietic growth factors. In tests using a range of growth factors, G-CSF was exceptional in that it consistently specifically inhibited megakaryocyte colony formation stimulated by EPO, TPO, or IL-6 but not that stimulated by IL-3. The mechanisms involved in this inhibitory action of G-CSF are unknown, but the inhibitory action could be of relevance for the dose-dependent lowering of platelet levels observed in some subjects injected with G-CSF.
There are only three known stimuli for eosinophil formation-GM-CSF, interleukin-5 (IL-5), and IL-... more There are only three known stimuli for eosinophil formation-GM-CSF, interleukin-5 (IL-5), and IL-3. Because mice with inactivation of the gene encoding the common β receptor chain for GM-CSF, IL-5, and IL-3 (βc -/mice) cannot respond to GM-CSF or IL-5 and do not produce IL-3, they should lack eosinophils. However, they produce reduced numbers of eosinophils, indicating the existence of at least one additional stimulatory factor. Use of βc -/mouse marrow cells failed to detect a novel eosinophil-stimulating factor in cell line-or organ-conditioned media, but stem cell factor (SCF) was found to have a previously unrecognized ability to stimulate the formation of eosinophil colonies or mixed neutrophil-eosinophil colonies. This action of SCF was also observable with marrow cells from other mouse strains and was enhanced by the addition of G-CSF but no other factor tested. Recloning of SCF-stimulated blast colonies showed that progenitors forming pure eosinophil or mixed neutrophil-eosinophil colonies can have a common ancestor but many appear to arise independently from different preprogenitor cells. The observed activity of SCF in marrow cultures was relatively weak, but its action may be stronger in vivo, and SCF needs to be added to GM-CSF, IL-5, and IL-3 in the list of cytokines able to stimulate eosinophil formation.
, SOCS-1, SOCS-2, SOCS-3, SOCS-5, SOCS-6, or SOCS-7 indicated that the inhibitory SOCS proteins, ... more , SOCS-1, SOCS-2, SOCS-3, SOCS-5, SOCS-6, or SOCS-7 indicated that the inhibitory SOCS proteins, with the possible exception of SOCS-3, were not involved in the G-CSF-initiated suppression of megakaryocyte colony formation. Stem Cells 2005;23:55-62
Proceedings of the National Academy of Sciences, 2013
Previous studies have shown that mouse bone marrow cells can produce mast cells when stimulated i... more Previous studies have shown that mouse bone marrow cells can produce mast cells when stimulated in vitro by stem cell factor (SCF) and interleukin-3 (IL-3). Experiments to define the marrow cells able to generate mast cells showed that the most active subpopulations were the Kit + Sca1progenitor cell fraction and the more ancestral Kit + Sca1 + blast colony-forming cell fraction. In clonal cultures, up to 64% of blast colony-forming cells were able to generate mast cells when stimulated by SCF and IL-3, and, of these, the most active were those in the CD34 -Flt3Rlong-term repopulating cell fraction. Basophils, identified by the monoclonal antibody mMCP-8 to mouse mast cell serine protease-8, were also produced by 50% of blast colony-forming cells with a strong concordance in the production of both cell types by individual blast colony-forming cells. Enriched populations of marrow-derived basophils were shown to generate variable numbers of mast cells after a further incubation with SCF and IL-3. The data extend the repertoire of lineage-committed cells able to be produced by multipotential hematopoietic blast colony-forming cells and show that basophils and mast cells can have common ancestral cells and that basophils can probably generate mast cells at least under defined in vitro conditions.
Proceedings of the National Academy of Sciences, 2009
units, spleen (CFU-S) supporting the possibility that the 2 cells may be identical. The colony po... more units, spleen (CFU-S) supporting the possibility that the 2 cells may be identical. The colony populations generated by BL-CFC-S were similar in their phenotype and proliferative capacity to progenitor cells in whole bone marrow but the progeny of BL-CFC-F were skewed with an abnormally high proportion of Kit ؊ Flt3R ؉ cells whose clonogenic cells tended to generate only macrophage progeny. Both blast colony populations had a high percentage of GR1 ؉ and Mac1 ؉ cells but BL-CFC-F colonies also contained a significant population of B220 ؉ and IL-7R ؉ cells relevant to the superior ability of BL-CFC-F colony cells to generate B lymphocytes and the known dependency of this process on Flt3 ligand and IL-7. The commitment events and phenotypic changes during the generation of differing progenitor cells in blast colonies can now be clonally analyzed in a convenient in vitro culture system. blast colonies ͉ lineage commitment ͉ hematopoietic stem cells
Murine Ba/F3 cells were transfected with cDNA for the ␣-chain of the murine interleukin-5 (IL-5) ... more Murine Ba/F3 cells were transfected with cDNA for the ␣-chain of the murine interleukin-5 (IL-5) receptor and cloned lines of these cells were able to proliferate in response to as little as 2.5 pg/ml of IL-5. The bioassay was demonstrated to be specific for IL-5 and was able to measure IL-5 produced in culture by organs from adult C57BL/6 and BALB/c mice. The highest levels of IL-5 were produced by lung tissue but thymus and bladder consistently produced IL-5 and more variable production was observed by the heart, spleen, muscle, bone shaft, uterus and testes. Bone marrow cells produced no detectable IL-5. Observed levels of production of IL-5 were similar when using organs from mice lacking high-affinity receptors for IL-5 and from nu/nu, RAG-1−/− and NOD/SCID mice lacking T lymphocytes. In inflammatory peritoneal exudates induced by the injection of casein plus bacteria, levels of induced IL-5 were higher if the mice lacked high-affinity receptors for IL-5. The data indicate that T lymphocytes are not the dominant cellular source of IL-5 in organ-conditioned media and that local IL-5 production can occur with a wide range of normal murine organs. Leukemia (2001) 15, 1248-1255.
Polycomb Repressive Complex 2 (PRC2) is a chromatin modifier that regulates stem cells in embryon... more Polycomb Repressive Complex 2 (PRC2) is a chromatin modifier that regulates stem cells in embryonic and adult tissues. Loss of function studies of PRC2 components has been complicated by early embryonic dependence on PRC2 activity and the partial functional redundancy of Ezh1 and Ezh2, which encode the enzymatic component of PRC2. Here, we investigated the role of PRC2 in hematopoiesis by conditional deletion of Suz12, a core component of PRC2. Complete loss of Suz12 resulted in failure of hematopoiesis, both in the embryo and the adult, with a loss of maintenance of hematopoietic stem cells (HSCs). In contrast, partial loss of PRC2 enhanced HSC self-renewal. Deletion in individual blood cell lineages revealed that, while Suz12 was required for lymphoid development, it was dispensable for the development of granulocytic, monocytic and megakaryocytic cells. Collectively, these data reveal the multi-faceted role of PRC2 in hematopoiesis, with divergent dose-dependent effects in HSC an...
Down syndrome (DS), with trisomy of chromosome 21 (HSA21), is the commonest human aneuploidy. Pre... more Down syndrome (DS), with trisomy of chromosome 21 (HSA21), is the commonest human aneuploidy. Pre-leukemic myeloproliferative changes in DS foetal livers precede the acquisition of GATA1 mutations, transient myeloproliferative disorder (DS-TMD) and acute megakaryocytic leukemia (DS-AMKL). Trisomy of the Erg gene is required for myeloproliferation in the Ts(17 16 )65Dn DS mouse model. We demonstrate here that genetic changes specifically attributable to trisomy of Erg lead to lineage priming of primitive and early multipotential progenitor cells in Ts(17 16 )65Dn mice, excess megakaryocyte-erythroid progenitors, and malignant myeloproliferation. Gene expression changes dependent on trisomy of Erg in Ts (17 16 )65Dn multilineage progenitor cells were correlated with those associated with trisomy of HSA21 in human DS hematopoietic stem and primitive progenitor cells. These data suggest a role for ERG as a regulator of hematopoietic lineage potential, and that trisomy of ERG in the context of DS foetal liver hemopoiesis drives the pre-leukemic changes that predispose to subsequent DS-TMD and DS-AMKL.
Marrow cells from mice lacking high-affinity receptors for granulocyte-macrophage colony-stimulat... more Marrow cells from mice lacking high-affinity receptors for granulocyte-macrophage colony-stimulating factor (GM-CSF; betac-/- mice) were shown to bind and internalize much less GM-CSF than cells from normal (betac+/+) mice. betac-/- mice were used to determine the effect of negligible receptor-mediated clearance on detectible GM-CSF responses to the intravenous injection of endotoxin or the intraperitoneal injection of casein plus microorganisms. Unlike the minor serum GM-CSF responses to endotoxin seen in betac+/+ mice, serum GM-CSF levels rose 30-fold to 9 ng/mL in betac-/- mice even though loss of GM-CSF in the urine was greater than in betac+/+ mice. Organs from betac-/- and betac+/+ mice had a similar capacity to produce GM-CSF in vitro, as did peritoneal cells from both types of mice when challenged in vitro by casein. However, when casein was injected intraperitoneally, betac-/- mice developed higher and more sustained levels of GM-CSF than did betac+/+ mice. The data indicat...
Upon ligand binding, RIPK1 is recruited to tumor necrosis factor receptor superfamily (TNFRSF) an... more Upon ligand binding, RIPK1 is recruited to tumor necrosis factor receptor superfamily (TNFRSF) and Toll-like receptor (TLR) complexes promoting prosurvival and inflammatory signaling. RIPK1 also directly regulates caspase-8-mediated apoptosis or, if caspase-8 activity is blocked, RIPK3-MLKL-dependent necroptosis. We show that C57BL/6 Ripk1(-/-) mice die at birth of systemic inflammation that was not transferable by the hematopoietic compartment. However, Ripk1(-/-) progenitors failed to engraft lethally irradiated hosts properly. Blocking TNF reversed this defect in emergency hematopoiesis but, surprisingly, Tnfr1 deficiency did not prevent inflammation in Ripk1(-/-) neonates. Deletion of Ripk3 or Mlkl, but not Casp8, prevented extracellular release of the necroptotic DAMP, IL-33, and reduced Myd88-dependent inflammation. Reduced inflammation in the Ripk1(-/-)Ripk3(-/-), Ripk1(-/-)Mlkl(-/-), and Ripk1(-/-)Myd88(-/-) mice prevented neonatal lethality, but only Ripk1(-/-)Ripk3(-/-)Cas...
Proceedings of the National Academy of Sciences, 2003
Injection of neonatal bone marrow cells from mice lacking the gene encoding suppressor of cytokin... more Injection of neonatal bone marrow cells from mice lacking the gene encoding suppressor of cytokine signaling 1 (SOCS1) into irradiated syngeneic 129͞Sv or C57BL͞6 mice led to a decreased survival, more rapidly occurring in 129͞Sv than in C57BL͞6 mice. Moribund mice did not exhibit the acute or chronic diseases developed by Socs1 ؊/؊ mice but developed a pathology characteristic of graft-versus-host disease with typical chronic inflammatory lesions in the liver, skin, lungs, and gut. The results indicate that cells derived from the Socs1 ؊/؊ bone marrow are autoaggressive but did not identify the cell types involved. Failure of the engrafted Socs1 ؊/؊ marrow cells to reproduce the tissue damage typical of Socs1 ؊/؊ disease indicates that loss of SOCS1 from target tissues may also be required for the development of the Socs1 ؊/؊ diseases, such as fatty degeneration of the liver, polymyositis, or corneal inflammation.
Proceedings of the National Academy of Sciences, 2002
Mice with inactivation of the gene encoding the suppressor of cytokine signaling-1 (SOCS-1) die i... more Mice with inactivation of the gene encoding the suppressor of cytokine signaling-1 (SOCS-1) die in neonatal life with an IFN-␥dependent inflammatory disease dominated by fatty degeneration and necrosis of the liver. To establish the long-term pathological consequences of loss of SOCS-1 in mice, where initial survival was made possible by also deleting the IFN-␥ gene, a comparison was made of the lifespan of groups of SOCS-1 ؊/؊ IFN-␥ ؊/؊ , SOCS-1 ؉/؉ IFN-␥ ؊/؊ and SOCS-1 ؉/؉ IFN-␥ ؉/؉ mice. Mice lacking the genes for both SOCS-1 and IFN-␥ exhibited an accelerated death rate compared with control groups. Disease states developing selectively in SOCS-1 ؊/؊ IFN-␥ ؊/؊ mice were polycystic kidneys, pneumonia, chronic skin ulcers, and chronic granulomas in the gut and various other organs. Mice of all three groups developed cataracts, but disease development was accelerated in the groups lacking IFN-␥. SOCS-1 ؊/؊ IFN-␥ ؊/؊ mice exhibited a slightly increased predisposition to the development of T lymphoid leukemia, either spontaneous or radiation-induced. The development of polycystic kidneys may be caused by a developmental defect in renal-tubule organization noted in neonatal SOCS-1 ؊/؊ mice. The chronic infections and granulomas of SOCS-1 ؊/؊ IFN-␥ ؊/؊ mice may be based on autoaggression of SOCS-1 ؊/؊ T lymphoid and related cells or a functional deficiency of these cells when lacking SOCS-1.
Proceedings of the National Academy of Sciences, 2006
Genetically primed adult C57BL mice were deleted of exon 5 of the gene encoding the transcription... more Genetically primed adult C57BL mice were deleted of exon 5 of the gene encoding the transcription factor PU.1 by IFN activation of Cre recombinase. After a 13-week delay, conditionally deleted (PU.1 ؊/؊ ) mice began dying of myeloid leukemia, and 95% of the mice surviving from early postinduction death developed transplantable myeloid leukemia whose cells were deleted of PU.1 and uniformly Gr-1 positive. The leukemic cells formed autonomous colonies in semisolid culture with varying clonal efficiency, but colony formation was enhanced by IL-3 and sometimes by granulocyte-macrophage colony-stimulating factor. Nine of 13 tumors analyzed had developed a capacity for autocrine IL-3 or granulocyte-macrophage colony-stimulating factor production, and there was evidence of rearrangement of the IL-3 gene. Acquisition of autocrine growth-factor production and autonomous growth appeared to be major events in the transformation of conditionally deleted PU.1 ؊/؊ cells to fully developed myeloid leukemic populations.
Proceedings of the National Academy of Sciences, 2007
Murine granulocytic cells, in becoming leukemic, need to acquire enhanced self-generation and a c... more Murine granulocytic cells, in becoming leukemic, need to acquire enhanced self-generation and a capacity for autocrine growth stimulation. Mice transplanted with bone marrow cells transduced with the Mixl1 homeobox gene develop a very high frequency of myeloid leukemia derived from the transduced cells. Preleukemic mice contained a high frequency of transduced clonogenic granulocytic cells. They exhibited an abnormally high capacity for self-replication and could generate immortalized granulocytic cell lines that remained absolutely dependent on either GM-CSF or IL-3 and were not leukemic. Organs from mice repopulated by marrow cells transduced either with Mixl1 or the control murine stem cell virus vector exhibited a capacity to produce IL-3 in vitro, activity being highest with the lungs, marrow, bladder, and thymus.
Proceedings of the National Academy of Sciences, 2007
In agar cultures of marrow cells from adult female BALB/c chimeric GATA-1 Plt13/؉ mice, a high fr... more In agar cultures of marrow cells from adult female BALB/c chimeric GATA-1 Plt13/؉ mice, a high frequency of unusual dispersed colonies was noted. Analysis showed that these were colonies of mast cells and that mast cell colony-forming cells (progenitors) could be detected in clonal cultures of adult marrow, neonatal marrow, or fetal liver if the combined stimulus of stem cell factor and interleukin-3 was used. Mast cell progenitors were in active cell cycle and showed an extensive capacity for self-generation. Mast cell colonies both from control GATA-1 ؉/؉ mice and GATA-1 Plt13/؉ mice could generate growth factor-dependent cloned cell lines that grew for >18 months. Surprisingly, the majority of the excessive numbers of mast cell progenitors in chimeric GATA-1 Plt13/؉ mice were transcribing the inactive Plt13 allele of GATA-1, suggesting that GATA-1 normally acts to restrict the emergence of committed mast cell progenitors. In sharp contrast, all eosinophil progenitors in these mice were transcribing the normal GATA-1 allele. No excess tissue mast cells were observed in GATA-1 Plt13/؉ mice, suggesting that the excess mast cell progenitors in these mice might be generating mast cells with a defective in vivo proliferative or tissue homing capacity. mast cell colonies ͉ mast cell lines ͉ eosinophil progenitors Author contributions: D.M., I.M., and W.S.A. designed research; D.M., I.M., S.M., and L.D.R. performed research; D.M., I.M., S.M., L.D.R., and W.S.A. analyzed data; and D.M. wrote the paper.
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