Showing papers on "Granulopoiesis published in 2012"

Lysine-specific demethylase 1 restricts hematopoietic progenitor proliferation and is essential for terminal differentiation.

Abstract: Lysine (K)-specific demethylase 1A (LSD1/KDM1A) has been identified as a potential therapeutic target in solid cancers and more recently in acute myeloid leukemia. However, the potential side effects of a LSD1-inhibitory therapy remain elusive. Here, we show, with a newly established conditional in vivo knockdown model, that LSD1 represents a central regulator of hematopoietic stem and progenitor cells. LSD1 knockdown (LSD1-kd) expanded progenitor numbers by enhancing their proliferative behavior. LSD1-kd led to an extensive expansion of granulomonocytic, erythroid and megakaryocytic progenitors. In contrast, terminal granulopoiesis, erythropoiesis and platelet production were severely inhibited. The only exception was monopoiesis, which was promoted by LSD1 deficiency. Importantly, we showed that peripheral blood granulocytopenia, monocytosis, anemia and thrombocytopenia were reversible after LSD1-kd termination. Extramedullary splenic hematopoiesis contributed to the phenotypic reversion, and progenitor populations remained expanded. LSD1-kd was associated with the upregulation of key hematopoietic genes, including Gfi1b, Hoxa9 and Meis1, which are known regulators of the HSC/progenitor compartment. We also demonstrated that LSD1-kd abrogated Gfi1b-negative autoregulation by crossing LSD1-kd with Gfi1b:GFP mice. Taken together, our findings distinguish LSD1 as a critical regulator of hematopoiesis and point to severe, but reversible, side effects of a LSD1-targeted therapy.

Functionally relevant neutrophilia in CD11c diphtheria toxin receptor transgenic mice

Abstract: Transgenic mice expressing the diphtheria toxin receptor (DTR) in specific cell types are key tools for functional studies in several biological systems. B6.FVB-Tg(Itgax-DTR/EGFP)57Lan/J (CD11c.DTR) and B6.Cg-Tg(Itgax-DTR/OVA/EGFP)1Gjh/Crl (CD11c.DOG) mice express the DTR in CD11c(+) cells, allowing conditional depletion of dendritic cells. We report that dendritic-cell depletion in these models caused polymorphonuclear neutrophil (PMN) release from the bone marrow, which caused chemokine-dependent neutrophilia after 6-24 h and increased bacterial clearance in a mouse pyelonephritis model. We present a transgenic mouse line, B6.Cg-Tg(Itgax-EGFP-CRE-DTR-LUC)2Gjh/Crl (CD11c.LuciDTR), which is unaffected by early neutrophilia. However, CD11c.LuciDTR and CD11c.DTR mice showed late neutrophilia 72 h after dendritic cell depletion, which was independent of PMN release and possibly resulted from increased granulopoiesis. Thus, the time point of dendritic cell depletion and the choice of DTR transgenic mouse line must be considered in experimental settings where neutrophils may be involved.

Interactions among HCLS1, HAX1 and LEF-1 proteins are essential for G-CSF-triggered granulopoiesis

Abstract: We found that hematopoietic cell-specific Lyn substrate 1 (HCLS1 or HS1) is highly expressed in human myeloid cells and that stimulation with granulocyte colony-stimulating factor (G-CSF) leads to HCLS1 phosphorylation. HCLS1 binds the transcription factor lymphoid-enhancer binding factor 1 (LEF-1), transporting LEF-1 into the nucleus upon G-CSF stimulation and inducing LEF-1 autoregulation. In patients with severe congenital neutropenia, inherited mutations in the gene encoding HCLS1-associated protein X-1 (HAX1) lead to profound defects in G-CSF-triggered phosphorylation of HCLS1 and subsequently to reduced autoregulation and expression of LEF-1. Consistent with these results, HCLS1-deficient mice are neutropenic. In bone marrow biopsies of the majority of tested patients with acute myeloid leukemia, HCLS1 protein expression is substantially elevated, associated with high levels of G-CSF synthesis and, in some individuals, a four-residue insertion in a proline-rich region of HCLS1 protein known to accelerate intracellular signaling. These data demonstrate the importance of HCLS1 in myelopoiesis in vitro and in vivo.

C/EBPβ Is Involved in the Amplification of Early Granulocyte Precursors during Candidemia-Induced “Emergency” Granulopoiesis

Abstract: Granulopoiesis is tightly regulated to meet host demands during both "steady-state" and "emergency" situations, such as infections. The transcription factor CCAAT/enhancer binding protein β (C/EBPβ) plays critical roles in emergency granulopoiesis, but the precise developmental stages in which C/EBPβ is required are unknown. In this study, a novel flow cytometric method was developed that successfully dissected mouse bone marrow cells undergoing granulopoiesis into five distinct subpopulations (#1-5) according to their levels of c-Kit and Ly-6G expression. After the induction of candidemia, rapid mobilization of mature granulocytes and an increase in early granulocyte precursors accompanied by cell cycle acceleration was followed by a gradual increase in granulocytes originating from the immature populations. Upon infection, C/EBPβ was upregulated at the protein level in all the granulopoietic subpopulations. The rapid increase in immature subpopulations #1 and #2 observed in C/EBPβ knockout mice at 1 d postinfection was attenuated. Candidemia-induced cell cycle acceleration and proliferation of hematopoietic stem/progenitors were also impaired. Taken together, these data suggest that C/EBPβ is involved in the efficient amplification of early granulocyte precursors during candidemia-induced emergency granulopoiesis.

Current insights into neutrophil homeostasis

Abstract: Neutrophil granulocytes represent the first immunologic barrier against invading pathogens, and neutropenia predisposes to infection. However, neutrophils may also cause significant collateral inflammatory damage. Therefore, neutrophil numbers are tightly regulated by an incompletely understood homeostatic feedback loop adjusting the marrow's supply to peripheral needs. Granulocyte colony-stimulating factor (G-CSF) is accepted to be the major determinant of neutrophil production, and G-CSF levels have, soon after its discovery, been described to be inversely correlated with neutrophil counts. A neutrophil sensor, or "neutrostat," has, therefore, been postulated. The prevailing feedback hypothesis was established in adhesion molecule-deficient mice; it includes macrophages and Th17 cells, which determine G-CSF levels in response to the number of peripherally transmigrated, apoptosing neutrophils. Recent work has deepened our understanding of homeostatic regulation of neutrophil granulopoiesis, but there are still inconsistent findings and unresolved questions when it comes to a plausible hypothesis, similar to the feedback control models of red cell or platelet homeostasis.

A hypomorphic mutation in the Gfi1 transcriptional repressor results in a novel form of neutropenia

Abstract: Using N-ethyl-N-nitrosourea-induced mutagenesis, we established a mouse model with a novel form of neutropenia resulting from a point mutation in the transcriptional repressor Growth Factor Independence 1 (Gfi1). These mice, called Genista, had normal viability and no weight loss, in contrast to mice expressing null alleles of the Gfi1 gene. Furthermore, the Genista mutation had a very limited impact on lymphopoiesis or on T- and B-cell function. Within the bone marrow (BM), the Genista mutation resulted in a slight increase of monopoiesis and in a block of terminal granulopoiesis. This block occurred just after the metamyelocytic stage and resulted in the generation of small numbers of atypical CD11b + Ly-6G int neutrophils, the nuclear morphology of which resembled that of mature WT neutrophils. Unexpectedly, once released from the BM, these atypical neutrophils contributed to induce mild forms of autoantibody-induced arthritis and of immune complex-mediated lung alveolitis. They additionally failed to provide resistance to acute bacterial infection. Our study demonstrates that a hypomorphic mutation in the Gfi1 transcriptional repressor results in a novel form of neutropenia characterized by a split pattern of functional responses, reflecting the distinct thresholds required for eliciting neutrophil-mediated inflammatory and anti-infectious responses.

The role of transcription factors in the guidance of granulopoiesis.

Abstract: In recent years, the prospective isolation of hematopoietic stem and progenitor cells has identified the hierarchical structure of hematopoietic development and lineage-commitment. Moreover, these isolated cell populations allowed the elucitation of the molecular mechansims associated with lineage choice and revealed the indispensable functions of transcription factors as lineage determinants. This review summarizes current concepts regarding adult murine granulopoiesis and illustrates the importance of the transcription factors C/EBPα, PU.1 and GATA-2 for the development of neutrophil, eosinophil and basophil granulocytes.

The potential of interleukin-17 to mediate hematopoietic response

Abstract: It has long been known that T cells have the potential to modulate hematopoietic response in different ways. More recently, the importance of interleukin (IL)-17-secreting Th17 cells in T-cell-mediated regulation of hematopoiesis was indicated by the line of evidence that IL-17 links T-cell function and hematopoiesis through stimulation of granulopoiesis and neutrophil trafficking. Furthermore, our data demonstrated that IL-17 also affects other cells of hematopoietic system, such as erythroid progenitors, as well as mesenchymal stem cells. In order to better understand the regulatory role of IL-17 in hematopoiesis, molecular mechanisms underlying the effects of IL-17 on hematopoietic and mesenchymal stem cells were also studied.

Early Systemic Bacterial Dissemination and a Rapid Innate Immune Response Characterize Genetic Resistance to Plague of SEG Mice

Abstract: Background Although laboratory mice are usually highly susceptible to Yersinia pestis, we recently identified a mouse strain (SEG) that exhibited an exceptional capacity to resist bubonic plague and used it to identify immune mechanisms associated with resistance Methods The kinetics of infection, circulating blood cells, granulopoiesis, lesions, and cellular populations in the spleen, and cytokine production in various tissues were compared in SEG and susceptible C57BL/6J mice after subcutaneous infection with the virulent Y pestis CO92 Results Bacterial invasion occurred early (day 2) but was transient in SEG/Pas mice, whereas in C57BL/6J mice it was delayed but continuous until death The bacterial load in all organs significantly correlated with the production of 5 cytokines (granulocyte colony-stimulating factor, keratinocyte-derived chemokine (KC), macrophage cationic peptide-1 (MCP-1), interleukin 1a, and interleukin 6) involved in monocyte and neutrophil recruitment Indeed, higher proportions of these 2 cell types in blood and massive recruitment of F4/801CD11b2 macrophages in the spleen were observed in SEG/Pas mice at an early time point (day 2) Later times after infection (day 4) were characterized in C57BL/6J mice by destructive lesions of the spleen and impaired granulopoiesis Conclusion A fast and efficient Y pestis dissemination in SEG mice may be critical for the triggering of an early and effective innate immune response necessary for surviving plague

Gene Expression-Based Detection of Radiation Exposure in Mice after Treatment with Granulocyte Colony-Stimulating Factor and Lipopolysaccharide

Abstract: In a large-scale nuclear incident, many thousands of people may be exposed to a wide range of radiation doses. Rapid biological dosimetry will be required on an individualized basis to estimate the exposures and to make treatment decisions. To ameliorate the adverse effects of exposure, victims may be treated with one or more cytokine growth factors, including granulocyte colony-stimulating factor (G-CSF), which has therapeutic efficacy for treating radiation-induced bone marrow ablation by stimulating granulopoiesis. The existence of infections and the administration of G-CSF each may confound the ability to achieve reliable dosimetry by gene expression analysis. In this study, C57BL/6 mice were used to determine the extent to which G-CSF and lipopolysaccharide (LPS, which simulates infection by gram-negative bacteria) alter the expression of genes that are either radiation-responsive or non-responsive, i.e., show potential for use as endogenous controls. Mice were acutely exposed to 60Co γ rays at eithe...

Radioprotective effects of (-)-epigallocatechin-3-gallate on human erythrocyte/granulocyte lineages.

Abstract: Epigallocatechin-3-gallate (EGCg) is widely recognised as a powerful antioxidant and free radical scavenger. This study examined the radioprotective effects of EGCg on human granulopoiesis and erythropoiesis. Highly purified human CD34(+) haematopoietic stem/progenitor cells were prepared from human placental/umbilical cord blood. The cells were exposed to X rays at a dose rate of ∼1 Gy min(-1) and then cultured in a medium supplemented with either granulocyte colony-stimulating factor (G-CSF) or erythropoietin (EPO). EGCg (100 nM) was added to the culture immediately before or after X-irradiation. The concentration of 100-nM EGCg was determined in the authors' previous study. The number of granulocyte and erythrocyte colonies generated by X-irradiated CD34(+) cells decreased in a dose-dependent manner. Although EGCg addition yielded an ∼2-fold increase in the proliferation of each haematopoietic progenitor, no significant protective effect was observed in the surviving fraction of granulocyte progenitors (G-CSF alone: D(0)=1.06 Gy, n=1.14). However, EGCg addition before or after irradiation conferred a significantly higher protective effect on erythrocyte colony formation compared with the control (EPO alone: D(0)=0.66 Gy, n=1.56; EGCg (before): D(0)=0.43 Gy, n=5.48). EGCg addition before irradiation significantly improved the survival of erythroid progenitors subjected to radiation of <1 Gy. These results suggest that EGCg is more protective of erythropoiesis than granulopoiesis from radiation damage.

Effects of IL-17 on erythroid progenitors growth: involvement of MAPKs and GATA transcription factors.

Abstract: Interleukin-17 is Th17 cell cytokine implicated in regulation of hematopoiesis and inflammation. Besides promoting granulopoiesis, we have previously shown that IL-17 also affects erythropoiesis stimulating the development of early erythroid progenitors, BFU-E, but suppressing, at least partly via p38 MAPK, the growth of late stage erythroid progenitors, CFU-E. The aim of the present study was to investigate the involvement of other MAPKs, JNK and ERK1/2, as well as GATA transcription factors, in IL-17-mediated effects on murine bone marrow erythroid progenitors. Data obtained by use of specific MAPKs inhibitors indicated that MEK1/2-ERK1/2 MAPK signaling mediates IL-17-induced CFU-E inhibition, as well as that JNK and/or MEK1/2-ERK1/2 MAPKs activation underlies IL-17-induced stimulation of BFU-E growth. Furthermore, Western blot analyses demonstrated no effect on early hematopoiesis transcription factor, GATA-2, and enhanced expression level of erythroid-specific factor GATA-1 in murine bone marrow cells after IL-17 stimulation, which in light of previous reports that GATA-1 overexpression inhibits erythroid differentiation, could be related to IL-17-mediated inhibition of CFU-E growth. Although, no contribution for p38, JNK and ERK MAPKs in IL-17-induced GATA-1 expression was shown, data obtained using specific inhibitors pointed to the role of JNK and MEK1/2-ERK1/2 in GATA-1 downregulation. Overall, obtained data gave an insight into the mechanisms by which IL-17 exerts its effects on erythropoiesis, implying the involvement of JNK and ERK MAPKs, as well as GATA-1, in IL-17-regulated growth of erythroid progentors.

Sodium caseinate induces mouse granulopoiesis.

Abstract: Objective and design Sodium caseinate (CasNa) induces differentiation and M-CSF production in mouse band granulocytes in vitro; however, it is not yet known if this molecule can also induce the proliferation and activation of the granulocyte lineage in vivo. In this work we evaluated the induction in vivo of granulopoiesis and the activation of granulocytes in mice treated with CasNa.

Clonal patterns of X-chromosome inactivation in peripheral blood cells of female patients with chronic idiopathic neutropenia.

Abstract: Chronic idiopathic neutropenia (CIN) is an acquired disorder of granulopoiesis characterized by prolonged, unexplained reduction in peripheral blood (PB) neutrophils due to accelerated apoptosis of the bone marrow (BM) granulocytic progenitors.[1][1] There has been growing evidence to suggest that

Interpretation of Maturing Trilineage Hematopoiesis

Abstract: Background: Normal adult bone marrow occupies the medullary spaces of large bones. The main function of bone marrow is the production of blood cells (hematopoiesis). Bone marrow is composed of a matrix requisite for hematopoiesis as well as erythriod precursors, myeloid precursors with monocyte macrophage system, megakaryocytes, lymphocytes, plasma cells, blood vessels and stroma. Interpretation of bone marrow maturing trilineage hematopoiesis includes the assessment of the maturation sequence and morphologic features for each lineage. Aim: This review presents detailed guidelines for interpretation of bone marrow maturing trilineage hematopoiesis according to the cell line and the pathologic condition. Keywords: Bone marrow, Trilineage hematopoiesis, Erythropoiesis, Granulopoiesis, Megakaryopoiesis