A Constitutively-Active ABL Family Kinase, TEL-ARG, Induces Lethal Mastocytosis through Sensitizing Hematopoietic Stem/Progenitor Cells to IL-3

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2828-2828
Author(s):  
Asumi Yokota ◽  
Hideyo Hirai ◽  
Tsukimi Shoji ◽  
Taira Maekawa ◽  
Keiko Okuda
Keyword(s):  
Mast Cell ◽  
Cell Differentiation ◽  
Mast Cells ◽  
Progenitor Cells ◽  
Myeloid Leukemia ◽  
Myeloid Differentiation ◽  
Dependent Manner ◽  
Hematopoietic Stem ◽  
Terminal Domain ◽  
Constitutively Active

Abstract ABL family kinases, ABL1 (ABL) and ABL2 (ARG), share functional domains such as SH2-, SH3- and kinase domains, and are highly homologous except their C-terminal domain. Fusions to TEL (ETV6), TEL-ABL and TEL-ARG, are constitutively-active kinases and have been reported in rare cases of human CML, AML or ALL. Although TEL-ABL is involved in leukemogenesis, the role of TEL-ARG has not been elucidated because this fusion protein has been always accompanied with other major translocations, such as PML-RARα. We have previously shown that although their kinase activities are comparable, TEL-ABL strongly transforms Ba/F3 cells, while TEL-ARG has a much lower transforming activity, and these differences are attributed to their distinct C-terminal domain (Okuda K and Hirai H, Open Journal of Blood Diseases 2013). At the last ASH annual meeting, we have shown that TEL-ABL induces myeloid leukemia in a short latency, whereas TEL-ARG induces lethal mastocytosis in a long latency in a mouse bone marrow (BM) transplantation model (Abstract number #2368, ASH 2014). Here we investigated the clonogenicity of mastocytosis and explored the detailed mechanism underlying the onset of mastocytosis induced by TEL-ARG. First, we performed a serial transplantation experiment to evaluate mastocytosis-initiating capacity of TEL-ARG-expressing cells. Hematopoietic stem/progenitor cells (HSPCs) from 5-FU-treated mice were retrovirally transduced with TEL-ARG and transplanted to the first recipient mice. BM cells from moribund mice due to mastocytosis were transplanted to the sublethally irradiated second recipients. On day 219 after transplantation, we detected mast cells circulating in the peripheral blood of these two recipients, and observed severe pancytopenia and body weight loss in one of them. In this mouse, mast cells engulfing blood cells were accumulated in the BM and spleen, and subcutaneous tissues were massively infiltrated by mast cells, all of which were characteristics of mastocytosis observed in the first recipients. These results indicate that TEL-ARG confers mastocytosis-initiating capacity on HSPCs. Next, we focused on the mechanisms why TEL-ARG induces mastocytosis, whereas TEL-ABL induces myeloid leukemia. HSPCs from 5-FU-treated mice were retrovirally transduced with TEL-ABL or TEL-ARG, and subjected to the in vitro mast cell differentiation assay in the presence of WEHI-conditioned medium, as a source of IL-3 (Figure). IL-3 enhanced differentiation and proliferation of empty-virus-transduced HSPCs toward mast cells in a dose-dependent manner. TEL-ARG induced mast cell differentiation in the absence of IL-3 to some extent, and IL-3 markedly increased mast cell number even at a lower concentration. TEL-ARG-expressing mast cells continue to proliferate for more than 4 months maintaining their phenotype as mast cells. In contrast, IL-3 did not enhance mast cell differentiation but support myeloid differentiation of TEL-ABL-expressing HSPCs. These data suggest that while TEL-ABL induces myeloid differentiation, TEL-ARG strongly promotes differentiation toward mast cells through sensitizing HSPCs to IL-3, an important factor for differentiation, survival and proliferation of mast cells. Furthermore, these results might account for differences in the phenotypes of diseases induced by TEL-ABL (myeloid leukemia) or TEL-ARG (mastocytosis). In conclusions, TEL-ABL strongly induces myeloid-skewed differentiation, whereas TEL-ARG promotes mast cell differentiation through increasing sensitivity to IL-3 and induces clonal mast cell disease. We are currently investigating the molecular mechanisms by which they activate distinct differentiation pathways toward myeloid cells or mast cells. We believe that further exploration of the underlying mechanisms will deepen our understanding of the molecular basis for ABL kinase-mediated leukemogenesis as well as mast cell disorders. Disclosures No relevant conflicts of interest to declare.

Murine Mast Cells Express Mpl, the Thrombopoietin Receptor, and Thrombopoietin Is a Potent Regulator of Mast Cell Differentiation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1335-1335
Author(s):  
Fabrizio Martelli ◽  
Giovanni Amabile ◽  
Barbara Ghinassi ◽  
Rodolfo Lorenzini ◽  
Alessandro M. Vannucchi ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mast Cell ◽  
Cell Differentiation ◽  
Mast Cells ◽  
Progenitor Cells ◽  
Peritoneal Cavity ◽  
Surface Protein ◽  
Wild Type ◽  
Hematopoietic Stem ◽  
Progenitor Cell Proliferation

Abstract Mast cells are hematopoietic cells localized in extramedullary sites where they engage themselves in the process of allergic response and in the immune reaction against parasites. Mast cells derive from multilineage c-KitlowCD34lowSca-1pos progenitor cells present in the marrow. These cells give rise to Linnegc-KitposSca-1neg T1/ST2pos mast cell restricted progenitor cells (MCP) whose futher maturation in the marrow remains limited under steady state conditions. MCP migrate through the blood in extramedullary sites were they mature into tissue-retricted c-KitposFceRIpos mast cells characterized by a specific mast cell protease (MMCP) profiling (dermal, mucosal and serosal mast cells in skin, gut and peritoneal cavity, respectively). The molecular mechanism that, in normal mice, restricts the mastocytopoietic potential of progenitor cells to the extramedullary sites, as well as the factors that guide the tissue-restricted differentiation of these cells, are unknown. Thrombopoietin (TPO)-Mpl interactions play an important role in the regulation of hematopoietic stem/progenitor cell proliferation and differentiation in the marrow. Here we report that mast cells, and their precursors, express Mpl (both as mRNA and cell surface protein) (see Table). Furthermore, targeted deletion of this gene (Mplnull mutation) decrease the number of MCP (by 1-log) and increases that of mast cells in dermis (by 3-fold), peritoneal cavity (by 3-fold), bone marrow (2-log) and spleen (2-log). Furthermore, because of their higher (by 2-log) MMCP-7 expression, serosal Mplnull mast cells resemble more wild-type dermal rather than serosal mast cells. On the other hand, either treatment of mice with TPO or addition of TPO to bone marrow-derived mast cell cultures induces mast cell apoptosis (by Tunel and Annexin staining) and severely hampers mast cell differentiation (by expression profiling). These data are consistent with a regulatory mechanism for murine mastocytopoiesis according to which TPO favours the transition from multilineage progenitors to CMP but blocks differentiation of MCP to mature mast cells. We propose TPO as the growth factor that restrict mast cell differentiation to extramedullaty sites and that control the switch between serosal vs dermal mast cell differentiation. Mpl expression mRNA 2-ΔCt Protein (AFU) Cy7-A Protein (AFU) Cy7-AMM2 AFU= arbitrary fluorescence intensity. p< 0.01 with respect to Cy7-A (irrilevant antibody) Wild type Marrow B cells (B220pos) b.d. 120±4 205±4 Wild type Marrow Megakaryocytes (CD61pos/CD41pos) 5.0±0.1 × 10-2 178±3 978±74* Wild type Marrow MCP (cKitpos/T1ST2pos) 1.3±0.01 × 10-2 139±16 1658±73* Wild-type Marrow Mast Cells (cKitpos/Fcε RIpos) 1.9±0.1 × 10-2 110±1 868±71* Serosal Mast Cells (cKitpos/FcεRIpos) 7.2±2.1 × 10-4 393±1 1374±25* Mplnull Marrow Megakaryocytes (CD61pos/CD41pos) b.d. 365±28 469±50 Mplnull Marrow Mast Cells (cKitpos/FcεRIpos) b.d 107±1 109±3

scholarly journals MicroRNA-29a and microRNA-142-3p are regulators of myeloid differentiation and acute myeloid leukemia

Blood ◽  
2012 ◽  
Vol 119 (21) ◽  
pp. 4992-5004 ◽  
Author(s):  
Xiao-Shuang Wang ◽  
Jia-Nan Gong ◽  
Jia Yu ◽  
Fang Wang ◽  
Xin-Hua Zhang ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Progenitor Cells ◽  
Myeloid Leukemia ◽  
Leukemia Cell ◽  
Myeloid Differentiation ◽  
Loss Of Function ◽  
Monocytic Differentiation ◽  
Granulocytic Differentiation ◽  
Hematopoietic Stem ◽  
Acute Myeloid

Abstract Although microRNAs (miRNAs) are increasingly linked to various physiologic processes, including hematopoiesis, their function in the myeloid development is poorly understood. We detected up-regulation of miR-29a and miR-142-3p during myeloid differentiation in leukemia cell lines and CD34+ hematopoietic stem/progenitor cells. By gain-of-function and loss-of-function experiments, we demonstrated that both miRNAs promote the phorbol 12-myristate 13-acetate–induced monocytic and all-trans-retinoic acid-induced granulocytic differentiation of HL-60, THP-1, or NB4 cells. Both the miRNAs directly inhibited cyclin T2 gene, preventing the release of hypophosphorylated retinoblastoma and resulting in induction of monocytic differentiation. In addition, a target of miR-29a, cyclin-dependent kinase 6 gene, and a target of miR-142-3p, TGF-β–activated kinase 1/MAP3K7 binding protein 2 gene, are involved in the regulation of both monocytic and granulocytic differentiation. A significant decrease of miR-29a and 142-3p levels and an obvious increase in their target protein levels were also observed in blasts from acute myeloid leukemia. By lentivirus-mediated gene transfer, we demonstrated that enforced expression of either miR-29a or miR-142-3p in hematopoietic stem/progenitor cells from healthy controls and acute myeloid leukemia patients down-regulated expression of their targets and promoted myeloid differentiation. These findings confirm that miR-29a and miR-142-3p are key regulators of normal myeloid differentiation and their reduced expression is involved in acute myeloid leukemia development.

scholarly journals Full-length but not truncated CD34 inhibits hematopoietic cell differentiation of M1 cells

Blood ◽  
1995 ◽  
Vol 85 (11) ◽  
pp. 3040-3047 ◽  
Author(s):  
MJ Fackler ◽  
DS Krause ◽  
OM Smith ◽  
CI Civin ◽  
WS May
Keyword(s):  
Cell Differentiation ◽  
Progenitor Cells ◽  
Myeloid Leukemia ◽  
Hematopoietic Cell ◽  
Full Length ◽  
Truncated Form ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells ◽  
The Difference ◽  
Negative Regulatory Role

CD34 is expressed on human and murine hematopoietic stem and progenitor cells and its clinical usefulness for isolation of stem/progenitor cells has been well established. Although expression of CD34 is regulated in a developmental stage-specific manner, the function of CD34 is not known. Recently we have shown that both a full-length and truncated form of CD34 protein is expressed by hematopoietic cells (Blood 84:691, 1994). To test whether failure to suppress either form of CD34 could affect terminal myeloid differentiation, we constitutively expressed these CD34 proteins in murine M1 myeloid leukemia cells, which can be terminally differentiated to macrophages by treatment with interleukin-6 of leukemia inhibitory factor. Surprisingly our results show that forced expression of the full-length but not the truncated form of CD34 impedes terminal differentiation by these agents. Because the difference between the two forms of CD34 protein resides in the length of their respective cytoplasmic tail domains, our findings strongly suggest that the cytoplasmic domain region of full-length CD34 is responsible for the observed maturation arrest phenotype. These findings suggest a potential negative regulatory role for full-length CD34 in hematopoietic cell differentiation and may explain, at least in part, the block in maturation observed in CD34+ acute myeloid leukemia.

scholarly journals Constitutive Activation of Shp2 Cooperates with MLL1 Fusion Proteins to Increase Expression of the Homeodomain Transcription Factors Cdx4, HoxA9 and HoxA10

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3537-3537
Author(s):  
Ling Bei ◽  
Chirag Shah ◽  
Hao Wang ◽  
Elizabeth Eklund
Keyword(s):  
Transcription Factors ◽  
Progenitor Cells ◽  
Tyrosine Phosphorylation ◽  
Dependent Manner ◽  
Hematopoietic Stem ◽  
Constitutive Activation ◽  
Myeloid Progenitor ◽  
Myeloid Progenitor Cells ◽  
Homeodomain Transcription Factors ◽  
Constitutively Active

Abstract Leukemias with chromosomal translocation or partial tandem duplications involving the MLL (mixed lineage leukemia) gene have exceptionally poor prognosis (referred to as 11q23-leukemias). At the molecular level, 11q23-leukemias are characterized by aberrant expression of a set of homeodomain transcription factors in hematopoietic stem cells (HSC) and differentiating myeloid progenitor cells. This transcription factor set includes HoxB3, B4, A7-11, Cdx2-4 and Meis1. Cdx and Hox proteins are involved in regulating hematopoiesis. Transcription of HOX and CDX genes decreases normal myelopoiesis, but is aberrantly sustained in 11q23-leukemias. Cdx4 activates transcription of the HOXA9 and HOXA10 genes, and HoxA10 activates CDX4 transcription. The events that break this feedback loop, permitting a decrease in Cdx4-expression during normal myelopoiesis, were previously undefined. In the current study, we find that HoxA9 represses CDX4 transcription in differentiating myeloid cells; antagonizing activation by HoxA10. We determine that tyrosine phosphorylation of HoxA10 impairs transcriptional activation of CDX4, but tyrosine phosphorylation of HoxA9 facilitates repression of this gene. Since HoxA9 and HoxA10 are phosphorylated during myelopoiesis, this provides a mechanism for differentiation-stage-specific Cdx4 expression. HoxA9 and HoxA10 are increased in cells expressing Mll-Ell, a leukemia associated MLL1 fusion protein. We find that Mll-Ell induces a HoxA10-dependent increase in Cdx4-expression in myeloid progenitor cells. However, expression of Cdx4 decreases in a HoxA9-dependent manner upon exposure of Mll-Ell-expressing cells to differentiating cytokines. Leukemia associated, constitutively active mutants of Shp2 block cytokine-induced tyrosine phosphorylation of HoxA9 and HoxA10. In comparison to cells expressing Mll-Ell alone, we find that co-expression of Mll-Ell plus constitutively active Shp2 increases CDX4 transcription and Cdx4 expression in myeloid progenitor cells. And, increased Cdx4 expression is sustained upon exposure of these cells to differentiating cytokines. Our results identify a mechanism for increased and sustained CDX4 transcription in leukemias co-overexpressing HoxA9 and HoxA10 in combination with constitutive activation of Shp2. We also demonstrate that inhibition of Shp2-PTP activity decreases Cdx4 expression in Hox-overexpressing human myeloid leukemias. Disclosures No relevant conflicts of interest to declare.

Differential context-dependent effects of friend of GATA-1 (FOG-1) on mast-cell development and differentiation

Blood ◽  
2008 ◽  
Vol 111 (4) ◽  
pp. 1924-1932 ◽  
Author(s):  
Daijiro Sugiyama ◽  
Makoto Tanaka ◽  
Kenji Kitajima ◽  
Jie Zheng ◽  
Hilo Yen ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Mast Cell ◽  
Cell Differentiation ◽  
Mast Cells ◽  
Molecular Mechanisms ◽  
Embryonic Stem ◽  
Experimental System ◽  
Cell Phenotype ◽  
Dependent Manner

Friend of GATA-1 (FOG-1) is a binding partner of GATA-1, a zinc finger transcription factor with crucial roles in erythroid, megakaryocytic, and mast-cell differentiation. FOG-1 is indispensable for the function of GATA-1 during erythro/megakaryopoiesis, but FOG-1 is not expressed in mast cells. Here, we analyzed the role of FOG-1 in mast-cell differentiation using a combined experimental system with conditional gene expression and in vitro hematopoietic induction of mouse embryonic stem cells. Expression of FOG-1 during the progenitor period inhibited the differentiation of mast cells and enhanced the differentiation of neutrophils. Analysis using a mutant of PU.1, a transcription factor that positively or negatively cooperates with GATA-1, revealed that this lineage skewing was caused by disrupted binding between GATA-1 and PU.1, which is a prerequisite for mast-cell differentiation. However, FOG-1 expression in mature mast cells brought approximately a reversible loss of the mast-cell phenotype. In contrast to the lineage skewing, the loss of the mast-cell phenotype was caused by down-regulation of MITF, a basic helix-loop-helix transcription factor required for mast-cell differentiation and maturation. These results indicate that FOG-1 inhibits mast-cell differentiation in a differentiation stage-dependent manner, and its effects are produced via different molecular mechanisms.

scholarly journals Mast Cells Differentiated in Synovial Fluid and Resident in Osteophytes Exalt the Inflammatory Pathology of Osteoarthritis

2022 ◽  
Vol 23 (1) ◽  
pp. 541
Author(s):  
Priya Kulkarni ◽  
Abhay Harsulkar ◽  
Anne-Grete Märtson ◽  
Siim Suutre ◽  
Aare Märtson ◽  
...  
Keyword(s):  
Mast Cell ◽  
Cell Differentiation ◽  
Mast Cells ◽  
Synovial Fluid ◽  
Subchondral Bone ◽  
Rna Seq ◽  
Proteomics Analysis ◽  
Hematopoietic Stem ◽  
Alpha Beta

Introduction: Osteophytes are a prominent feature of osteoarthritis (OA) joints and one of the clinical hallmarks of the disease progression. Research on osteophytes is fragmentary and modes of its contribution to OA pathology are obscure. Aim: To elucidate the role of osteophytes in OA pathology from a perspective of molecular and cellular events. Methods: RNA-seq of fully grown osteophytes, collected from tibial plateau of six OA patients revealed patterns corresponding to active extracellular matrix re-modulation and prominent participation of mast cells. Presence of mast cells was further confirmed by immunohistochemistry, performed on the sections of the osteophytes using anti-tryptase alpha/beta-1 and anti-FC epsilon RI antibodies and the related key up-regulated genes were validated by qRT-PCR. To test the role of OA synovial fluid (SF) in mast cell maturation as proposed by the authors, hematopoietic stem cells (HSCs) and ThP1 cells were cultured in a media supplemented with 10% SF samples, obtained from various grades of OA patients and were monitored using specific cell surface markers by flow cytometry. Proteomics analysis of SF samples was performed to detect additional markers specific to mast cells and inflammation that drive the cell differentiation and maturation. Results: Transcriptomics of osteophytes revealed a significant upregulation of mast cells specific genes such as chymase 1 (CMA1; 5-fold) carboxypeptidase A3 (CPA3; 4-fold), MS4A2/FCERI (FCERI; 4.2-fold) and interleukin 1 receptor-like 1 (IL1RL1; 2.5-fold) indicating their prominent involvement. (In IHC, anti-tryptase alpha/beta-1 and anti- FC epsilon RI-stained active mast cells were seen populated in cartilage, subchondral bone, and trabecular bone.) Based on these outcomes and previous learnings, the authors claim a possibility of mast cells invasion into osteophytes is mediated by SF and present in vitro cell differentiation assay results, wherein ThP1 and HSCs showed differentiation into HLA-DR+/CD206+ and FCERI+ phenotype, respectively, after exposing them to medium containing 10% SF for 9 days. Proteomics analysis of these SF samples showed an accumulation of mast cell-specific inflammatory proteins. Conclusions: RNA-seq analysis followed by IHC study on osteophyte samples showed a population of mast cells resident in them and may further accentuate inflammatory pathology of OA. Besides subchondral bone, the authors propose an alternative passage of mast cells invasion in osteophytes, wherein OA SF was found to be necessary and sufficient for maturation of mast cell precursor into effector cells.

scholarly journals Mast Cell Involvement in Fibrosis in Chronic Graft-Versus-Host Disease

2021 ◽  
Vol 22 (5) ◽  
pp. 2385
Author(s):  
Ethan Strattan ◽  
Gerhard Carl Hildebrandt
Keyword(s):  
Wound Healing ◽  
Mast Cell ◽  
Mast Cells ◽  
Graft Versus Host Disease ◽  
Chronic Gvhd ◽  
Fibrotic Disease ◽  
Acute Leukemias ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host

Allogeneic hematopoietic stem cell transplantation (HSCT) is most commonly a treatment for inborn defects of hematopoiesis or acute leukemias. Widespread use of HSCT, a potentially curative therapy, is hampered by onset of graft-versus-host disease (GVHD), classified as either acute or chronic GVHD. While the pathology of acute GVHD is better understood, factors driving GVHD at the cellular and molecular level are less clear. Mast cells are an arm of the immune system that are known for atopic disease. However, studies have demonstrated that they can play important roles in tissue homeostasis and wound healing, and mast cell dysregulation can lead to fibrotic disease. Interestingly, in chronic GVHD, aberrant wound healing mechanisms lead to pathological fibrosis, but the cellular etiology driving this is not well-understood, although some studies have implicated mast cells. Given this novel role, we here review the literature for studies of mast cell involvement in the context of chronic GVHD. While there are few publications on this topic, the papers excellently characterized a niche for mast cells in chronic GVHD. These findings may be extended to other fibrosing diseases in order to better target mast cells or their mediators for treatment of fibrotic disease.

scholarly journals Calcium Pyrophosphate Dihydrate Crystals Increase the Granulocyte/Monocyte Progenitor (GMP) and Enhance Granulocyte and Monocyte Differentiation In Vivo

2020 ◽  
Vol 22 (1) ◽  
pp. 262
Author(s):  
Nobuyuki Onai ◽  
Chie Ogasawara
Keyword(s):  
Cell Differentiation ◽  
Peripheral Blood ◽  
Calcium Pyrophosphate ◽  
Hematopoietic Progenitor Cell ◽  
Calcium Pyrophosphate Dihydrate ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Hematopoietic Stem ◽  
Cppd Crystal ◽  
Pyrophosphate Dihydrate

Calcium pyrophosphate dihydrate (CPPD) crystals are formed locally within the joints, leading to pseudogout. Although the mobilization of local granulocytes can be observed in joints where pseudogout has manifested, the mechanism of this activity remains poorly understood. In this study, CPPD crystals were administered to mice, and the dynamics of splenic and peripheral blood myeloid cells were analyzed. As a result, levels of both granulocytes and monocytes were found to increase following CPPD crystal administration in a concentration-dependent manner, with a concomitant decrease in lymphocytes in the peripheral blood. In contrast, the levels of other cells, such as dendritic cell subsets, T-cells, and B-cells, remained unchanged in the spleen, following CPPD crystal administration. Furthermore, an increase in granulocytes/monocyte progenitors (GMPs) and a decrease in megakaryocyte/erythrocyte progenitors (MEPs) were also observed in the bone marrow. In addition, CPPD administration induced production of IL-1β, which acts on hematopoietic stem cells and hematopoietic progenitors and promotes myeloid cell differentiation and expansion. These results suggest that CPPD crystals act as a “danger signal” to induce IL-1β production, resulting in changes in course of hematopoietic progenitor cell differentiation and in increased granulocyte/monocyte levels, and contributing to the development of gout.

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