High Levels of Thrombopoietin Impair the Promotion of Myelofibrosis by Monocyte/Macrophage Dysfunctions Do
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Monocyte/Macrophage Dysfunctions Do Not Impair the Promotion of Myelofibrosis by High Levels of Thrombopoietin This information is current as Orianne Wagner-Ballon, Hédia Chagraoui, Eric Prina, of September 28, 2021. Micheline Tulliez, Geneviève Milon, Hana Raslova, Jean-Luc Villeval, William Vainchenker and Stéphane Giraudier J Immunol 2006; 176:6425-6433; ; doi: 10.4049/jimmunol.176.11.6425 Downloaded from http://www.jimmunol.org/content/176/11/6425 References This article cites 45 articles, 26 of which you can access for free at: http://www.jimmunol.org/content/176/11/6425.full#ref-list-1 http://www.jimmunol.org/ Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists by guest on September 28, 2021 • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2006 by The American Association of Immunologists All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Monocyte/Macrophage Dysfunctions Do Not Impair the Promotion of Myelofibrosis by High Levels of Thrombopoietin1 Orianne Wagner-Ballon,*† He´dia Chagraoui,2* Eric Prina,2‡ Micheline Tulliez,§ Genevie`ve Milon,‡ Hana Raslova,* Jean-Luc Villeval,* William Vainchenker,* and Ste´phane Giraudier3*† Several lines of evidence indicate that the megakaryocyte/platelet lineage is crucial in myelofibrosis induction. The demonstration that NOD/SCID mice with functionally deficient monocytes do not develop fibrotic changes when exposed to thrombopoietin (TPO) also suggests an important role for monocyte/macrophages. However, in this animal model, the development of myelofibrosis is dependent on the level of TPO. This study was conducted to investigate whether NOD/SCID mice exposed to high TPO levels Downloaded from mediated by a retroviral vector would be refractory to the development of bone marrow fibrosis. We show that TPO and TGF-1 in plasma from NOD/SCID and SCID mice engrafted with TPO-overexpressing hemopoietic cells reach levels similar to the ones reached in immunocompetent mice, and all animals develop a myeloproliferative disease associated with a dense myelofibrosis at 8 wk posttransplantation. Monocytes in NOD/SCID mice are functionally deficient to secrete cytokines such as IL-1␣ in response to stimuli, even under TPO expression. Surprisingly, the plasma of these mice displays high levels of IL-␣, which was demonstrated to originate from platelets. Together, these data suggest that completely functional monocytes are not required to develop my- http://www.jimmunol.org/ elofibrosis and that platelets are able, under TPO stimulation, to synthesize inflammatory cytokines, which may be involved in the pathogenesis of myelofibrosis and osteosclerosis. The Journal of Immunology, 2006, 176: 6425–6433. ibrosis is a prominent clinical complication in several dis- proteases that inhibit enzymes involved in the degradation of the orders and occurs particularly in the lung, kidney, heart, or extracellular matrix. TGF-1 is secreted by numerous cell lin- F liver. In human hemopoietic tissues, myelofibrosis is less eages, but mainly by monocytes/macrophages and megakaryocytes frequent but is typically associated with two distinct clinical enti- (MKs)/platelets. Several lines of evidence obtained both from 4 ties: chronic idiopathic myelofibrosis (IM) with myeloid metapla- studies of patients with IM (5, 6) or murine models ending with sia and acute panmyelosis with myelofibrosis (1). Bone marrow myelofibrosis (7–9) are in favor of a crucial role of MK in my- by guest on September 28, 2021 fibrosis occurs as a cytokine-mediated secondary response to a elofibrosis induction. However, other studies suggested a central clonal malignant event originating in a multipotent hemopoietic role for the monocyte/macrophage lineage. This theory is based on stem cell (2, 3) and is characterized by excessive deposits of ex- two different observations: 1) Rameshwar et al. (10) reported that tracellular matrix proteins (4). In vivo and in vitro studies have monocytes from patients with IM are spontaneously activated and involved several cytokines in the aberrant stromal reaction, with a secreted abnormally TGF-1; and 2) Frey et al. (11) reported that strong emphasis on the pleiotropic cytokine TGF-1. Through fi- thrombopoietin (TPO) overexpression in SCID mice led to myelo- broblast stimulation, this cytokine is able to produce extracellular fibrosis, but not in NOD/SCID mice. Because NOD/SCID mice matrix, induce cell adhesion proteins, and enhance expression of differ from SCID mice by impaired mononuclear phagocyte func- tions (12, 13), the authors postulated that monocytes and macro- *Institut National de la Sante et de la Recherche Medicale (INSERM) U790, Pavillon phages must be required for promotion of myelofibrosis in TPO- de Recherche 1, Institut Gustave Roussy, Villejuif, France; †Laboratoire overexpressing mice. However, these experiments were performed ‡ d’He´matologie, Hoˆpital Henri Mondor, Cre´teil, France; Unite´Immunophysiologie et with a human TPO cDNA vectorized in an adenovirus construct Parasitisme Intracellulaire, Institut Pasteur, Paris, France; and §Laboratoire d’Anatomopathologie, Hoˆpital Cochin, Paris, France that leads to a relatively low elevation of plasma TPO. In the Received for publication September 29, 2005. Accepted for publication March present study, to ensure a high and sustained TPO level, we used 2, 2006. a retroviral vector encoding a mouse TPO cDNA to overexpress The costs of publication of this article were defrayed in part by the payment of page TPO in NOD/SCID, SCID, and immunocompetent wild-type charges. This article must therefore be hereby marked advertisement in accordance high with 18 U.S.C. Section 1734 solely to indicate this fact. (WT) mice. All TPO mice invariably developed a myelopro- 1 This work was supported by grants from INSERM and La Ligue Nationale contre liferative syndrome associated with a myelofibrosis and osteoscle- le Cancer E´ quipe Labellise´e 2000, fellowships from Association pour la Recherche rosis. At any time during the follow-up, TPO and TGF-1 plasma sur la Cancer and INSERM (Poste d’Accueil) (to O.W.-B.), and the Research Min- levels were substantially and similarly elevated in the three groups istry (to H.C.). of mice. Our data provide evidence that the monocyte/macrophage 2 H.C. and E.P. contributed equally to this work. lineage does not play a crucial role in the development of the 3 Address correspondence and reprint requests to Dr. Ste´phane Giraudier, Hoˆpital Henri Mondor, Laboratoire d’He´matologie, 51 Avenue du Mare´chal de Lattre de myelofibrosis induced by TPO. In addition, platelets and MKs Tassigny, 94000 Cre´teil, France. E-mail address: [email protected] when stimulated are able to synthesize IL-1␣ and thus to replace paris.fr or [email protected] monocytes in the synthesis of inflammatory cytokines, which 4 Abbreviations used in this paper: IM, idiopathic myelofibrosis; MK, megakaryocyte; TPO, thrombopoietin; WT, wild type; CFC, colony-forming cells; FISH, fluorescent may be involved in the pathogenesis of myelofibrosis and in situ hybridization. osteosclerosis. Copyright © 2006 by The American Association of Immunologists, Inc. 0022-1767/06/$02.00 6426 MYELOFIBROSIS IN TPOhigh NOD/SCID MICE Materials and Methods pressed as the percentage of apoptotic MKs among the total MK Mice population. All procedures were approved by the local Institut Gustave Roussy ethics Production of bone marrow-derived monocytes committee. WT C57BL/6J mice (Janvier) were maintained at the Institute Bone marrow cells were harvested from femurs and tibia of age- and sex- Gustave Roussy animal facility under specific pathogen-free conditions. high SCID mutant mice (C57BL/6J-scid/scid) (14) (Charles River Laboratories) matched control or TPO mice (WT and NOD/SCID). Cells were cul- ϫ 6 and double NOD/SCID mutant mice (15) (NOD/LtSz-scid/scid obtained tured (1.6 10 cells/ml), as described previously (18), in DMEM sup- from J. Dick (University Health Network, Toronto, Ontario, Canada)) were plemented with antibiotics, 10% FBS, and 10% CSF-1-conditioned bred under sterile conditions and maintained in isolators. All mice used in medium (provided by M. Lebastard, Unite´Immunophysiologie et Parasit- this study have the same C57BL/6J genetic background. isme Intracellulaire, Institut Pasteur, Paris, France) with and without 10 U/ml IFN-␥. Bone marrow cells were cultured at 37°C in a humidified Infection procedure incubator (5% CO2) with an addition of fresh medium at day 3. After 5 days in culture, nonadherent cells were removed and centrifuged for 10 min Six- to 8-wk-old WT, SCID, or NOD/SCID male mice were used as bone at 400 ϫ g. Pellets were suspended in DMEM with antibiotics and 10% marrow donors. Seven- to 10-wk-old WT, SCID, or NOD/SCID female FBS by flushing through, successively, 25-, 27-, and 30-gauge needles, and mice were recipients. The infection procedure was performed as described single-cell suspensions were produced. Cells (2 ϫ 105 cells/ml) were previously (7, 16). Briefly, 4 days after 5-fluorouracil treatment (one in- seeded in fresh medium alone (DMEM supplemented with antibiotics, 10% jection of 150 mg/kg administered i.p.), total bone marrow cells were FBS, and 10% CSF-1-conditioned medium) or in fresh medium containing cocultured with 1 ϫ 106 MPZenTPO virus-producing GPϩE-86 cells in Escherichia coli LPS at 10 g/ml. After a 24-h incubation period, culture DMEM supplemented with antibiotics, 20% FBS and murine IL-3 (100 supernatants were harvested and used for determination of IL-1␣ level.