The TRPM4 Channel Controls Monocyte and Macrophage, but Not Neutrophil, Function for Survival in Sepsis This information is current as Nicolas Serafini, Albert Dahdah, Gaëtan Barbet, Marie of September 29, 2021. Demion, Tarik Attout, Grégory Gautier, Michelle Arcos-Fajardo, Hervé Souchet, Marie-Hélène Jouvin, François Vrtovsnik, Jean-Pierre Kinet, Marc Benhamou, Renato C. Monteiro and Pierre Launay J Immunol published online 29 August 2012 Downloaded from http://www.jimmunol.org/content/early/2012/08/29/jimmun ol.1102969 Supplementary http://www.jimmunol.org/content/suppl/2012/08/29/jimmunol.110296 http://www.jimmunol.org/ Material 9.DC1 Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! 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Published August 29, 2012, doi:10.4049/jimmunol.1102969 The Journal of Immunology The TRPM4 Channel Controls Monocyte and Macrophage, but Not Neutrophil, Function for Survival in Sepsis Nicolas Serafini,*,†,‡ Albert Dahdah,*,†,‡,1 Gae¨tan Barbet,*,†,‡,1 Marie Demion,*,†,‡ Tarik Attout,*,†,‡ Gre´gory Gautier,*,†,‡ Michelle Arcos-Fajardo,*,†,‡ Herve´ Souchet,*,†,‡ Marie-He´le`ne Jouvin,x Franc¸ois Vrtovsnik,*,‡,{ Jean-Pierre Kinet,x Marc Benhamou,*,‡ Renato C. Monteiro,*,‡ and Pierre Launay*,†,‡,{ A favorable outcome following acute bacterial infection depends on the ability of phagocytic cells to be recruited and properly activated within injured tissues. Calcium (Ca2+) is a ubiquitous second messenger implicated in the functions of many cells, but the mechanisms involved in the regulation of Ca2+ mobilization in hematopoietic cells are largely unknown. The monovalent 2+ cation channel transient receptor potential melastatin (TRPM) 4 is involved in the control of Ca signaling in some hematopoietic Downloaded from cell types, but the role of this channel in phagocytes and its relevance in the control of inflammation remain unexplored. In this study, we report that the ablation of the Trpm4 gene dramatically increased mouse mortality in a model of sepsis induced by cecal ligation and puncture. The lack of the TRPM4 channel affected macrophage population within bacteria-infected peritoneal cavities and increased the systemic level of Ly6C+ monocytes and proinflammatory cytokine production. Impaired Ca2+ mobili- zation in Trpm42/2 macrophages downregulated the AKT signaling pathway and the subsequent phagocytic activity, resulting in 2+ bacterial overgrowth and translocation to the bloodstream. In contrast, no alteration in the distribution, function, or Ca http://www.jimmunol.org/ mobilization of Trpm42/2 neutrophils was observed, indicating that the mechanism controlling Ca2+ signaling differs among phagocytes. Our results thus show that the tight control of Ca2+ influx by the TRPM4 channel is critical for the proper functioning of monocytes/macrophages and the efficiency of the subsequent response to infection. The Journal of Immunology, 2012, 189: 000–000. epsis, which is a major cause of mortality worldwide, is neutrophils in the control of septic peritonitis remain unclear. In characterized by excessive inflammation in response to the circulation, monocytes show functional heterogeneity and are infection (1). During septic peritonitis, neutrophil recruit- distinguished based on their expression of specific surface mark- S by guest on September 29, 2021 ment within the peritoneal cavity is followed by the extravasation ers. In mice, two main subsets have been characterized, as follows: of monocytes, which differentiate into inflammatory macrophages patrolling (or Ly6C2) monocytes (5, 6) and inflammatory (or (2). Macrophages play a major role in the maintenance of tissue Ly6C+) monocytes (7). Ly6C+ monocytes contribute to the control homeostasis as they contribute, together with neutrophils, to the of pathogen proliferation during infection (8). In contrast, Ly6C2 clearance of microorganisms and dying cells (3). In septic monocytes are believed to be involved in healing and tissue repair patients, macrophage dysregulation has been associated with ad- (9). Although macrophages and monocytes are important for the verse prognosis (4). However, the mechanisms regulating the appropriate response to infection, their regulation in the context of functioning of macrophages and their role compared with that of bacteria-mediated inflammatory reactions remains to be fully understood. One potential mechanism involved in the regulation of these † *INSERM U699, Paris F-75018, France; Equipe Avenir INSERM, Paris F-75018, myeloid cells is the control of their intracellular Ca2+ concentra- France; ‡Universite´ Paris 7-Denis Diderot, Faculte´ de Me´decine, Site Xavier Bichat, x 2+ 2+ Paris F-75018, France; Department of Pathology, Beth Israel Deaconess Medical tion ([Ca ]i). Intracellular Ca governs a wide range of cellular Center, Harvard Medical School, Boston, MA 02215; and {Assistance Publique- functions, such as proliferation, cytokine secretion, phagocytosis, Hoˆpitaux de Paris, Hoˆpital Bichat, Service de Ne´phrologie, Paris F-75018, France and apoptosis (10, 11). Ca2+ mobilization through Ca2+ release- 1 A.D. and G.B. contributed equally to this work. activated Ca2+ (CRAC) channels has been observed in macro- Received for publication October 13, 2011. Accepted for publication August 1, 2012. phages (12, 13) and neutrophils (14), and seems to be a key de- This work was supported by grants to P.L. from Fondation pour la Recherche Me´d- terminant for the activation of these phagocytes. However, the icale (FRM) and Agence Nationale de la Recherche (ANR-Genopath). N.S. was mechanisms regulating Ca2+ homeostasis in these innate immunity supported by Ministe`re de l’Enseignement Supe´rieur et de la Recherche, Universite´ Paris 7, and FRM. A.D. was supported by ANR. G.B. was supported by la Ligue cells are largely unknown. Recently, the protein ORAI1 (15-17) Nationale Contre le Cancer; G.G. was supported by FRM. and its activator STIM1 (18, 19) have been identified as essential Address correspondence and reprint requests to Dr. Pierre Launay, Equipe Avenir, components of the CRAC channel, whose activity is regulated at INSERM U699, Faculte´ de Me´decine X. Bichat, Paris F-75018, France. E-mail multiple levels (20–23). address: [email protected] Two Ca2+-activated nonselective (CAN) channels, transient The online version of this article contains supplemental material. receptor potential melastatin (TRPM) 4 and TRPM5, have been Abbreviations used in this article: BMDC, bone marrow-derived dendritic cell; BMM, bone marrow-derived macrophage; [Ca2+] , intracellular Ca2+ concentration; characterized in nonexcitable cells (24, 25). These channels are i 2+ CAN, Ca2+-activated nonselective; CLP, cecal ligation and puncture; CRAC, Ca2+ involved in the regulation of Ca homeostasis by reducing the 2+ release-activated Ca ; DC, dendritic cell; NMDG, N-methyl-D-glucamine; PI, pro- driving force of Ca2+ influx through the CRAC channel (26). In- pidium iodide; TRPM, transient receptor potential melastatin. deed, under physiological conditions, the opening of TRPM4 by 2+ + Copyright Ó 2012 by The American Association of Immunologists, Inc. 0022-1767/12/$16.00 high [Ca ]i allows massive sodium (Na ) entry and membrane www.jimmunol.org/cgi/doi/10.4049/jimmunol.1102969 2 TRPM4 REGULATES MACROPHAGE-MEDIATED INFLAMMATION depolarization, thereby decreasing further Ca2+ influx (26). TRPM4 gentle massage of peritoneum. Peritoneal fluids were collected using a 21-g 2+ needle attached to a 10-ml syringe and then passed through a 100 mm filter thus acts in concert with the CRAC channel to control [Ca ]i and subsequent physiological responses in T cells, mast cells, and and washed. Cells were labeled at 4˚C in PBS with 0.5% BSA. Fcg receptors were blocked with the 2.4G2 mAb before cell staining for specific surface dendritic cells (DCs) (26–28). The molecular targeting of the markers (15 min). Surface staining was performed with fluorochrome- Trpm4 gene in the mouse does not result in perinatal death or in conjugated mAbs to mouse Ags, as follows: CD3 (145-2C11), CD19 an overt spontaneous phenotype. TRPM4-deficient mice are thus (1D3), Siglec-F (E50-2440), CD4 (L3T4), CD8 (53-6-7), IA/IE (M5/ valuable tools for the detailed study of the impact of Ca2+ influx 114.15.2), CD11b (M1/70), Gr1 (RB6-8C5), Ly-6G (1A8), and Ly-6C (AL21; all from BD Biosciences); CD115 (AFS98; eBioscience); and on cellular functions under pathological conditions. F4/80 (C1:A3-1; Serotec). The viability of the cells was assessed using the 2+ In this study, we addressed the importance of Ca regulation in LIVE/DEAD Aqua cell stain (Molecular Probes), and neutrophils were the control of the acute inflammatory reaction induced by cecal gated as population with both high side and high forward light scatter.