Chronic Exposure to Glucocorticoids Shapes Gene Expression and Modulates Innate and Adaptive Activation Pathways in Macrophages with Distinct Changes in This information is current as Leukocyte Attraction of September 29, 2021. Martijn D. B. van de Garde, Fernando O. Martinez, Barbro N. Melgert, Machteld N. Hylkema, René E. Jonkers and Jörg Hamann J Immunol published online 6 January 2014 Downloaded from http://www.jimmunol.org/content/early/2014/01/05/jimmun ol.1302138 http://www.jimmunol.org/ Supplementary http://www.jimmunol.org/content/suppl/2014/01/05/jimmunol.130213 Material 8.DCSupplemental Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision by guest on September 29, 2021 • No Triage! Every submission reviewed by practicing scientists • 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 © 2014 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. Published January 6, 2014, doi:10.4049/jimmunol.1302138 The Journal of Immunology Chronic Exposure to Glucocorticoids Shapes Gene Expression and Modulates Innate and Adaptive Activation Pathways in Macrophages with Distinct Changes in Leukocyte Attraction Martijn D. B. van de Garde,*,1 Fernando O. Martinez,†,1 Barbro N. Melgert,‡,x || Machteld N. Hylkema,x,{ Rene´ E. Jonkers, and Jo¨rg Hamann* Glucocorticoids (GCs) have been used for more than 50 y as immunosuppressive drugs, yet their efficacy in macrophage-dominated disorders, such as chronic obstructive pulmonary disease, is debated. Little is known how long-term GC treatment affects macrophage responses in inflammatory conditions. In this study, we compared the transcriptome of human macrophages, matured in the presence or absence of fluticasone propionate (FP), and their ability to initiate or sustain classical activation, mimicked using acute LPS and chronic IFN-g stimulation, respectively. We identified macrophage gene expression networks, modulated by FP long-term exposure, Downloaded from and specific patterns of IFN-g– and LPS-induced genes that were resistant, inhibited, or exacerbated by FP. Results suggest that long-term treatment with GCs weakens adaptive immune signature components of IFN-g and LPS gene profiles by downmodulating MHC class II and costimulatory molecules, but strengthens innate signature components by maintaining and increasing expression of chemokines involved in phagocyte attraction. In a mouse model of chronic obstructive pulmonary disease, GC treatment induced higher chemokine levels, and this correlated with enhanced recruitment of leukocytes. Thus, GCs do not generally suppress macrophage effector functions, but they cause a shift in the innate–adaptive balance of the immune response, with distinct changes http://www.jimmunol.org/ in the chemokine–chemokine receptor network. The Journal of Immunology, 2014, 192: 000–000. acrophages are innate immune cells with well-established of a developing concept: macrophage plasticity (3). Plasticity is roles in tissue homeostasis, primary response to patho- naturally the basis of macrophage heterogeneity in basal and M gens, coordination of adaptive immunity, and even wound inflammatory conditions; it is also the basis of worldwide efforts repair (1, 2). Macrophages accomplish these varied roles by adapting to treat diseases by subverting aberrant macrophage activation. their gene and protein expression programs in response to endoge- Macrophage-mediated inflammation is increasingly recognized nous and exogenous environmental cues, such as cytokines and as contributing to chronic inflammatory disorders, such as chronic pathogen-associated molecular patterns. The ability of macrophages obstructive pulmonary disease (COPD), severe asthma, rheumatoid by guest on September 29, 2021 to change their gene and protein signatures falls under the umbrella arthritis, and multiple sclerosis (4). The best-characterized macrophage activation pathway has been called classical macrophage activation, and it is triggered by *Department of Experimental Immunology, Academic Medical Center, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands; †Sir William Dunn School of the recognition of pathogen-associated molecular patterns by TLRs, Pathology and Kennedy Institute of Rheumatology, University of Oxford, Oxford supported by further elicitation of IFN pathways (2). Classically ‡ OX1 3RE, United Kingdom; Department of Pharmacokinetics, Toxicology and activated macrophages produce microbicidal enzymes, such as Targeting, Groningen Research Institute for Pharmacy, University of Groningen, 9713 AV Groningen, The Netherlands; xGroningen Research Institute for Asthma inducible NO synthase, inflammatory cytokines, such as TNF, IL-6, and COPD, University Medical Center Groningen, University of Groningen, IL-1b, IL-12, and IFN-b, various chemoattractants, and matrix 9713 GZ Groningen, The Netherlands; {Department of Pathology, University Medical Center Groningen, University of Groningen, 9713 GZ Groningen, The metallopeptidases (MMPs). These mediators support IFN-g–mediated Netherlands; and ||Department of Pulmonology, Academic Medical Center, University Th1 responses, which are important for sustained antimicrobial of Amsterdam, 1105 AZ Amsterdam, The Netherlands activity but are deleterious in chronic diseases, because they lead to 1 M.D.B.v.d.G. and F.O.M. contributed equally to this work. tissue damage and chronic inflammation. Received for publication August 14, 2013. Accepted for publication November 25, To limit macrophage activation and restore homeostasis, endog- 2013. enous glucocorticoids (GCs) induce anti-inflammatory programs via This work was supported by a grant from the J.K. de Cock Stichting and Prof. Dirkje Postma through a Spinoza grant from the Dutch Government. the GC receptor (GCR) (5, 6). GCR agonists are prescribed for a wide range of inflammatory disorders, and their use is crucial to Address correspondence and reprint requests to Dr. Fernando O. Martinez or Dr. Jo¨rg Hamann, Kennedy Rheumatology Institute, Nuffield Department of Orthopaedics, treat diseases of the respiratory tract, especially asthma (6). Drugs Rheumatology and Musculoskeletal Sciences, Botnar Research Centre, University such as prednisolone, dexamethasone, budesonide, and fluticasone of Oxford, Windmill Road, Headington, Oxford, OX37 LD, U.K. (F.O.M.) or De- partment of Experimental Immunology, Academic Medical Center, University of propionate (FP) share great GCR specificity albeit different phar- Amsterdam, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands (J.H.). E-mail macodynamics (7). The remarkable efficacy of GCs in suppressing addresses: [email protected] (F.O.M.) or j.hamann@amc. uva.nl (J.H.) inflammation and decreasing lymphocyte activation, proliferation, The online version of this article contains supplemental material. and survival is well appreciated. However, our knowledge of the interactions of GC and inflammatory pathways in human mac- Abbreviations used in this article: COPD, chronic obstructive pulmonary disease; FP, fluticasone propionate; GC, glucocorticoid; GCR, GC receptor; IPA, Ingenuity Path- rophages remains limited (8), and recent studies have shown that GCs ways Analysis; MDM, monocyte-derived macrophage; MEV, Multiple Experiment may have little effect in controlling inflammation in macrophage- Viewer; MMP, matrix metalloproteinase. dominated diseases, and their long-term use is associated with Copyright Ó 2014 by The American Association of Immunologists, Inc. 0022-1767/14/$16.00 persisting complications (9–16). www.jimmunol.org/cgi/doi/10.4049/jimmunol.1302138 2 GLUCOCORTICOID ACTION ON MACROPHAGES In this study, we investigated from a whole genome point of view study we focused our analysis on data generated on human cells backed the effects of chronic GCR ligation on the maturation of human up by experimental evidence. Data compliant for Minimum Information macrophages and on their ability to initiate and sustain classical About a Microarray Experiment for the datasets used in this study are deposited in Gene Expression Omnibus (www.ncbi.nlm.nih.gov/geo/) pro- activation that is important for Th1 inflammation, dissected by files dataset GSE49852. This dataset is part of a superseries—GSE35495— acute and chronic stimulation with the TLR4 ligand LPS and IFN-g. that covers microarrays over the spectrum of macrophage activation, in- We show that long-term treatment with FP modulates classical cluding IFN-g, IL-4, IL-10, IL-13, and dexamethasone (18). activation in human inflammatory macrophage models in a com- For comparison, CEL files generated by Ehrchen et al. (19) in a microarray analysis of monocytes stimulated with FP for 16 h were summarized as plex and not solely suppressive manner by maintaining or even Robust Multiarray Averaging, and quantiles were normalized using Affy increasing the expression of chemokines involved in leukocyte package, R. recruitment, while clearly affecting their ability to initiate adaptive