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Immune Complex–Driven Generation of Human Macrophages with Anti-Inflammatory and Growth-Promoting Activity

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Immune Complex–Driven Generation of Human Macrophages with Anti-Inflammatory and Growth-Promoting Activity Immune Complex−Driven Generation of Human Macrophages with Anti-Inflammatory and Growth-Promoting Activity This information is current as of September 30, 2021. Elizabeth Dalby, Stephen M. Christensen, Jingya Wang, Kajal Hamidzadeh, Prabha Chandrasekaran, V. Keith Hughitt, Wagner Luiz Tafuri, Rosa Maria Esteves Arantes, Ismael Alves Rodrigues, Ronald Herbst, Najib M. El-Sayed, Gary P. Sims and David M. Mosser Downloaded from J Immunol published online 20 May 2020 http://www.jimmunol.org/content/early/2020/05/19/jimmun ol.1901382 http://www.jimmunol.org/ Supplementary http://www.jimmunol.org/content/suppl/2020/05/19/jimmunol.190138 Material 2.DCSupplemental Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision by guest on September 30, 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 © 2020 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. Published May 20, 2020, doi:10.4049/jimmunol.1901382 The Journal of Immunology Immune Complex–Driven Generation of Human Macrophages with Anti-Inflammatory and Growth-Promoting Activity Elizabeth Dalby,*,1 Stephen M. Christensen,*,1 Jingya Wang,† Kajal Hamidzadeh,* Prabha Chandrasekaran,* V. Keith Hughitt,*,‡ Wagner Luiz Tafuri,x Rosa Maria Esteves Arantes,x Ismael Alves Rodrigues,{ Ronald Herbst,† Najib M. El-Sayed,*,‡ Gary P. Sims,† and David M. Mosser* To maintain homeostasis, macrophages must be capable of assuming either an inflammatory or an anti-inflammatory phenotype. To better understand the latter, we stimulated human macrophages in vitro with TLR ligands in the presence of high-density immune Downloaded from complexes (IC). This combination of stimuli resulted in a broad suppression of inflammatory mediators and an upregulation of molecules involved in tissue remodeling and angiogenesis. Transcriptomic analysis of TLR stimulation in the presence of IC pre- dicted the downstream activation of AKT and the inhibition of GSK3. Consequently, we pretreated LPS-stimulated human mac- rophages with small molecule inhibitors of GSK3 to partially phenocopy the regulatory effects of stimulation in the presence of IC. The upregulation of DC-STAMP and matrix metalloproteases was observed on these cells and may represent potential biomarkers for this regulatory activation state. To demonstrate the presence of these anti-inflammatory, growth-promoting macrophages in a http://www.jimmunol.org/ human infectious disease, biopsies from patients with leprosy (Hanseniasis) were analyzed. The lepromatous form of this disease is characterized by hypergammaglobulinemia and defective cell-mediated immunity. Lesions in lepromatous leprosy contained mac- rophages with a regulatory phenotype expressing higher levels of DC-STAMP and lower levels of IL-12, relative to macrophages in tuberculoid leprosy lesions. Therefore, we propose that increased signaling by FcgR cross-linking on TLR-stimulated macrophages can paradoxically promote the resolution of inflammation and initiate processes critical to tissue growth and repair. It can also contribute to infectious disease progression. The Journal of Immunology, 2020, 205: 000–000. he phenotypic changes that macrophages undergo sub- production (2–7). We hypothesized that these cells may play a sequent to their encounter with pathogen-associated mo- role in dampening inflammation during the resolution of immune by guest on September 30, 2021 T lecular patterns have been well described (1). The cell responses. surface receptors and downstream signaling molecules that acti- The first reprogramming stimulus identified in the murine system vate transcription factors to drive inflammatory responses have was immune complexes (IC) that signal through the macrophage been thoroughly studied. These so-called M1 (inflammatory) mac- FcgR (6). In macrophages, IC induce the clustering of stimulatory rophages make important contributions to the initiation of immune FcgR, resulting in the phosphorylation of Syk (8, 9). This initiates responses, but their myriad inflammatory secretory products can a powerful signaling cascade, leading to actin remodeling, PI3K cause tissue damage when left uncontrolled. Working in the murine activation, and Ca+ mobilization (10). The most widely studied system, our laboratory characterized a population of regulatory consequences of these signaling pathways in macrophages are the macrophages that were generated by TLR stimulation in the induction of phagocytosis and the generation of the respiratory presence of a second “reprogramming” stimulus. Paradoxically, burst, both important components of host defense. We previously the increased signaling strength through the dual stimuli resulted demonstrated that FcgR ligation on TLR-stimulated murine macro- in reduced inflammatory and increased anti-inflammatory cytokine phages can also result in the rapid and prolonged hyperphosphorylation *Department of Cell Biology and Molecular Genetics, University of Maryland, S.M.C., N.M.E.-S., G.P.S., and D.M.M. conceptualized and designed the research; College Park, MD 20742; †Department of Respiratory, Inflammation, and Autoimmu- E.D., S.M.C., N.M.E.-S., and D.M.M. wrote the paper. nity, AstraZeneca, Gaithersburg, MD 20878; ‡Center for Bioinformatics and Computa- x Address correspondence and reprint requests to Dr. Gary P. Sims or Dr. David M. tional Biology, University of Maryland, College Park, MD 20742; Department of Mosser, Department of Respiratory, Inflammation, and Autoimmunity, AstraZeneca, General Pathology, Federal University of Minas Gerais, Belo Horizonte 31270-901, { One MedImmune Way, Gaithersburg, MD 20878 (G.P.S.) or Department of Cell Biol- Brazil; and Instituto Metropolitano de Ensino Superior, Ipatinga 35164-253, Brazil ogy and Molecular Genetics, University of Maryland, 3102 Bioscience Research Build- 1E.D. and S.M.C. contributed equally to this work. ing, College Park, MD 20742 (D.M.M.). E-mail addresses: [email protected] (G.P.S.) or [email protected] (D.M.M.) ORCIDs: 0000-0003-1343-7255 (P.C.); 0000-0003-0787-9559 (V.K.H.); 0000-0001- 7970-3312 (N.M.E.-S.); 0000-0003-2796-0485 (G.P.S.); 0000-0002-9503-4187 The online version of this article contains supplemental material. (D.M.M.). Abbreviations used in this article: DC-STAMP, dendritic cell–specific transmembrane Received for publication November 26, 2019. Accepted for publication April 22, protein; DTH, delayed-type hypersensitivity; FC, fold change; HKLM, heat-killed 2020. Listeria monocytogenes; IC, immune complex; IPA, Ingenuity Pathway Analysis; LPS + IC, LPS in the presence of high-density IC; LPS + IC-Mf, macrophages This work was supported in part by National Institutes of Health National Institute of stimulated with LPS in the presence of high-density IC; MMP, matrix metallopro- Allergy and Infectious Diseases Grant NIH R01 GM 102589. tease; Poly I:C, polyinosinic-polycytidylic acid; RNA-seq, RNA sequencing. E.D., S.M.C., J.W., P.C., K.H., W.L.T., R.M.E.A., and I.A.R. performed experiments; E.D., S.M.C., J.W., P.C., K.H., V.K.H., W.L.T., N.M.E.-S., R.H., G.P.S., and D.M.M. Copyright Ó 2020 by The American Association of Immunologists, Inc. 0022-1767/20/$37.50 analyzed results; E.D., S.M.C., P.C., K.H., and D.M.M. constructed the figures; E.D., www.jimmunol.org/cgi/doi/10.4049/jimmunol.1901382 2 IDENTIFICATION OF HUMAN IMMUNOREGULATORY MACROPHAGES of ERK, triggering chromatin remodeling at the IL-10 promoter, the University of California, Santa Cruz genome browser (20) (http:// leading to increased IL-10 transcription (11). genome.ucsc.edu) using TopHat (v 2.0.13) (21), with parameters matching A similar regulatory phenotype has not been described in previous work (22). The abundance of reads mapped to coding features was determined using HTSeq (23). Quantile normalization and log2 human macrophages, and no biomarkers exist to identify these transformation was applied to all samples (24). Limma was used to cells. Furthermore, our understanding of how to manipulate this conduct differential expression analyses (25). The voom module was activation state is limited. In this study, we use high-throughput used to transform the databased on observational level weights derived approaches to characterize human regulatory macrophages from the mean-variance relationship prior to statistical modeling. Ex- perimental batch effects were adjusted for by including experimental generated in response to IC (R-Mf-IC) with the aim of iden- batch as a covariate in our statistical model. Differentially expressed tifying biomarkers that could be used for their identification genesweredefinedasgeneswithalog2 fold change (FC) .1and in tissue. By providing a global picture of the behavior and Benjamini–Hochberg multiple-testing adjusted p value ,0.05. All function of these human macrophages, our studies revealed that components of the statistical pipeline, named cbcbSEQ, can be accessed IC can dampen inflammatory responses and inhibit
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