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Soluble IL-7Rα (sCD127) Inhibits IL-7 Activity and Is Increased in HIV Infection Angela M. Crawley, Sylvie Faucher and Jonathan B. Angel This information is current as J Immunol 2010; 184:4679-4687; Prepublished online 19 of September 25, 2021. March 2010; doi: 10.4049/jimmunol.0903758 http://www.jimmunol.org/content/184/9/4679 Downloaded from References This article cites 51 articles, 19 of which you can access for free at: http://www.jimmunol.org/content/184/9/4679.full#ref-list-1 Why The JI? Submit online. http://www.jimmunol.org/ • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication *average by guest on September 25, 2021 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 © 2010 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Soluble IL-7Ra (sCD127) Inhibits IL-7 Activity and Is Increased in HIV Infection Angela M. Crawley,*,† Sylvie Faucher,‡ and Jonathan B. Angel*,†,x Soluble CD127 (sCD127) appears to play an important role in the immunopathogenesis of several chronic infections, multiple scle- rosis, and various cancers. The function of sCD127 and whether it influences IL-7 bioavailability or activity is unknown. In this study, we demonstrated that recombinant and native sources of sCD127 significantly inhibited IL-7–mediated STAT5 and Akt phosphorylation in CD8+ T cells. IL-7–mediated proliferation and Bcl-2 expression were similarly reduced by sCD127. In each case, native sCD127 inhibited IL-7 activity to a greater degree than rsCD127. Anti–IL-7 activity was inherent to human plasma and could be reversed by depletion of CD127, revealing for the first time the biological activity of naturally occurring sCD127. Plasma sCD127 concentrations were increased in HIV+ individuals compared with HIV2 controls, correlated with IL-7 levels, and + remained unchanged in HIV individuals following 1 y of effective antiretroviral therapy. Determining the regulation and function Downloaded from of sCD127 may be critical for understanding both the pathogenesis of diseases in which IL-7 likely has a role (e.g., HIV infection, cancer) and its potential impact on IL-7 as a therapeutic approach. The Journal of Immunology, 2010, 184: 4679–4687. t has recently been suggested that soluble CD127 (sCD127) modulating activity of their cytokine ligands, soluble cytokine plays a role in the immunopathogenesis of several diseases receptors have been shown to be of prognostic value in a number I such as multiple sclerosis (MS), acute lymphoblastic leuke- of diseases and have also been developed as therapeutic agents in http://www.jimmunol.org/ mia, and HIV infection (1–3). Genetic evidence has revealed an the treatment of a number of inflammatory diseases (5). To date, association between soluble isoforms of CD127 and chronic a number of soluble cytokine receptors have been identified, and progressive MS, and increased levels of sCD127 transcripts have their mechanisms of release, unique functions, and putative been documented in children with acute lymphoblastic leukemia. pathophysiological roles have been described (4, 5). The first In HIV infection, individuals lacking HIV-specific cytotoxic ac- identified soluble cytokine receptor was IL-2Ra (sIL-2Ra), and it tivity have increased concentrations of sCD127 (3). Although has been associated with leukemia disease progression and is increased levels of sCD127 have been detected in a number of a marker of poor prognosis in children with Hodgkin’s disease (6). disease states and have been associated with disease activity, the The IL-2Ra gene region, a known autoimmune susceptibility lo- biological function of sCD127 and its clinical relevance remains cus, possesses significant allelic heterogeneity related to either by guest on September 25, 2021 to be established. susceptibility and risk to MS and type 1 diabetes, and these allelic Soluble cytokine receptors have the ability to influence the variants independently correlate with plasma sIL-2Ra concen- extracellular bioavailability of their ligand and have been shown to trations (7). Antagonistic soluble receptors including TNFR I and play important roles in several aspects of the immune response II can inhibit TNF signaling and have been shown to prevent TNF- including inflammation, cellular proliferation, and apoptosis. In mediated tumor lysis (8). The IL-6R subunit gp130 has also been many cases, soluble cytokine receptors act as competitive inhib- shown to inhibit the proinflammatory activity of IL-6 (9). sIL- itors, or, alternatively, they can enhance the activity of the cytokine. 15Ra is an example of an agonistic cytokine receptor. Pre- The former effect has led to the idea that the release of soluble complexed IL-15–sIL-15Ra increases NK and CD8+ T cell pro- cytokine receptors is a physiological mechanism for regulating the liferation and antitumor activity up to 50 times compared with IL- activity of cytokines, as production of the soluble receptor often 15 alone (10, 11). parallels synthesis of the ligand (4). In addition to their role in The biological role of sCD127 is not yet known, although it is plausible that sCD127 could bind circulating IL-7, thereby de- creasing IL-7 bioavailability. In HIV infection, there may be causal *Ottawa Hospital Research Institute; †Department of Biochemistry, Microbiology and Immunology, University of Ottawa; ‡National HIV Immunology Laboratory, National links between the increase in plasma IL-7 levels and higher con- HIV and Retrovirology Labs, Centre for Infectious Disease Prevention and Control centrations of sCD127. Therefore, we have investigated the impact x Health Canada; and Division of Infectious Diseases, Ottawa Hospital-General Campus, of sCD127 on IL-7 activity in human CD8+ T cells and quantified Ottawa, Ontario, Canada plasma sCD127 concentration in HIV infection to better un- Received for publication November 23, 2009. Accepted for publication February 20, 2010. derstand its role in immunopathogenesis. This work was supported by Grant ROGB131 from the Ontario HIV Treatment Network (OHTN), Grant HOP84649 from the Canadian Institutes of Health Re- Materials and Methods search, and Canadian Foundation for AIDS Research Grant 019014. A.M.C. is a re- Sample collection and cell isolation cipient of an OHTN fellowship and J.B.A. is an OHTN Career Scientist. Address correspondence and reprint requests to Dr. Jonathan B. Angel, Division of All research conducted using blood from human subjects was approved by Infectious Diseases, Ottawa Hospital-General Campus, 501 Smyth Road, Room the Ottawa Hospital and Ottawa Hospital Research Institute’s Research G-12, Ottawa, ON K1H 8L6, Canada. E-mail address: [email protected] Ethics Board (Ottawa, Ontario, Canada). Plasma was collected from Fi- coll-Paque gradients of heparinized blood from both HIV2 and HIV+ in- Abbreviations used in this paper: MFI, mean fluorescence intensity; MS, multiple 2 + sclerosis; pSTAT5, phosphorylated STAT5; sCD127, soluble CD127; WI-sup, super- dividuals and stored at 80˚C. The HIV individuals studied were natants of WI-26VA4 cell. recruited from the HIV clinic at the Ottawa Hospital. Blood was drawn, and plasma was isolated from individuals either naive to antiretroviral Copyright Ó 2010 by The American Association of Immunologists, Inc. 0022-1767/10/$16.00 therapy or off antiretroviral therapy for .6 mo at the time of sampling. www.jimmunol.org/cgi/doi/10.4049/jimmunol.0903758 4680 ROLE OF SOLUBLE IL-7Ra IN IL-7 ACTIVITY AND HIV Plasma from an additional 14 HIV+ individuals was obtained in the context of Buckinghamshire, U.K.). Densitometry analysis of protein bands was a previously reported clinical trial of highly active antiretroviral therapy (12). performed using AlphaEase Fc Software 6.0 (a Innotech, San Leandro, For in vitro tissue culture experiments, PBMCs from healthy HIV seroneg- CA), and relative changes in protein expression of treated cells were cal- ative volunteers were isolated by Ficoll-Paque PLUS (Pharmacia Fine culated compared with unstimulated controls. Chemicals, Piscataway, NJ) gradient separation, washed twice in PBS, and resuspended in RPMI medium 1640 supplemented with penicillin-strepto- Bcl-2 expression mycin and 10% FCS at a concentration of 1 3 106 cells/ml. Isolation of CD8+ After 48 h of culture, intracellular Bcl-2 staining was performed using the T cells from PBMCs was conducted using a MACS CD8+ T Cell Isolation kit BD Biosciences FITC-conjugated Bcl-2 Ab reagent set, following the (Miltenyi Biotec, Auburn, CA) in accordance with the manufacturer’s in- manufacturer’s protocol (BD Biosciences). Cells were washed with PBS structions, achieving a purity of .98–99% as verified by flow cytometry. and resuspended in fixation buffer (100 ml/1 3 105 cells; eBioscience, San Purified CD8+ T cells were resuspended at 106 cells/ml in RPMI 1640 sup- Diego, CA) and incubated at room temperature for 20 min. Following plemented with penicillin-streptomycin and 20% FCS. incubation, cells were washed, resuspended in permeabilization buffer (100 ml/105 cells) with FITC-conjugated Bcl-2 Ab (2 ml/1 3 105 cells; BD IL-7 and IL-7R reagents Biosciences), and incubated at room temperature for 20 min. Samples were 5 Recombinant human IL-7 (Sigma-Aldrich, Oakville, Ontario, Canada), washed and then resuspended in flow staining buffer (0.5 ml/1 3 10 cells; a lyophilized product, was resuspended in PBS at a concentration of 10 mg/ eBioscience). Analysis was performed by flow cytometry, collecting ml and stored at 220˚C.