Exacerbated Staphylococcus aureus Foot Infections in Obese/Diabetic Mice Are Associated with Impaired Germinal Center Reactions, Ig Class Switching, and Humoral This information is current as Immunity of October 1, 2021. Christopher W. Farnsworth, Eric M. Schott, Abigail Benvie, Stephen L. Kates, Edward M. Schwarz, Steven R. Gill, Michael J. Zuscik and Robert A. Mooney J Immunol 2018; 201:560-572; Prepublished online 1 June Downloaded from 2018; doi: 10.4049/jimmunol.1800253 http://www.jimmunol.org/content/201/2/560 http://www.jimmunol.org/ Supplementary http://www.jimmunol.org/content/suppl/2018/05/31/jimmunol.180025 Material 3.DCSupplemental References This article cites 46 articles, 20 of which you can access for free at: http://www.jimmunol.org/content/201/2/560.full#ref-list-1 Why The JI? Submit online. by guest on October 1, 2021 • 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 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 © 2018 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Exacerbated Staphylococcus aureus Foot Infections in Obese/Diabetic Mice Are Associated with Impaired Germinal Center Reactions, Ig Class Switching, and Humoral Immunity Christopher W. Farnsworth,*,† Eric M. Schott,*,† Abigail Benvie,*,† Stephen L. Kates,‡ Edward M. Schwarz,† Steven R. Gill,x Michael J. Zuscik,† and Robert A. Mooney*,† Obese patients with type 2 diabetes (T2D) are at an increased risk of foot infection, with impaired immune function believed to be a critical factor in the infectious process. In this study, we test the hypothesis that humoral immune defects contribute to exacerbated foot infection in a murine model of obesity/T2D. C57BL/6J mice were rendered obese and T2D by a high-fat diet for 3 mo and were compared with controls receiving a low-fat diet. Following injection of Staphylococcus aureus into the footpad, obese/T2D mice had greater foot swelling and reduced S. aureus clearance than controls. Obese/T2D mice also had impaired humoral immune responses as indicated by lower total IgG levels and lower anti–S. aureus Ab production. Within the draining popliteal lymph Downloaded from nodes of obese/T2D mice, germinal center formation was reduced, and the percentage of germinal center T and B cells was decreased by 40–50%. Activation of both T and B lymphocytes was similarly suppressed in obese/T2D mice. Impaired humoral immunity in obesity/T2D was independent of active S. aureus infection, as a similarly impaired humoral immune response was demonstrated when mice were administered an S. aureus digest. Isolated splenic B cells from obese/T2D mice activated normally but had markedly suppressed expression of Aicda, with diminished IgG and IgE responses. These results demonstrate impaired humoral immune responses in obesity/T2D, including B cell–specific defects in Ab production and class-switch recombination. http://www.jimmunol.org/ Together, the defects in humoral immunity may contribute to the increased risk of foot infection in obese/T2D patients. The Journal of Immunology, 2018, 201: 560–572. besity and type 2 diabetes (T2D) are among the greatest index ,30) and avoiding skin ulceration. There is currently no risk factors for infection (1). Obese T2D patients have a known medical treatment that proactively improves immune re- O 25% lifetime risk of developing a foot ulcer (2), and sponses and lessens infection risk in this population. Importantly, ∼56% of these become infected (3, 4). Although infections are the consequences of a foot infection are catastrophic, causing by guest on October 1, 2021 common among those with obesity/T2D, the only preventative substantial physical, emotional, and financial stress. Twenty percent treatments are reducing body weight to a healthy range (body mass of foot infections result in amputation, the most feared consequence of a foot ulcer (5). Because obese/T2D patients are disproportion- ally affected by foot infection, new therapies are needed to proac- *Department of Pathology and Laboratory Medicine, University of Rochester, tively reduce the risk of infection in this population. Rochester, NY 14642; †Center for Musculoskeletal Research, University of Roches- ter, Rochester, NY 14642; ‡Department of Orthopaedic Surgery, Virginia Common- It is generally accepted that diabetes increases the risk of foot x wealth University, Richmond, VA 23298; and Department of Microbiology and ulcers via two primary mechanisms: impaired vascular flow and Immunology, University of Rochester, Rochester, NY 14642 peripheral neuropathy (2). However, these factors alone do not ORCIDs: 0000-0001-7169-3850 (C.W.F.); 0000-0002-0160-1890 (A.B.); 0000-0002- fully explain the increased risk of infection in this population. 4854-9698 (E.M. Schwarz); 0000-0002-6651-4747 (R.A.M.). Multiple studies have shown obesity to be a risk factor for in- Received for publication February 22, 2018. Accepted for publication May 7, 2018. fection in surgical sites (1, 6). Our studies have shown that obesity, This work was supported by AOTrauma Research (Clinical Priority Program: Bone Infection) and by National Institutes of Health Grants P30 AR069655, P50 but not hyperglycemia, impairs humoral immune responses to AR072000, UL1 TR000042, and T32 AR053459. orthopedic Staphylococcus aureus infection (7). Other studies All authors contributed to the study design. C.W.F., E.M. Schott, and A.B. contrib- have demonstrated impaired innate (8, 9) and adaptive immune uted to the acquisition of data. C.W.F., E.M. Schwarz, S.L.K., S.R.G., M.J.Z., and responses in obesity (7, 10). However, the underlying mechanisms R.A.M. were responsible for drafting and editing the manuscript. All authors read and approved the submitted version. that alter immune function in obesity remain elusive. Much effort has focused on macrophages and neutrophils, demonstrating im- The RNA sequencing data presented in this article have been submitted to the Gene Expression Omnibus (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE113075) paired phagocytosis and bacterial killing despite increased cell under accession number GEO113075. recruitment (8, 9, 11). We have previously demonstrated impaired Address correspondence and reprint requests to Dr. Robert A. Mooney, Department Ab production in obese/T2D mice in a model of orthopedic im- of Pathology and Laboratory Medicine, University of Rochester, Box 626, 601 plant–associated infection, a result that was confirmed in patients Elmwood Avenue, Rochester, NY 14642. E-mail address: robert_mooney@urmc. rochester.edu infected with S. aureus (7). Recent studies have demonstrated The online version of this article contains supplemental material. reduced T cell response upon challenge in obese patients despite Abbreviations used in this article: AID, activation-induced cytidine deaminase; CSR, increased prechallenge activation (12). Others have demonstrated class-switch recombination; GC, germinal center; miR-155, microRNA 155; PLN, reduced Ab titers following vaccination in obese individuals (13, popliteal lymph node; PNA, peanut agglutinin; RNAseq, RNA sequencing; T2D, type 14), an effect that results in increased mortality following infec- 2 diabetes/diabetic; TfH, T follicular helper. tion in mice (15). However, the effect of obesity/T2D on humoral Copyright Ó 2018 by The American Association of Immunologists, Inc. 0022-1767/18/$35.00 immune responses is largely understudied. www.jimmunol.org/cgi/doi/10.4049/jimmunol.1800253 The Journal of Immunology 561 In the absence of infection, the immune system of the obese host overnight at 4˚C. Anti-fluorescein conjugated to Alexa Fluor 488 (Thermo is in a state of chronic inflammation (16, 17). Importantly, Fisher Scientific) or mouse IgG kappa binding protein conjugated to B lymphocytes are known to participate in this chronic inflam- CruzFluor 594 (mIgGk BP-CFL 594) (Santa Cruz Biotechnology, Dallas, TX) were used at 1:200 for 1 h in TBS, followed by DAPI staining (Thermo matory state. B lymphocytes have been shown to produce IgG Fisher Scientific) and fixation using ProLong Gold Antifade Mounting autoantibodies (16) and secrete proinflammatory cytokines (18) Reagent (Life Technologies, Eugene, OR). All slides were visualized using within the adipose tissue of high fat–fed mice, propagating in- an Olympus VS120 Virtual Slide Scanning Microscope with Olympus flammation and contributing to insulin resistance. How chronic OlyVIA software. Quantification was as follows: germinal centers (GCs) were quantitated by a blinded observer by counting the number of PNA+ activation of the immune system in obesity/T2D, particularly in- B220+ areas within the lymph node. GC size and anti-AID staining were volving B cells, impacts normal B lymphocyte function during an determined using Visiopharm software (Brookfield, CO). infection is largely unknown. Together, these previous studies suggest that obesity