Dendritic Cells in Chronic Mycobacterial Granulomas Restrict Local Anti-Bacterial T Cell Response in a Murine Model Heidi A. Schreiber1,2*, Paul D. Hulseberg1,2, JangEun Lee1¤a, Jozsef Prechl1¤b, Peter Barta3, Nora Szlavik4, Jeffrey S. Harding1,2, Zsuzsanna Fabry1,2, Matyas Sandor1,2 1 Department of Pathology and Laboratory Medicine, University of Wisconsin, School of Medicine and Public Health, Madison, Wisconsin, United States of America, 2 Cellular and Molecular Pathology Training Program, University of Wisconsin, Madison, Wisconsin, United States of America, 3 Department of Pulmonology, Semmelweis University, Budapest, Hungary, 4 Sejtdiagnosztika Kft, Hospital Bajcsy Zsilinszky, Budapest, Hungary Abstract Background: Mycobacterium-induced granulomas are the interface between bacteria and host immune response. During acute infection dendritic cells (DCs) are critical for mycobacterial dissemination and activation of protective T cells. However, their role during chronic infection in the granuloma is poorly understood. Methodology/Principal Findings: We report that an inflammatory subset of murine DCs are present in granulomas induced by Mycobacteria bovis strain Bacillus Calmette-guerin (BCG), and both their location in granulomas and costimulatory molecule expression changes throughout infection. By flow cytometric analysis, we found that CD11c+ cells in chronic granulomas had lower expression of MHCII and co-stimulatory molecules CD40, CD80 and CD86, and higher expression of inhibitory molecules PD-L1 and PD-L2 compared to CD11c+ cells from acute granulomas. As a consequence of their phenotype, CD11c+ cells from chronic lesions were unable to support the reactivation of newly-recruited, antigen 85B- specific CD4+IFNc+ T cells or induce an IFNc response from na¨ıve T cells in vivo and ex vivo. The mechanism of this inhibition involves the PD-1:PD-L signaling pathway, as ex vivo blockade of PD-L1 and PD-L2 restored the ability of isolated CD11c+ cells from chronic lesions to stimulate a protective IFNc T cell response. Conclusions/Significance: Our data suggest that DCs in chronic lesions may facilitate latent infection by down-regulating protective T cell responses, ultimately acting as a shield that promotes mycobacterium survival. This DC shield may explain why mycobacteria are adapted for long-term survival in granulomatous lesions. Citation: Schreiber HA, Hulseberg PD, Lee J, Prechl J, Barta P, et al. (2010) Dendritic Cells in Chronic Mycobacterial Granulomas Restrict Local Anti-Bacterial T Cell Response in a Murine Model. PLoS ONE 5(7): e11453. doi:10.1371/journal.pone.0011453 Editor: Delia Goletti, National Institute for Infectious Diseases L. Spallanzani, Italy Received February 21, 2010; Accepted June 4, 2010; Published July 6, 2010 Copyright: 2010 Schreiber et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by National Institutes of Health funding R01-A1048087 and R21-A1072638 (M. Sandor). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: msandor@wisc.edu €a Current address: Department of Medicine - Division of Rheumatology, University of California San Francisco, San Francisco, California, United States of America €b Current address: Department of Immunology, Lora´nd Eo¨ tvo¨ s University, Budapest, Hungary Introduction The formation of a granuloma in response to a pathogen creates an immunological foci that contains the antigen. During infection with mycobacteria, the granuloma contains the bacteria, prevents dissemination, and localizes immune responses to limit tissue damage. Though it protects the host, the granuloma also facilitates bacterial survival, which may eventually allow for disease tansmission. Acute mycobacteria-induced granulomas formed early in infection are large lesions with high bacterial burden [1]. They are characterized by the presence of a high proportion of IFNc-producing CD4+ T cells, which are critical activators of microbicidal pathways in bacteria-containing macrophages [2]. Chronic granulomas are smaller, more structured lesions with a lower bacterial load and reduced killing–they contain bacteria and prevent dissemination, but are ultimately unable to sterilize the lesion. In this way the chronic granuloma provides a home for bacterial latency, in which reactivation can occur decades later after immune stress from AIDS, old age, or anti-TNFa therapy, for instance. Reactivated bacteria grow, disseminate and often result in fatality [3,4]. Mutant mycobacteria strains that induce poor granuloma formation result in increased bacterial growth [5–9]. Virulent mycobacteria species have actually evolved gene- specific strategies to promote early granuloma formation and ensure their own long-term survival so as to increase chances of disease transmission. While the function of acute granulomas is the focus of intense research, much less is known about chronic granulomas. DCs are critical in the initiation of immune responses since they are the only antigen presenting cell capable of activating na¨ıve T cells and efficiently initiating a recall T cell response [10]. Following mycobacterial infection, DCs are required for initiation PLoS ONE | www.plosone.org 1 July 2010 | Volume 5 | Issue 7 | e11453