Antigen Load Governs the Differential Priming of CD8 T Cells in Response to the Bacille Calmette Guérin Vaccine or Mycobacterium tuberculosis Infection This information is current as of September 25, 2021. Anthony A. Ryan, Jonathan K. Nambiar, Teresa M. Wozniak, Ben Roediger, Elena Shklovskaya, Warwick J. Britton, Barbara Fazekas de St. Groth and James A. Triccas J Immunol 2009; 182:7172-7177; ; doi: 10.4049/jimmunol.0801694 Downloaded from http://www.jimmunol.org/content/182/11/7172 References This article cites 40 articles, 23 of which you can access for free at: http://www.jimmunol.org/ http://www.jimmunol.org/content/182/11/7172.full#ref-list-1 Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists by guest on September 25, 2021 • 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 © 2009 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Antigen Load Governs the Differential Priming of CD8 T Cells in Response to the Bacille Calmette Gue´rin Vaccine or Mycobacterium tuberculosis Infection1 Anthony A. Ryan,*‡ Jonathan K. Nambiar,*‡ Teresa M. Wozniak,* Ben Roediger,* Elena Shklovskaya,* Warwick J. Britton,*† Barbara Fazekas de St. Groth,* and James A. Triccas2*‡ One reason proposed for the failure of Mycobacterium bovis bacille Calmette Gue´rin (BCG) vaccination to adequately control the spread of tuberculosis is a limited ability of the vaccine to induce effective CD8 T cell responses. However, the relative capacity of the BCG vaccine and virulent Mycobacterium tuberculosis to induce activation of CD8 T cells, and the factors that govern the initial priming of these cells after mycobacterial infection, are poorly characterized. Using a TCR transgenic CD8 T cell transfer Downloaded from model, we demonstrate significant activation of Ag-specific CD8 T cells by BCG, but responses were delayed and of reduced magnitude compared with those following infection with M. tuberculosis. The degree of CD8 T cell activation was critically dependent on the level of antigenic stimulation, as modifying the infectious dose to achieve comparable numbers of BCG or M. tuberculosis in draining lymph nodes led to the same pattern of CD8 T cell responses to both strains. Factors specific to M. tuberculosis infection did not influence the priming of CD8 T cells, as codelivery of M. tuberculosis with BCG did not alter the /magnitude of BCG-induced T cell activation. Following transfer to RAG-1؊/؊ recipients, BCG and M. tuberculosis-induced CD8 http://www.jimmunol.org T cells conferred equivalent levels of protection against M. tuberculosis infection. These findings demonstrate that BCG is able to prime functional CD8 T cells, and suggest that effective delivery of Ag to sites of T cell activation by vaccines may be a key requirement for optimal CD8 T cell responses to control mycobacterial infection. The Journal of Immunology, 2009, 182: 7172–7177. ational design of vaccines requires an understanding of M. tuberculosis (7). Therefore an understanding of the factors that the host determinants necessary for resistance to patho- facilitate the activation of CD8 T cells is important for the devel- R gens. An effective vaccine against Mycobacterium tuber- opment of effective strategies to control mycobacterial infections. culosis should mimic the natural immune response to infection, M. tuberculosis is a facultative intracellular pathogen that re- by guest on September 25, 2021 generating a large repertoire of both CD4 and CD8 T cells re- sides within phagosomes of infected cells, thereby promoting Ag sponding to protective mycobacterial Ags. Although CD4 T cells entry into the MHC-class II presentation pathway. M. tuberculosis are known to play a central role in protection against M. tubercu- infection of both humans and mice, however, leads to the gener- losis (1), there is mounting evidence that CD8 T cells are also ation of pathogen-specific CD8 T cell responses (8). Cross-pre- important in antimycobacterial immunity. Mice deficient in CD8 T sentation of mycobacterial Ags by dendritic cells (DCs)3 may play cells succumb rapidly to infection with M. tuberculosis (2), and a role, as evidenced by the demonstration that transfer of apoptotic CD8 T cells are both expanded during M. tuberculosis infection vesicles from infected macrophages to bystander DCs can stimu- and recruited to sites of bacterial burden (2, 3). Vaccines that elicit late CD8 T cell responses (9, 10). Early reports demonstrated that CD8 T cells, such as viruses encoding mycobacterial Ags, can M. tuberculosis could directly deliver Ag to the class I processing induce high levels of protective immunity (4, 5), and subunit vac- pathway, while M. bovis bacille Calmette Gue´rin (BCG) was less cines inducing CD8 T cells can protect mice deficient in CD4 T able to activate CD8 T cells in these studies (11). This suggested cells from M. tuberculosis (6). The protective effect of CD8 T cells egress into the cytoplasm from the endosomal compartment may is most apparent late in tuberculosis infection in mice, suggesting be a property of M. tuberculosis infection that promotes CD8 T this subset may be important in the control of latent infection with cell activation. Indeed, a recent report suggests that virulent M. tuberculosis and Mycobacterium leprae can translocate into the cytosol of infected DCs, possibly allowing Ags to be directly ‡Microbial Pathogenesis and Immunity Group, Discipline of Infectious Diseases, presented on MHC class I molecules, a property that was not *Centenary Institute of Cancer Medicine and Cell Biology, and †Discipline of Med- icine, University of Sydney, Australia shared by BCG (12). Differences in CD8 T cell activation by different mycobacterial strains may also be influenced by other Received for publication May 27, 2008. Accepted for publication March 23, 2009. factors associated with infection. M. tuberculosis infection elic- The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance its a substantial inflammatory response, characterized by the with 18 U.S.C. Section 1734 solely to indicate this fact. release of numerous inflammatory cytokines and chemokines 1 This work was supported by the National Health and Medical Research Council of (13). Proinflammatory cytokines play an important role in Australia. J.A.T. is supported by an NHMRC Career Development Award, and B.F. de S.G. is supported by an NHMRC Principal Research Fellowship. A.A.R., B.R., and J.K.N. are supported by Australian Postgraduate Awards and T.M.W. is the recipient of the University of Sydney Faculty of Medicine Postgraduate Scholarship. 3 Abbreviations used in this paper: DC, dendritic cell; BCG, M. bovis bacille Calmette 2 Address correspondence and reprint requests to Dr. James A. Triccas, Discipline of Gue´rin; DLN, draining lymph node; RD1, region of deletion 1. Infectious Diseases and Immunology, University of Sydney, Sydney, Australia. E-mail address: [email protected] Copyright © 2009 by The American Association of Immunologists, Inc. 0022-1767/09/$2.00 www.jimmunol.org/cgi/doi/10.4049/jimmunol.0801694 The Journal of Immunology 7173 shaping the proliferation and maintenance of CD8 T cells in software (TreeStar). The CFSE profile of dividing cells was analyzed as models of viral and bacterial infection (reviewed in Ref. 14). It previously described (19, 20). is yet to be determined if inflammatory processes or mecha- Detection of cytokine-producing cells nisms specific to M. tuberculosis infection contribute to the initiation of CD8 T cell response to mycobacteria. Single cell suspensions were prepared, resuspended at a concentration of 3 ϫ 106 cells/ml, and either left unstimulated or stimulated overnight with To delineate the factors that influence early priming of CD8 T the OVA peptide SIINFEKL (10 ␮g/ml) and brefeldin A (10 ␮g/ml). Cell cells upon encounter of mycobacteria by the host immune re- suspensions were washed with FACS buffer (2% FCS, PBS) and surface sponse, we have compared the ability of the BCG vaccine or vir- stained with CD8, CD45.1, or CD45.2 fluorochrome conjugates (BD ulent M. tuberculosis expressing a recombinant Ag to induce the Pharmingen). Following the surface staining, cells were washed with activation of Ag-specific CD8 T cells after infection of mice. We FACS buffer and permeabilized with Cytofix/Cytoperm buffer (BD Pharm- ingen). Cells were then washed in FACS buffer and intracellular cytokines found that BCG was able to induce specific activation of CD8 T were detected with anti-IFN-␥ FITC and anti-TNF-PE conjugates (BD cells, but the response was delayed and of a lesser magnitude when Pharmingen). compared with infection with M. tuberculosis. This reduced acti- Ϫ Ϫ T cell transfer to RAG-1 / mice and survival studies vation of T cells by BCG was associated with reduced bacterial load in the draining lymph nodes (DLN) at the time of T cell C57BL/6 mice (n ϭ 5) were infected s.c. with 1 ϫ 107 CFU BCG or 5 ϫ 4 priming. Ag load was the key determinant of CD8 T activation 10 CFU M. tuberculosis. Four weeks postinfection, the DLN were har- vested and CD8 T cells were purified by positive selection using MACS after mycobacterial infection, as delivery of equivalent numbers of separation (Miltenyi Biotec).