Interferon-Inducible CXC Chemokines Directly Contribute to Host Defense against Inhalational Anthrax in a Murine Model of Infection

et al. (2010) Interferon-Inducible CXC Chemokines Directly Contribute to Host Defense against Inhalational Anthrax in a Murine Model of Infection. PLoS Pathog 6(11): e1001199. doi:10.1371/journal.ppat.1001199 Interferon-Inducible CXC Chemokines Directly Contribute to Host Defense against Inhalational Anthrax in a Murine Model of Infection Matthew A. Crawford 0 Marie D. Burdick 0 Ian J. Glomski 0 Anne E. Boyer 0 John R. Barr 0 Borna Mehrad 0 Robert M. Strieter 0 Molly A. Hughes 0 Theresa Koehler, The University of Texas-Houston Medical School, United States of America 0 1 Department of Medicine, Division of Infectious Diseases, University of Virginia, Charlottesville, Virginia, United States of America, 2 Department of Medicine, Division of Pulmonary & Critical Care Medicine, University of Virginia, Charlottesville, Virginia, United States of America, 3 Department of Microbiology, University of Virginia, Charlottesville, Virginia, United States of America, 4 Centers for Disease Control and Prevention , Atlanta, Georgia , United States of America Chemokines have been found to exert direct, defensin-like antimicrobial activity in vitro, suggesting that, in addition to orchestrating cellular accumulation and activation, chemokines may contribute directly to the innate host response against infection. No observations have been made, however, demonstrating direct chemokine-mediated promotion of host defense in vivo. Here, we show that the murine interferon-inducible CXC chemokines CXCL9, CXCL10, and CXCL11 each exert direct antimicrobial effects in vitro against Bacillus anthracis Sterne strain spores and bacilli including disruptions in spore germination and marked reductions in spore and bacilli viability as assessed using CFU determination and a fluorometric assay of metabolic activity. Similar chemokine-mediated antimicrobial activity was also observed against fully virulent Ames strain spores and encapsulated bacilli. Moreover, antibody-mediated neutralization of these CXC chemokines in vivo was found to significantly increase host susceptibility to pulmonary B. anthracis infection in a murine model of inhalational anthrax with disease progression characterized by systemic bacterial dissemination, toxemia, and host death. Neutralization of the shared chemokine receptor CXCR3, responsible for mediating cellular recruitment in response to CXCL9, CXCL10, and CXCL11, was not found to increase host susceptibility to inhalational anthrax. Taken together, our data demonstrate a novel, receptor-independent antimicrobial role for the interferon-inducible CXC chemokines in pulmonary innate immunity in vivo. These data also support an immunomodulatory approach for effectively treating and/or preventing pulmonary B. anthracis infection, as well as infections caused by pathogenic and potentially, multi-drug resistant bacteria including other spore-forming organisms. - Funding: This work was supported by the Virginia Commonwealth Health Research Board and by the National Institutes of Health/National Institute of Allergy and Infectious Diseases grant R21 AI072469 (MAH). Support was also provided by National Institutes of Health grant T32 AI055432-08, Biodefense Research Training and Career Development (MAC). 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. The pulmonary airways represent a major site of interaction between the mammalian host and microbial pathogens. Infection resulting from the exposure of the respiratory tract to a variety of microorganisms is opposed by pulmonary innate immunity, a complex host response that protects against infection by directly mediating initial host defense in the airspace while helping to shape the activation of adaptive immunity [1,2]. Among the primary components of innate immunity are secreted mediators including chemokines, small proteins produced mainly by epithelial and phagocytic cells in response to pattern-recognition receptor engagement and pro-inflammatory cytokines [3]. Chemokines were originally recognized for their ability to induce directed migration of leukocytes and facilitate controlled cellular accumulation and activation during an inflammatory response through receptordependent interactions between chemokines and their specific Gprotein-coupled receptor(s) expressed by responsive cells [4]. In addition to their role in cellular recruitment, a number of chemokines have been found to mediate direct antimicrobial effects against a broad range of Gram-positive and Gram-negative bacteria in vitro [58]. While the mechanistic details of these effects remain undefined, antimicrobial activity is thought to result from interactions between positively-charged regions present at the chemokine C-terminus and negatively-charged moieties at the microbial cell surface, resulting in cell lysis [8]. Although chemokines have been shown to be central components of the host response to pulmonary infection [9], these molecules have primarily been viewed in the context of receptor/ligand interactions, without consideration for direct ligand-mediated antimicrobial activity. As such, the biological relevance of receptor-independent, chemokine-mediated antimicrobial activity in host defense in vivo remains to be established. The disease anthrax is caused by the Gram-positive, spore forming bacterium Bacillus anthracis. The infectious B. anthracis spore consists of distinct, concentric layers that encase the spores Innate immunity is critical to host defense and plays a central role in protecting the lungs from respiratory pathogens. Among the mediators important in the innate host response to pulmonary infection are chemokines, proteins originally described for their ability to regulate immune cell trafficking during an inflammatory response. More recently, chemokines have been found to exert direct antimicrobial activity against a broad range of bacteria and fungi in vitro. While these observations suggest chemokines may contribute to host defense by killing microorganisms at local sites of infection through activities not associated with cellular chemokine receptors, the biological relevance of direct chemokine-mediated antimicrobial activity in vivo has not been established. Here we show that the murine chemokines CXCL9, CXCL10, and CXCL11 exert direct antimicrobial effects against B. anthracis in vitro and that neutralization of these CXC chemokines, but not their shared receptor CXCR3, increases host susceptibility to pulmonary B. anthracis infection in vivo. These data provide unique insight into the host mediators important in host-pathogen interaction and pathogenesis of disease and support the emerging concept that host chemokines mediate efficient, pleiotropic roles that include receptor-independent promotion of host defense in vivo. genomic material and provide protection (...truncated)