Arrows indicate neutrophils. pathogenesis of several clinical isolates on wounded cutaneous tissue was investigated. We demonstrated that neutrophil depletion enhances bacterial burden using colony forming unit determinations. Also, mAb 1A8 reduces global measurements of wound healing in infection PRT 062070 (Cerdulatinib) alter pro-inflammatory cytokine release leading to severe microbial disease. Our findings provide a better understanding of the impact of these innate immune cells in controlling skin infections. has become an increasingly prevalent cause of hospital-acquired infections during the last 15 years (Howard et al., 2012). This pathogen is a frequent cause of pneumonia and has been identified as the etiologic agent of complicated infections, especially wound infections (Johnson et al., 2007). For instance, the organism causes 2.1% of PRT 062070 (Cerdulatinib) intensive care units-acquired skin/soft tissue infections (Gaynes et al., 2005) and was isolated from 30% of combat victims with open tibial fractures in the Middle East (Johnson et al., 2007). Moreover, the majority of clinical isolates display high-level resistance to antimicrobials, which severely compromises our capacity to care for patients with disease (Mihu and Martinez, 2011; Howard et al., 2012). Despite its clinical importance, little is known about the cellular and molecular mechanisms of host defense against PRT 062070 (Cerdulatinib) cutaneous infection. Neutrophils play an important role in early control of acute bacterial infections by killing bacteria through powerful oxidative and non-oxidative mechanisms and the production of pro-inflammatory cytokines (Mantovani et al., 2011). Clinical studies have shown that is one of the most frequently isolated gram-negative bacteria in neutropenic febrile patients in nosocomial settings, (Karim et al., PRT 062070 (Cerdulatinib) 1991; Fukuta et al., 2013; Yadegarynia et al., 2013; Kim et al., 2014) particularly after prolonged hospitalization (Wisplinghoff et al., 2004). Previous studies have also shown that neutrophils (van Faassen et al., 2007; Qiu et al., 2009) and neutrophil-recruiting chemokines (Zhao et al., 2011) are present at the site of infection, and neutrophil granule extract is bactericidal to other species of (Loeffelholz and Modrzakowski, 1988). However, the contribution of neutrophils in host resistance to cutaneous infection has not been directly investigated. Most of our current knowledge about neutrophil function in the setting of infection originates from mice treated with cyclophosphamide, (Qiu et al., 2009; Lin et al., 2012; Manepalli et al., 2013; Thompson et al., 2014; Bruhn et al., 2015) a cytotoxic alkylating agent widely used for the treatment of neoplastic and severe autoimmune diseases. Cyclophosphamide suppresses myelopoiesis resulting in neutrophil depletion in murine models (Zuluaga et al., 2006). Moreover, cyclophosphamide inhibits a suppressor response that normally prevents activation of effector T cells (Yasunami and Bach, 1988). The exacerbation of inflammatory responses and blockade of suppressive activity after cyclophosphamide treatment is consistent with the suggestion that this agent preferentially depletes suppressor or regulatory T cells (Yasunami and Bach, 1988; Ghiringhelli et al., 2004). Additionally, cyclophosphamide reduces the number of peripheral and circulating macrophages, (Santosuosso et al., 2002) phagocytic cells that are capable of detecting and eliminating as well as initiating a host early immune response (Qiu et al., 2012). Nevertheless, while cyclophosphamide is useful to study immunosuppression in rodents challenged with establishes infections in a murine model of pneumonia. Here, the Ly-6G-specific monoclonal antibody (mAb), 1A8, has been used to deplete neutrophils in mice and investigate the role of these cells in host defense (Dovi et al., 2003). We hypothesized that depletion of neutrophils would increase severity of disease in an experimental murine wound model. We showed that neutrophil depletion increases bacterial load in cutaneous tissue and alters the host immune response using distinct clinical isolates. Our findings provide a deeper understanding of the impact of neutrophils in controlling skin infections which may lead to the development of more effective therapeutic strategies. Materials and Methods Acinetobacter baumannii A total of 7 clinical isolates (0057, 1422, 1611, 2098, 2231, 3559, and 7405) were included in the study. They were isolated from blood and wound cultures at Rabbit polyclonal to PIWIL2 PRT 062070 (Cerdulatinib) the Walter Reed Medical Center, Washington, DC, USA and Montefiore Medical Center, Bronx, NY, USA. The antimicrobial susceptibility profile for each clinical isolate tested in this study was previously published (Orsinger-Jacobsen et al., 2013). The strains were stored at C80C in brain heart infusion (BHI; Becton Dickinson (BD) Biosciences, Franklin Lakes, NJ, USA) broth with 40% glycerol until use. Test organisms were grown in a Tryptic Soy broth (TSB; MP Biomedicals, LLC, Solon, OH, USA) overnight at 37C using a rotary shaker set at 150 rpm. Growth was monitored by measuring the optical density at 600 nm using a microtiter plate reader (OD600; Bio-Tek,.