A single-cell suspension was prepared. Rabbit Polyclonal to MED26 tumor necrosis factor (TNF)- and interleukin (IL)-6 production, decreased IL-10 production, and enhanced bacterial clearance. Conclusions PD-L1 blockade exerts a protective effect on sepsis at least partly by inhibiting lymphocyte apoptosis and reversing monocyte dysfunction. Anti-PD-L1 antibody administration may be a promising therapeutic strategy for sepsis-induced immunosuppression. Introduction Sepsis, a systemic inflammatory response to infection, results in the death of more than 210,000 people in the United States annually [1]; it remains the leading cause of death in critical ill patients [2]. Because critical care treatment is becoming expensive, understanding the molecular mechanisms underlying the development of sepsis is important in identifying new therapeutic strategies. Protracted immunosuppression caused by impaired pathogen clearance after primary infection or susceptibility to secondary infection may contribute to the high rates of morbidity and mortality associated with sepsis [3,4]. Accumulating evidence [5-7] suggests the pivotal role of apoptosis in sepsis-induced immunosuppression. Numerous studies have shown that the numbers of peripheral and splenic lymphocytes are reduced during sepsis in both humans and animals [8,9]. Apoptosis is known to be mainly responsible for decreased lymphocyte Chlorcyclizine hydrochloride numbers, and the extent of lymphocyte apoptosis correlates with the severity of sepsis [10]. In multiple animal models of sepsis, survival rates have been remarkably improved by inhibiting lymphocyte apoptosis by using selective caspase inhibitors [11,12]; altering proapoptotic/antiapoptotic protein expression [13,14]; treatment with survival promoting cytokines such as interleukin (IL)-7 [15] and/or IL-15 [16]; and modulating costimulatory receptors [17,18]. Monocytes play an essential role in innate immune defense against microbial infection. rapidly exhibit an impaired production of proinflammatory cytokines in response to additional bacterial challenge [19], and a reduced antigen presentation capacity likely due to their decreased expression of human leukocyte antigen(locus)DR (HLA-DR) [20]. Such monocytic deactivation indicates a state of globally impaired immune functions and correlates with poor clinical outcome in critically ill patients. Programmed death-1 (PD-1) is a newly defined co-inhibitory receptor whose expression can be induced, primarily on the cell surface of activated CD4 and CD8 T cells. PD-1 has two main ligands: PD-L1 (B7-H1) and PD-L2 (B7-DC). PD-L1 is broadly expressed on hematopoietic and non-hematopoietic cells, including T cells, B cells, dendritic cells (DCs), macrophages, endothelial cells, epithelial cells, pancreatic islet cells, and fibroblastic reticular cells [21]. PD-1 and its ligand exert inhibitory effects in the setting of persistent antigenic stimulation by regulating the balance among T cell activation, tolerance, and immunopathology. The PD-1/PD-L1 pathway plays a critical role in the regulation of autoimmunity, tumor immunity, transplantation immunity, allergy, immune privilege, and ischemia/reperfusion injury [22]. Recent findings suggest that the PD-1/PD-L1 pathway plays an important role in the interaction between host and pathogenic microbes that evolved to resist immune responses. Those pathogens include viruses [23], certain bacteria [24], fungi [25], and some worms [26]. Studies using PD-L1-knockout mice support the finding that PD-L1 is the primary regulatory counter receptor for the inhibitory function of PD-1 [27]. Many studies showed that PD-L1 antagonism Chlorcyclizine hydrochloride can block the interaction of PD-1 and PD-L1 [28-31]. Hence, we hypothesized that the blockade of PD-L1 using anti-PD-L1 antibody would improve survival in sepsis. The purpose of this study was to elucidate the effect of PD-L1 blockade caused by an antagonistic antibody to PD-L1 on survival in a murine cecal ligation and puncture (CLP) model of sepsis. In addition, this study attempted to determine the potential mechanism underlying the putative beneficial effect of PD-L1 antagonism in sepsis. Materials and methods CLP model of sepsis All experiments were approved by the Institutional Animal Care and Use Committee. Adult 8- to 10-week-old (22 to 30 g) C57BL/6 male mice were purchased from the Animals Experimentation Center of Second Military Medical University. CLP-induced polymicrobial sepsis was performed as described previously [15]. Briefly, mice were anesthetized with isofluorane and a Chlorcyclizine hydrochloride midline abdominal incision was made. The cecum was mobilized, ligated Chlorcyclizine hydrochloride below the ileocecal valve, and punctured twice with a 22 gauge needle to induce polymicrobial peritonitis. The abdominal wall was closed in two layers. Sham-operated mice underwent the same procedure, including opening the peritoneum and exposing the bowel, but without ligation.