P1Impaired innate and adaptive immunity of accelerated-aged Klotho mice in sepsisS Inoue, K Suzuki-Utsunomiya, K Suzuki-Utsunomiya, T Sato, T Chiba, K HozumiTokai University, Kanagawa, JapanCritical Care 2012, 16(Suppl 1):P1 (doi: 10.1186/cc10608)Introduction Sepsis is primarily a disease of the aged and 60% of sepsis occurs in patients older than 65 years, 80% of deaths due to sepsis occur in this age group. Klotho knockout mice (Klotho mice) develop a syndrome resembling human aging, and exhibit shortened life spans (8 weeks); however, details regarding the immunity of and immunological changes in Klotho mice after sepsis are still unclear. The purpose of the study is to elucidate the immunological changes that occur in Klotho mice after sepsis in order to identify therapeutic targets for sepsis that occurs in aged individuals.Methods (1) Survival study: cecum ligation puncture (CLP) was performed to Klotho and wild-type (WT) mice and 4-day survivals were compared. (2) Cell analysis study: mice were sacrifi ced at 8 hours post CLP or sham surgery. Spleens, thymus, and serum were harvested for FACS analysis using caspase 3 as a marker for apoptosis, and blood for serum cytokine assay. Bacterial colony count in peritoneal lavage was also analyzed.Results (1) Klotho septic mice started to die from 8 to 12 hours after CLP, and fi nal survival of Klotho mice with CLP was signifi cantly lower than that of WT with CLP (0% vs. 100%, P <0.01). (2) Increased bacterial count in peritoneal cavity and decreased recruitment of neutrophils and macrophages to the peripheral cavity were observed in Klotho-CLP mice. Serum concentration of IL-6, TNF, and IL-10 were signifi cantly higher in Klotho-CLP mice than those in the WT-CLP mice. A dramatically increased caspase 3 positive proportion in Klotho-CLP mice was observed in both fl ow cytometric and immunohistological analysis (P <0.01).Conclusion Poor survival in Klotho-septic mice may be associated with impaired bacterial clearance with decreased recruitment of neutrophils/macrophages in peritoneal cavity, elevated cytokines in serum, and increased apoptosis in thymus and spleen, following to impaired innate and adaptive immunity.P2IL-17A rs1974226 GG genotype is associated with increased susceptibility to Gram-positive infection and mortality of severe sepsisT Nakada, J Russell, J Boyd, K WalleyUniversity of British Columbia, Vancouver, CanadaCritical Care 2012, 16(Suppl 1):P2 (doi: 10.1186/cc10609)Introduction IL-17A plays a key role in host defense against microbial infection including Gram-positive bacteria. Genetic factors contribute to the host defense. Whether genetic variation of IL-17A is associated with altered clinical outcome of severe sepsis is unknown.Methods We tested for genetic association of IL-17A SNPs with susceptibility to infection and clinical outcome of severe sepsis using two cohorts of European ancestry (St Paul’s Hospital (SPH) derivation cohort, n = 679; Vasopressin and Septic Shock Trial (VASST) validation cohort n = 517). The primary outcome variable was susceptibility to Gram-positive bacterial infection. The secondary outcome variable was 28-day mortality.Results Of four tested tag SNPs (rs4711998, rs8193036, rs2275913, rs1974226) in the IL-17A gene, rs1974226 SNP was associated with altered susceptibility to Gram-positive bacterial infection in the derivation cohort (corrected P = 0.014). Patients who have the GG genotype of the rs1974226 SNP were more susceptible to Gram-positive bacterial infection, compared to the AG/AA genotype in the two cohorts of severe sepsis (SPH, P = 0.0036; VASST, P = 0.011) and in the subgroup having lung infection (P = 0.017). Furthermore, the G allele of the IL-17A rs1974226 SNP was associated with increased 28-day mortality in two cohorts (SPH, adjusted OR 1.44, 95% CI 1.04 to 2.02, P = 0.029; VASST, adjusted OR 1.67, 95% CI 1.17 to 2.40, P = 0.0052).Conclusion IL-17A genetic variation is associated with altered suscepti-bility to Gram-positive infection and 28-day mortality of severe sepsis.References1. Puel A, et al.: Chronic mucocutaneous candidiasis in humans with inborn errors of interleukin-17 immunity. Science 2011, 332:65-68.2. Cho JS, et al.: IL-17 is essential for host defense against cutaneous Staphylococcus aureus infection in mice. J Clin Invest 2010, 120:1762-1773.P3Prevalence of TLR4 single nucleotide polymorphisms (ASP299GLY, THR399ILE) in healthy subjects and septic patients, and association with outcomeT Mohovic1, R Salomao2, E Nogueira21Albert Einstein Hospital, São Paulo, Brazil; 2UNIFESP, São Paulo, BrazilCritical Care 2012, 16(Suppl 1):P3 (doi: 10.1186/cc16010)Introduction Our study aimed to determine the prevalence of functional SNPs (Asp299Gly, Thr399Ile) of TLR4 receptors, in healthy volunteers and septic patients in a Brazilian population and to correlate the presence of these polymorphisms in septic patients with clinical outcome.Methods We verifi ed the presence of polymorphisms ASP299GLY, THR399 ILE by PCR-restriction fragment length polymorphism followed by digestion with enzymes NcoI for SNP 299 and HinfI for SNP399 followed by electrophoresis for identifi cation of alleles.Results We observed a statistically signifi cant diff erence between the genotypes of the Thr399Ile polymorphism and respiratory dysfunction, indicating a higher frequency than wild-type genotype in subjects with respiratory dysfunction than those without this condition (P = 0.001). We also observed a statistically signifi cant diff erence between genotype groups formed by the Asp299Gly and Thr399Ile polymorphisms and respiratory dysfunction more often featuring group 299Selv/399Selv grupo299Het/399Het and less frequently in individuals with respiratory dysfunction than those without this condition (P = 0.003).© 2010 BioMed Central Ltd32nd International Symposium on Intensive Care and Emergency MedicineBrussels, Belgium, 20-23 March 2012Published: 20 March 2012MEETING ABSTRACTSCritical Care 2012, Volume 16 Suppl 1 http://ccforum.com/supplements/16/S1© 2012 BioMed Central Ltd
