Broad defects in the energy metabolism of leukocytes underlie immunoparalysis in sepsis.

The acute phase of sepsis is characterized by a strong inflammatory reaction. At later stages in some patients, immunoparalysis may be encountered, which is associated with a poor outcome. By transcriptional and metabolic profiling of human patients with sepsis, we found that a shift from oxidative phosphorylation to aerobic glycolysis was an important component of initial activation of host defense. Blocking metabolic pathways with metformin diminished cytokine production and increased mortality in systemic fungal infection in mice. In contrast, in leukocytes rendered tolerant by exposure to lipopolysaccharide or after isolation from patients with sepsis and immunoparalysis, a generalized metabolic defect at the level of both glycolysis and oxidative metabolism was apparent, which was restored after recovery of the patients. Finally, the immunometabolic defects in humans were partially restored by therapy with recombinant interferon-γ, which suggested that metabolic processes might represent a therapeutic target in sepsis.

The Selective Sirtuin 1 Activator SRT2104 Reduces Endotoxin-Induced Cytokine Release and Coagulation Activation in Humans.

OBJECTIVES: Sirtuin 1 influences gene expression and other cellular functions through deacetylation of histone and nonhistone proteins. We here sought to determine the effects of a small molecule sirtuin 1 activator, SRT2104, on inflammation and coagulation induced by lipopolysaccharide in humans. DESIGN: A randomized, double-blind, placebo-controlled study. SETTING: An academic hospital. SUBJECTS: Twenty-four healthy humans. INTERVENTIONS: All subjects received an intravenous injection with lipopolysaccharide. Subjects were randomized to one of three groups (n=8 per group): 1) pretreatment with oral SRT2104 for 7 days (2 g/d), 2) pretreatment with a single SRT2104 dose (2 g), or 3) placebo. MEASUREMENTS AND MAIN RESULTS: SRT2104 attenuated lipopolysaccharide-induced release of the cytokines interleukin-6 (mean peak levels of 58.8% [p<0.05] and 80.9% [p=0.078] after single and repeated SRT2104 administration, respectively, relative to those measured after placebo treatment) and interleukin-8 (mean peak levels of 57.0% [p<0.05 vs placebo] and 77.1% [p<0.05 vs placebo] after single and repeated SRT2104 ingestion, respectively, while not affecting tumor necrosis factor-α and interleukin-10 release). SRT2104 also reduced the lipopolysaccharide-induced acute phase protein response (C-reactive protein). SRT2104 inhibited activation of coagulation, as reflected by lower plasma levels of the prothrombin fragment F1+2 (mean peak levels 57.9% [p<0.05] and 64.2% [p<0.05] after single and repeated SRT2104 administration, respectively, relative to those measured after placebo treatment). Activation of the vascular endothelium (plasma von Willebrand levels) and the fibrinolytic system (plasma tissue-type plasminogen activator and plasminogen activator inhibitor type I) was not influenced by SRT2104. CONCLUSIONS: This is the first human study to demonstrate biological anti-inflammatory and anticoagulant responses consistent with the activation of sirtuin 1 by a small molecule.

Myeloid-related protein-14 deficiency promotes inflammation in staphylococcal pneumonia.

Staphylococcus aureus has evolved as an important cause of pneumonia in both hospital and community settings. Staphylococcal lung infection can lead to overwhelming pulmonary inflammation. During infection, neutrophils release complexes of myeloid-related protein (MRP)8 and MRP14 (MRP8/14). MRP8/14 has been shown to exert pro-inflammatory and chemotactic activity, and to assist in the killing of S. aureus. In the current study we sought to determine the role of MRP8/14 in the host response during S. aureus pneumonia.Pneumonia was induced in wildtype and MRP14-deficient mice (mice unable to form MRP8/14) by intranasal inoculation of 1×10(7) CFU of S. aureus USA300. Mice were sacrificed at 6, 24, 48 or 72 h after infection for analyses.S. aureus pneumonia was associated with a strong rise in MRP8/14 in bronchoalveolar lavage fluid and lung tissue. Surprisingly, MRP14 deficiency had a limited effect on bacterial clearance and was associated with increased cytokine levels in bronchoalveolar lavage fluid and aggravated lung histopathology. MRP14 deficiency in addition was associated with a diminished transmigration of neutrophils into bronchoalveolar lavage fluid at late time-points after infection together with reduced release of nucleosomes.MRP8/14 serves in an unexpected protective role for the lung in staphylococcal pneumonia.

Protease-activated receptor-2 deficient mice have reduced house dust mite-evoked allergic lung inflammation.

Protease-activated receptor-2 (PAR2) is abundantly expressed in the pulmonary compartment. House dust mite (HDM) is a common cause of allergic asthma and contains multiple PAR2 agonistic proteases. The aim of this study was to determine the role of PAR2 in HDM-induced allergic lung inflammation. For this, the extent of allergic lung inflammation was studied in wild type (Wt) and PAR2 knockout (KO) mice after repeated airway exposure to HDM. HDM exposure of Wt mice resulted in a profound influx of eosinophils in bronchoalveolar lavage fluid (BALF) and accumulation of eosinophils in lung tissue, which both were strongly reduced in PAR2 KO mice. PAR2 KO mice demonstrated attenuated lung pathology and protein leak in the bronchoalveolar space, accompanied by lower BALF levels of the anaphylatoxins C3a and C5a. This study reveals, for the first time, an important role for PAR2 in allergic lung inflammation induced by the clinically relevant allergens contained in HDM.

Neutrophil extracellular traps in the host defense against sepsis induced by Burkholderia pseudomallei (melioidosis).

BACKGROUND: Neutrophil extracellular traps (NETs) are a central player in the host response to bacteria: neutrophils release extracellular DNA (nucleosomes) and neutrophil elastase to entrap and kill bacteria. We studied the role of NETs in Burkholderia pseudomallei infection (melioidosis), an important cause of Gram-negative sepsis in Southeast Asia. METHODS: In a prospective observational study, circulating nucleosomes and neutrophil elastase were assayed in 44 patients with Gram-negative sepsis caused by B. pseudomallei (melioidosis) and 82 controls. Functional assays included human neutrophil stimulation and killing assays and a murine model of B. pseudomallei infection in which NET function was compromised using DNase. Specified pathogen-free 8- to 12-week-old C57BL/6 mice were sacrificed post-infection to assess bacterial loads, inflammation, and pathology. RESULTS: Nucleosome and neutrophil elastase levels were markedly elevated in patients compared to controls. NETs killed B. pseudomallei effectively, and neutrophils stimulated with B. pseudomallei showed increased elastase and DNA release in a time- and dose-dependent matter. In mice, NET disruption with intravenous DNase administration resulted in decreased nucleosome levels. Although DNase treatment of mice resulted in diminished liver inflammation, no differences were observed in bacterial dissemination or systemic inflammation. CONCLUSION: B. pseudomallei is a potent inducer of NETosis which was reflected by greatly increased levels of NET-related components in melioidosis patients. Although NETs exhibited antibacterial activity against B. pseudomallei, NET formation did not protect against bacterial dissemination and inflammation during B. pseudomallei-induced sepsis.