Neutrophils and extracellular traps in abdominal sepsis
Forskningsoutput: Avhandling › Doktorsavhandling (sammanläggning)
Abdominal sepsis is associated with changes in systemic inflammation response and coagulation homeostasis. Excessive neutrophil activation is a key regulator in both pro-inflammatory immune response and septic lung injury. Neutrophil-derived DNA may form extracellular traps in response to invading pathogens. To examine the role of neutrophil extracellular traps for septic lung injury and systemic coagulation. We hypothesized that neutrophil extracellular traps (NETs) from neutrophils might regulate systemic inflammation and thrombin generation in severe abdominal sepsis. Abdominal sepsis was induced by cecal ligation and puncture (CLP) in C57BL/6 mice. CLP increased plasma levels of cell-free DNA and DNA-histone complexes, suggesting potential NET formation in septic animals. Treatment with rhDNAse decreased CLP-induced formation of CXC chemokines, IL-6, and high-mobility group box 1 (HMGB1) in plasma, as well as CXC chemokines and pro-inflammatory cytokines in the lung. Moreover, administration of DNAse not only eliminated NET formation but also reduced lung tissue damage and elevated total thrombin generation in plasma from CLP animals. In vitro, we found that NETs derived from phorbol myristate acetate (PMA) stimulated mouse neutrophils markedly triggered secretion of CXCL2, TNF-α, and HMGB1 from alveolar macrophages. Isolated NETs increased thrombin generation, and co-incubation with DNAse abolished NET-induced formation of thrombin. Interestingly, thrombin generation triggered by NETs was inhibited by blocking factor XII and abolished in factor XII-deficient plasma but intact in factor VII-deficient plasma. Importantly, PMA stimulation of neutrophils was also found to generate large amount of neutrophil-derived MPs. Co-incubation with wild-type annexin V, but not a mutant annexin V, abolished NET-induced thrombin generation. We conclude that NETs regulate pulmonary infiltration of neutrophils and tissue injury via formation of proinflammatory compounds, and induce thrombin generation via the intrinsic pathway of coagulation and that neutrophil-derived MPs might play a key role in NET-dependent coagulation.
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Ämnesklassifikation (UKÄ) – OBLIGATORISK