Effect of heparin pretreatment on the level of neutrophil extracellular traps of serum and lung tissue in septic mice / 中华危重病急救医学

ABSTRACT

Objective To investigate the influence of heparin pretreatment on serum and lung tissue level of neutrophil extracellular traps (NETs) in septic mice model and its molecular mechanism.Methods Ninety male C57BL/6J mice were randomly divided into control group (n = 30), lipopolysaccharides (LPS) group (n = 30, 30 mg/kg LPS in 100μL normal saline was intraperitoneally injected) and LPS+heparin group (n = 30, 8 U of heparin in 20μL normal saline was subcutaneously injected 30 minutes before the injection of LPS). Six hours later of LPS injection, blood was collected and lung tissue was harvested. Enzyme linked immunosorbent assay (ELISA) was used to assess the concentration of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and histones 2AX (H2AX), neutrophil elastase (NE), which reflected NETs concentration. PicoGreen fluorescent dyes was used to detect serum circulating free DNA (cf-DNA/NETs) concentration. The protein expression levels of H2AX and NE in lung tissue were examined by Western Blot.Results The serum concentrations of TNF-α, IL-6, H2AX, NE, cf-DNA/NETs, and the protein expression levels of H2AX and NE in lung tissue of septic mice were significantly higher than those of control group [TNF-α (ng/L) 133.0±14.1 vs. 2.7±1.0, IL-6 (ng/L) 3911.2±189.2 vs. 298.9±52.5, H2AX (ng/L) 545.5±40.0 vs. 21.9±8.3, NE (μg/L) 6.48±0.12 vs. 0.47±0.15, cf-DNA/NETs (μg/L) 846.3±137.5 vs. 152.7±36.4, H2AX protein (gray value) 1.14±0.09 vs. 0.68±0.04, NE protein (gray value) 0.56±0.03 vs. 0.32±0.04, allP 0.05).Conclusion Heparin pretreatment could significantly decrease the level of NETs in serum and lung tissue, and can be the potential mechanism of its organ protection in sepsis.