In vivo effect of IAIP and HMW-HA in a mouse model of histone challenge. (A) Effect of IAIP and HMW-HA on histone-induced production of proinflammatory cytokines and chemokines. TNFα, IL-6, KC, IL-1β, monocyte chemoattractant protein-1 (MCP-1), and IL-10 levels in mouse plasma collected 3 hours after challenge with 50 mg/kg histones alone (n = 5) or preincubated with 50 mg/kg IAIP (n = 5) or 90 mg/kg HMW-HA (n = 5). Data are presented as mean ± SEM and analyzed with one-way ANOVA with Bonferroni’s multicomparison test. ****P < .0001, ***P < .001, **P < .01, and *P < .05 compared with the histones challenge group. (B) Immunostaining of neutrophil elastase in the lungs of mice injected with histones alone, histones incubated with IAIP or HMW-HA, and controls. Nuclei are shown in blue. Magnification bars: 100 µm. (C) Quantitation of neutrophils in the lungs; histogram data are shown as mean ± SEM for 10 microscopic fields collected from at least 3 different animals per condition. One-way ANOVA with Bonferroni’s multicomparison test was used; ***P < .001 compared with the histone challenge group. (D) Measurement of MPO activity in plasma from mice injected with histones only compared with controls and histones preincubated with IAIP or HMW-HA. Data are shown as mean ± SEM from 3 independent experiments using one-way ANOVA with Bonferroni’s multicomparison test. ***P < .001 and **P < .01 compared with the histone challenge group.