FIGURE 3.

(A) Neutrophils from healthy subjects, pretreated for 2 min with increasing concentrations of RNO (0–1,000 nMoles/L), were exposed to endotoxin and allowed to adhere on fibrinogen-coated surfaces for 18 h in the presence or in the absence of CFTRinh-172 (10 µMoles/L). Unstimulated neutrophils, pretreated with increasing concentrations of RNO (0–1,000 nMoles/L), were incubated in parallel. At the end of the incubation, free DNA was quantitated. Results are mean ± SEM of experiments performed with cells from 9–11 different donors in duplicates. *p < 0.05 (ANOVA, Dunnett test), RNO-treated vs untreated samples; **p < 0.05 (Student’s t-test), CFTRinh-172–treated vs untreated samples. (B) Neutrophils from individuals with CF, pretreated with increasing concentrations of RNO (0–1,000 nMoles/L), were incubated with endotoxin and allowed to adhere on fibrinogen-coated surfaces for 18 h. Unstimulated neutrophils, pretreated with increasing concentrations of RNO (0–1,000 nMoles/L), were incubated in parallel. Results are mean ± SEM of experiments performed with cells from seven different patients with CF (see Table 1 for patients’ characteristics). *p < 0.05 (ANOVA, Dunnett test) vs untreated samples. The presence of citrullinated histone H3 in neutrophils from three healthy donors (C) or two people with CF (D) was analyzed after 18 h of incubation. Samples were then subjected to Western blot analysis using a monoclonal antibody which specifically recognizes citrullinated histone H3. Representative Western blots are shown. The right image in panel C reports a densitometric analysis from n = 3. *p < 0.05 (Student’s t-test).