Figure 6. Fibrosis inhibits NE release, ROS and H₂O₂ production, and neutrophil chemotaxis.

(A) NE release by bone marrow neutrophils (n = 8). Cells from 2–3 mice per group. Data are representative of 2 independent experiments. (B) ROS production by bone marrow neutrophils (saline n = 4, bleomycin n = 3, saline + MRSA n = 8, bleomycin + MRSA n = 6). Cells from 4–6 mice per group. Data from 2 combined experiments. (C) H₂O₂ production by bone marrow neutrophils (uninfected n = 6, saline + MRSA n = 12, bleomycin + MRSA n = 11). Cells from 2–3 mice per group. Data are representative of 2 independent experiments. (A–C) Dots represent technical replicates of pooled cells. Statistical analysis by 1-way ANOVA with Tukey’s multiple comparisons. Data represent the means ± SD. **P < 0.01, ***P < 0.001, ****P < 0.0001. (D) Under agarose chemotaxis assay measuring bone marrow neutrophil migration toward formyl peptide receptor (FPR) agonist WKYMVm (100 nM) for 3 hours with 20-second time intervals. Representative endpoint images of the migrating neutrophils from saline or bleomycin-treated mice. Five out of 6 experiments performed showed inhibited migration in neutrophils from fibrotic mice compared with neutrophils from nonfibrotic mice, while 1 out of 6 experiments showed the opposite effect. (E and F) Bone marrow neutrophils from saline- and bleomycin-treated mice were allowed to migrate under agarose toward WKYMVm (100 nM) for 1.5 to 4 hours. Plots showing the number of neutrophils that left the well (E) and distance traveled by the chemotaxing neutrophils (F) are presented. Data represent mean ± SEM from 4–5 independent experiments. Data points with identical shape represent data collected from the same experiment with matched migration time. *P < 0.05 when compared with saline group (paired 2-tailed Student’s t test).