Figure 6.

Effect of NADPH oxidase (NOX) inhibition on radiation fibrosis and type II airway epithelial cells (AECII) senescence. C57/Bl6NCr mice (n = 8 per group) were treated with diphenyleneiodonium (DPI; 1mg/kg in phosphate-buffered saline) or vehicle 5 days per week for 16 weeks, beginning immediately after irradiation (IR) of the thorax to 5×6 Gy or 0 Gy. A) Masson trichrome staining of lung tissue collected at 16 weeks after IR (0 Gy or 5×6 Gy). Lungs of mice treated to 5×6 Gy with vehicle develop extensive fibrotic foci, whereas lungs from mice receiving the same radiation dose with DPI develop only minimal collagen deposition. Collagen: blue; nuclei: purple; cytoplasm/epithelia: pink. Collagen content in the R lung at 16 weeks after irradiation was assessed by sircol assay (B) and hydroxyproline assay (C). Treatment with DPI reduced collagen content after IR compared with vehicle-treated controls. D) Frozen sections of lung of mice treated with or without IR and treated with vehicle or DPI were collected at 16 weeks, and dihydroethidium (DHE) oxidation was assessed. E) The number of AECII that were senescent was scored in each group at 16 weeks. F) The number of AECII per alveolus was scored and is reported as the percentage relative to the vehicle-treated, unirradiated control samples at 16 weeks. Columns represent the mean. Error bars represent the standard deviation. Brackets indicate P < .05 by analysis of variance. All statistical tests were two-sided. Scale bar = 40 μm.