Figure 7
From: Deficiency of α7 Nicotinic Acetylcholine Receptor Attenuates Bleomycin-Induced Lung Fibrosis in Mice
Deficiency of α7 nAChR reduced lung fibrosis but did not change lung inflammatory parameters. (A-E) Changes of BAL parameters and flow cytometry analysis of BAL monocytes and neutrophils. The mice were intratracheally challenged with BLM (0.5 mg/kg) and euthanized at 21 d. BAL was collected to measure (A) protein levels and (B) cell counts. The BAL cells were also labeled with fluorescent anti-Ly6C and Ly6G antibodies. The whole cell population and percentage of Ly6ClowLy6Ghi neutrophils and Ly6Chily6Glow monocytes was determined by flow cytometry. The number of neutrophils and monocytes in the BAL was calculated by multiplying the percentage of these two cell populations by total BAL cell counts. N = 3–5 in each group. **p < 0.01, tested by two-way ANOVA. (F) Deficiency of α7 nAChR prevents body-weight loss during bleomycin-induced lung fibrosis. The mice were intratracheally challenged with BLM (0.5 mg/kg) and body weight was followed for 21 d. N = 10–11 in each group. Data were pooled from two independent experiments. *p < 0.05, tested by two-way ANOVA. (G) Measurements of lung hydroxyproline levels in bleomycin-induced lung fibrosis. At 21 d after 0.5 mg/kg intratracheal challenge, the mice were euthanized and lungs were homogenized. The supernatant of lung homogenate was used to measure lung hydroxyproline levels. N = 4–5 in each group. **p < 0.01 in compared groups. (H) Detection of lung TGF-β levels in bleomycin-induced lung fibrosis. At 21 d after 0.5 mg/kg intratracheal challenge, the supernatant of lung homogenate was used to measure TGF-β levels by ELISA. N = 4 in each group. *p < 0.05 in compared groups. (I, J) Western blotting analysis of α-SMA in bleomycin-induced lung fibrosis. At 21 d after 0.5 mg/kg intratracheal challenge, the supernatant of lung homogenate was used to determine expression of α-SMA by western blotting. (H) Western blotting bands in compared groups. (I) Ratio of expression of α-SMA to GAPDH analyzed by Image J software. N = 2–5 in each group. *p < 0.05 in compared groups. Data are mean ± SD. (K-N) Changes of lung profibrotic and inflammatory genes in bleomycin-induced lung fibrosis. At 21 d after 0.5 mg/kg intratracheal challenge, lungs were removed and homogenized for extracting RNA. Lung Acta2, Il6, Mcp1 and Cxcl2 mRNA were detected by real-time PCR. N = 3–5 in each group. *p < 0.05, **p < 0.01 in compared groups. Data are mean ± SD.