Fig. 8

Immunofluorescent localization of macrophage markers. a Anti-Iba-1 was used to detect total macrophages, and anti-CD163 was used to detect M2-biased macrophages. Representative images showing DAPI (blue), Iba-1 (Green, AlexaFlour488), and CD 163 (red, AlexaFlour594). b Quantitation of Iba-1-positive, CD163-negative cells (all macrophages except M2 macrophages). c Quantitation of Iba-1-positive, CD163-positive cells (M2-biased macrophages). The graphs show the percentage obtained by dividing the number of macrophages of the target phenotype by the total number of cells (measured by DAPI staining). There was no difference in the number of Iba-1-positive and CD163-negative cells between the groups with and without hydrogen inhalation; however, the number of Iba-1-positive and CD163-positive cells was significantly reduced by inhaling hydrogen for 7 days. SA, saline administration and air inhalation, n = 6; SH, saline administration and hydrogen inhalation, n = 6; BA, bleomycin administration and air inhalation, n = 15, BH, bleomycin administration and hydrogen inhalation, n = 15. NS: not significant, *p < 0.05, **p < 0.01. Error bars: 95% CI