Fig. 4

The histidine-pyridine-histidine ligand derivative HPH-15 inhibits leukocyte infiltration and suppresses Smad3 phosphorylation in bleomycin (BLM)-injected skin. a–d Back skin samples of mice treated with HPH-15 or vehicle were harvested after 14 days of daily BLM injections. Skin sections were incubated with anti-F4/80 antibody (a) and anti-CD3 antibody (b). Representative images are shown on the left. Scale bar = 50 μm. Quantitative analysis is shown in the bar graph on the right. c Skin sections were stained for p-Smad3. Scale bar = 50 μm. Quantitative analysis of p-Smad3-positive cells in the dermis is shown in the bar graph. d Cryosections of the skin were subjected to immunofluorescent staining for 4′,6-diamidino-2-phenylindole (DAPI) (nuclear, blue), p-Smad3 (nuclear, green), and F4/80 (cell surface, red). Dashed box inset indicates the location of zoomed image. Scale bar = 30 μm. All values represent mean ± SEM; n = 5 per group. * p ≤ 0.05. HPF High-power field