Fig. 3

Ccr6 gene silencing restores BLM-induced vascular hyperpermeability in mice. Wild-type mice were injected with Ccr6 siRNA or scrambled non-silencing RNA (SCR), followed by 1-week and 4-week bleomycin (BLM) challenge. A The leakage of Evans blue dye was macroscopically evaluated in the skin after 1-week BLM injection. B In vitro transwell permeability assay showed decreased permeability in the monolayer of HDMECs treated with CCR6 siRNA relative to the control cells treated with SCR. C–F After 4-week BLM injection, CCR6 was visualized by immunohistochemistry (left photographs of C). CCR6 knockdown was also confirmed by qRT-PCT with the whole skin sections (a right graph of C). α-smooth muscle actin (D), NG2 (E), and VE-cadherin (F) were visualized by immunohistochemistry. Apoptotic cells were visualized by staining cleaved caspase 3 (G). Skin sections of angiosarcoma were used as positive controls (a bottom panel of G). Unbroken arrows, arrow heads, and dashed arrows in each figure represent venules, capillaries, and inflammatory cells, respectively. Each graph indicates mean ± SEM of the indicated parameters. A scale bar is 25 μm. AU, arbitrary unit. Representative results of 6 independent experiments were shown