Histologic assessment of ears during induction of CHS and response to DEX. Paraformaldehyde-fixed serial sections of mouse ear were prepared from female Vegfr2-luc mice 2 days after challenge. (A) H&E staining (original magnification ×400). Normal mouse ear has blood vessels (red arrows) in normal dermis. Blue arrows point to lymphatic vessels. After treatment with oxazolone, the ear was thickened because of the infiltration of inflammatory cells and edema. Dilated lymphatics (blue arrows) are also present after oxazolone treatment. Dermal vessels (red arrows) contain red blood cells and white blood cells. Ear cartilage, denoted by the letter C within this panel, is surrounded by dermal inflammatory infiltrate. An intraepidermal abscess (green arrow) is present, and the surrounding epidermis is hyperplastic. Yellow arrows identify sebaceous glands. Sensitized mice exposed to oxazolone and treated with DEX have minimal dermal inflammatory cell infiltration. (B) Giemsa staining, (original magnification × 400). Normal control skin has dermal mast cells (brown arrows) in association with the connective tissue surrounding ear cartilage. The ear treated with oxazolone is markedly thickened with prominent enlarged mast cells (brown arrows), dilated lymphatics (blue arrows), and dermal blood vessels (red arrows) in a hypercellular dermis containing a mixed population of inflammatory cells. An intraepidermal abscess (green arrow) is present, and the adjacent epidermis is hyperplastic. The ear treated with oxazolone and DEX resembles normal ear. (C) Staining with an anti-VEGFR2 antibody (original magnification × 400). Normal control ear shows positive peroxidase staining (indicated by the brown tint) in cells in the epidermis (black arrow) and sebaceous gland (yellow arrow). Ears treated with oxazolone show increased VEGFR2 staining in endothelial cells of blood vessels in the dermis (red arrow) and epidermis (black arrow). The oxazolone-treated ears also show some staining for VEGFR2 protein in skeletal muscle cells (purple arrow). Some dermal inflammatory cells also show staining. Epidermal hyperplasia is clearly shown. Generally, increased staining for VEGFR2 is found in the ears after oxazolone treatment; however, this is not specific for the endothelial cells. Ears treated with oxazolone and DEX have VEGFR2 staining in the epidermis (black arrow), endothelial cells of blood vessels (red arrow), and skeletal muscle cells (purple arrows). Overall VEGFR2 staining is reduced in ear tissues of DEX-treated mice.