Induction of hair follicle growth and melanogenesis after intradermal administration of AdShh. AdShh, AdNull, or PBS was administered to the dorsal skin of postnatal day 19 C57BL/6 mice as in Figure 1c, and analyses were performed on postnatal day 26. (a) Histologic evaluation 7 days after vector administration. The AdShh-injected skin has increased depth of the dermal layer, increased follicle length, and increased follicle area compared with the naive, PBS, and AdNull controls. The changes associated with AdShh injection were limited to the area of injection (injection site vs. distant site [control]). (b) Histologic evaluation of the border of the injection site 7 days after vector administration. Hair follicles in the injection site are in late anagen; hair follicles in adjacent skin are in early anagen. A corresponding change in skin thickness was noted. (c) Quantitation of follicle area as a percentage of the total dermal/epidermal area using digital imaging and pixel area integration. Each data point represents an area measurement from a representative field. Data from 3 fields are shown per mouse; data from 2 mice are shown per treatment. (d) Northern analysis showing enhanced expression of the hair-specific keratin ghHb-1 gene induced by AdShh in vivo relative to the naive, PBS, and AdNull controls. Equal RNA loading shown by probing for GAPDH mRNA. (e and f) Melanin expression in hair follicle. Accumulation of melanin was evaluated by bright-field microscopy of unstained paraffin sections. Skin at the site of injection of AdShh (f), but not AdNull-injected skin (e), showed increased melanin in hair follicle bulbs. Skin of PBS-injected and naive control mice was similar to skin of AdNull-injected mice (not shown). (g) Northern analysis showing upregulation of melanogenesis-related tyrosinase gene expression after administration of AdShh relative to the naive, PBS, and AdNull controls. Equal RNA loading was confirmed by probing for GAPDH mRNA. Scale bars: 200 μm.