Figure 1. Methods for hair follicle regeneration.

(a) Schematic representation of the methods used for the generation and transplantation of a bioengineered hair follicle germ. (b) Phase-contrast images of murine embryonic back skin, tissues, dissociated single cells and bioengineered hair follicle germ, which was reconstituted using the organ germ method, with a nylon thread guide (arrow head). Scale bars, 200 μm. (c) Histological analyses of a natural vibrissa isolated from an adult mouse. Macroscopic and H & E staining of whole vibrissa is shown in left two panels. Dashed lines (red) via macro-morphological observations (left) and H & E staining (right) indicate the interface of the bulge and SB region. Boxed area in the left panels that is H & E stained to indicate the bulge, and SB area is shown at a higher magnifi cation in the right panel. The bulge region was immune-stained with anti-CD49f (red, left) and anti-CD34 (red, centre) antibodies and with Hoechst 33258 dye (blue). Black broken lines in high-magnified H & E indicate the interface of epithelium of the hair follicle. IF, infundibulum; RW, ringwulst; 1/2, half portion of hair follicle. Scale bars, 100 μm. (d) Histological and ALP analyses of the bulb area of a natural vibrissa and of primary cultures of DP cells. Hair bulb (left two) and cultured DP cells (right two) were anlaysed via enzymatic staining of ALP. The red dashed line indicates Auber?s line. Scale bars, 100 μm. (e) Longitudinal sections of bioengineered pelage during eruption and growth processes mediated by an inter-epithelial tissue-connecting plastic device (with guide). The counterpart is shown as cyst formation with a bioengineered pelage follicle at 14 days after intracutaneous transplantation (without guide). H & E staining (upper) and fluorescent microscopy (lower) of the bioengineered hair follicles at 0, 3 and 14 days after transplantation. Scale bars, 100 μm. (f) Macro-morphological observations of the bioengineered hairs during the eruption and growth of bioengineered pelages (upper) and vibrissae (lower), including just after transplantation at day 0 (left), the healing of the wound at day 3 (centre), eruption of the hair shaft and growth at day 14 in the bioengineered pelage and 37 days in the vibrissa (right). Scale bars, 1.0 mm.