L2.      Long-term culture of mouse vibrissal dermal papilla cells and hair follicle-inducing ability.
Aki Osada, Tokiro Iwabachi, Jiro Kishimoto*, Tatsuo Hamazaki, Hitoshi Okochi; Department of Tissue Regeneration, Research Institute, International Medical Center of Japan, *Skin Biology Research Labs, Life Science Research Center, Shisheido Co Ltd, Japan.

Large numbers of dermal papilla cells are needed for transplantation with epidermal cells to induce de novo hair follicles. However, dermal papilla cells under normal culture conditions do not proliferate well and lose their hair follicle-inducing capacity after more than 10 passages. Therefore, simpler and more stable culture methods for dermal papilla cells have been long desired. When explants of mouse vibrissa dermal papillae were cultured with 10% FBS-DMEM including bFGF, the outgrowth of dermal papilla cells was markedly stimulated. Moreover, the proliferation of dermal papilla cells was maintained during serial cultivations, and more than 30 passage cultivations were established. To examine their follicle-inducing ability, these established dermal papilla cells were mixed with epidermal cells and injected subcutaneously into nude mice. New hair follicles were induced when dissociated dermal papilla cells at earlier passages (under passage 4) were injected with epidermal cells isolated from newborn or embryonic mice skin, but the cells from higher passages could not induce follicles, as previously reported. Next, we generated spheres by aggregating dermal papilla cells in advance and injected them with epidermal cells. Surprisingly, the spheres made from the higher passage cells (maximum 26 passages) did induce new hair follicles. The expression of several genes specific for dermal papillae in vivo was elevated in the spheres compared with that in adhered dermal papilla cells. These results suggest that bFGF is essential for dermal papilla cell culture and that sphere formation partially represents the intact dermal papilla, resulting in hair follicle induction even by highly passaged cells.