title: Protocols for Cryopreservation of Intact Hair Follicle That Maintain Pluripotency of Nestin-Expressing Hair-Follicle-Associated Pluripotent (HAP) Stem Cells.
abstract: Hair follicles contain nestin-expressing pluripotent stem cells, the origin of which is above the bulge area, below the sebaceous gland. We have termed these cells hair-follicle-associated pluripotent (HAP) stem cells. Cryopreservation methods of the hair follicle that maintain the pluripotency of HAP stem cells are described in this chapter. Intact hair follicles from green fluorescent protein (GFP) transgenic mice were cryopreserved by slow-rate cooling in TC-Protector medium and storage in liquid nitrogen. After thawing, the upper part of the hair follicle was isolated and cultured in DMEM with fetal bovine serum (FBS). After 4 weeks culture, cells from the upper part of the hair follicles grew out. The growing cells were transferred to DMEM/F12 without FBS. After 1 week culture, the growing cells formed hair spheres, each containing approximately 1 × 10(2) HAP stem cells. The hair spheres contained cells which could differentiate to neurons, glial cells, and other cell types. The formation of hair spheres by the thawed and cultured upper part of the hair follicle produced almost as many pluripotent hair spheres as fresh follicles. The hair spheres derived from cryopreserved hair follicles were as pluripotent as hair spheres from fresh hair follicles. These results suggest that the cryopreservation of the whole hair follicle is an effective way to store HAP stem cells for personalized regenerative medicine, enabling any individual to maintain a bank of pluripotent stem cells for future clinical use.
authors.affiliation: Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kajiura
authors.affiliation: Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan. miisumi@med.kitasato-u.ac.jp.
authors.forename: Sumiyuki
authors.initials: S
authors.lastname: Mii
authors.affiliation: Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan.
authors.forename: Ryoichi
authors.initials: R
authors.lastname: Aki
authors.affiliation: Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan.
authors.forename: Yuko
authors.initials: Y
authors.lastname: Hamada
authors.affiliation: Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan.
authors.forename: Nobuko
authors.initials: N
authors.lastname: Arakawa
authors.affiliation: Department of Physiology, Kitasato University School of Medicine, Sagamihara, Japan.|Department of Cellular and Molecular Physiology, Kitasato University Graduate School Medical Science, Sagamihara, Japan.
authors.forename: Katsumasa
authors.initials: K
authors.lastname: Kawahara
authors.affiliation: AntiCancer, Inc., San Diego, CA, USA.
authors.forename: Lingna
authors.initials: L
authors.lastname: Li
authors.affiliation: Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan.
authors.forename: Kensei
authors.initials: K
authors.lastname: Katsuoka
authors.affiliation: AntiCancer, Inc., San Diego, CA, USA.|Department of Surgery, University of California, San Diego, San Diego, CA, USA.
authors.forename: Robert M
authors.initials: RM
authors.lastname: Hoffman
authors.affiliation: Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan.|AntiCancer, Inc., San Diego, CA, USA.|Department of Surgery, University of California, San Diego, San Diego, CA, USA.
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Amoh
mesh_terms: D000818:Animals; D015415:Biomarkers; D002454:Cell Differentiation; D000066673:Cell Self Renewal; D002478:Cells, Cultured; D015925:Cryopreservation / Q000379:methods; D015870:Gene Expression; D017930:Genes, Reporter; D018859:Hair Follicle / Q000166:cytology*; D007150:Immunohistochemistry; D008822:Mice, Transgenic; D064231:Nestin / Q000235:genetics* / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism*; D014738:Vibrissae
chemical_list: D015415:Biomarkers; D064231:Nestin
keywords: Cryopreservation; HAP stem cell; Hair follicle; Hair spheres; Nestin; Pluripotent