title: Rapid isolation of integrin rich multipotent stem cell pool and reconstruction of mouse epidermis equivalent.
abstract: We describe here epidermis reconstruction using multipotent mouse epidermal stem cells (EpSCs) enriched from keratinocyte isolates exploting exclusively the stem cell-adhesive property. This method excluded flowcytometry and was swift. Percent enrichment was measured by the uptake of Propidium iodide and Hoechst-33342 dye using flowcytometry to determine EpSCs yield. The sorted cells were characterized by analysis of stem cell markers using immunocytochemistry and immunoblotting techniques. Epidermis was reconstructed using the identified seeding density of EpSCs and the airlift tissue culture. Histology of natural vs reconstructed mammalian epidermis was also compared. Results showed a radical improvement of near 99% in the yield of integrin overexpressing EpSCs. The enriched EpSCs tested positive for biomarkers namely cytokeratin K-15 and, K-14, p63, beta-1-integrin, CD34 and could be passaged for longer durations. Adhesion sorted cells reconstructed the epidermis. The process of tissue reconstruction was faster using the adhesion sorted cells than the FACS sorted EpSCs. The product bioengineered using multipotent EpSCs was histologically similar to normal epidermis. Features like strata basalae, spinosum, granulosum, and corneum were alike real epidermis. The reconstructed epidermis displayed normal homeostasis, which can be considered an approximating actual product for investigative dermatology, toxicology, therapeutic research, regenerative medicine, and tissue engineering.
authors.affiliation: Environmental Carcinogenesis Division, CSIR-Indian Institute of Toxicology Research Mahatma Gandhi Marg, P Box 80, Lucknow-226001, India.
authors.forename: Sushil
authors.initials: S
authors.lastname: Kumar
authors.affiliation: Environmental Carcinogenesis Division, CSIR-Indian Institute of Toxicology Research Mahatma Gandhi Marg, P Box 80, Lucknow-226001, India.
authors.forename: Shiv
authors.initials: S
authors.lastname: Poojan
authors.affiliation: Environmental Carcinogenesis Division, CSIR-Indian Institute of Toxicology Research Mahatma Gandhi Marg, P Box 80, Lucknow-226001, India.
authors.forename: Vikas
authors.initials: V
authors.lastname: Verma
authors.affiliation: Environmental Carcinogenesis Division, CSIR-Indian Institute of Toxicology Research Mahatma Gandhi Marg, P Box 80, Lucknow-226001, India.
authors.forename: Mukesh K
authors.initials: MK
authors.lastname: Verma
authors.affiliation: Department of Biotechnology, Integral University Lucknow-226026, India.
authors.forename: Mohatashim
authors.initials: M
authors.lastname: Lohani
keywords: Integrin; mouse epidermis equivalent; multipotent stem cell; reconstruction