2003 Volume 113 Issue 14 Pages 2033-2038
In recent years, cultured autologous epidermal keratinocytes have been successfully applied to covering skin defects caused by burn injury. Instead of epidermal keratinocytes, the authors tried to culture outer root sheath (ORS) cells from plucked hair follicles, which are more accessible than epidermis. This report describes a simple method for culturing ORS cells using a non-collagen-coated dish and serum-free medium. We carried out the explant culture of ORS cells using plucked human scalp hair follicles, and studied the influence on growth of ORS cells of several factors, including age, sex, and plucking regions. After plucking the hair follicles from 11 males aged from 7 to 83 years, and 12 females aged from 9 to 81 years, ORS cells were cultured. There was no significant differences found with regard to age, sex, or plucked region. Four types of explant culture method were compared as follows : ① plating the hair follicles simply on a culture dish, ② plating the hair follicles after treatment with 0.3% trypsin and 0.4% EDTA, ③ fixing the hair follicles with glass coverslips, and ④ fixing hair follicles with glass coverslips after treatment with 0.3% trypsin and 0.4% EDTA. We found that fixing hair follicles with glass coverslips on culture dish gave us good results ; ORS cells could be cultured easily. In this study, a rather simple method was established for culturing the stratified ORS cell sheet from plucked hair follicles. First, the hair follicles were placed on non-collagencoated dish, and then fixed with glass colverslips. Next, the hair follicles were cultured in DMEM supplemented with 10% fetal calf serum (high Ca medium). After the colony of ORS cells was recognized, the culture medium was changed to low Ca medium. When the monolayer cells become confluent, the culture medium was changed back to a high Ca medium, and a stratified ORS cell sheet was finally obtained. Using this culture system, cultured autologous ORS cells could easily be repeatedly established from the patient’s follicles, avoiding additional wounding in skin donor sites, and cultured ORS cells could be utilized as epidermal grafts on skin defects.
