DMEM/F-12

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In Vitro Assay. As shown in Fig. 1A, cells were prepared according to our previously described method (19). Usually, the E:D ratio in a piece of back skin from a newborn mouse is about 1:9 when we dissociate the back skin into single cells. For the preparation of adult cells, the skins (n ≥ 3) from 2-mo-old mice, in which hair follicles are at refractory telogen phase, were peeled off before the hair fibers were plucked through waxing. Then the s.c. fat was removed from the skins by scissors, and the skins were floated on a 0.25% trypsin solution at 4 °C for overnight digestion. The epidermis was scrapped off the dermis by a scalpel. Then epidermis and dermis were dissociated into single cells as in the preparation of newborn cells. The dissociated epidermal cell and dermal cells were mixed at a ratio of 1:9 and were dropped onto to a Transwell culture insert (Fisher Scientific) that was put in a six-well culture plate. The lower part of the culture insert was filled with 1.5 mL DMEM/F12 (1: 1) (Gibco) culture medium containing 10% FBS (Gibco). The cells were cultured in a humidified atmosphere containing 5% CO2 at 37 °C, with the culture medium being changed every other day. All animal procedures were performed upon approval of the University of Southern California (USC) Institutional Animal Care and Use Committee.

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