Fig. 4

Induced keratinocytes (iKCs) can regenerate normal skin in vivo. (a) iKCs or mouse embryonic fibroblasts (MEFs) were transplanted in the back skin of nude mice. Nude mice transplanted with iKCs showed de novo hair formation at the graft site after 6 weeks (a), whereas hair development was not observed on mice transplanted with MEFs (b). All layers of the epidermis, including de novo hair follicles and sebaceous glands, are apparent in H&E stained skin sections from iKC-transplanted mice (c) but not MEF-grafted skin (d). EYFP staining (green) marks iKCs and de novo iKC-generated tissue (e), which is not present in MEF-transplanted epidermis (f). Scale bars = 50 μm (c and d) and 10 μm (e and f). (b) Epidermal thickness comparing iKC-derived vs MEF-transplanted skin. Blindfolded comparison was performed by measuring epidermal thickness in 25 images (two measurements/image) captured from five independent tissue sections (five images/section) from iKC-derived or MEF-derived skin. Note a nearly twofold increase in iKC-generated epidermis. Error bars represent standard error amongst 50 independent measurements for each sample. (c) Immunofluorescence staining for K14 (a marker of undifferentiated keratinocytes), K10 (an intermediate differentiation marker), and loricrin (a terminal differentiation marker). Tissue sections were stained for different keratinocyte stratification/differentiation markers in iKC- or MEF-transplanted tissue sections. IKC-transplanted epidermis expressed both K14 (green) and K10 (red), whereas MEF-transplanted tissue only displayed some K14-positive cells, and no K10 staining (b). Loricrin (red) was expressed on the outer epidermal layer of the iKC-derived skin (c) and indicates the presence of iKC-derived terminally differentiated cells. In contrast, MEF-derived skin was negative for loricrin expression (d). Scale bars = 10 μm