Abstract: Objective To explore the differentiation potency of ES cell-derived epidermal stem cells compounded by dermal analogs reconstructed with bone MSCs in hypodermis. Methods The dermal analogs were reconstructed with rat bone MSCs and compound gel-gelatin sponge. E14-ES cells, labeled with Hoechst 33342, were cocultured with human amnion. Four days later, epidermal stem cell clones were formed. The skin analogs, reconstructed with ES cell-derived epidermal stem cells and dermal analogs, were transplanted into 129 mice hypodermis. The differentiation tissue of skin analogs was sampled at 2w, 4w, 6w, 8w. The sections were observed with HE staining, immunohistochemical and di-labeled immunofluorescence methods to test the expression of β1 integrin, CK15, CK19, CEA, CK18. Results The sections were showed tubular or follicular like structures formed with simple or stratified epithelium at 2 and 4 weeks. Keratinized stratified squamous epithelium, sweat glands-like, sebaceous glands-like and hair follicles-like structures were observed at 6 week and 8 week after transplantation. The cells labeled by Hoechst 33342, formed tubular or follicular like structures, expressedβ1 integrin, CK15, CK19, CEA and CK18 positive respectively at 2w and 4w. The sweat glands-like structure expressed CEA and CK18 positive respectively at 6w and 8w. There were more sebaceous glands-like structures. Conclusion ES cell-derived epidermal stem cells compounded by dermal analogs reconstructed with bone MSCs can differentiate into keratinized stratified squamous epithelium, sweat glands-like, sebaceous glands-like and hair follicles-like structures in hypodermis.

Key words: ES cells, epidermal stem cells, skin appendages, committed differentiation