Abstract
De novo skin regeneration with human keratinocytes amplified in culture is a life-saving procedure for patients with extensive skin loss and chronic wounds. It also provides a valuable platform for gene function and therapeutic assessments. Nevertheless, tissues generated in this manner lack hair follicles that are important for skin homeostasis, barrier function, and repair. In this study, we generated skin tissues with human keratinocytes combined with dermal papilla (DP) cells isolated from mouse whisker hair. For this, cultured keratinocytes and mouse DP (mDP) cells were mixed at 10:1 ratio and seeded onto devitalized human dermal matrix derived from surgically discarded human abdominoplasty skin. After 1 week in submerged culture, the cell/matrix composites were grafted onto the skin wound beds of immunocompromised NSG.SCID mice. Histological analysis of 6-week-old skin grafts showed that tissues generated with the addition of mDP cells contained Sox2-positive dermal condensates and well-differentiated folliculoid structures that express human keratinocyte markers. These results indicate that cultured mDP cells can induce hair follicle neogenesis in the de novo regenerated skin tissues. Our method offers a new experimental system for mechanistic studies of hair follicle morphogenesis and tissue regeneration and provides insights to solving an important clinical challenge in generation of fully functional skin with a limited source of donor cells.