Superficial Dermal Fibroblasts Enhance Basement Membrane and Epidermal Barrier Formation in Tissue-Engineered Skin: Implications for Treatment of Skin Basement Membrane Disorders

Abstract

Basement membrane is a highly specialized structure that binds the dermis and the epidermis of the skin, and is mainly composed of laminins, nidogen, collagen types IV and VII, and the proteoglycans, collagen type XVIII and perlecan, all of which play critical roles in the function and resilience of skin. Both dermal fibroblasts and epidermal keratinocytes contribute to the development of the basement membrane, and in turn the basement membrane and underlying dermis influence the development and function of the epidermal barrier. Disruption of the basement membrane results in skin fragility, extensive painful blistering, and severe recurring wounds as seen in skin basement membrane disorders such as epidermolysis bullosa, a family of life-threatening congenital skin disorders. Currently, there are no successful strategies for treatment of these disorders; we propose the use of tissue-engineered skin as a promising approach for effective wound coverage and to enhance healing. Fibroblasts and keratinocytes isolated from superficial and deep dermis and epidermis, respectively, of tissue from abdominoplasty patients were independently cocultured on collagen-glycosaminoglycan matrices, and the resulting tissue-engineered skin was assessed for functional differences based on the underlying specific dermal fibroblast subpopulation. Tissue-engineered skin with superficial fibroblasts and keratinocytes formed a continuous epidermis with increased epidermal barrier function and expressed higher levels of epidermal proteins, keratin-5, and E-cadherin, compared to that with deep fibroblasts and keratinocytes, which had an intermittent epidermis. Further, tissue-engineered skin with superficial fibroblasts and keratinocytes formed better basement membrane, and produced more laminin-5, nidogen, collagen type VII, compared to that with deep fibroblasts and keratinocytes. Overall, our results demonstrate that tissue-engineered skin with superficial fibroblasts and keratinocytes forms significantly better basement membrane with higher expression of dermo-epidermal adhesive and anchoring proteins, and superior epidermis with enhanced barrier function compared to that with deep fibroblasts and keratinocytes, or with superficial fibroblasts, deep fibroblasts, and keratinocytes. The specific use of superficial fibroblasts in tissue-engineered skin may thus be more beneficial to promote adhesion of newly formed skin and wound healing, and is therefore promising for the treatment of patients with basement membrane disorders and other skin blistering diseases.

Advancing biomaterials of human origin for tissue engineering

“…Further elucidation of the potential roles of entactin and specific laminin chains not only will clarify important aspects of BM regeneration but also may provide a better understanding of BM disorders ( e.g. , hepatic cirrhosis, skin-blistering diseases and inherited kidney syndromes) and even suggest therapeutic approaches [380–383]. …”

Keratinocyte Microvesicles Regulate the Expression of Multiple Genes in Dermal Fibroblasts

“…Keratinocytes and fibroblasts collectively contribute proteins to the basement membrane zone (Varkey, Ding and Tredget, 2014). Cytokines and chemokines are important regulators of wound healing (Barrientos et al, 2008).…”

Biology and Principles of Scar Management and Burn Reconstruction

“…In tissue-engineered models of skin containing deep or superficial fibroblasts, deep fibroblasts are profibrotic and keratinocytes exert paracrine effects that have antifibrotic properties. 57,59,60 Therefore, early recognition of deep dermal burn injuries allows early resur-facing and improved quality of wound healing. 56,57,59 Using scanning laser Doppler, thermography, and other instruments, many investigators have demonstrated acceptable levels of accuracy in the prediction of deep burn wounds that can be targeted for early skin graft surgery to prevent the development of HTS that would occur if the wounds were allowed to heal spontaneously.…”

“…57,59,60 Therefore, early recognition of deep dermal burn injuries allows early resur-facing and improved quality of wound healing. 56,57,59 Using scanning laser Doppler, thermography, and other instruments, many investigators have demonstrated acceptable levels of accuracy in the prediction of deep burn wounds that can be targeted for early skin graft surgery to prevent the development of HTS that would occur if the wounds were allowed to heal spontaneously. 61,62 These instruments are common components of surgical decision-making in many burn centers that provide objective information in identifying deep dermal wounds in addition to wound observation and judgment, which have been demonstrated to be subjective and of limited accuracy.…”