Development of a bilayered living skin construct for clinical applications

Abstract

An in vitro construct of human skin (living skin equivalent, LSE) has been engineered using serially passaged human epidermal keratinocytes and human dermal fibroblasts with a matrix of type I collagen. Cells are obtained from neonatal foreskin. LSE is cast, cultured, and shipped in a single culture insert. The size and shape of the insert determines the size and shape of the LSE. The dermal matrix consists of dermal fibroblasts within a condensed collagen lattice. The overlying epidermis is developed at the air-liquid interface to generate a protective cornified layer. Serum was not necessary for development of the epidermis. LSE for graft (Graftskin) has handling characteristics similar to split-thickness skin allowing it to be meshed, stapled, and sutured. LSE was cryopreserved using 65% glycerol an rapid freezing. Viability and in vivo performance on athymic mice were similar to fresh LSE. Cells derived from human eccrine gland were able to invade and form tubules rudimentary appendages may be possible.

Clinical Evaluation of NIKS-Based Bioengineered Skin Substitute Tissue in Complex Skin Defects: Phase I/IIa Clinical Trial Results

“…[42][43][44][45][46][47] The precise mechanism for this is unclear, but it may be related to the fact that cultured skin substitute tissue lacks many of the cell types (e.g., endothelial or Langerhans cells) that play a role in cadaver allograft rejection, while still expressing minor histocompatibility antigens on the allogeneic cells. 43,48,49 …”

Collagen: Applications of a Natural Polymer in Regenerative Medicine

“…It has shown to, decrease hospital stay compared to conventional treatment (silvazine) (Amani et al 2006), prevent frequent and painful dressing changes, autografting, and decreases the time to wound closure compared to alternative treatments (Biobrane alone, silvazine) (Kumar et al 2004). Apligraf (Organogenesis Inc., MA and Novartis Pharmaceutical Corp., NJ, US) (Wilkins et al 1994) is a commercially available skin equivalent consisting of a bovine collagen (type I) hydrogel seeded with allogeneic human neonatal foreskin fibroblasts and keratinocytes (Bell et al 1981a;Bell et al 1981b). The mechanical properties of the dermal component are improved by allowing collagen to contract for 6 days after which keratinocytes are seeded and cultured for 4 days under submerged conditions and terminally differentiated into a stratified epidermis for an additional 7 days at the air/liquid interface.…”

Cryopreservation of Skin Tissues for Skin Grafts

“…These preservation solutions are mainly used for the transport of kidneys, liver, or pancreas. However, they cannot preserve the organs for longer than 24 hours at 4℃ (40)(41)(42)(43)(44). In light of the increasing demand for donor organs and tissues, preservation solutions with better preserving abilities and the research and development of such solutions are urgently needed.…”

“…time limitation on the medical team (4)(5)(6). In addition, through advances in tissue engineering, cultured skin and cultured cartilage have reached the level of clinical application and demanded long term storage techniques for optimum utilization.…”