# Bioengineered Skin Substitutes: Advances and Future Trends
## 3 Cell-Based Skin Substitutes
### Bioengineered Dermo-Epidermal Skin Substitutes (Under Development)  
Treatment of extraordinarily large deep skin defects remains a great clinical challenge. Today's \"gold standard\" approaches to cover such skin defects are primarily split- and full-thickness skin autografts, as well as skin flaps, skin expansion techniques, and dermal substitutes [62; 63; 64; 65; 66]. Laboratory-grown skin substitutes offer a novel promising treatment option for patients suffering from severe, full-thickness skin injuries [20; 24; 67; 68]. Those patients need an artificial skin substitute due to the shortage of healthy donor skin sites for autografts.  
Therefore, our laboratory-Tissue Biology Research Unit (TBRU) at the University of Zurich, Switzerland, and several laboratories worldwide have developed dermo-epidermal skin substitutes (DESSs) containing both--dermal and epidermal skin layers [12; 66; 67; 68; 69; 70; 71; 72; 73; 74]. Our hypothesis is that a skin substitute closely resembling native human skin can yield more satisfactory clinical results both functionally and cosmetically. Of note, the addition of the missing dermal component significantly enhanced the mechanical stability and mesenchymal-epithelial interaction of those skin substitutes.  
Moreover, our laboratory also investigated the mechanical modifications of the dermal compartment for better surgical handling [75; 71]. Consequently, we developed a plastically compressed hydrogel based on a collagen type I matrix serving as a dermal template for DESSs [71]. Uncompressed collagen hydrogels are fragile and fold when manipulated with forceps, but after plastic compression, they are significantly more stable, which improves handling. Importantly, this dermal template has been successfully used for the establishment of large DESSs (7.5 \(\times\) 7.5 cm) and was eventually tested in a pig animal model [36; 37; 71; 72; 73; 74; 75; 76; 77].  
Recently, these autologous human DESSs (DenovoSkin) were successfully used in phase I clinical trial on 10 children at the University Children's Hospital, Zurich [75]. We recently started a phase II clinical trial in various burn and reconstructive surgery centers in Switzerland (University Children's Hospital Zurich, University Hospital Zurich) and Europe using those autologous skin grafts [75].  
However, the prevalent challenge facing this innovative approach is the lack of sufficient vascularization to support the survival and viability of the above-mentioned DESSs after transplantation. This aspect is of utmost importance for the treatment of non-healing chronic wounds.  
In a pilot study, we overcame these hurdles by the in vitro generation of capillary networks in DESSs using endothelial and mesenchymal progenitors derived from the stromal vascular fraction (SVF) of human adipose tissue (Figure 2) [12; 13; 20]. This approach holds great promise for future clinical applications as adipose tissue represents a convenient, abundant, and easily accessible cell source [78]. Moreover, this concept was previously successfully used in the engineering of precisely sized osteogenic constructs, increasing the efficiency and uniformity of bone tissue formation [79].  
Importantly, the rapid onset of blood infiltration in the skin substitutes had efficient effects on promoting epithelial and dermal tissue repair in vivo by (1) increased collagen type I expression, (2) elevated cell proliferation rate of both dermis and epidermis, (3) improved graft take rate, (4) reduced expression of wound healing markers such as cytokeratin 16 (CK16) and cytokeratin 17 (CK17), and (5) rapid achievement of epidermal homeostasis. These results confirm that such a co-culture-based pre-vascularization strategy of tissue-engineered skin grafts emerges as an efficient method to noticeably improve wound healing, cell engraftment, and skin function after transplantation [20].