Abstract

Current tissue engineered skin fails to meet the need for skin replacement in full-thickness wounds. Bioprinting technology allows for fabrication of full-thickness skin with multiple cell types organized into biomimetic layers in vitro. The purpose of this study is to determine if bioprinted skin will integrate, form an epidermal barrier, and vascularize when implanted into full-thickness wounds on mice. Cells were isolated from human skin, expanded in vitro, suspended in a fibrinogen bioink, bioprinted to form a biomimetic tri-layer skin construct, and implanted onto 2.5 x 2.5cm full-thickness excisional wounds on mice. A significant difference in total wound closure was found between bioprinted skin and non-treated wounds, as well as an accelerated time to wound closure (14.8 vs. 24.5 days, p<0.001). By day 90, H&E and Masson’s trichrome staining revealed rete ridge formation in the bioprinted skin comparable to native human epidermis, compared with a flat epidermis in controls. Picrosirius red stained samples demonstrated a normal basket weave collagen orientation in bioprinted skin-treated wounds, and hypertrophic scar-like, parallel collagen alignment in controls (41.3% vs. 80.0% alignment, p<0.001). Immunohistochemical staining confirmed the presence of human cells in the regenerated skin, the formation of epidermal rete ridges, and incorporation of human endothelial cells in infiltrating host blood vessels. Measurement of blood vessels demonstrated an increase in small diameter blood vessels in bioprinted skin treated wounds compared to control (8.4 vs. 4.3 blood vessels, p<0.01). In conclusion, bioprinted skin accelerates full-thickness wound closure, with implanted cells laying down a healthy, basket-weave collagen matrix. The remodeled skin forms epidermal rete ridges and a vascular network composed of both graft and infiltrating host vessels. Ultimately, this technology could translate into a new treatment for full-thickness wounds in human patients.

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