Design and in vivo evaluation of a molecularly defined acellular skin construct: reduction of early contraction and increase in early blood vessel formation.
Skin substitutes are of great benefit in the treatment of patients with full thickness wounds, but there is a need for improvement with respect to wound closure with minimal contraction, early vascularisation, and elastin formation. In this study we designed and developed an acellular double-layered skin construct, using matrix molecules and growth factors to target specific biological processes. The epidermal layer was prepared using type I collagen, heparin and fibroblast growth factor 7 (FGF7), while the porous dermal layer was prepared using type I collagen, solubilised elastin, dermatan sulfate, heparin, fibroblast growth factor 2 (FGF2) and vascular endothelial growth factor (VEGF). The construct was biochemically and morphologically characterised and evaluated in vivo using a rat full thickness wound model. The results were compared with the commercial skin substitute IntegraDRT and untreated wounds. The double-layered construct was prepared according to the design specifications. The epidermal layer was about 40 μm thick, containing 9% heparin and 0.2 μg FGF7 mg per layer, localised at the periphery. The dermal layer was 2.5 mm thick, had rounded pores and contained 10% dermatan sulfate+heparin, and 0.7 μg FGF2+VEGF mg per layer. The double-layered skin construct was implanted in a skin defect and on day 7, 14, 28 and 112 the (remaining) wound area was photographed, excised and (immuno) histologically evaluated. The double-layered skin construct showed more cell influx, significantly less contraction and increased blood vessel formation at early time points in comparison with IntegraDRT and/or the untreated wound. On day 14 the double-layered skin construct also had the fewest myofibroblasts present. On day 112 the double-layered skin construct contained more elastic fibres than IntegraDRT and the untreated wound. Structures resembling hair follicles and sebaceous glands were found in the double-layered skin construct and the untreated wound, but hardly any were found in IntegraDRT. The results provide new opportunities for the application of acellular skin constructs in the treatment of surgical wounds.
Department of Biochemistry 280, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, P.O. Box 9101, 6500 HB Nijmegen, The Netherlands.
S T M
ST
Nillesen
G
G
Lammers
R G
RG
Wismans
M M
MM
Ulrich
E
E
Middelkoop
P H
PH
Spauwen
K A
KA
Faraj
J
J
Schalkwijk
W F
WF
Daamen
T H
TH
van Kuppevelt
D000818:Animals; D001808:Blood Vessels / Q000254:growth & development* / Q000378:metabolism; D003094:Collagen / Q000378:metabolism; D004353:Drug Evaluation, Preclinical; D004549:Elastin / Q000378:metabolism; D008297:Male; D008855:Microscopy, Electron, Scanning; D051381:Rats; D017208:Rats, Wistar; D012867:Skin / Q000254:growth & development* / Q000378:metabolism; D017768:Skin, Artificial
D003094:Collagen; D004549:Elastin