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Identification of functional markers in a self-assembled skin substitute in vitro

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Abstract

Some functional parameters were identified and assessed in a tissue-engineered self-assembled skin substitute. This skin substitute was produced using fibroblasts and keratinocytes isolated from adult human skin. Keratinocytes were seeded on a dermal layer, composed of two fibroblast sheets cultured for 35 d. The epidermal cells formed a stratified and cornified epidermis and expressed differentiation markers, notably involucrin and transglutaminase. Interestingly and for the first time, the receptor for vitamin D3 was detected in all of the epidermal cell layers of the skin substitute, as it is reported for normal human skin. This observation suggests that keratinocytes retain key receptors during their differentiation in the skin model. A network of collagen fibers was observed by electron microscopy in the dermal layer of the model. In the dermis, collagen fibers remodeling and assembly is dependent on enzymes, notably prolyl-4-hydroxylase. For the first time in a skin construct, the expression of prolyl-4-hydroxylase was detected in dermal fibroblasts by in situ hybridization. The secretion of collagenases by the cells seeded in our skin substitute was confirmed by zymography. We conclude that the self-assembly approach allows the maintenance of several functional activities of human skin cells in a skin model in vitro.

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Acknowledgements

The authors thank Alexandre Deschambeault for his photographic assistance. This work was supported by the Canadian Institutes of Health Research/CIHR (no. 49478, F. Goulet; no. 12087, L. Germain), the Canada Foundation for Innovation/CFI, the Fonds de la Recherche en Santé du Québec (FRSQ) and the Régie régionale de la santé et des services sociaux du Québec (F. Goulet, N. Islam et al). B. Cvetkovska was recipient of a fellowship from Parke-Davis Cie (USA) and from the department of Pharmacy of Université Laval. L. Germain is recipient of the Canadian Research Chair from CIHR on stem cells and tissue engineering. F. Goulet is recipient of a Career Award from The Arthritis Society.

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Authors and Affiliations

  1. Laboratoire de Recherche des Grands Brûlés/LOEX, Hôpital du Saint-Sacrement du CHA, Université Laval, Quebec City, QC, Canada

    Bisera Cvetkovska & Lucie Germain

  2. Laboratoire de Génie Tissulaire/LOEX, Hôpital de l’Enfant-Jésus du CHA, Université Laval, Quebec City, QC, Canada

    Nazrul Islam & Francine Goulet

  3. Département de Chirurgie, Université Laval, Quebec City, QC, Canada

    Bisera Cvetkovska & Lucie Germain

  4. Département de Réadaptation, Université Laval, Quebec City, QC, Canada

    Nazrul Islam & Francine Goulet

  5. Laboratoire de Recherche des Grands Brûlés/LOEX, Hôpital du Saint-Sacrement du CHA, 1050 Chemin Sainte-Foy, Sainte-Foy, QC, Canada, GIS 4L8

    Lucie Germain

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  1. Bisera Cvetkovska
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  2. Nazrul Islam
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  3. Francine Goulet
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  4. Lucie Germain
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Correspondence to Lucie Germain.

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Editor: J. Denry Sato

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Cvetkovska, B., Islam, N., Goulet, F. et al. Identification of functional markers in a self-assembled skin substitute in vitro. In Vitro Cell.Dev.Biol.-Animal 44, 444–450 (2008). https://doi.org/10.1007/s11626-008-9140-y

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  • DOI: https://doi.org/10.1007/s11626-008-9140-y

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