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Personne :
Pouliot, Roxane

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Pouliot

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Roxane

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Université Laval. Faculté de pharmacie

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ncf11849049

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  • PublicationRestreint
    Reconstructed human skin produced in vitro and grafted on athymic mice
    (Ovid, 2002-06-15) Li, Hui; Germain, Lucie; Xu, Wen; Larouche, Danielle; Juhász, Julianna; Auger, François A.; Pouliot, Roxane
    Background. The best alternative to a split-thickness graft for the wound coverage of patients with extensive burns should be in vitro reconstructed autologous skin made of both dermis and epidermis and devoid of exogenous extracellular matrix proteins and synthetic material. We have designed such a reconstructed human skin (rHS) and present here its first in vivo grafting on athymic mice. Methods. The rHS was made by culturing newborn or adult keratinocytes on superimposed fibrous sheets obtained after culturing human fibroblasts with ascorbic acid. Ten days after keratinocyte seeding, reconstructed skins were either cultured at the air-liquid interface or grafted on athymic mice. We present the macroscopic, histologic, and phenotypic properties of such tissues in vitro and in vivo after grafting on nude mice. Results. After maturation in vitro, the reconstructed skin exhibited a well-developed human epidermis that expressed differentiated markers and basement membrane proteins. Four days after grafting, a complete take of all grafts was obtained. Histological analysis revealed that the newly generated epidermis of newborn rHS was thicker than that of adult rHS after 4 days but similar 21 days after grafting. The basement membrane components (bullous pemphigoid antigens, laminin, and type IV and VII collagens) were detected at the dermo-epidermal junction, showing a continuous line 4 days after grafting. Ultrastructural studies revealed that the basement membrane was continuous and well organized 21 days after transplantation. The macroscopic aspect of the reconstructed skin revealed a resistant, supple, and elastic tissue. Elastin staining and elastic fibers were detected as a complex network in the rHS that contributes to the good elasticity of this new reconstructed tissue. Conclusions. This new rHS model gives supple and easy to handle skins while demonstrating an adequate wound healing on mice. These results are promising for the development of this skin substitute for permanent coverage of burn wounds.
  • PublicationRestreint
    Tissue-engineered skin substitutes: from in vitro constructs to in vivo applications
    (Wiley, 2004-06-01) Germain, Lucie; Auger, François A.; Berthod, François; Moulin, Véronique; Pouliot, Roxane
    The field of skin tissue engineering is a paradigm for the various efforts towards the reconstruction of other tissues and organ substitutes. As skin replacement, this biotechnological approach has evolved from simple cultured autologous epidermal sheets to more complex bilayered cutaneous substitutes. The various types of such substitutes are herein presented with their intended use. However, two integrative characteristics are analysed more specifically because of their critical role: neovascularization and re-innervation. Furthermore, the in vitro use of these various skin substitutes has shed light on various physiological and pathological phenomena. Thus, not only the in vivo application of these skin substitutes as grafts, but also their in vitro value as skin models, are presented.
  • PublicationRestreint
    Multistep production of bioengineered skin substitutes : sequential modulation of culture conditions.
    (2000-02-01) Germain, Lucie; Noël, Patricia; Juhász, Julianna; Auger, François A.; Guignard, Rina; Goulet, Francine; Tremblay, Nathalie; Pouliot, Roxane
    Many studies are being conducted to define the role of growth factors in cutaneous physiology in order to add cytokines in a timely fashion for optimal tissue engineering of skin. This study is aimed at developing a multistep approach for the production of bioengineered skin substitutes, taking into account the effects of various growth factors according to the culture time. The use of a serum-supplemented medium throughout the whole culture period of skin substitutes was compared to the sequential use of specific additives at defined culture steps. Histological analysis revealed that serum was necessary for keratinocyte proliferation and migration on dermal substitutes during the first 2 d after their seeding. However, the serum-free medium presented some advantages when supplemented with different additives at specific culture steps. Interestingly, ascorbic acid added to the dermal substitutes before and after keratinocyte seeding maintained their cuboïdal morphology in the basal epidermal layer. In the absence of serum, collagen matrix degradation slowed down, and a better multilayered epidermal organization was obtained, notably with retinoic acid. Stratum corneum formation was also enhanced by fatty acids. Thus, sequential addition of exogenous factors to the medium used to produce skin substitutes can improve their structural features and functional properties in vitro.