Proliferation, differentiation and barrier function of the human skin equivalent culture model

Nowadays, human skin constructs (HSCs) are required for biomaterials, pharmaceuticals and cosmetics in vitro testing and for the development of complex skin wound therapeutics. In vitro three-dimensional (3D) dermal-epidermal based interfollicular, full-thickness, human skin equivalent (HSE) was here developed, recapitulating skin morphogenesis, epidermal differentiation, ultra-structure, tissue architecture, and barrier function properties of human skin. Different 3D cell culture conditions were tested to optimize HSE maturation, using various commercially available serum/animal component-free and/or fully defined media, and air-liquid interface (ALI) culture. Optimized culture conditions allowed the production of HSE by culturing normal human dermal fibroblasts (NHDFs) for 5–7 days in CELLnTEC-Prime Fibroblast (CnT-PR-F) medium and then culturing normal human epidermal keratinocytes (NHEKs) for 3 days in CELLnTEC-Prime Epithelial culture (CnT-PR) medium on them. Co-culture was then submerged overnight in CELLnTEC-Prime-3D barrier (CnT-PR-3D) medium to stimulate cell-cell contact formation and finally placed at ALI for 15–20 days using CnT-PR-3D medium. Histological analysis revealed uniform distribution of NHDFs in the dermal layer and their typical elongated morphology with filopodia. Epidermal compartment showed a multi-layered structure, consisting of stratum basale, spinosum, granulosum, and corneum. NHDFs and keratinocytes of basal layer were positive for the proliferation marker Kiel 67 (Ki-67) demonstrating their active state of proliferation. The presence of typical epidermal tissue proteins (keratins, laminins, filaggrin, loricin, involucrin, and β-tubulin) at their correct anatomical position was verified by immunohistochemistry (IHC). Moreover, transmission electron microscopy (TEM) analyses revealed basement membrane with lamina lucida, lamina densa, hemidesmosomes and anchoring fibers. The epidermal layers showed abundant intracellular keratin filaments, desmosomes, and tight junction between keratinocytes. Scanning electron microscopy (SEM) analyses showed the interwoven network of collagen fibers with embedded NHDFs and adjacent stratified epidermis up to the stratum corneum similar to native human skin. HSE physiological static contact angle confirmed the barrier function. The developed HSE represents a fundamental in vitro tool to assess biocompatibility of biomaterials, pharmacotoxicity, safety and effectiveness of cosmetics, as well as to investigate skin biology, skin disease pathogenesis, wound healing, and skin infection.

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Desmosomal distribution in the epidermis of a non-contracting human skin equivalent

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100124884 ◽

1992 ◽

Vol 50 (1) ◽

pp. 896-897

Author(s):

L.X. Oakford ◽

S.D. Dimitrijevich ◽

R. Gracy

Keyword(s):

Human Skin ◽

Basal Layer ◽

Skin Equivalent ◽

Tissue Component ◽

Intact Skin ◽

Similar Sequence ◽

Sequence Of Events ◽

Suprabasal Keratinocytes ◽

Human Skin Equivalent

In intact skin the epidermal layer is a dynamic tissue component which is maintained by a basal layer of mitotically active cells. The protective upper epidermis, the stratum corneum, is generated by differentiation of the suprabasal keratinocytes which eventually desquamate as anuclear comeocytes. A similar sequence of events is observed in vitro in the non-contracting human skin equivalent (HSE) which was developed in this lab (1). As a part of the definition process for this model of living skin we are examining its ultrastructural features. Since desmosomes are important in maintaining cell-cell interactions in stratified epithelia their distribution in HSE was examined.

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Transformative Encounters and Uncanny Intimacies: Biotechnology and Contemporary Art – A Practitioner's Approach and Perspective

Somatechnics ◽

10.3366/soma.2012.0061 ◽

2012 ◽

Vol 2 (2) ◽

pp. 263-283 ◽

Cited By ~ 1

Author(s):

Svenja J. Kratz

Keyword(s):

Tissue Culture ◽

Human Skin ◽

Contemporary Art ◽

Skin Equivalent ◽

Biomedical Innovation ◽

Equivalent Models ◽

Science Projects ◽

Insight Into ◽

Human Skin Equivalent

Abstract: Presented from an ArtScience practitioner's perspective, this paper provides an overview of Svenja Kratz's experience working as an artist within the area of cell and tissue culture at QUT's Institute of Health and Biomedical Innovation (IHBI). Using The Absence of Alice, a multi-medium exhibition based on the experience of culturing cells, as a case study, the paper gives insight into the artist's approach to working across art and science and how ideas, processes, and languages from each discipline can intermesh and extend the possibilities of each system. The paper also provides an overview of her most recent artwork, The Human Skin Equivalent/Experience Project, which involves the creation of personal jewellery items incorporating human skin equivalent models grown from the artist's skin and participant cells. Referencing this project, and other contemporary bioart works, the value of ArtScience is discussed, focusing in particular on the way in which cross-art-science projects enable an alternative voice to enter into scientific dialogues and have the potential to yield outcomes valuable to both disciplines.

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