Development of Technologies to Support the Robust and Reproducible Growth of Complex Bioengineered Human Skin Equivalents

Abstract

Native human skin is a diverse and complex organ that confers many important functions necessary for human survival, such as preventing transdermal water loss and pathogen infiltration. The literature often over simplifies the importance of the extracellular matrix (ECM) in applications related to the generation of representative epidermal skin equivalents and for treatment of skin wounding. It is of value to generate a complex and physiologically representative dermal matrix solution derived from the ECM of dermal skin equivalents which is fully human in origin. It was hypothesised that the development of a robust dermal construct containing fibroblasts and extracellular matrix using DMEM with serum will provide a stable platform capable of supporting an overlying epidermis and a effective supply of dermal derived ECM. Subsequently, the dermal skin equivalent only was upscaled and methodology optimised for the extraction and refinement of the ECM present within the dermal matrix. This dermal extract was then characterised through mass spectroscopy analysis and found to contain hundreds of components from ECM proteins to cytokines. This dermal derived extracellular matrix solution was then re-applied in vitro to functionally validate it though epidermal model generation and investigations into wound repair in vitro. Finally, looking into the effects of tape stripping as an act of exfoliation to enhance topical absorption of the dermal matrix solution it was found that tape stripping is an effective method of exfoliation and resulted in increased topical absorbance of the dermal extract. The objectives of this project are to firstly develop a methodology to produce a robust dermal compartment using a standardised base medium, determining the appropriate conditions for optimal ECM production and enhancement of ECM production utilising a novel bioreactor system. Secondly to develop and optimise the methodologies for the extraction and refinement of this dermal derived ECM matrix and its characterisation. Finally, to demonstrate the application and benefits of this human dermal extract to enhance wound repair in a damaged skin equivalent and to enhance growth and development of ageing skin equivalents.