Novel three-dimensional live skin-like in vitro composite for safety tests

We describe the construction of novel three-dimensional (3D) skin-like cultures of spontaneously immortalised normal keratinocytes derived from human HaCaT cells using a non-woven membrane of silica fibre in differentiation medium. Analyses of skin differentiation markers in a 3D-skin-like in vitro composite (SLIC) revealed the expression of keratin 2, keratin 10, filaggrin, and involucrin, indicating mature skin characteristics. We further engineered SLIC to incorporate real-time bioluminescence monitoring driven by the IL-8 and UBC genes. We also monitored the time- and dose-dependent responses of SLIC to UV stress, and to hydrophilic chemicals, and hydrophobic ones which are difficult to assay so far as they do not disperse in water. We propose that SLIC could serve as a novel alternative to using animals to test the safety of various chemicals on skin for medical, cosmetic purposes.