Assessment of skin toxicity using living skin equivalents / 대한피부과학회지

BACKGROUND: There are various alternative in vitro irritancy tests for substituting animal experiment. However, there is no standardized method or guideline for assessing in vitro cytotoxicity. OBJECTIVES: In order to predict toxicity on human skin, an experimental model was evaluated for the cutaneous cytotoxicity using living skin equivalents(LSE). METHODS: After applying test substances (SLS, various plant extracts) on LSE, morphological examination and MTT reduction assay was done. RESULTS: The results showed that cytotoxicity of SLS using LSE was well correlated with the concentration of SLS. Furthermore, characteristic histologic findings were observed according to the increasing concentrations of test materials. CONCLUSION: These results showed that LSE can be used as an alternative model to replace animal model for cytotoxicity testing.

Growth of Human Melanocytes in Human Epidermis Reconstructed by Culture / 대한피부과학회지

BACKGROUND: Melanocytes grown in pure monolayer culure lack many of the cellular interactions that exist in vivo. This can be partially overcome by growing melanocytes together with other epidermal cells in skin equivalent models. OBJECTIVE: The objective of the present study was to grow human melanocytes in human epidermis reconstructed on dermal substrates in vitro and to examine their response to UV radiation. METHODS: The skin equivalents were prepared by seeding cultured human keratinocytes together with cultured human melanocytes(in a ratio of 5%) onto de-epidermized dermis. After 7 days of culture, they were exposed to UVB irradiation(total 150m J/cm over 5days). On day 12 of air exposure the sections of the skin equivalents were prepared for histology. The structure of the skin equivalents was studied following staining with hematoxylin and eosin. Melanocytes were characterized by DOPA staining and by immunohistochemistry. RESULTS: Melanocytes were localized singly within the basal layer of the reconstructs. Melanin was also visible both in the melanocytes and in neighboring keratinocytes. There was an increase in melanocyte size and dendricity following UV irradiation. Melanocytes became positive to staining with HMB-45 antibody following UV irradiation. CONCLUSION: Our results indicate that melanocytes grown in reconstructed human epidermis are functional and capable of responding to UV irradiation.