KeraskinTM?VM is a reconstructed human epidermis (RHE) model developed in Korea. Considering its use for development of skin sensitization or irritation alternative methods, basal expression level of cytokines or chemokines associated with inflammatory signals in skin was investigated. Furthermore, predictability for skin irritant or non-irritant chemicals was examined under double-blinded condition.
Culture supernatants of Keraskin™-VM incubated for 24h or 48h were used for analyzing protein expression levels; interleukin (IL)-18, IL-6, IL-1α, tumor necrosis factor-alpha(TNF-α), monocyte chemotactic protein-1(MCP-1), granulocyte macrophage colony-stimulating factor(GM-CSF), macrophage inflammatory protein-1β(MIP-1β), regulated on activation normal T cell expressed and secreted(RANTES), vascular endothelial growth factor(VEGF) by a luminescence detection system. In addition, the tissue lysates collected at 48h were also analyzed for the expression of the same cytokines and chemokines. Skin irritation test using 20 reference chemicals recommended at the Organization for Economic Cooperation and Development test guideline No. 439 was performed. The test chemicals were applied onto the surface of Keraskin™-VM for 45min and rinsed out, then incubated for additional 42h. Cell viability for deciding skin irritancy was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay.
Key mediators for skin inflammation including IL-18, IL-6, IL-1α, and TNF-α were well expressed in both the supernatants and the lysates. Except IL-18, the 24h supernatants demonstrated higher expression levels than the 48h supernatants. Major chemokines in the skin such as MCP-1, RANTES, and VEGF were more produced at 48h than 24h, wherein GM-CSF and MIP-1β were barely detected. Predictability test for categorizing 20 skin irritant or non-irritant chemicals resulted in 80% sensitivity and 50% specificity leading to 65% accuracy.
In conclusion, KeraskinTM-VM could be a valuable reconstructed human epidermis for developing skin sensitization alternative method. In addition, more consideration on the low specificity of KeraskinTM-VM skin irritation assay should be made for successful development of KeraskinTM-VM skin irritation assay as an alternative test for screening skin irritant chemicals.