Phototoxicity in Reconstructed Human Epidermal (RhE) Models

The 3-D Phototoxicity assay uses  reconstructed human epidermal (RhE) tissue constructs to evaluate the dermal phototoxicity potential of a test material. Toxicity is determined by measuring cytotoxicity [based on the relative conversion of MTT (3-[4,5 – dimethylthiazol-2-yl] – 2,5 – diphenyltetrazolium bromide)] in cultures treated with the test material in the presence and absence of UVA light.

An advantage of using 3-D tissues to evaluate phototoxicity potential is that, unlike the 96-well phototoxicity assay, test materials are applied topically. As such,  solids, undiluted final formulations, and insoluble test materials can be tested. RhE tissue constructs are prepared using human epidermal cells  cultured on specially designed inserts. The cells differentiate to form a fully differentiated epidermis, complete with a functional stratum corneum (see picture below).

MTT, a vital dye, is actively taken up by the tissues and subsequently reduced in the mitochondria of living cells. This chemical reaction produces a purple-colored formazan within the cells, causing the live tissues to turn deep purple in color. The colored dye can be extracted from the cells and the absorbance read spectrophotometrically.

Quick Facts

Assay Model: organotypic RhE tissue constructs

Endpoint: a 30% reduction in relative viability in treated tissues in the presence of UVA exposure relative to treated tissues in the absence of UVA exposure is predictive of phototoxic potential.

Specialized protocols may be prepared as requested through consultation with an IIVS Study Director.

 

3D Tissue incubation

3D Tissue incubation


Epidermal 3-D Tissue Construct Histology

Epidermal 3-D Tissue Construct Histology

Topical Application of Test Material onto RhE model