Characterization of ultraviolet radiation-induced damage to keratinocytes in a skin equivalent in vitro

The human skin equivalent (HSE) provides a convenient model for studying the dermatological effects of exposure to ultraviolet (UV) radiation. HSEs, constructed by overlaying a collagen-fibroblast matrix with epidermal cells, were maintained submerged for 1 week after the addition of epidermal cells and then raised to the air-liquid interface for an additional 3 weeks. HSEs were exposed to sublethal doses of UV radiation ranging from 0 to 500 J/m2, incubated up to 48 h in medium containing 3H-thymidine and fixed for ultrastructural and autoradiographic analysis. Exposure to radiation doses greater than 50 J/m2 led to vacuolation of the cornified envelopes and enlargement of intercellular spaces. These doses also led to the formation of dense cytoplasmic bodies, and separation and vesiculation of the nuclear envelope in the basal cells. DNA synthesis in the basal cells was analyzed autoradiographically. Maximal numbers of labeled basal cells were observed 24 h after exposure to UV radiation at 50 J/m2. Although the proportions of labeled cells varied among different epidermal donors, the maximal responses and time-course of 3H-thymidine incorporation remained consistent, supporting the usefulness of the HSE in studying the effects of UV irradiation on human skin.