DNA repair in cultured keratinocytes

Most of our understanding of DNA repair mechanisms in human cells has come from the study of these processes in cultured fibroblasts. The unique properties of keratinocytes and their pattern of terminal differentiation led us to a comparative examination of their DNA repair properties. We have examined the relative repair capabilities of the basal cells and the differentiated epidermal keratinocytes as well as possible correlations of DNA repair capacity with respect to age of the donor. In addition, since portions of human skin are chronically exposed to sunlight, we have assessed the repair response to ultraviolet (UV) irradiation (254 nm) when the cells are conditioned by chronic low-level UV irradiation. The methods of Liu and Karasek were used to grow pure keratinocytes on collagen gels following their isolation from abdominal skin of newborns and adults at autopsy. Density labeling with 5-bromodeoxyuridine was used to resolve repair replication from the semiconservative mode. We found similar repair characteristics in human epidermal keratinocytes to those previously reported for cultured fibroblasts. However, the DNA repair response in basal cells was much greater than that in differentiated cells from the same skin preparation. Our comparative studies of DNA repair in keratinocytes from infant and aged donors have revealed no significant age-related differences for repair of UV-induced damage to DNA. Sublethal UV conditioning of cells from infant skin had no appreciable effect on either the repair or normal replication response to higher, challenge doses of UVL. However, such conditioning resulted in attenuated repair in keratinocytes from adult skin after UV doses above 25 J/m2. In addition, a surprising enhancement in replication was seen in conditioned cells from adult following challenge UV doses.