Measurement of radiation-induced DNA double-strand breaks in human diploid fibroblasts from keloid and normal skin by single-cell gel electrophoresis.

Abstract

Cell culture experiments recently have shown that different populations of fibroblasts may exist in keloid patients. Moreover, several studies have strongly suggested heterogeneity in cell response of skin fibroblasts in the same individual. The purpose of this study was to evaluate the radiation sensitivity of skin fibroblasts from keloid and normal skin. Single-cell gel electrophoresis was employed in detecting amounts of DNA double-strand breaks generated in individual fibroblasts primarily cultured from normal skin and from keloid tissue after various doses of gamma-irradiation (0, 25, 50, and 100 Gy aerobically) from a 137Cs radiation generator. For the repair study, cells were incubated at 37 degrees C for 0 and 15 minutes and 1 and 2 hours after irradiation. Following neutral lysis and electrophoresis, DNA double-strand breaks were then detected by propidium iodide staining and determined by fluorescence microscope and quantitative image analysis. No significant difference in the extent of DNA double strand breaks in fibroblasts from normal skin or keloid after various doses of irradiation was observed. However, the residual DNA double-strand breaks remaining after various periods of incubation were demonstrated to be significantly increased in fibroblasts from keloid (p < 0.01). In conclusion, different repair capacities were found in fibroblasts from normal skin and keloid scar; this finding may play a role in the treatment of the disease entity.