The method of DNA alkaline elution was applied to a study of the formation and resealing of DNA single-strand breaks after irradiation of human fibroblasts with ultraviolet light (UV). The general features of the results were consistent with current concepts of DNA excision repair, in that breaks appeared rapidly after UV, and resealed slowly in normal fibroblasts, whereas breaks did not appear in those cells of patients with xeroderma pigmentosum (XP) that are known to have defects in DNA repair synthesis. The appearance of breaks required a short post-UV incubation, consistent with the expected action of an endonuclease. Cells of the variant form of XP characterized by normal DNA repair synthesis exhibited normal production of breaks after UV, but were slower than normal cells in resealing these breaks. This difference was enhanced by caffeine. A model is proposed to relate this finding with a previously described defect in post-replication repair in these XP variant cells. DNA crosslinking appears to cause an underestimate in the measurement of DNA breakage after UV.The major photochemical damage produced in DNA by ultraviolet light (UV) consists of pyrimidine dimers. The efficiency of dimer production by UV appears to be the same in all mammalian cells (1). Major differences, however, exist in the DNA repair events that follow UV damage [recently reviewed by Cleaver (2)]. Of particular interest are the defects in DNA repair that have been noted in patients with xeroderma pigmentosum, a rare inherited disease characterized by extreme photosensitivity and the formation of multiple skin cancers [recently reviewed by Robbins et al. (3)].Pyrimidine dimers are thought to be removed from the DNA of mammalian cells by the process of excision repair (2). This process begins with a single-strand scission near the site of the dimer. The dimer is then removed, along with a substantial segment of the adjacent DNA. The resulting gap in the DNA strand is filled by synthesis of new DNA which is finally joined to the end of the preexisting strand (4, 5). The occurrence of DNA repair synthesis, the filling of gaps, and the joining to preexisting strands have all been demonstrated in mammalian cells (2).The excision model predicts the transient appearance of single-strand breaks in the DNA (2). Such breaks have been difficult to detect in eukaryotes, however, because their frequency is so low that the resulting single-strand segments are too long to be readily measured by standard techniques. The technique of DNA alkaline elution, recently developed in our laboratory, has helped us to approach this problem (6).We have examined several lines of xeroderma pigmento- Products, Inc., Calif.). Prior to irradiation, the cells were washed with warm phosphate-buffered saline (PBS). The PBS was decanted and the cells were irradiated at 370. Fresh nonradioactive medium, containing 10% serum and buffered with 0.01 M N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (Hepes) buffer, was quickly added. The cells were incubated at ...
Abstract. When paroxysmal nocturnal hemoglobinuria (PNH) erythrocytes were exposed to H202 they lysed excessively and formed greater than normal quantities of lipid peroxides when compared to red cells of normal subjects and patients with most types of hematologic disease. It was also shown that lytic sensitivity to acidified serum was related to the enhanced lytic sensitivity to H202. If the lipid of PNH cells was first extracted then exposed to ultraviolet radiation more lipid peroxides were formed than in extracts of normal red blood cells. The possible explanations for these findings and their relationship to the PNH hemolytic mechanism are discussed.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.