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Detection of single-strand breaks and formamidopyrimidine-DNA glycosylase-sensitive sites in DNA of cultured human fibroblasts. 1995

S Czene, and M Harms-Ringdahl
Department of Molecular Virology, Cancer Research Institute, Bratislava, Slovak Republic.

Under oxidative stress 7,8-dihydro-8-oxo-2'-deoxyguanosine (8-oxodG), a damaged base with mutagenic potential, and single-strand breaks (SSB) are formed in DNA. Both lesions are frequently used as a parameter for oxidative damage of DNA. Here we report on results from the evaluation of a modified nick translation assay, where 8-oxodG and SSB formation in cellular DNA of cultured human fibroblasts were simultaneously detected. The assay is based on a method previously described by others, with several modifications in reaction conditions and type of substrate. We used formamidopyrimidine-DNA glycosylase (FPG) in our assay in order to measure the formation of FPG-sensitive lesions (which include 8-oxodG) in DNA of human fibroblasts in response to ionising radiation. The quantification of the DNA damage was based on calibration experiments with plasmid DNA pUC19. Dose-response curves of SSB and FPG-sensitive lesion formation in human fibroblasts VH-10 were established. A very low background level of 8-oxodG was detected in unirradiated fibroblasts (approx. 500 residues per cell).

UI MeSH Term Description Entries
D009699 N-Glycosyl Hydrolases A class of enzymes involved in the hydrolysis of the N-glycosidic bond of nitrogen-linked sugars. Glycoside Hydrolases, Nitrogen-linked,Hydrolases, N-Glycosyl,Nucleosidase,Nucleosidases,Nucleoside Hydrolase,Nitrogen-linked Glycoside Hydrolases,Nucleoside Hydrolases,Glycoside Hydrolases, Nitrogen linked,Hydrolase, Nucleoside,Hydrolases, N Glycosyl,Hydrolases, Nitrogen-linked Glycoside,Hydrolases, Nucleoside,N Glycosyl Hydrolases,Nitrogen linked Glycoside Hydrolases
D003849 Deoxyguanosine A nucleoside consisting of the base guanine and the sugar deoxyribose.
D004247 DNA A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine). DNA, Double-Stranded,Deoxyribonucleic Acid,ds-DNA,DNA, Double Stranded,Double-Stranded DNA,ds DNA
D004249 DNA Damage Injuries to DNA that introduce deviations from its normal, intact structure and which may, if left unrepaired, result in a MUTATION or a block of DNA REPLICATION. These deviations may be caused by physical or chemical agents and occur by natural or unnatural, introduced circumstances. They include the introduction of illegitimate bases during replication or by deamination or other modification of bases; the loss of a base from the DNA backbone leaving an abasic site; single-strand breaks; double strand breaks; and intrastrand (PYRIMIDINE DIMERS) or interstrand crosslinking. Damage can often be repaired (DNA REPAIR). If the damage is extensive, it can induce APOPTOSIS. DNA Injury,DNA Lesion,DNA Lesions,Genotoxic Stress,Stress, Genotoxic,Injury, DNA,DNA Injuries
D005347 Fibroblasts Connective tissue cells which secrete an extracellular matrix rich in collagen and other macromolecules. Fibroblast
D005821 Genetic Techniques Chromosomal, biochemical, intracellular, and other methods used in the study of genetics. Genetic Technic,Genetic Technics,Genetic Technique,Technic, Genetic,Technics, Genetic,Technique, Genetic,Techniques, Genetic
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000080242 8-Hydroxy-2'-Deoxyguanosine Common oxidized form of deoxyguanosine in which C-8 position of guanine base has a carbonyl group. 2'-Deoxy-7,8-Dihydro-8-Oxoguanosine,2'-Deoxy-8-Hydroxyguanosine,2'-Deoxy-8-Oxo-7,8-Dihydroguanosine,2'-Deoxy-8-Oxoguanosine,7,8-Dihydro-8-Oxo-2'-Deoxyguanosine,7-Hydro-8-Oxodeoxyguanosine,8-Hydroxydeoxyguanosine,8-Oxo-2'-Deoxyguanosine,8-Oxo-7,8-Dihydro-2'-Deoxyguanosine,8-Oxo-7,8-Dihydrodeoxyguanosine,8-Oxo-7-Hydrodeoxyguanosine,8-Oxo-Deoxyguanosine,8OHdG,8-OH-dG,8-oxo-dG,8-oxo-dGuo,8-oxodG,8-oxodGuo,2' Deoxy 7,8 Dihydro 8 Oxoguanosine,2' Deoxy 8 Hydroxyguanosine,2' Deoxy 8 Oxo 7,8 Dihydroguanosine,2' Deoxy 8 Oxoguanosine,7 Hydro 8 Oxodeoxyguanosine,7,8 Dihydro 8 Oxo 2' Deoxyguanosine,8 Hydroxy 2' Deoxyguanosine,8 Hydroxydeoxyguanosine,8 Oxo 2' Deoxyguanosine,8 Oxo 7 Hydrodeoxyguanosine,8 Oxo 7,8 Dihydro 2' Deoxyguanosine,8 Oxo 7,8 Dihydrodeoxyguanosine,8 Oxo Deoxyguanosine
D012588 Scintillation Counting Detection and counting of scintillations produced in a fluorescent material by ionizing radiation. Scintillation Counters,Counter, Scintillation,Counters, Scintillation,Counting, Scintillation,Scintillation Counter
D013462 Sulfur Radioisotopes Unstable isotopes of sulfur that decay or disintegrate spontaneously emitting radiation. S 29-31, 35, 37, and 38 are radioactive sulfur isotopes. Radioisotopes, Sulfur

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