Assessment of DNA Damage

Implicit in the discussion of the chemistry of formation for DNA adducts in vitro is the generation of DNA adducts in vivo. The previously described pyrimidopuri-none, propano, and etheno adducts are found in human tissue samples and cultured human cell lines. The estimated levels of these adducts range from one to 10 adducts/108 bp DNA. Owing to the relatively low abundance of these lesions amid a preponderance of impurities, highly sensitive and specific analytical 

Xanthine oxidoreductase and aldehyde oxidase are the principal enzymes involved in the oxidation of exocydic DNA adducts. Their role in purine catabolism and in the oxidation of nitrogen-rich heterocycles is well documented [143-145]. 

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Castano, A Becerril, C. (2004) In vitro assessment of DNA damage after short and long-term exposure to benzo(a)pyrene using RAPD and the RTG-2 fish cell line. Mutation Reseatch 552 141-151. 

Santos, F. V. Coins, I. M. S. Silva, M. A. Vilegas, W. Varanda, E. A., Assessment of DNA damage by extracts and fractions of Strychnos pseudoquina, a Brazilian medicinal plant with antiulcerogenic activity. Food Chem. Tox/co/., 2006, 44, 1585-1589. 

Ozcagli, E., Sardas, S., and Biri, A. (2005) Assessment of DNA damage in postmenopausal women under hormone replacement therapy. Maturitas, 51, 280-285. 

Derivatives for nucleic acids have been reviewed [48]. The application of TMS and d9-labelled TMS derivatives is useful in the structure elucidation of newly discovered nucleosides [49]. Recent examples include the GC-MS (SIM) of trimethylsilylated nucleosides in the assessment of DNA damage [50] and the identification of a novel nucleoside, l,N -dimethyladenosine, in human cancer urine [51]. 

Adequate body reserves and normal metabolic integrity. This is the (possibly untestable) goal. Both immune function and rninirnization of DNA damage offer potential methods of assessing optimum micronutrient status, but both are affected by a variety of different nutrients and other factors (Fenech, 2001). 

It is well known that lipid peroxidation, DNA singlestrand breaks, and other forms of DNA damage occur in response to oxidative stress (Ames et al, 1982). Depletion of reduced glutathione also commonly precedes or accompanies lipid peroxidation and oxidative stress (Muldoon and Stohs, 1991 Omar et al, 1990). In an in vivo study, the effects of ricin administered orally on hepatic lipid peroxidation, nonprotein sulfhydryl content, and DNA singlestrand breaks were assessed in mice (Muldoon et al, 1992). The incidence of hepatic DNA damage increased 2.9-, 2.8-, and 2.4-fold relative to control values at 24, 36, and 48 h post-treatment with ricin, respectively. Hepatic nonprotein sulfhydryl concentration decreased significantly from 51 to 65% to control values at 24, 36, and 48 h post-treatment (Figures 25.2 and 25.3). 

Lyras L, Cairns NJ, Jeriner A, Jeimer P, Halliwell B (1997) An assessment of oxidadve damage to proteins, lipids, and DNA in brain from paderits widi Alzheimer s disease. J Neurochem 68 2061—2069. 

The Assessment of DNA Crosslinking by Alkaline Elution. DNA damage, that is, interstrand crosslinks, DNA-protein crosslinks, and strand breaks, was determined using the alkaline elution technique (7, ). Cells labeled with C-thymidine for 20-24 hr were deposited on a membrane filter and lysed with a detergent-containing solution. An alkaline solution (pH 12.1-12.2) was then slowly pumped through the filter, and fractions were collected to determine the rate of release of DNA from the filter. For assay of crosslinks, the cells were exposed to x-ray at 0 C prior to deposition on the filter. In order to improve quantitation, control cells labeled with H-thymidine and x-irradiated at 0 were mixed with the experimental Relabeled cells prior to deposition on the filters. The elution of H-DNA serves as an internal reference for normalization of the elution of C-DNA. 

Ochratoxin A was shown to induoe DNA strand breaks as assessed by comet assay in liver, kidney and spleen of Fisoher 344 rats given doses of 0, 0.25, 0.50, 1.0 or 2.0 mg/kg bw per day for 2 weeks, 5 days/week. In liver and kidney, the extent of DNA damage was further enhanced in a dose-dependent manner in the presence of the repair enzyme Fpg, which converts oxidative DNA damage into strand breaks, suggesting the presence of oxidative DNA damage (Mally et al., 2005b). 

MORRIS ID, ILOTT s, DIXON L and BRisoN DR (2002) The spectrum of DNA damage in human sperm assessed by single cell gel electrophoresis (Comet assay) and its relationship to fertilization and embryo development. Hum Reprod, 17,990-998. 

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