Oxidative DNA damage and formalin-fixation procedures. Issue 5 (1st July 2014)

• Record Type: Journal Article
• Authors: Peluso, Marco E. M.; Munnia, Armelle; Tarocchi, Mirko; Giese, Roger W.; Annaratone, Laura; Bussolati, Gianni; Bono, Roberto
• Is Part Of: Toxicology research. Volume 3:Issue 5(2014:Sep.)
• Page Start: 341
• Page End: 349
• Publication Date: 2014-07-01
• Subjects: Toxicology -- Periodicals; 615.9005
• DOI: 10.1039/c4tx00046c
• Languages: English
• ISSNs: 2045-452X

Abstract

Graphical Abstract: Abstract: Formaldehyde is the most commonly used fixative for the preparation of formalin-fixed paraffin-embedded tissues (FFPETs) in the "reduction rooms" of pathology wards. Therefore, we analysed for the generation of 3-(2-deoxy-β-d -erythro-pentafuranosyl)pyrimido[1, 2- α ]purin-10(3 H )-one adducts (M1 dG), an exocyclic DNA adduct considered to be a biomarker of oxidative stress and lipid peroxidation, in DNA extracted from the FFPETs of liver and lung samples of six C57BL/6 control mice and from the FFPETs of four cancer patients with respect to paired flash-frozen tissues by 32 P-postlabeling. When the experimental animals were examined, the percentage of M1 dG adducts was about 4–6 fold greater with the FFPET mouse samples as compared to flash-frozen mouse samples. Specifically, 4.75 M1 dG ± 0.21 (SE) per 10 6 normal nucleotides (nn) were detected in the FFPET liver samples, and 1.07 M1 dG ± 0.08 (SE) per 10 6 nn in the flash-frozen liver samples ( p = 0.02). Then, 3.80 M1 dG ± 0.73 (SE) per 10 6 nn were measured in the FFPET lung samples, and 1.02 M1 dG ± 0.07 (SE) per 10 6 nn in the flash-frozen lung samples ( p = 0.02). Also, significantly increased levels of oxidatively damaged DNA were detected in the human colon DNA from the FFPETs with respect to the flash-frozen tissues. There were 30.2 M1 dG ± 7.7 (SE) per 10 8 nn in the colon mucosa DNA from the FFPETs and 4.4 M1 dG ± 0.7 (SE) per 10 8 nn in the corresponding flash-frozen human tissues ( p = 0.016). Formalin penetration through cell membrane components induces excessive oxidative stress, causing both direct oxidation of DNA and increased lipid peroxidation, which in turn produce M1 dG adducts, a kind of DNA damage that can partially block DNA synthesis and induce error prone translesion synthesis. An excess of exocyclic DNA adducts in formalin-fixed specimens can stall DNA polymerases and contribute to the induction of artefactual sequence alterations during PRC amplification.