Journal Article

Induction of direct adducts, apurinic/apyrimidinic sites and oxidized bases in nuclear DNA of human HeLa S3 tumor cells by tetrachlorohydroquinone

Po-Hsiung Lin, Jun Nakamura, Shuji Yamaguchi, David K. La, Patricia B. Upton and James A. Swenberg

in Carcinogenesis

Volume 22, issue 4, pages 635-639
Published in print April 2001 | ISSN: 0143-3334
Published online April 2001 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/22.4.635
Induction of direct adducts, apurinic/apyrimidinic sites and oxidized bases in nuclear DNA of human HeLa S3 tumor cells by tetrachlorohydroquinone

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DNA damage induced by tetrachlorohydroquinone (Cl4HQ), the quinonoid metabolite of pentachlorophenol (PCP), was investigated in human HeLa S3 tumor cells. Formation of one major and two minor DNA adducts in cells treated with Cl4HQ (50–300 μM) was detected by 32P-post-labeling assay and the adducts accumulated over the course of the experiment (0.5–2 h), with total adduct levels estimated to be 3–6 per 108 nucleotides. These adducts did not correspond to those derived from calf thymus DNA treated with tetrachloro-1,4-benzoquinone. Results from the apurinic/apyrimidinic (AP) sites assay indicated that the number of AP sites was 2-fold greater in cells exposed to Cl4HQ (300 μM) than the corresponding control. Further characterization of the AP sites confirmed that Cl4HQ induced predominantly (75%) putrescine-excisable AP sites in HeLa S3 cells. In parallel, the concentration of 8-hydroxy-2′-deoxyguanosine (8-HO-dG) in cells treated with Cl4HQ for 0.5 and 2 h was increased 2- and 5-fold, respectively, compared with the control. The extent of oxidative DNA damage induced by Cl4HQ was approximately two orders of magnitude greater than those of direct DNA adducts. Overall, it appears that reactive oxygen species mediate the parallel formation of AP sites and 8-HO-dG in HeLa S3 cells following treatment with Cl4HQ and that the contribution of depurination/depyrimidination of direct DNA adducts is relatively insignificant compared with the formation of oxidized AP sites. We conclude that putrescine-excisable AP sites represent a major type of ROS-mediated oxidative DNA damage in cellular DNA induced by Cl4HQ and may play a role in PCP-induced clastogenicity in mammalian cells.

Keywords: AP, apurinic/apyrimidinic; DMEM, Dulbecco's modified Eagle's medium; ESA, electrochemical array detector; BHT, butylated hydroxytolvene; HPLC-ECD, HPLC/electrochemical detection; 8-HO-dG, 8-hydroxy-2′-deoxyguanosine; PCP, pentachlorophenol; PEI, polyethyleneimine; ROS, reactive oxygen species; RAL, relative adduct levels; Cl4HQ, tetrachlorohydroquinone.

Journal Article.  3459 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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