DNA Adducts In Livers Of Mice Research Articles

Dietary curcumin post-treatment enhances the disappearance of B(a)P-derived DNA adducts in mouse liver and lungs

To study the post-treatment effects of dietary curcumin on the levels of benzo(a)pyrene [B(a)P]-induced DNA adducts, mice were administered oil or B(a)P and randomized into 7 subgroups after 24 h. One of the subgroups from both the oil and B(a)P groups was killed at 24 h while the remaining 6 subgroups were shifted to powdered control or 0.05% curcumin diet and killed after 24, 72 and 120 h (experiment 1), and 7, 14, and 28 days (experiment 2). Quantitative comparisons of BPDE-DNA nuclear adducts (area and intensity) in immunohistochemically stained lungs and liver sections was carried out by IHC profiler. A time-dependent decrease in the levels of adducts in B(a)P-treated animals was further enhanced by curcumin exposure compared to the levels in time-matched controls. To assess the contribution of apoptosis and cell proliferation in observed curcumin-mediated enhanced decrease of BPDE-DNA adducts, comparative evaluation of apoptosis and cell proliferation markers was undertaken. Results suggested enhancement of B(a)P-induced apoptosis in liver and lungs by curcumin during 24–120 h while no such enhancement was observed at 7–28 days. Results suggest curcumin-mediated enhancement in apoptosis (experiment 1) and adduct dilution (experiment 2) to be the reason for the observed higher decrease of BPDE-DNA adducts.

Detection and Quantitation of Benzo[a]pyrene-Derived DNA Adducts in Mouse Liver by Liquid Chromatography−Tandem Mass Spectrometry: Comparison with 32P-Postlabeling

The polycyclic aromatic hydrocarbon, benzo[a]pyrene (B[a]P) is a proven animal carcinogen that is potentially carcinogenic to humans. B[a]P is an ubiquitous environmental pollutant and is also present in tobacco smoke, coal tar, automobile exhaust emissions, and charred food. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method using electrospray ionization and selected reaction monitoring (SRM) has been developed for the detection of 10-(deoxyguanosin-N(2)-yl)-7,8,9-trihydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene (B[a]PDE-N(2)dG) adducts formed in DNA following the metabolic activation of B[a]P to benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (B[a]PDE). The method involves enzymatic digestion of the DNA sample to 2'-deoxynucleosides following the addition of a stable isotope internal standard, [(15)N(5)]B[a]PDE-N(2)dG, and then solid phase extraction to remove unmodified 2'-deoxynucleosides prior to analysis by LC-MS/MS SRM. The limit of detection of the method was 10 fmol (approximately 3 B[a]PDE-N(2)dG adducts per 10(8) 2'-deoxynucleosides) using 100 microg of calf thymus DNA as the matrix. Calf thymus DNA reacted with B[a]PDE in vitro and mouse liver DNA samples at different time points following dosing intraperitoneally with 50, 100, and 200 mg/kg B[a]P was analyzed. Three stereoisomers of the B[a]PDE-N(2)dG adduct were detected following the reaction of calf thymus DNA with B[a]PDE in vitro. The levels of B[a]PDE-N(2)dG DNA adducts in the mice livers were found to increase in a dose-dependent manner with adducts reaching maximal levels at 1-3 days and then gradually decreasing over time but still detectable after 28 days. A very good correlation (r = 0.962, p < 0.001) was observed between the results obtained for the mouse liver DNA samples using LC-MS/MS SRM as compared to those obtained using a (32)P-postlabeling method. However, the levels of adducts observed following (32)P-postlabeling using butanol enrichment were approximately 3.7-fold lower. The LC-MS/MS method allowed the more precise quantitation of DNA adduct levels that were structurally characterized, in addition to a reduction in the time taken to perform the analysis when compared with the (32)P-postlabeling method.

The Different Genotoxicity of P-Dichlorobenzene in Mouse and Rat: Measurement of the in Vivo and in Vitro Covalent Interaction with Nucleic Acids

Twenty-two hours after i.p. injection to male Wistar rats and BALB/c mice para-dichlorobenzene (p-DCB) is bound covalently to DNA from liver, kidney, lung and stomach of mice but not of rats. DNA adducts in mouse liver are repaired in seventy-two hours. The covalent binding index value, calculated on the labelling of mouse liver DNA, classifies p-DCB as a weak initiator with an oncogenic activity lower than that of chlorobenzene. The labelling of RNA and proteins from the different organs of both species is, however, low. In vitro interaction with calf thymus DNA mediated by mouse and rat microsomes from liver and lung did occur. Binding extent was strongly reduced by addition of 2-diethylaminoethyl-2,2-diphenylvalerate hydrochloride (SKF 525-A) to the microsomal standard incubation mixture, whereas it was enhanced by adding GSH. Cytosolic fractions from kidney and lung were able to induce binding of p-DCB to DNA to a lower extent with respect to microsome-mediated binding. These results indicate that microsomal mixed function oxidase system and microsomal GSH-transferases can be involved in overall activating metabolism whereas cytosolic GSH-transferases play a minor role. This study, which is a part of a structure-activity relationship approach on benzene and its haloderivatives, provides the first evidence of genotoxicity of p-DCB in mammalian cell. It allows to partly explain variations of susceptibility of different species to hepatocarcinogenesis and of hepatotoxicity of different isomers.

DNA damage and repair in mouse liver.

The formation of DNA adducts in mouse liver has been demonstrated for numerous chemicals including members of most major classes of carcinogens. Considerably less is known about the persistence and repair of DNA adducts in mouse liver. Likewise, major gaps in present knowledge exist regarding the molecular dosimetry of DNA adducts and their potential for miscoding during continuous exposure to high versus low doses of carcinogens. A prime example of this is 2-acetylaminofluorene (2-AAF), the carcinogen used in the ED01 megamouse study. There are no molecular dosimetry studies on the DNA adducts of 2-AAF, even though such a unique data base exists for the dose-response relationship of mouse liver tumors. Reviewing the pertinent literature, identifying deficiencies, and conducting the required research will hopefully permit a better determination of the relevance of mouse liver tumors to man.