Hypomethylation of DNA and the insulin-like growth factor-II gene in dichloroacetic and trichloroacetic acid-promoted mouse liver tumors
Introduction
Dichloroacetic acid (DCA) and trichloroacetic acid (TCA) are mouse liver carcinogens (Bull et al., 1990, Pereira, 1996, Pereira and Phelps, 1996, Pereira et al., 1997, Boorman, 1999) that are found in finished drinking water being formed as by-products during chlorine disinfection (Uden and Miller, 1983, Chen and Weisel, 1998, Boorman, 1999). DCA and TCA have been shown to induce hepatocellular adenomas and carcinomas (Herren-Freund et al., 1987, Bull et al., 1990, Pereira, 1996, Boorman, 1999) and to promote N-methyl-N-nitrosourea (MNU)-initiated foci of altered hepatocytes and liver tumors in B6C3F1 mice (Pereira and Phelps, 1996, Pereira et al., 1997). Both chemicals have been reported to increase cell proliferation in mouse liver (Pereira, 1996, Stauber and Bull, 1997, Ge et al., 2001b). Increased hepatic cell proliferation is associated with increased expression of growth-related genes including insulin-like growth factor-II (IGF-II) (Werner and Le Roith, 2000, Furstenberger and Senn, 2002).
IGF-II has both mitogenic and anti-apoptotic activity in the liver being important in the regulation of cell proliferation, death and differentiation (Werner and Le Roith, 2000, Furstenberger and Senn, 2002). In mouse liver, IGF-II is imprinted, that is the parental allele is expressed while the maternal allele is methylated and silent (Khandwala et al., 2000, Scharf et al., 2001). Loss of imprinting and increased expression of IGF-II have been observed in a variety of malignancies including liver tumors (Khandwala et al., 2000, Scharf et al., 2001).
DNA methylation is a naturally occurring modification of eukaryotic DNA that involves an addition of a methyl group to the 5-position carbon of the cytosine ring to form 5-methylcytosine (5-MeC) at CpG dinucleotides or CpNpG trinucleotides, which are frequently clustered in regions of about 1−2 kb in length, called CpG islands, in or near the promoter and first coding exon of genes (Baylin et al., 2001). Decreased DNA methylation (DNA hypomethylation) is a common event in many cancers including liver cancer. Liver tumors induced by a choline-methionine deficient diet in mice and rats exhibited DNA hypomethylation and decreased methylation of protooncogenes including H-ras, c-myc and c-fos (Wainfan and Poirier, 1992, Counts et al., 1996, Henning and Swendseid, 1996). We have previously demonstrated that the protooncogenes, c-jun and c-myc were hypomethylated in mouse liver tumors initiated by N-methyl-N-nitrosourea (MNU) and promoted by either DCA or TCA (Tao et al., 1998, Tao et al., 2000b). Furthermore, short-term treatment with either chloroacetic acid as well as other non-genotoxic carcinogens including trichloroethylene, chloroform and other trihalomethanes, and peroxisome proliferators induced hypomethylation of the c-jun and c-myc genes in mouse liver (Tao et al., 1998, Tao et al., 1999, Coffin et al., 2000, Tao et al., 2000a; Ge et al., 2001a, Ge et al., 2001b, Ge et al., 2002). We report here that the DNA and the DMR-2 of the IGF-II gene are hypomethylated in mouse liver tumors that were initiated by MNU and promoted by DCA and TCA to a greater degree than in non-involved liver tissues. We also reported that the mRNA expression of the IGF-II gene is increased in the tumors.
Section snippets
Chemicals and reagents
Ribonuclease A type III−A, Sephadex G-50, sodium metabisulfite, hydroquinone, and proteinase K were from Sigma Chemical Company, Inc. (St. Louis, MO). TA cloning kit with pCR 2.1 vector was from Invitrogen Corp. (Carlsbad, CA). Mouse monoclonal primary antibody specific for 5-MeC was purchased from the Eurogentec Company (Belgium). Horseradish peroxidase (HRP)-conjugated anti-mouse IgG was from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). Pfu DNA polymerase was from Stratagene Corp. (La
Methylation of DNA
A mouse monoclonal primary antibody specifically against 5-MeC for dot–blotting analysis was used to determine the level of 5-MeC in DNA. Specificity of the anti-5-MeC antibody was determined by the ability of 5-MeC but not cytosine to block the antibody reaction with liver DNA. On a microgram basis, the anti-5-MeC antibody was blocked by an equal amount of 5-MeC, while even a 10-fold greater amount of cytosine had no affect on the reaction of the antibody with DNA.
The reaction of the 5-MeC
Discussion
DCA and TCA are by-products of the chlorination process of drinking water (Uden and Miller, 1983, Chen and Weisel, 1998, Boorman, 1999) and metabolites of trichloroethylene and tetrachloroethylene (Ware, 1988, IARC, 1995). Both chloroacetic acids are mouse liver carcinogens (Herren-Freund et al., 1987, Bull et al., 1990, Pereira, 1996, Boorman, 1999) and liver tumor promoters (Pereira and Phelps, 1996, Pereira et al., 1997), that have been classified by the US Environmental Protection Agency as
Acknowledgments
This research was supported in part by US Environmental Protection Agency Grant No. R-82808301-0.
References (38)
- et al.
Liver tumor induction in B6C3F1 mice by dichloroacetate and trichloroacetate
Toxicology
(1990) - et al.
Hypomethylation of DNA: a nongenotoxic mechanism involved in tumor promotion
Toxicol. Lett.
(1995) - et al.
Insulin-like growth factors and cancer
Lancet Oncol.
(2002) - et al.
The carcinogenicity of trichloroethylene and its metabolites, trichloroacetic acid and dichloroacetic acid, in mouse liver
Toxicol. Appl. Pharmacol.
(1987) - et al.
In vivo MRI measurements of tumor growth induced by dichloroacetate: implications for mode of action
Toxicology
(2000) Carcinogenic activity of dichloroacetic acid and trichloroacetic acid in the liver of female B6C3F1 mice
Fund. Appl. Toxicol.
(1996)- et al.
Promotion by mixtures of dichloroacetic acid and trichloroacetic acid of N-methyl-N-nitrosourea-initiated cancer in the liver of female B6C3F1 mice
Cancer Lett.
(1997) - et al.
Promotion by dichloroacetic acid and trichloroacetic acid of N-methyl-N-nitrosourea-initiated cancer in the liver of female B6C3F1 mice
Cancer Lett.
(1996) - et al.
Differences in phenotype and cell replicative behavior of hepatic tumors induced by dichloroacetate (DCA) and trichloroacetate (TCA)
Toxicol. Appl. Pharmacol.
(1997) - et al.
Effect of dichloroacetic acid and trichloroacetic acid on DNA methylation in liver and tumors of female B6C3F1 mice
Toxicol. Sci.
(1998)
Hypomethylation and overexpression of c-jun and c-myc protooncogenes and increased DNA methyltransferase activity in dichloroacetic and trichloroacetic acid-promoted mouse liver tumors
Cancer Lett.
Aberrant patterns of DNA methylation, chromatin formation and gene expression in cancer
Hum. Mol. Genet.
Drinking water disinfection byproducts: review and approach to toxicity evaluation
Environ. Health Perspect.
Concentration changes of halogenated disinfection by-products in a drinking water distribution system
J. Am. Water Works Assoc.
Effect of trihalomethanes on cell proliferation and DNA methylation in female B6C3F1 mouse liver
Toxicol. Sci.
Cell proliferation and global methylation status changes in mouse liver after phenobarbital and/or choline-devoid, methionine-deficient diet administration
Carcinogenesis
Effect of peroxisome proliferators on the methylation and protein level of the c-myc protooncogene in B6C3F1 mice liver
J. Biochem. Mol. Toxicol.
Cited by (30)
Epigenetic alterations induced by genotoxic occupational and environmental human chemical carcinogens: An update of a systematic literature review
2022, Mutation Research - Reviews in Mutation ResearchCitation Excerpt :Several studies utilized trichloroethylene metabolites, dichloroacetic acid, and trichloroacetic acid. These studies concluded that trichloroethylene metabolite exposure induced DNA hypomethylation [236,237] and increased expression of c-jun and c-myc genes [238] within liver tumors. Dichloroacetic acid-associated hypomethylation was prevented by methionine co-administration [239].
DNA methyltransferase expression and DNA hypomethylation status in human hepatocytes following trichloroacetic acid exposure
2019, Biochemical and Biophysical Research CommunicationsCitation Excerpt :TCA has been shown to increase cell proliferation and induced global DNA hypomethylation in mouse liver [11]. TCA has also been shown to up-regulate many protooncogenes, such as, H-ras, K-ras, Raf, C-fos, C-jun, IGF-II and C-myc, and lead to hypomethylation around the promoters of above genes [12]. Thus, epigenetic mechanisms play an important role in the toxic effect of TCA.
Synthesis, characterization and mechanism analysis of modified crayfish shell biochar possessed ZnO nanoparticles to remove trichloroacetic acid
2017, Journal of Cleaner ProductionCitation Excerpt :It has been proved that TCAA could lead to carcinogenic, reproductive and developmental problems when its content in the water exceeds a certain value (Tao et al., 1998, 2000; Veeramachaneni et al., 2001). The toxicity of TCAA can be manifested in the following aspects: 1) Destroy DNA structure (Tao et al., 1998, 2004, 2000); 2) Influence peroxisome (Deangelo et al., 1989; Parrish et al., 1996); 3) Increase the incidence of cancer (Herrenfreund et al., 1987); 4) Interfere reproductive organs and cells (Veeramachaneni et al., 2001). Therefore, it has become a hot topic in the field of water treatment to deal with the disinfection by-products such as TCAA by an economic and environmental-friendly method.
Hypermethylation of repetitive DNA elements in livers of mice fed an atherogenic diet
2012, NutritionCitation Excerpt :Total RNA was extracted using EasyBlue reagent (Intron Biotechnology, Seongnam, Republic of Korea) by the procedure specified by the manufacturer. The dot-blot analysis using monoclonal antibody against 5-MeC was performed to assess levels of global DNA methylation in livers of experimental mice [10]. Genomic DNA (2 mg) was denatured by heating to 100°C for 10 min in a denaturation solution containing 0.4 N NaOH and 10 mM EDTA.