s-adenosylhomocysteine and Precancerous-Conditions

Folic acid (FA) supplementation during carcinogenesis is controversial. Considering the impact of liver cancer as a public health problem and mandatory FA fortification in several countries, the role of FA supplementation in hepatocarcinogenesis should be elucidated. We evaluated FA supplementation during early hepatocarcinogenesis. Rats received daily 0.08 mg (FA8 group) or 0.16 mg (FA16 group) of FA/100 g body weight or water (CO group, controls). After a 2-week treatment, animals were subjected to the "resistant hepatocyte" model of hepatocarcinogenesis (initiation with diethylnitrosamine, selection/promotion with 2-acetylaminofluorene and partial hepatectomy) and euthanized after 8 weeks of treatment. Compared to the CO group, the FA16 group presented: reduced (p < 0.05) number of persistent and increased (p < 0.05) number of remodeling glutathione S-transferase (GST-P) positive preneoplastic lesions (PNL); reduced (p < 0.05) cell proliferation in persistent GST-P positive PNL; decreased (p < 0.05) hepatic DNA damage; and a tendency (p < 0.10) for decreased c-myc expression in microdissected PNL. Regarding all these parameters, no differences (p > 0.05) were observed between CO and FA8 groups. FA-treated groups presented increased hepatic levels of S-adenosylmethionine but only FA16 group presented increased S-adenosylmethionine/S-adenosylhomocysteine ratio. No differences (p > 0.05) were observed between experimental groups regarding apoptosis in persistent and remodeling GST-P positive PNL, and global DNA methylation pattern in microdissected PNL. Altogether, the FA16 group, but not the FA8 group, presented chemopreventive activity. Reversion of PNL phenotype and inhibition of DNA damage and of c-myc expression represent relevant FA cellular and molecular effects.

Topics: Animals; Apoptosis; Body Weight; Cell Proliferation; Cell Transformation, Neoplastic; Chemoprevention; Dietary Supplements; DNA Damage; DNA Methylation; Folic Acid; Gene Expression; Genes, myc; Glutathione Transferase; Liver; Liver Neoplasms, Experimental; Male; Organ Size; Precancerous Conditions; Rats; Rats, Wistar; S-Adenosylhomocysteine; S-Adenosylmethionine

Low dietary folate intake is associated with an increased risk for colon cancer; however, relevant genetic animal models are lacking. We therefore investigated the effect of targeted ablation of two folate transport genes, folate binding protein 1 (Folbp1) and reduced folate carrier 1 (RFC1), on folate homeostasis to elucidate the molecular mechanisms of folate action on colonocyte cell proliferation, gene expression, and colon carcinogenesis. Targeted deletion of Folbp1 (Folbp1(+/-) and Folbp1(-/-)) significantly reduced (P < 0.05) colonic Folbp1 mRNA, colonic mucosa, and plasma folate concentration. In contrast, subtle changes in folate homeostasis resulted from targeted deletion of RFC1 (RFC1(+/-)). These animals had reduced (P < 0.05) colonic RFC1 mRNA and exhibited a 2-fold reduction in the plasma S-adenosylmethionine/S-adenosylhomocysteine. Folbp1(+/-) and Folbp1(-/-) mice had larger crypts expressed as greater (P < 0.05) numbers of cells per crypt column relative to Folbp1(+/+) mice. Colonic cell proliferation was increased in RFC1(+/-) mice relative to RFC1(+/+) mice. Microarray analysis of colonic mucosa showed distinct changes in gene expression specific to Folbp1 or RFC1 ablation. The effect of folate transporter gene ablation on colon carcinogenesis was evaluated 8 and 38 weeks post-azoxymethane injection in wild-type and heterozygous mice. Relative to RFC1(+/+) mice, RFC1(+/-) mice developed increased (P < 0.05) numbers of aberrant crypt foci at 8 weeks. At 38 weeks, RFC1(+/-) mice developed local inflammatory lesions with or without epithelial dysplasia as well as adenocarcinomas, which were larger relative to RFC1(+/+) mice. In contrast, Folbp1(+/-) mice developed 4-fold (P < 0.05) more lesions relative to Folbp1(+/+) mice. In conclusion, Folbp1 and RFC1 genetically modified mice exhibit distinct changes in colonocyte phenotype and therefore have utility as models to examine the role of folate homeostasis in colon cancer development.

Topics: Animals; Azoxymethane; Carcinogens; Carrier Proteins; Cell Cycle; Cell Transformation, Neoplastic; Colon; Colonic Neoplasms; Folate Receptors, GPI-Anchored; Gene Expression Profiling; Gene Silencing; Genetic Predisposition to Disease; Kidney; Male; Membrane Transport Modulators; Membrane Transport Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Oligonucleotide Array Sequence Analysis; Precancerous Conditions; Receptors, Cell Surface; Reduced Folate Carrier Protein; Reverse Transcriptase Polymerase Chain Reaction; S-Adenosylhomocysteine; S-Adenosylmethionine

Several epidemiological studies have suggested a modulatory effect of dietary folate intake on the risk of colorectal cancer. The molecular basis for this inverse association is not clearly understood, but may involve alterations in DNA methylation. In this study, we examined the levels of methylation intermediates [S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH)] and of global DNA methylation in the pre-neoplastic small intestine of Min (multiple intestinal neoplasia) mice. We also studied the effect of folate/choline deficiency on these parameters and on tumor multiplicity in this animal model. In folate-adequate Min mice, we identified positive linear correlations between SAM or SAH and tumor numbers (R(2) = 0.38, P < 0.005; R(2) = 0.26, P = 0.025, respectively). A positive correlation between global DNA hypomethylation and tumor multiplicity was also observed (R(2) = 0.29, P = 0.014). These three biochemical determinants (SAM, SAH and DNA hypomethylation) may, therefore, serve as early markers of cell transformation. Folate/choline deficiency, however, did not produce a consistent effect on tumor numbers in three separate experiments. As an increase in tumor numbers was observed only in folate- and choline-deficient mice with low levels of SAM and DNA hypomethylation, the modulatory role of folate may be dependent on the transformation state of the cell.

Topics: Animals; Biomarkers; Choline Deficiency; Diet; DNA Methylation; Folic Acid Deficiency; Intestinal Neoplasms; Methionine; Mice; Neoplasms; Precancerous Conditions; S-Adenosylhomocysteine; S-Adenosylmethionine

The development of gamma-glutamyltranspeptidase (GGT)-positive foci, in Wistar rats, initiated with diethylnitrosamine and subjected to selection according to 'resistant hepatocyte' protocol, was coupled, 7 weeks after initiation, with liver DNA hypomethylation and with a fall in S-adenosylmethionine/S-adenosylhomocysteine (SAM/SAH) ratio, and in 5-methylthio-adenosine (MTA) content. A 15-day treatment with SAM, started 1 week after selection, caused a dose-dependent decrease in the development of GGT-positive foci, recovery of liver SAM/SAH ratio and MTA level, and liver DNA methylation. A 12-day treatment with 20 mumol/kg per day of 5-azacytidine (AzaC), starting 1 week after selection, enhanced growth of GGT-positive foci, caused strong DNA hypomethylation, and partially counteracted the inhibition of GGT-positive foci growth, without affecting recovery of SAM/SAH ratio and MTA level, induced by SAM. These results suggest a role of DNA methylation in the antipromoting effect of SAM.

Topics: Adenosine; Animals; Azacitidine; Biomarkers, Tumor; Deoxyadenosines; Diethylnitrosamine; gamma-Glutamyltransferase; Liver; Liver Neoplasms, Experimental; Male; Precancerous Conditions; Rats; Rats, Inbred Strains; S-Adenosylhomocysteine; S-Adenosylmethionine; Thionucleosides

S-adenosylmethionine:S-adenosylhomocysteine (SAM/SAH) ratio, 5-methylcytosine (5mC) DNA content, and methylation and expression of c-myc, c-Ha-ras and c-Ki-ras have been studied in liver nodules, induced by diethylnitrosamine according to the 'resistant hepatocyte' model, and in regenerating liver (RL) between 0.5 and 72 h after partial hepatectomy (PH). Nodules, 11, 13 and 21 weeks after initiation, grew actively, showed a low tendency to remodel (persistent nodules), and did not exhibit carcinomatous changes. They underwent extensive remodeling after a 1-week SAM treatment (64 mumol/kg/day), and decreased in size and number after a 3-11-week treatment. A low SAM/SAH ratio was coupled, in nodules, with a high labeling index (LI), 2-fold fall in 5mC DNA content, increase in c-myc, c-Ha-ras and c-Ki-ras expression and hypomethylation of CCGG sequences in the DNA hybridizing with the three protooncogenes. In RL a low SAM/SAH ratio, overall DNA hypomethylation and enhanced c-myc expression were first observed 0.5 h after PH, reached a peak at 5 h and progressively returned to pre-PH levels later on. Maximum expression of c-Ha-ras and c-Ki-ras occurred 24-30 h after PH, roughly coincident with the LI peak. However, no great modifications of the methylation pattern of protooncogene CCGG sequence occurred at any time after PH, indicating the presence of hypomethylated genes and/or DNA sequences different from those investigated in this paper. SAM injection to nodule-bearing rats, for 1-11 weeks before killing, and to hepatectomized rats, 2 days before PH and then up to killing, largely prevented decrease in the SAM/SAH ratio and overall DNA methylation and inhibited LI and protooncogene expression. In nodules these effects were proportional to the treatment length and coupled with methylation of CpG residues in the CCGG sequence of the three protooncogenes studied. SAM treatment left the methylation pattern of these genes unchanged in RL. Kinetics of increase in protooncogene expression suggest a role in the regulation of cell cycle in RL. However, decrease in the SAM/SAH ratio, protooncogene hypomethylation and enhanced expression are apparently stable in nodules 11-21 weeks after initiation and could be implicated in continuous nodule growth and progression. Control of DNA methylation and gene expression by exogenous SAM could be a mechanism of the SAM anti-progression effect.

Topics: Animals; Diethylnitrosamine; Genes, ras; Homocysteine; Immunoblotting; Liver; Liver Neoplasms, Experimental; Liver Regeneration; Male; Methylation; Precancerous Conditions; Proto-Oncogenes; Rats; Rats, Inbred F344; RNA; S-Adenosylhomocysteine; S-Adenosylmethionine

Prior studies by our laboratory, utilizing the 1,2-dimethylhydrazine experimental model of colonic cancer, had shown that administration of this procarcinogen for 5 weeks was found to increase phospholipid methyltransferase activity and the fluidity of rat distal colonic brush-border membranes. The present studies were conducted to further explore these 'premalignant' colonic phenomena. Male albino rats of the Sherman strain were subcutaneously injected with dimethylhydrazine (20 mg/kg body weight per week) or diluent for 5 weeks. Animals from each group were killed, distal colonic tissue harvested and the levels of S-adenosylmethionine, S-adenosylhomocysteine and decarboxylated S-adenosylmethionine measured by high performance liquid chromatography. The activity of methionine adenosyltransferase was also examined in these tissues. Additionally, brush-border membranes were isolated from the distal colonocytes of control and treated-animals and examined and compared with respect to their phospholipid methylation activities as well as their lipid fluidity as assessed by the rotational mobilities of the probes 1,6-diphenyl-1,3,5-hexatriene and DL-12-(9-anthroyl)stearic acid and translational mobility of the fluorophore pyrenedecanoic acid. The results of these studies demonstrated: (1) phospholipid methyltransferase activity in rat colonic plasma membranes was increased concomitantly with increases in the cellular levels of S-adenosylmethionine and the S-adenosylmethionine/S-adenosylhomocysteine ratio in the distal colonic segment of treated-animals; and (2) the lateral diffusion of rat distal colonic brush-border membrane lipids, as assessed by the ratio of excimer/monomer fluorescence intensities of the fluorophore pyrenedecanoate, was also increased after dimethylhydrazine administration to these animals for 5 weeks.

Topics: 1,2-Dimethylhydrazine; Animals; Colonic Neoplasms; Decanoic Acids; Diffusion; Dimethylhydrazines; Homocysteine; Membrane Lipids; Methionine Adenosyltransferase; Methylhydrazines; Precancerous Conditions; Rats; S-Adenosylhomocysteine; S-Adenosylmethionine