s-adenosylhomocysteine and Cell-Transformation--Neoplastic

Topics: Acetylserotonin O-Methyltransferase; Animals; Biopterins; Cell Transformation, Neoplastic; Female; gamma-Aminobutyric Acid; Gonads; Indoles; Male; Mammals; Melatonin; Models, Biological; Neopterin; Neurophysins; Norepinephrine; Pineal Gland; Pituitary Hormones, Anterior; S-Adenosylhomocysteine; S-Adenosylmethionine; Seasons; Vasotocin

Topics: Adenosine; Animals; Azacitidine; Carcinogens; Cell Line; Cell Transformation, Neoplastic; Cell Transformation, Viral; Choline; DNA; DNA (Cytosine-5-)-Methyltransferases; DNA, Viral; Ethionine; Gene Expression Regulation; Homocysteine; Humans; Methionine; Methylation; Methyltransferases; Neoplasms; Oncogenes; Oncogenic Viruses; S-Adenosylhomocysteine; S-Adenosylmethionine; Simian virus 40

The Long-Evans Cinnamon (LEC) rat shows age-dependent hepatic manifestations that are similar to those of Wilson's disease (WD). The pathogenic process in the brain has, however, not been evaluated in detail due to the rarity of the neurological symptoms. However, copper accumulation is noted in LEC rat brain tissue from 24 weeks of age, which results in oxidative injuries. The current study investigated the gene expression profiles of LEC rat brains at 24 weeks of age in order to identify the important early molecular changes that underlie the development of neurological symptoms in WD. Biological ontology-based analysis revealed diverse altered expressions of the genes related to copper accumulation. Of particular interest, we found altered expression of genes connected to mitochondrial respiration (Sdhaf2 and Ndufb7), calcineurin-mediated cellular processes (Ppp3ca, Ppp3cb, and Camk2a), amyloid precursor protein (Anks1b and A2m) and alpha-synuclein (Snca). In addition to copper-related changes, compensatory upregulations of Cp and Hamp reflect iron-mediated neurotoxicity. Of note, reciprocal expression of Asmt and Bhmt is an important clue that altered S-adenosylhomocysteine metabolism underlies brain injury in WD, which is directly correlated to the decreased expression of S-adenosylhomocysteine hydrolase in hepatic tissue in LEC rats. In conclusion, our study indicates that diverse molecular changes, both variable and complex, underlie the development of neurological manifestations in WD. Copper-related injuries were found to be the principal pathogenic process, but Fe- or adenosylhomocysteine-related injuries were also implicated. Investigations using other animal models or accessible human samples will be required to confirm our observations.

Topics: alpha-Synuclein; Animals; Antimicrobial Cationic Peptides; Brain; Cell Transformation, Neoplastic; Cluster Analysis; Copper; Disease Models, Animal; Gene Expression Profiling; Gene Expression Regulation; Hepatolenticular Degeneration; Hepcidins; Humans; Iron; Liver; Mitochondria; Neurons; Oligonucleotide Array Sequence Analysis; Rats; Rats, Inbred LEC; Real-Time Polymerase Chain Reaction; Reproducibility of Results; S-Adenosylhomocysteine; Time Factors; Visual Pathways

Chronic inflammation is an underlying risk factor for colon cancer. Tumor necrosis factor alpha (TNF-α) plays a critical role in the development of inflammation-induced colon cancer in a mouse model. S-adenosylmethionine (SAMe) and its metabolite methylthioadenosine (MTA) can inhibit lipopolysaccharide-induced TNF-α expression in macrophages. The aim of this work was to examine whether SAMe and MTA are effective in preventing inflammation-induced colon cancer and if so identify signaling pathways affected. Balb/c mice were treated with azoxymethane (AOM) and dextran sulfate sodium to induce colon cancer. Two days after AOM treatment, mice were divided into three groups: vehicle control, SAMe or MTA. Tumor load, histology, immunohistochemistry, gene and protein expression were determined. SAMe and MTA treatment reduced tumor load by ∼40%. Both treatments raised SAMe and MTA levels but MTA also raised S-adenosylhomocysteine levels. MTA treatment prevented the induction of many genes known to play pathogenetic roles in this model except for TNF-α and inducible nitric oxide synthase (iNOS). SAMe also had no effect on TNF-α or iNOS and was less inhibitory than MTA on the other genes. In vivo, both treatments induced apoptosis but inhibited proliferation, β-catenin, nuclear factor kappa B activation and interleukin (IL) 6 signaling. Effect of SAMe and MTA on IL-6 signaling was examined using Colo 205 colon cancer cells. In these cells, SAMe and MTA inhibited IL-6-induced IL-10 expression. MTA also inhibited IL-10 transcription and signal transducer and activator of transcription 3 activation. In conclusion, SAMe and MTA reduced inflammation-induced colon cancer and inhibited several pathways important in colon carcinogenesis.

Topics: Animals; Apoptosis; Azoxymethane; beta Catenin; Cell Proliferation; Cell Transformation, Neoplastic; Chemoprevention; Colonic Neoplasms; Dextran Sulfate; Inflammation; Interleukin-10; Interleukin-6; Male; Mice; Mice, Inbred BALB C; NF-kappa B; Nitric Oxide Synthase Type II; Proto-Oncogene Proteins c-akt; Purine-Nucleoside Phosphorylase; S-Adenosylhomocysteine; S-Adenosylmethionine; Signal Transduction; STAT3 Transcription Factor; Transcriptional Activation; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

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

C --> T transitions at CpG sites are the most prevalent mutations found in the p53 tumor suppressor gene in human colon tumors and in the germline (Li-Fraumeni syndrome). All of the mutational hot spots are methylated to 5-methylcytosine, and it has been hypothesized that the majority of these mutations are caused by spontaneous hydrolytic deamination of this base to thymine. We have previously reported that bacterial methyltransferases induce transition mutations at CpG sites by increasing the deamination rate of C --> U when the concentration of the methyl group donor S-adenosylmethionine (AdoMet) drops below its Km, suggesting an alternative mechanism to create these mutations. Unrepaired uracil pairs with adenine during replication, completing the C --> T transition mutation. To determine whether this mechanism could contribute to the development of human colon cancer, we examined the level of DNA (cytosine-5)-methyltransferase (MTase) expression, the concentration of AdoMet, and the activity of uracil-DNA glycosylase in human colon tissues, and searched for the presence of mutations in the MTase gene. Using reverse transcription-PCR methods, we found that average MTase mRNA expression levels were only 3.7-fold elevated in tumor tissues compared with surrounding normal mucosa from the same patient. Also, no mutations were found in conserved regions of the gene in 10 tumors sequenced. High-performance liquid chromatographic analysis of extracts from the same tissues showed that AdoMet concentrations were not reduced below the Km value for the mammalian enzyme, and the concentration ratio of AdoMet:S-adenosylhomocysteine, the breakdown product of AdoMet and the competitive MTase inhibitor, did not differ significantly. Finally, extracts from the tumor tissue efficiently removed uracil from DNA. Therefore, biochemical conditions favoring a mutagenic pathway of C --> U --> T were not found in a target tissue known to undergo a high rate of C --> T transitions at CpG sites.

Topics: Base Sequence; Cell Transformation, Neoplastic; Colonic Neoplasms; Cytosine; DNA; DNA (Cytosine-5-)-Methyltransferases; DNA Damage; DNA Glycosylases; DNA Repair; DNA, Complementary; DNA, Neoplasm; Escherichia coli; Humans; Intestinal Mucosa; Methylation; Molecular Sequence Data; Mutagenesis; Mutagenesis, Site-Directed; N-Glycosyl Hydrolases; Polymerase Chain Reaction; RNA, Messenger; RNA, Neoplasm; S-Adenosylhomocysteine; S-Adenosylmethionine; Thymidine; Uracil-DNA Glycosidase

The tumorigenic cell line termed "MCA Cl 16" was derived from C3H/10T1/2 clone (Cl) 8 cells by chemical transformation in the presence of 3-methylcholanthrene [(MCA) CAS: 56-49-5]. Transformed (Cl 16) cells were more sensitive toward the cytotoxic effect of methotrexate (MTX) than their normal counterpart Cl 8 cells. The disposition of endogenous L-homocysteine (Hcy) was investigated in these two cell lines after MTX exposure. Both nonmalignant and transformed cells exported Hcy into the extracellular medium, and only small amounts were retained within the cells. The Hcy efflux from the malignant cells was markedly increased after MTX exposure (0.5-10 microM), and this effect was almost completely prevented by 5-formyl-tetrahydrofolate (THF), whereas treatment with thymidine plus hypoxanthine did not inhibit the MTX-dependent Hcy efflux. Cytotoxic concentration of MCA reduced rather than increased the Hcy efflux from these cells. High concentrations of MTX (greater than 10 microM) were required to increase the release of Hcy from nonmalignant cells. The enhancement of Hcy export from the malignant cells in the presence of MTX was not associated with cellular build-up of S-adenosyl-L-homocysteine (AdoHcy), indicating that the amount of intracellular Hcy was kept below the level required for inhibition or reversion of the AdoHcy hydrolase reaction. MTX-dependent Hcy efflux probably reflects cellular deficiency of 5-methyl-THF required for the salvage of Hcy to methionine and may therefore be a measure of lack of this reduced folate relative to the metabolic demand.

Topics: Animals; Cell Division; Cell Line; Cell Survival; Cell Transformation, Neoplastic; Chromatography, Ion Exchange; Culture Media; Fibroblasts; Homocysteine; Methotrexate; Methylcholanthrene; Mice; Mice, Inbred C3H; Proteins; S-Adenosylhomocysteine; S-Adenosylmethionine

S-Adenosylhomocysteine (AdoHcy) is catabolized to adenosine and homocysteine through the action of AdoHcy hydrolase, and this reaction is the only known source of L-homocysteine in vertebrates. The disposition of endogenously formed L-homocysteine was investigated in isolated rat hepatocytes and nontransformed and malignant C3H/10T1/2 mouse embryo fibroblasts exposed to 3-deazaaristeromycin or D-eritadenine, compounds which are potent inhibitors of AdoHcy hydrolase. Cells in suspension release large amounts of L-homocysteine into the extracellular medium whereas small amounts are retained within the intracellular compartment. The L-homocysteine egress is inhibited by 3-deazaaristeromycin or D-eritadenine in a manner which closely parallels the inhibitory effect on AdoHcy catabolism, suggesting that L-homocysteine egress may be coupled to its formation from AdoHcy. In liver cells, the accumulation of AdoHcy exceeded the inhibition of L-homocysteine egress, whereas in the fibroblasts inhibition of egress equalled the accumulation of AdoHcy. Inhibition of AdoHcy catabolism was associated with an increase in both free and protein bound L-homocysteine in liver cells, whereas depletion of intracellular L-homocysteine occurred in the mouse embryo fibroblasts under these conditions. These data suggest that some properties of nucleoside analogues may be related to their effects on L-homocysteine metabolism. Furthermore, L-homocysteine is exported into the extracellular medium in proportion to the formation from AdoHcy, and extracellular L-homocysteine may be a measure of the balance between L-homocysteine formation and utilization.

Topics: Adenine; Adenosine; Adenosine Triphosphate; Adenosylhomocysteinase; Animals; Cell Line; Cell Transformation, Neoplastic; Fibroblasts; Homocysteine; Hydrolases; In Vitro Techniques; Liver; Methionine; Rats; S-Adenosylhomocysteine

The 5.8S rRNA of normal tissues contains a partially 2'-O-methylated uridylic acid residue which is methylated in the cytoplasm and undermethylated in rapidly growing neoplastic tissues (R. N. Nazar, T. O. Sitz, and K. D. Somers, J. Mol. Biol., 142: 117-121, 1980). This difference in methylation was further characterized by examining the effect of cell age or cell culture passage number on the level of methylation of 5.8S RNAs from normal and malignant cell lines and simultaneous changes in intracellular pools of S-adenosylmethionine and S-adenosylhomocysteine. The results indicate that the level of methylation decreases continuously with cell culture passage number as the cells become aneuploid, transformed, or tumorigenic, but there is no direct correlation with the intracellular pools of S-adenosylmethionine or S-adenosylhomocysteine. In contrast, there is a dramatic but inverse increase in the S-adenosylmethionine:S-adenosylhomocysteine ratio which correlates with the decreasing levels of 2'-O-methylation. The significance of these changes in substrate levels to the hypomethylation of 5.8S and other RNAs during oncogenesis is discussed.

Topics: Animals; Cell Cycle; Cell Line; Cell Transformation, Neoplastic; Child; Fibroblasts; HeLa Cells; Humans; Kidney; Kinetics; Liver; Liver Regeneration; Male; Methylation; Osteosarcoma; Rats; Rats, Inbred Strains; RNA, Ribosomal; S-Adenosylhomocysteine; S-Adenosylmethionine

Topics: Animals; Cell Line; Cell Transformation, Neoplastic; Kidney; NAD; Niacinamide; Rats; S-Adenosylhomocysteine; S-Adenosylmethionine; Tubercidin

A correlation was found between inhibition of protein methylase I and inhibition of virus-induced cell transformation by structural analogs of S-adenosylhomocysteine; all good inhibitors of this enzyme are also good inhibitors of Rous sarcoma virus-induced chicke embryo fibroblast transformation. The inhibitory effect of these analogs was similar on enzymes from normal and transformed cells; no significant variation of the inhibition constants was observed after purification of protein methylase I. From the kinetic constants obtained, a structure-activity relationship can be established for protein methylase I.

Topics: Animals; Avian Sarcoma Viruses; Cell Transformation, Neoplastic; Cells, Cultured; Chick Embryo; Fibroblasts; Homocysteine; Protein Methyltransferases; Protein-Arginine N-Methyltransferases; S-Adenosylhomocysteine; Structure-Activity Relationship

A high increase in the amount of methylated tRNA bases was found in vivo in Rous sarcoma virus infected and transformed chick embryo fibroblasts in comparison with normal cells, tRNA methylases extracted from transformed cells showed also higher activity in vitro with a heterologous substrate. 5'-deoxy-5'-S-isobutyl adenosine, (a structural analogue of S-adenosyl-L homocysteine), which inhibits virus-induced cell transformation, inhibits also the increase of incorporation of labelled methyl groups into tRNA in infected and transformed cells. When normal cells are grown in the presence of this inhibitor, undermethylated tRNAs are obtained. The effect of the drug is different in normal, infected and transformed cells. The methylation of the different bases is inhibited in vitro and in vivo to various extent. The effect of this substance on tRNA methylation may be the cause of its inhibitory effect on cell transformation.

Topics: Avian Sarcoma Viruses; Biological Transport; Cell Line; Cell Transformation, Neoplastic; Cell Transformation, Viral; Cell-Free System; Deoxyadenosines; Dose-Response Relationship, Drug; Methionine; RNA, Transfer; S-Adenosylhomocysteine; Thionucleosides; tRNA Methyltransferases

Topics: Adenosine; Adenosine Deaminase; Amino Acid Oxidoreductases; Animals; Avian Sarcoma Viruses; Cell Line; Cell Transformation, Neoplastic; Homocysteine; Hydrolases; Methylation; Neoplasm Proteins; Proteins; S-Adenosylhomocysteine; Tumor Virus Infections