Effect of administration of the carcinogen dimethylnitrosamine on urinary 7-methylguanine.
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ABSTRACT:
SUBMITTER: Craddock VM
PROVIDER: S-EPMC1270604 | biostudies-other | 1967 Aug
REPOSITORIES: biostudies-other
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Project description:Dimethylnitrosamine is metabolized to form an alkylating intermediate, which may have significance for its carcinogenic action. However, certain other compounds that are known to be highly mutagenic, including nitrous acid and hydroxylamine, might also be formed. Owing to the general reactivity of these compounds, it would be difficult to detect their formation in the intact animal. Instead, the nucleic acids of carcinogen-treated animals were examined for products of reaction with nitrous acid and hydroxylamine, i.e. for deamination of adenine and guanine, and formation of N(6)-hydroxycytosine, respectively. A double-labelling technique was used to detect very small amounts of the abnormal bases. The purine moieties of DNA in adult rat liver were labelled either with (14)C or with (3)H, by treating the neonatal animals with [(14)C]formate or with [(3)H]formate, and then allowing a period for normal growth. During this time, in liver, the labels were largely lost by metabolic turnover from cell components other than DNA. The pyrimidine moieties in DNA were labelled by treating the neonatal animals with [(14)C]orotate. The purine constituents of RNA of adult rat liver were labelled by repeated administration of [(14)C]- or [(3)H]-formate to the adult rats. The [(14)C]nucleic acid-labelled rat could then be treated with the carcinogen, and the [(3)H]nucleic acid-labelled animal could be used as a control. By this means the experimental and control tissues could be homogenized together in a single preparation, and the nucleic acids from the two tissues could be isolated, hydrolysed and analysed in a single sample. It was therefore possible to have an internal control for artifacts due to changes taking place in the nucleic acid bases during the experimental procedures. With this technique, the formation in vivo of 7-methylguanine in rat-liver DNA and RNA after administration of dimethylnitrosamine was confirmed, and no evidence was found for the formation of xanthine, hypoxanthine, N(6)-hydroxycytosine, or any other abnormal base.
| S-EPMC1265207 | biostudies-other
Project description:1. BD-IV rats were given labelled dimethylnitrosamine (2 mg/kg) by stomach tube on weekdays (Monday to Friday) for up to 24 weeks. The rats killed after 2, 4, 8, 16 and 24 weeks of treatment (72 h after the final dimethylnitrosamine gavage) and DNA was isolated from the pooled livers, kidneys and lungs. Purine bases were released from the DNA by mild acid hydrolysis and separated by Sephadex G-10 chromatography. 2. Throughout the experiment, the content of 7-methylguanine in liver DNA was approx. 16 times that in kidney and lung. The amount of this product increased in the DNA of all three tissues up to 16 weeks, but by 24 weeks had decreased by 20% in the liver and 46% in the other tissues. 3. O6-Methylguanine was not detected in liver DNA, but was easily measured in kidney and lung DNA after 4 weeks of dimethylnitrosamine administration. The amount of O6-methylguanine in kidney and lung DNA increased relative to that of 7-methylguanine, and by 24 weeks was 60% of the 7-methylguanine content in both tissues. 4. Incorporation of radioactive C1 breakdown products of dimethylnitrosamine into normal purines in DNA increased continuously in all three tissues. 5. The results are discussed with respect to the specific hepatocarcinogenic effect of chronic administration of dimethylnitrosamine and the possible contribution of increased DNA repair and DNA synthesis.
| S-EPMC1164928 | biostudies-other
Project description:After treatment with NN-di[14C]methylnitrosamine, samples of DNA were isolated from rat livers by a conventional phenol procedure and examined for the presence of phosphotriesters. A method of capable of detecting relatively small amounts of 14C-labelled phosphotriesters was developed and used to establish that these products account for 10-12% of the total methylation pattern found after treatment with this agent in vitro. The significance of the presence of phosphotriesters in DNA is discussed.
| S-EPMC1165247 | biostudies-other
Project description:1. [(14)C]Dimethylnitrosamine was administered to rats, and the formation of free and protein-bound S-methylcysteine was investigated. 2. Very little S-methylcysteine was found in liver acid-soluble fraction, in liver protein or in urine. 3. The methylhistidines formed in liver protein were found to have largely disappeared by 6 days after injection of dimethylnitrosamine. 4. The significance of these results in the carcinogenic action of dimethylnitrosamine is discussed.
| S-EPMC1206513 | biostudies-other
Project description:1. To assess the significance of the methylation of nucleic acids known to be caused by certain carcinogens, the metabolic fate of 7[(14)C]-methylguanine was studied, with special reference to its possible incorporation into RNA and DNA. 2. The major part (approx. 95%) of the dose was excreted unchanged in the urine. A small amount of N-demethylation took place, as evidenced by the formation of radioactive adenine and guanine, and expiration of (14)C-labelled carbon dioxide. No evidence was obtained for the direct incorporation of 7-methylguanine into systems synthesizing nucleic acids, i.e. RNA in liver, DNA in intestine or in the foetus.
| S-EPMC1186754 | biostudies-other
Project description:1. The amounts of 7-methylguanine and O(6)-methylguanine present in the DNA of liver and kidney of rats 4h and 24h after administration of low doses of dimethylnitrosamine were measured. 2. O(6)-Methylguanine was rapidly removed from liver DNA so that less than 15% of the expected amount (on the basis of 7-methylguanine found) was present within 4h after doses of 0.25mg/kg body wt. or less. Within 24h of administration of dimethylnitrosamine at doses of 1mg/kg or below, more than 85% of the expected amount of O(6)-methylguanine was removed. Removal was most efficient (defined in terms of the percentage of the O(6)-methylguanine formed that was subsequently lost within 24h) after doses of 0.25-0.5mg/kg body wt. At doses greater or less than this the removal was less efficient, even though the absolute amount of O(6)-methylguanine lost during 24h increased with the dose of dimethylnitrosamine over the entire range of doses from 0.001 to 20mg/kg body wt. 3. Alkylation of kidney DNA after intraperitoneal injections of 1-50mug of dimethylnitrosamine/kg body wt. occurred at about one-tenth the extent of alkylation of liver DNA. Removal of O(6)-methylguanine from the DNA also took place in the kidney, but was slower than in the liver. 4. After oral administration of these doses of dimethylnitrosamine, the alkylation of kidney DNA was much less than after intraperitoneal administration and represented only 1-2% of that found in the liver. 5. Alkylation of liver and kidney DNA was readily detectable when measured 24h after the final injection in rats that received daily injections of 1mug of [(3)H]dimethylnitrosamine/kg for 2 or 3 weeks. After 3 weeks, O(6)-methylguanine contents in the liver DNA were about 1% of the 7-methylguanine contents. The amount of 7-methylguanine in the liver DNA was 10 times that in the kidney DNA, but liver O(6)-methylguanine contents were only twice those in the kidney. 6. Extracts able to catalyse the removal of O(6)-methylguanine from alkylated DNA in vitro were isolated from liver and kidney. These extracts did not lead to the loss of 7-methylguanine from DNA. 7. The possible relevance of the formation and removal of O(6)-methylguanine in DNA to the risk of tumour induction by exposure to low concentrations of dimethylnitrosamine is discussed.
| S-EPMC1185839 | biostudies-other
Project description:1. The effects of various doses of dimethylnitrosamine on the hydrocortisone induction of tryptophan pyrrolase were studied. A single LD(50) dose of dimethylnitrosamine inhibits the synthesis of the enzyme if given within the first 5hr. of the induction. A quarter of this dose also inhibits the synthesis of the enzyme, but, in addition, retards the rate of decay of the enzyme. 2. A single small dose of dimethylnitrosamine significantly inhibits the hydrocortisone induction of tryptophan pyrrolase for at least 14 days without causing widespread damage to liver substructure. 3. The inhibition by dimethylnitrosamine of induced tryptophan pyrrolase synthesis is probably independent of any action of the toxin on the synthesis of cofactors necessary for full expression of the enzyme's activity. Dimethylnitrosamine appears to act directly on the synthesis of the enzyme protein. 4. The synthesis of that form of the enzyme which is most sensitive to hydrocortisone is apparently also most susceptible to the action of dimethylnitrosamine. 5. It is suggested that the inhibition of protein synthesis by dimethylnitrosamine may be a result of methylation of messenger RNA, which then is unable to code effectively for amino acid polymerization.
| S-EPMC1198860 | biostudies-other
Project description:Carcinogenic tobacco-specific nitrosamines (TSNAs) such as 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) are found only in tobacco and derived products. Food and Drug Administration of the United States (US FDA) lists NNK as one of the 93 harmful and potentially harmful constituents (HPHCs) found in tobacco products and tobacco smoke. The aim of this study was to use the urinary concentration of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), a major metabolite of NNK, to quantitatively estimate exposure to NNK in the US general population. In 2011-2012, the Centers for Disease Control and Prevention's National Health and Nutrition Examination Survey (NHANES) collected urine and serum samples from a representative sample of US residents. We used a serum cotinine cutoff of 10 ng/ml with combination of questionnaire data to select non-users from cigarette users and used self-reported data to determine different tobacco product user groups. We estimated the absorbed total daily dose of NNK using a probabilistic method based on a two-compartment model. The geometric mean (GM) for the daily dose of NNK among smokers aged 12-16 years was significantly higher than that for non-users at the same age stage exposed to second-hand smoke (SHS) (P<0.001). Among those exposed to SHS, the GM for daily dose of NNK in young children (6-11 years) was nearly three times of those for adults in the age range 21-59 years. Among cigarette users, non-Hispanic Whites had the highest NNK daily dose and Mexican Americans had the lowest levels. Exclusive snuff or chewing product users had significantly higher daily dose of NNK than did cigarette smokers. Our study found that the maximum daily dose of NNK for children aged from 6 to 11 years and that for a significant percentage of cigarette users, chewing product and snuff users were higher than an estimated provisional "reference" risk level.
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Project description:1. Syrian golden hamster liver ribosomal RNA was isolated up to 96 h after administration of [14C]dimethylnitrosamine at 25 mg/kg or 2.5 mg/kg body weight. 2. The chemical alkyation products, 7-methylguanine, 3-methylcytosine, O6-methylguanosine and 1-methyladenosine, were measured after acidic or enzymic hydrolysis of the RNA to bases or mononucleosides followed by ion-exchange chromatography. 3. Between 7 and 96 h, the relative amounts of alkylation products did not change with time even though the absolute amounts fell by approx. 80% and 51% after the high and low doses respectively. 4. The results suggest that base specific excision repair does not exist for RNA alkylation products in this experimental system.
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Project description:Loss of retinoid-containing lipid droplets upon hepatic stellate cell (HSC) activation is one of the first events in the development of liver disease leading to hepatocellular carcinoma. Although retinoid stores are progressively lost from HSCs during the development of hepatic disease, how this affects hepatocarcinogenesis is unclear. To investigate this, we used diethylnitrosamine (DEN) to induce hepatic tumorigenesis in matched wild-type (WT) and lecithin:retinol acyltransferase (LRAT) knockout (KO) mice, which lack stored retinoid and HSC lipid droplets. Male 15-day-old WT or Lrat KO mice were given intraperitoneal injections of DEN (25 mg/kg body wt). Eight months later, Lrat KO mice showed significantly less liver tumor development compared with WT mice, characterized by less liver tumor incidence and smaller tumor size. Two days after DEN injection, lower serum levels of alanine aminotransferase and decreased hepatic levels of cyclin D1 were observed in Lrat KO mice. Lrat KO mice also exhibited increased levels of retinoic acid-responsive genes, including p21, lower levels of cytochrome P450 enzymes required for DEN bioactivation and higher levels of the DNA repair enzyme O(6)-methylguanine-DNA methyltransferase (MGMT), both before and after DEN treatment. Our results indicate that Lrat KO mice are less susceptible to DEN-induced hepatocarcinogenesis due to increased retinoid signaling and higher expression of p21, which is accompanied by altered hepatic levels of DEN-activating enzymes and MGMT in Lrat KO mice also contribute to decreased cancer initiation and suppressed liver tumor development.
| S-EPMC3271263 | biostudies-literature