Biomedical subjects
H Bartsch
Publications and source records attributed to H Bartsch.
Opisthorchis viverrini infestation and endogenous nitrosamines as risk factors for cholangiocarcinoma in Thailand.
Cholangiocarcinoma (CCA) is one of the most common cancers in north-east Thailand and has been associated with infestation by the liver fluke Opisthorchis viverrini (OV). Two samples of 12-hr overnight urine (after dosing with proline and ascorbic acid or proline alone) were collected from 20 inhabitants from each of 5 contrasting incidence areas for CCA. The incidence of CCA was not correlated with either the amount of NPRO or other nitrosamino acids, endogenous nitrosation potential (difference in NPRO levels between proline dose and proline and ascorbic dose), or nitrate level. However, when urinary levels of nitrosamino acids were compared in subjects living in high-risk areas, subjects who were positive for OV antibody excreted significantly more (p less than 0.01) NPRO (12.3 +/- 18.7 micrograms/12 hr) after proline ingestion than those who were negative (3.5 +/- 3.2 micrograms/12 hr). After ingestion of ascorbic acid, the NPRO levels in the positive subjects were significantly reduced (p less than 0.01) to 2.4 +/- 2.0 micrograms/12 hr, suggesting that endogenous nitrosation of proline was inhibited. Thus, endogenous nitrosation potential estimated from the difference between NPRO and the sum of nitrosamino acids excreted in the 2 urine samples was significantly higher in subjects positive for the OV antibody. Small amounts of pre-formed nitrosamines were found in fermented fish and pork food items, which are consumed frequently in the high-risk area for CCA. These results suggest that the interaction between chemical carcinogens, especially nitrosamines, and OV infestation may play a role in the development of cholangiocarcinoma in Thailand.
Urinary excretion of S-benzylmercapturic acid as an indicator of N-nitroso-N-methylbenzylamine exposure.
The excretion of S-benzylmercapturic acid (SBzMA) in the urine of rats treated with N-nitroso-N-methylbenzylamine (NMBzA) was determined by gas chromatography-mass spectrometry (GC-MS). The identity of SBzMA in the urine was confirmed by full scan GC-MS. The amount of urinary SBzMA varied with the dose of NMBzA (up to 5 mg/kg) and with rat strain. For the three strains investigated, most of a 2.5 mg/kg dose of SBzMA was excreted within 24 h. Comparison of the levels of this SBzMA excreted by rats treated with equivalent doses of either NMBzA or benzaldehyde indicates that urinary SBzMA is derived mainly from benzylating species resulting from the hydroxylation of the methyl group of NMBzA.
Production of polyclonal and monoclonal antibodies for specific detection of nitrosation-proficient denitrifying bacteria in biological fluids.
Polyclonal and monoclonal antibodies were raised against the nitrosating enzyme previously isolated and purified from a denitrifying bacteria Pseudomonas aeruginosa. Using the polyclonal antibodies, a preliminary ELISA test was set up which allowed the detection of the nitrosating enzyme in 2 ml urine samples with a minimal total bacteria count of greater than or equal to 10(5) cells/ml. The use of such a rapid immunological screening test in clinical settings should ascertain whether individual subjects at higher risk for cancer of the stomach or bladder harbour more nitrosation-proficient microorganisms in their microflora. The availability of monoclonal antibodies provides a tool for studying the mechanisms of bacteria-mediated nitrosamine formation from precursor amines.
Depression of serum melatonin in patients with primary breast cancer is not due to an increased peripheral metabolism.
Serum melatonin and its main metabolic product 6-sulfatoxymelatonin were determined in 17 patients with breast cancer (BC) with either a fresh primary tumor (nine) or a secondary tumor (eight) as well as in four patients with untreated benign breast disease (controls). Circadian rhythms were detected in all groups with acrophases around 2 AM for melatonin and around 3 AM for 6-sulfatoxymelatonin. The nocturnal melatonin and 6-sulfatoxymelatonin concentrations were significantly depressed in the group of patients with primary breast cancer compared with controls (P less than 0.01, P less than 0.025). The circadian amplitudes of melatonin and 6-sulfatoxymelatonin were also depressed by 81% (P less than 0.01) and 63% (P less than 0.01). In contrast, patients with secondary BC had nocturnal melatonin and 6-sulfatoxymelatonin concentrations and amplitudes similar to controls. These results demonstrate that the depression of circulating melatonin in patients with primary BC is not due to an enhanced degradation to 6-sulfatoxymelatonin in the liver but must be due to a reduced activity of the pineal gland.
Smoking and peripheral type of cancer are related to high levels of pulmonary cytochrome P450IA in lung cancer patients.
A specific member of the cytochrome P450 superfamily of enzymes, designated P450IA (including 2 isozymes, P450IA1 and P450IA2), which is involved in the metabolic activation of polycyclic aromatic hydrocarbons and aromatic amines, was studied in lung tissue from 25 lung cancer patients by immunohistochemistry. The pulmonary activity of a P450IA1-dependent enzyme, aryl hydrocarbon hydroxylase (AHH), from the same patients was also measured. Cytochrome P450IA was localized principally in the peripheral airways in alveolar epithelium of types I and II and in ciliated columnar and cuboidal bronchiolar epithelium. The amount of P450IA in the bronchial wall was minimal and was localized mainly in the capillary endothelium and the epithelium of the bronchial glands. Smoking was the most important factor related to the presence of P450IA and the AHH activity in lung tissue. None of the 10 ex-smokers, but all except I of the current smokers had detectable level of P450IA. The localization of the cancer was also correlated with the presence of cytochrome P450IA. Peripheral lung tissue stained positively in all patients with a peripheral adenocarcinoma who currently smoked (8/8) but in less than half of those with a bronchial cancer who were smokers (3/7). Our data suggest that the smokers who have an inducible cytochrome P450IA are especially at increased risk of developing lung cancer of the peripheral adenocarcinomatous type.
[Studies on the chemistry of O,N- and S,N-containing heterocylics. 10. Synthesis and biologic activity of 2-substituted 2-ethylbenzoxazoles].
2-Vinylbenzoxazole (3) is obtained by condensation of acrylic acid with 2-aminophenol in a mixture of phosphorus pentoxide/methansulfonic acid. Interaction of 3 with N-, O-, and C-nucleophiles yields 2-substituted 2-ethyl-benzoxazoles. Compounds 8a, 8b and 8e show analgetic activity in mice. Moreover, fungicide, insecticide and nematocide activity of some compounds is found.
Role of cytochrome P-450 in ochratoxin A-stimulated lipid peroxidation.
The role of cytochrome P-450 in the stimulation of lipid peroxidation by the nephrotoxic mycotoxin ochratoxin A has been investigated. Ochratoxin A was previously shown to markedly stimulate lipid peroxidation in a reconstituted system consisting of phospholipid vesicles, NADPH-cytochrome P-450 reductase, Fe3+, ethylenediaminetetraacetic acid (EDTA), and reduced nicotinamide adenine dinucleotide phosphate (NADPH). We now show that purified cytochrome P-450IIB1 could effectively replace EDTA in stimulating lipid peroxidation suggesting that it could mediate the transfer of electrons from NADPH to Fe3+. Cobalt protoporphyrin is known to cause an extensive and long-lasting depletion of hepatic cytochrome P-450 in rats, and it has been used to evaluate the role of hepatic cytochrome P-450 in xenobiotic metabolism and toxicity. We have observed that microsomes isolated from livers of cobalt protoporphyrin-pretreated rats underwent ochratoxin A-dependent lipid peroxidation much more slowly than control microsomes. Also, the level of ethane exhaled (an index of in vivo lipid peroxidation) on ochratoxin A administration was much lower in cobalt protoporphyrin-pretreated rats than in control rats. Taken together, these results provide evidence for the stimulatory role of cytochrome P-450 in ochratoxin A-induced lipid peroxidation in a reconstituted system and strongly implicate its role in microsomal and in vivo ochratoxin A-induced lipid peroxidation.
Evaluation of frequency of micronucleated oral mucosa cells as a marker for genotoxic damage in chewers of betel quid with or without tobacco.
The frequency of micronucleated cells (MNC) derived from exfoliated human oral mucosal cells has been measured to assess genotoxic damage in chewers of betel quid with tobacco (BQT) and tobacco with lime (T). Significantly elevated frequencies of MNC were observed in the exposed groups (BQT = 4.83 +/- 0.70; T = 5.20 +/- 0.66 per 1000 cells) compared to the control group (C = 2.59 +/- 0.37) although the levels observed were lower than those reported in the literature. No correlation was seen between age, duration and frequency of habits and the frequency of MNC in the 2 habit groups. Clastogenic agents in betel quid possibly involved in micronucleus formation are discussed.
[Role of the pineal body in reproduction and in gynecologic tumors].
According to the current view, the main function of the pineal gland and its hormone melatonin is to convey information on environmental photoperiods to the endocrine, central nervous and immune systems, allowing their optimal adjustment to the time of the day and season. Within the endocrine system, the gonads are a major target; maturation of the female reproductive organs during pubertal development is inhibited by the pineal hormone. During the reproductive phase, melatonin seems to act as a coordinating signal for the menstrual cycle by timing the LH-peak and supporting progesterone production. A gradual decline of circulating melatonin with age suggests a role in age-associated endocrine changes, e.g. during menopause. Due to its important role in the female reproductive system, the pineal gland has been implicated in the aetiology of human breast cancer. Serum melatonin is depressed in patients with primary mammary cancer and experimental tumours are inhibited, a finding suggesting a therapeutic potential of melatonin. In addition, other potent antineoplastic substances exist in the pineal gland, which may become useful tools for future anti-cancer therapies.
Endogenous nitrosation of L-proline by dietary-derived nitrate.
Two studies were conducted to assess the availability of normal dietary sources of nitrate for endogenous nitrosation of an amino acid substrate, L-proline, and to investigate the potential for dietary ascorbic acid to inhibit such nitrosation. In the first study, 16 subjects consumed a salad meal (containing about 172 mg nitrate) with and without a loading dose of proline. A significant increase (10.8 compared with 2.7 micrograms/24 hrs, p less than 0.0001) in mean urinary N-nitrosoproline (NPRO) excretion following meal plus substrate ingestion was indicative of intragastric proline nitrosation by meal-derived nitrate. In the second study (19 different subjects), the mean urinary NPRO level was significantly decreased (15.8 micrograms compared with 28.4, p = 0.022) by inclusion of ascorbic acid sources in the meal. This demonstrated inhibition of proline nitrosation by dietary sources of the vitamin. Large interindividual variation in nitrosating ability was apparent that was associated with variation in meal nitrate to salivary nitrite conversion. Although meals containing fresh vegetables, as tested here, could provide sufficient nitrate to result in endogenous formation of N-nitroso compounds, there will be considerable inter- and intraindividual variation in the extent of this process.
Levels of nitrite, nitrate, N-nitroso compounds, ascorbic acid and total bile acids in gastric juice of patients with and without precancerous conditions of the stomach.
To determine the relevance of gastric juice factors to gastric carcinogenesis, 56 patients with unoperated stomachs undergoing endoscopy for dyspepsia had gastric juice aspirated and analysed for pH, ascorbic acid, total bile acids, nitrite, nitrate and total nitroso compounds (NOCs). Plasma was obtained for vitamin C estimation. Antral and body biopsies were assessed for gastritis, Helicobacter pylori, atrophy and intestinal metaplasia (IM). Patients with chronic atrophic gastritis (n = 17) had lower gastric juice ascorbic acid concentrations (P less than 0.001), higher pH (P less than 0.05) and higher incidence of H. pylori infection (P less than 0.001) than normal subjects (n = 12). Patients with reflux gastritis (n = 9) had higher total bile acids (P less than 0.01). Patients with chronic gastritis and IM (n = 11) had higher gastric juice pH (P less than 0.01) and total bile acid concentrations (P less than 0.05), and lower gastric ascorbic acid concentrations (P less than 0.01) than those with chronic gastritis and no IM (n = 24). In chronic gastritis, high nitrite concentrations were associated with high pH (P less than 0.01). However, there were no significant differences in plasma vitamin C or gastric nitrite, nitrate or total NOC concentrations in relation to gastric histology. We conclude that the premalignant condition IM is associated with H. pylori infection, low gastric ascorbic acid levels and elevated total bile acids, but not to elevation in nitrite or total NOCs in fasting gastric juice.
Bacterial formation of N-nitroso compounds from administered precursors in the rat stomach after omeprazole-induced achlorhydria.
The role of bacteria in catalysing intragastric formation of N-nitrosothiazolidine-4-carboxylic acid and N-nitrosomorpholine was investigated in a rat model of omeprazole-induced achlorhydria. Omeprazole-treated rats gavaged with nitrosation-proficient bacteria were treated with nitrosamines and/or precursors and compared to control animals that received no omeprazole treatment/no bacteria. Rats given thiazolidine-4-carboxylic acid, nitrate and 10(11) cells of Escherichia coli, had a five times higher endogenous formation of N-nitrosothiazolidine-4-carboxylic acid as compared to controls. Endogenous formation of N-nitrosomorpholine was quantified by measuring its urinary metabolite N-nitroso-(2-hydroxyethyl)glycine; when rats were given morpholine and nitrite together with E. coli or Pseudomonas aeruginosa endogenous N-nitrosomorpholine formation was increased approximately 2.5-fold as compared to controls. In the same experiment, a higher excretion of unchanged N-nitrosomorpholine was also observed in omeprazole-treated rats receiving bacteria as compared to controls. Rats given morpholine, nitrate and E. coli or P. aeruginosa, excreted three times higher levels of N-nitrosomorpholine as compared to controls. These results conclusively demonstrate that nitrosation-proficient bacteria are capable of increasing intragastric formation of N-nitrosothiazolidine-4-carboxylic acid and N-nitrosomorpholine. These N-nitrosamines are formed from nitrate (or nitrite) and the respective amino precursor via reduction of nitrate into nitrite and bacterial nitrosation catalysis.
Mechanisms of fat-related modulation of N-nitrosodiethylamine-induced tumors in rats: organ distribution, blood lipids, enzymes and pro-oxidant state.
Groups of rats, either dosed with N-nitrosodiethylamine (NDEA) for 10 weeks (from the age of 7 to 17 weeks) or untreated, were fed diets containing either 2% (low fat, LF) or 30% polyunsaturated fat (high fat, HF) on an equicaloric basis from 5 weeks until rats were 43 weeks old. Biochemical parameters were measured during and at the end of the experiment in various organs, blood, urine and exhaled air, for correlation with the presence or absence of tumors. The HF diet tended to increase the number of hepatic tumors induced by NDEA, while the number of extrahepatic tumors was higher in rats fed on the LF diet; also the overall tumor incidence was higher in the LF group. In the HF/NDEA group, only two benign extrahepatic tumors were found. Plasma total and free cholesterol and triglyceride concentrations were lower in the HF than the LF group without NDEA treatment. In animals bearing liver and/or extrahepatic tumors all plasma lipid concentrations were lower than in tumor-free animals. Only minor or no changes were detected in blood catalase activity, malondialdehyde level, reduced glutathione (GSH) level or GSH-related enzymes and excretion of thioethers in the urine due to dietary modulation or NDEA. Changes in the liver that were associated with the HF diet were: (i) increased amounts of some polyunsaturated fatty acids and of total phospholipids in liver microsomes; (ii) an enhanced level of lipid peroxidation in liver; (iii) a decrease in liver glutathione levels during NDEA treatment, with a simultaneous adaptive increase in superoxide dismutase levels, and a decrease in renal glutathione levels in both treated and untreated groups; (iv) enhanced microsomal induction of aminopyrine N-demethylase and epoxide hydrolase activities by NDEA, and (v) decreased hexose monophosphate shunt (HMS) activity. All mono-oxygenase activities were lower, and the activities of epoxide hydrolase, UDP-glucuronosyltransferase and HMS were higher, in liver tumors than in non-tumorous liver of similarly-treated rats. Neither diet nor NDEA had a major effect on drug-metabolizing enzyme activities in lung and kidney. HF diet significantly increased ethane exhalation (an indicator of the whole-body pro-oxidant state) over those on the LF diet: in rats on either diet, it was further increased when NDEA was given. Ethane exhalation was still elevated 30 weeks after the cessation of NDEA treatment. Our results suggest an association between the observed changes in biochemical parameters, notably oxidative stress, due to dietary modulation and the altered tumor incidence and organ distribution of tumors induced by NDEA.
32P Postlabelling analysis of urinary mutagens from smokers of black tobacco implicates 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) as a major DNA-damaging agent.
When mutagens extracted from the urine of two smokers of black tobacco were reacted with DNA in vitro in the presence of a metabolic activation system, several DNA adducts were detected by 32P-postlabelling analysis. Some of these adducts were also visible, but only faintly, on the autoradiogram for a non-smoker's urine. DNA adducts produced in vitro by 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline or 2-amino-1-methyl-6-phenylimidazo[3,5-b]pyridine could not account for the adduct pattern produced by the urinary mutagens. However, three or four 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-related DNA adducts were present among the five or six adducts observed for smokers in the autoradiograms of urinary mutagen-adducted nucleotides. Mutagenicity testing combined with HPLC fractionation of urinary extracts also supported the postlabelling data which implicates PhIP as a mutagen in the urine of smokers of black tobacco.
Unusually high levels of carcinogenic tobacco-specific nitrosamines in Sudan snuff (toombak).
The aim of these studies was to determine the levels of carcinogenic tobacco specific nitrosamines (TSNA) in Sudanese oral snuff (toombak) as recent retrospective epidemiological studies suggested an association between the use of toombak and subsequent development of oral cancer. We have analyzed the TSNA levels in 20 samples of Sudanese toombak, of four different quality levels, collected from five different vendors. Using GC coupled with thermal energy analysis, four TSNA were quantified in snuff extracts: N'-nitrosonornicotine (NNN), N'-nitrosoanatabine (NAT), N'-nitrosoanabasine (NAB) and 4-(methyl-nitrosamino)-1-(3-pyridyl)-1-butanone (NNK). Unusually high levels of these TSNA (mean; range, mg/g snuff, dry wt) were detected; NNN (1.13; 0.50-3.08); NAT (0.08; 0.02-0.29); NAB (0.22; 0.02-2.37); and NNK (2.31; 0.62-7.87). Previously, the highest levels of NNN and NNK reported in any snuff were 0.154 and 0.014 mg/g dry wt respectively. In comparison, the levels in Sudanese toombak were up to 20 and 560 times higher respectively. As the public health implications of these findings are significant, attempts should be made to reduce exposure to TSNA in oral snuff users in Sudan.
Comparison of pulmonary DNA adduct levels, measured by 32P-postlabelling and aryl hydrocarbon hydroxylase activity in lung parenchyma of smokers and ex-smokers.
In order to compare pulmonary DNA adducts and aryl hydrocarbon hydroxylase (AHH) activity, we have measured these two parameters in non-neoplastic surgical lung parenchymal samples from four ex-smokers and 19 smokers, out of 20 patients operated for lung cancer, and three for nonmalignant lung diseases. DNA adducts were determined by scintillation counting after 32P-postlabelling analysis. The microsomal fractions of the same lung specimen were assayed for AHH activity by a fluorometric method. Autoradiograms of DNA adducts found in lungs of smokers revealed two distinct diagonal radioactive zones that were absent in ex-smokers. The smokers had significantly higher levels (1.68-13.4 DNA adducts/10(8) nucleotides; mean +/- SD 5.38 +/- 3.19) than ex-smokers (0.23-2.21; 1.09 +/- 0.84). AHH activity in smokers ranged from 0.01 to 0.69 pmol/min/mg. This activity was significantly (P less than 0.05) higher in smokers (0.26 +/- 0.26) who had smoked until 1 week before surgery than in those who had stopped smoking for greater than 7 days (0.11 +/- 0.11). A positive linear correlation between DNA adduct levels and AHH activity (r = 0.69; P less than 0.001; n = 19) was found in smokers. This relationship could explain why AHH inducibility appears to be a crude marker for lung cancer risk in smokers.
Cytochrome P-450 isozyme pattern is related to individual susceptibility to diethylnitrosamine-induced liver cancer in rats.
Differences in susceptibility to chemical carcinogenesis between rodent strains and species have been linked to variations in genetically-determined mixed function oxidase activities. In order to verify whether such variations also determine the susceptibility of individual animals of the same strain to a chemical carcinogen, outbred male Wistar rats were administered diethylnitrosamine (DEN) (1, 2, or 3 mg/kg) five times a week for 20 weeks. The relationship was examined between the outcome (i.e., presence or absence of liver tumors, and latency period) and the hepatic activities of mixed function oxidases and conjugating enzymes, as well as of O6-methylguanine-DNA-methyltransferase, measured before the carcinogen treatment. In addition, the metabolic profiles of two model drugs, antipyrine and disopyramide, in the urine were analyzed and correlated with the carcinogen susceptibility. The length of the latency period of hepatocellular tumors in individual rats was negatively related to the activities of hepatic dimethylnitrosamine N-demethylase, aryl hydrocarbon hydroxylase and epoxide hydrolase and positively related to the amount of microsomal protein. Consistent relationships between the other 10 measured parameters and the susceptibility to DEN-induced carcinogenesis were not detected. Long-term treatment with DEN slightly decreased the proportion of metabolism of antipyrine into norantipyrine, and increased the share of 4-hydroxyantipyrine; a decrease in the metabolism of disopyramide to N-deisopropyldisopyramide was also detected. It is concluded that the pattern of cytochrome P-450 isoenzymes is related to differences in individual susceptibility to nitrosamine-induced carcinogenesis. The relationship was most marked at low dose levels, which are the levels at which nitrosamine exposures of humans are known to occur.