Biomedical subjects

I R Rowland

Publications and source records attributed to I R Rowland.

At least 37 records · Page 2Linked to original sources

PubMed1997

Shift from a dairy product-rich to a dairy product-free diet: influence on cytotoxicity and genotoxicity of fecal water--potential risk factors for colon cancer.

Several epidemiologic studies have suggested that dairy product intake is associated with a decreased incidence of colon cancer. To determine whether the cytotoxicity and genotoxicity of the aqueous portion of human stool (two potential risk markers for the disease) were affected by a change in dairy product intake, 18 healthy male and female volunteers were randomly divided into two groups. In a crossover design, the volunteers shifted from their normal dairy product-rich diet to a dairy product-free diet. Nutritional analysis of the food consumed during the study period showed a significant decrease in energy intake from 9000 to 7866 kJ/d because of a decreased intake of protein and fat. Carbohydrate and fiber intakes remained unchanged during the intervention. Calcium intake decreased significantly from 1488 to 372 mg/d, with similar significant decreases in phosphate and vitamin D intakes. Cytotoxicity of fecal water, analyzed by the HT-29 cytotoxicity assay, indicated a significant decrease in cell survival from 34% to 20% when dairy products were excluded from the participants' diets. Single-cell gel electrophoresis (COMET assay), used to analyze genotoxicity of fecal waters, indicated no differences brought about by the dietary intervention. In conclusion, our findings indicate that a shift from a dairy product-rich to a dairy product-free diet resulted in a significant effect on an accepted risk marker for colon cancer and may suggest that the mechanism by which dairy products are protective is at the level of tumor promotion rather than initiation.

Adult↗

PubMed1997

Genotoxic activity in human faecal water and the role of bile acids: a study using the alkaline comet assay.

Human faecal waters from 35 healthy non-smoking volunteers (23 from England and 12 from Sweden) consuming their habitual diet were screened for genotoxicity by the single-cell gel electrophoresis (comet) assay using a human colon adenocarcinoma cell line (CACO-2) as the target. Hydrogen peroxide induced DNA damage was categorized as low, intermediate or high for tail moments greater than 5, 17 and 32, respectively: 11 samples were highly genotoxic, four were intermediate, one was low and 19 showed no activity. Endonuclease III treatment significantly increased DNA damage for all except the non-genotoxic faecal waters, suggesting that faecal water genotoxicity may be due, at least in part, to oxidative damage. Faecal water cytotoxicity has previously been attributed to the bile and fatty acid content. In the comet assay no DNA damage was induced by deoxycholate or lithocholate at normal physiological concentrations, suggesting that the genotoxicity of faecal water was due to other substances. Both bile acids induced DNA damage above 300 microM, levels often found in patients with colonic polyps and there was a significant increase in genotoxicity after endonuclease III treatment indicative of oxidative DNA damage.

Adult↗

PubMed1997

Influence of diets containing high and low risk factors for colon cancer on early stages of carcinogenesis in human flora-associated (HFA) rats.

Germ-free rats colonised with a human intestinal flora were fed diets containing high risk (HR) or low risk (LR) factors for colorectal cancer, and putative biomarkers were evaluated in the colonic mucosa; (i) proliferation, (ii) 1,2-dimethylhydrazine (DMH)-induced aberrant crypt foci and (iii) DMH-induced DNA damage. The HR diet was high in fat (45% of calories) and low in calcium and fibre, reflecting levels characteristic of typical western diets. The LR diet was low in fat (<5% of calories), and high in calcium and fibre. The nutrient/energy ratio of the two diets were similar. Mucosal crypt cell proliferation, assessed after microdissection, was higher on the LR diet (mean number of mitoses per crypt was 2.65 on the LR diet, and 1.62 on the HR diet; P < 0.05). Aberrant crypt foci (ACF) were assessed in the mucosa 12 weeks after DMH treatment. On the HR diet there were significantly more small ACF with 1 and 2 crypts per focus, but fewer ACF with 3, 5 and 7 or more crypts per focus. There was no significant difference in total ACF or the total number of crypts. The effect of diet on DNA damage in the colon was assessed in vivo by the comet assay. Animals were fed a HR or LR diet for 12 weeks before treatment with DMH or saline. For carcinogen-treated animals, DNA damage was significantly higher in colon cells from animals on the HR diet. On the LR diet both DNA damage and the induction of small ACF were reduced despite an increase in cell proliferation. The increase in large ACF on the LR diet may be attributable to elevated crypt cell proliferation possibly increasing crypt fission rates.

1,2-Dimethylhydrazine↗

PubMed1996

Effect of dietary arginine on urinary nitrate excretion in germ-free rats.

The effect of dietary arginine on the endogenous synthesis of nitrate in germ-free rats was investigated. The animals were fed, for up to 18 days, purified diets containing either casein or lactalbumin (proteins differing in arginine content) with or without additional free arginine. Urine was collected from all animals for 4 days, and the animals were then dosed ip with Escherichia coli lipopolysaccharide (LPS) to stimulate nitrate synthesis and urine collected for a further 4 days. The urine samples were analysed for nitrate by ion chromatography. Although the excretion of nitrate was markedly stimulated by LPS treatment, there were no significant effects of protein source or arginine supplementation of the diet.

Administration, Oral↗

PubMed1996

The effect of lactulose on DNA damage induced by DMH in the colon of human flora-associated rats.

Germ-free rats were fed purified diets containing sucrose (0.3%, wt/wt) as control or the synthetic disaccharide lactulose (0.3% wt/wt) and were then dosed orally with a human fecal suspension. After five weeks on the diets, these "human flora-associated" rats were transferred to diets in which the sucrose or lactulose level was raised to 3% (wt/wt). Four weeks later, the rats from each dietary group were dosed orally with saline (controls) or 1,2-dimethylhydrazine dihydrochloride (DMH, 15 mg/kg). Sixteen hours after the carcinogen dose, the rats were killed, colon cells were isolated, and the degree of DNA damage in the cells was assessed using the single cell microgel electrophoresis (Comet) assay. Samples of cecal contents were also removed from the saline-treated rats to determine changes in composition of the gut microflora. No significant diet-related differences in cecal bacterial numbers were observed, although there was a trend for the numbers of lactobacilli to be higher in the lactulose-fed animals. DNA damage in colon cells was quantified by measurement of the length of the cellular DNA image after electrophoresis, and the evaluated cells were assigned to classes according to the degree of damage: undamaged (< 40 microns), moderately damaged (40-80 microns), or severely damaged (> 80 microns). There were no significant diet-related differences in DNA damage in colon cells from rats treated with saline. Treatment with DMH induced a highly significant increase in DNA damage in sucrose- and lactulose-fed rats. However, in the rats treated with DMH, the degree of DNA damage was significantly (p < 0.05) less in the lactulose-fed animals than in those given the sucrose diet, as evidenced by a decrease in the percentage of cells with severe damage. In the sucrose-fed rats, severely damaged cells accounted for 33% of the total compared with only 12.6% in the lactulose-fed DMH-treated animals. The results of this study, which was performed in human flora-associated rats to provide data of relevance to humans, indicate that lactulose consumption offered a degree of protection from the genotoxic effects in the colonic mucosa of a known colon carcinogen.

1,2-Dimethylhydrazine↗

PubMed1996

An assessment of three selective media for bifidobacteria in faeces.

Three previously described media enumerating Bifidobacterium spp. in faeces were compared with respect to their selectivity and quantitative recovery. The results of this study indicate that of the three media studied, Beerens' agar is the most suitable medium for isolation and enumeration of Bifidobacterium spp. from the gut microflora.

Bifidobacterium↗

PubMed1995

Estimation of the fermentability of dietary fibre in vitro: a European interlaboratory study.

Five European laboratories tested a simple in vitro batch system for dietary fibre fermentation studies. The inoculum was composed of fresh human faeces mixed with a carbonate-phosphate buffer complex supplemented with trace elements and urea. Five dietary fibre sources (cellulose, sugarbeet fibre, soyabean fibre, maize bran and pectin) were used by each laboratory on three occasions to determine pH, residual non-starch polysaccharides (NSP) and short-chain fatty acid production during fermentation. Cellulose and maize bran degradabilities were very low (7.2(SE 10.8) and 6.2 (SE 9.1)% respectively after 24 h), whereas pectin and soyabean fibre were highly degraded (97.4 (SE 4.4) and 91.1 (SE 3.4)% respectively after 24 h). Sugarbeet fibre exhibited an intermediate level of degradability (59.5 (SE 14.9)%). Short-chain fatty acid production was closely related to NSP degradation (r 0.99). Although each variable was ranked similarly by all laboratories, some differences occurred with respect to absolute values. However, the adaptation of donors to the experimental substrates was not an influential factor. Interlaboratory differences could be reduced either by adding less substrate during incubations or using less-diluted inocula. In vitro fermentations with inocula made from human faeces and from rat caecal contents gave similar results. There was a close correspondence between the data obtained in the present experiment and those previously published in in vivo studies in the rat using the same fibres. The in vitro batch system tested during the present study provides a rapid means of obtaining quantitative estimates of the fermentation and the estimation of the energy content of new sources of dietary fibre.

Animals↗

PubMed1995

Dietary caffeine reduces the genotoxicity of MeIQx in the host-mediated assay in mice.

The influence of dietary caffeine on the genotoxicity of the cooked food mutagen 2-amino-3,8-dimethylimidazo[4,5-f]-quinoxaline (MeIQx) was evaluated using the host-mediated assay in mice. For four weeks, BALB/c mice were fed a purified diet with or without caffeine (0.01% wt/wt in the diet). In the host-mediated assay, Salmonella typhimurium TA98 was given intravenously immediately before an oral dose of MeIQx (1.5 mg/kg body wt). After one hour, the mice were killed, the Salmonellae were recovered from the liver, and the number of mutants (his+ revertants) were determined. Consumption of caffeine led to a 47% reduction in the number of mutants induced by MeIQx (p < 0.001). Subsequent in vitro experiments using S. typhimurium TA98 revealed that the capacity of hepatic S-9 fractions from the caffeine-fed mice to covert MeIQx to an active mutagen was reduced by approximately 35%. This effect was not attributable to caffeine in the S-9 preparation. These data suggest that consumption of caffeine modifies MeIQx mutagenicity by altering the spectrum of enzymes involved in its activation.

Animals↗

PubMed1993

Influence of dietary fat on DNA binding by 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in the mouse liver.

Female BALB/c mice were fed either a low (1%)-fat or one of three high-fat diets (containing an additional 25% (w/w) beef fat, hydrogenated vegetable oil or non-hydrogenated vegetable oil) for 4 wk. They were then orally treated with 10 mg 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx)/kg body weight and killed 6 hr later. Consumption of the hydrogenated vegetable oil was accompanied by increased DNA adduct formation in mice. The abilities of hepatic S-9 preparations from mice fed the various diets to convert MeIQx to an active bacterial mutagen was assessed using Salmonella typhimurium TA98. Preparations from mice fed the high-fat diets exhibited significantly greater capacity to activate MeIQx than did those from low-fat-fed mice. The greatest increases were seen with S-9 from animals fed either beef fat or hydrogenated vegetable oil.

Administration, Oral↗

PubMed1993

Conversion of IQ to 7-OHIQ by gut microflora.

The rates of conversion of 2-amino-3-methyl-3H-imidazo[4,5-f]quinoline (IQ) to its reportedly mutagenic 7-keto derivative (7-OHIQ) by intestinal bacteria from humans, mice, and rats were compared. IQ was metabolized faster by cecal contents from rats or mice than by human fecal samples (113 and 87 mumol 7-OHIQ formed/hr/g cecal contents, respectively, vs. 12.3 mumol/hr/g feces). Cecal contents from germ-free rats colonized with human fecal bacteria [human flora-associated (HFA) rats] converted IQ to 7-OHIQ at rates generally lower than contents from rats colonized with their native flora. Diet had a marked effect on IQ metabolism by HFA rat cecal contents. The rate of IQ conversion to 7-OHIQ was increased in rats fed a diet high in beef dripping compared with that in rats fed a low-fat control diet. A diet high in olive oil, however, did not produce an increase in the IQ conversion rate. Addition of fiber to a purified diet increased the rate of IQ metabolism in the following order: sugar beet fiber > wheat bran > oat bran fiber > fiber-free diet. In a further study, HFA rats were fed human diets altered independently in their fat, fiber (wheat bran), and beef contents. The high-fiber diet produced the greatest increase in IQ conversion rate, followed by the high-fat diet. The diet with a high beef content and the control diet (low levels of all 3 macrocomponents) produced similarly low rates of IQ conversion. Material from incubations of IQ with HFA rat cecal contents, assumed to be 7-OHIQ on the basis of chromatographic behavior, was confirmed to be directly mutagenic, producing approximately 800 His+ revertants per microgram with S. typhimurium TA98.

Adult↗

PubMed1993

Effects of risk-associated human dietary macrocomponents on processes related to carcinogenesis in human-flora-associated (HFA) rats.

Dietary fat, protein and fibre have been shown to modulate cancer risk in humans and the present study examined the biological effects in human-flora-associated (HFA) rats of altering intake levels within the normal human range. Two control groups, one HFA and the other germfree (GF), consumed a human diet low in fat, fibre and beef for 4 weeks; three other groups consumed human diets similar except for independent 3-fold increases in fat, beef protein or fibre. After 2 weeks on the diets, magnetically recoverable microcapsules were given orally to the rats and subsequently recovered from the faeces to assess endogenous cross-linking agents. After 4 weeks, measurements were made of gut microfloral enzyme activities, hepatic activation of dietary mutagens and hepatic DNA adducts by 32P-postlabelling. Activation in vitro of the dietary mutagens 2-amino-3-methyl-3H-imidazo[4,5-f]quinoline (IQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) by hepatic S9, formation of endogenous hepatic DNA adducts in vivo and the beta-glucuronidase activity of caecal contents were all increased in the sequence high fat > high fibre > high beef = control. Of the two DNA adducts found in all HFA rats, only one was present in GF controls, indicating that the human gut microflora (subject to human dietary modulation) either releases a DNA-adducting product able to act outside the gastrointestinal tract, or stimulates the generation of such a product by mammalian processes. Caecal nitrate reductase activity was highest in rats fed the high beef diet, whilst entrapment of cross-linking agents was highest in those fed the high fibre diet. These results show that risk-related components of human diets interact with human gut microflora to modulate the production of endogenous DNA-adducting and cross-linking substances.

Animals↗

PubMed1993

The effects of transgalactosylated oligosaccharides on gut flora metabolism in rats associated with a human faecal microflora.

The effect on various caecal bacteria and their metabolic activities of feeding diet containing transgalactosylated oligosaccharides (TOS) with or without Bifidobacterium breve (administered in the drinking water) was investigated in rats colonized with a human faecal microflora. TOS (5% w/w in diet) or TOS plus B. breve, given for 4 weeks, induced increases in caecal concentration of total anaerobic bacteria, lactobacilli and bifidobacteria, and decreases in numbers of enterobacteria. Caecal pH was significantly reduced by feeding TOS, as were the activities of beta-glucuronidase and nitrate reductase. In contrast, beta-glucosidase activity was increased in TOS-fed rats. Dietary TOS was also associated with decreased conversion, by caecal contents, of the dietary carcinogen 2-amino-3-methyl-3H-imidazo[4,5-f] quinoline (IQ) to its genotoxic 7-hydroxy derivative.

Animals↗

PubMed1992

Inhibition of the metabolism of mutagens occurring in food by arachidonic acid.

Hepatic microsomal fractions (microsomes) were prepared from male Sprague-Dawley rats. The effect of arachidonic acid on the conversion of the heterocyclic aromatic amine 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) to its genotoxic metabolites was investigated using a modified bacterial mutation assay (indicator: Salmonella typhimurium TA98). Arachidonic acid inhibited the mutagenicity of IQ without effect on the uptake of the active metabolites and/or on the DNA-repair processes within the bacterial cell. The activation of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and aflatoxin B1 (AFB1) was also inhibited by this polyunsaturated fatty acid.

Aflatoxin B1↗

PubMed1992

Effect of hepatic cytochrome P-450 inducing agents on mutagen activity in the host-mediated assay.

Intraperitoneal treatment of female BALB/c mice with either phenobarbitone or beta-naphthoflavone led to the induction of various hepatic enzymes associated with xenobiotic metabolism and to increased abilities of hepatic S9 fractions to convert the dietary carcinogen 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) to an active bacterial mutagen. In the case of another carcinogen, aflatoxin B1 an increase in in vitro hepatic activation was seen only in mice treated with phenobarbitone. In contrast, pretreatment with either phenobarbitone or beta-naphthoflavone reduced the in vivo activity of both aflatoxin B1 and MeIQx in the host mediated bacterial mutation assay. These data indicate that, for some carcinogens at least, the host-mediated assay may be used to predict the carcinogenic consequences of hepatic enzyme induction.

Aflatoxins↗

PubMed1992

Effect in vitro of arachidonic acid, eicosapentaenoic acid and docosahexaenoic acid on the hepatic activation of dietary genotoxins by rat post-mitochondrial fractions.

The effect of arachidonic acid, eicosapentaenoic acid and docosahexaenoic acid on the conversion of the heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) to its genotoxic metabolites was investigated using a modified bacterial mutation assay. The assay used Salmonella typhimurium TA98 as an indicator of the mutagenicity and hepatic post-mitochondrial fractions (S-9) from male Sprague-Dawley rats as the activating system. All three fatty acids inhibited the mutagenicity of IQ without effect on the uptake of the active metabolites and/or on the DNA repair processes within the bacterial cell. The activation of three other food mutagens, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and aflatoxin B1 (AFB1) was also inhibited by these fatty acids.

Animals↗

PubMed1992

In vivo and in vitro models of the human colonic flora.

The study of colonic flora composition and metabolism presents considerable methodological problems. Attempts to circumvent these problems have led to the development of numerous in vitro and in vivo models to simulate the human colon and its microbial population. In terms of in vivo models, conventional laboratory animals have many limitations. Data of greater relevance to man can be obtained by using germ-free rodents associated with human colonic bacteria. The applications of such animals to studies of toxicity of chemicals and gastrointestinal infections are discussed. The advantages and disadvantages of the various in vitro systems for studying gut microflora and its metabolic activity (from simple static cultures to the more sophisticated continuous and semicontinuous flow models) are reviewed. The apparatus involved is described together with practical information on media, running conditions, and sampling. The bacteriological and metabolic criteria for establishing the similarity of the models to the in situ colonic flora are also discussed. The final sections of the review are devoted to the major applications (current and future) of the models, including fermentation studies on dietary fiber, metabolism of nutrients and foreign compounds (including carcinogens) in food, and the investigation of colonization resistance.

Anaerobiosis↗

PubMed1992

Development changes to gut microflora metabolism in mice.

Developmental changes in the activities of bacterial nitrate reductase, nitroreductase and beta-glucuronidase and their response to fermentable dietary fibre, were investigated in caecal contents from suckling mice (2-week-old) and in mice aged 4-24 weeks fed either a purified fibre-free diet or that diet supplemented with 5% (w/w) pectin. There was no apparent age-related trend common to the three enzymes studied. Nitrate reductase activity in the mice fed the fibre-free diet did not markedly alter with age. Pectin administration, however, was associated with a significant increase in nitrate reductase activity, particularly in 4-week-old mice. Nitroreductase activity exhibited an overall upward trend in mice from 2 to 12 weeks and thereafter decreased. Caecal beta-glucuronidase activity in mice increased sharply between 2 weeks and 4 weeks of age, thereafter not changing significantly until the 24th week. Pectin feeding had no consistent effect on activities either of nitroreductase or beta-glucuronidase. The changes in enzyme activities with age were not related to the concentration of bacteria in the caecum, which was highest in the 2-week-old mice. We conclude that the weaning is a period in which marked changes in caecal bacterial enzyme activities can occur.

Aging↗

PubMed1991

Interaction with microsomal lipid as a major factor responsible for S9-mediated inhibition of 1,8-dinitropyrene mutagenicity.

1,8-Dinitropyrene (1,8-DNP), present in polluted air, is a rodent carcinogen and a potent, direct-acting mutagen in salmonella typhimurium TA98. This mutagenicity is markedly reduced in the presence of mammalian hepatic S9 or microsomes. We demonstrate that at least a substantial part of this effect is attributable to non-enzymatic processes. The microsomal-dependent inhibition was unaffected by omission of an NADPH-generating system or when the cytochrome P-450 inhibitor, SKF-525A, or the cytochrome P-448 inhibitor, ellipticine, was incorporated in the metabolic activation system, suggesting that mixed function oxidases are not involved. Heat inactivation partially decreased the ability of induced S9 to reduce DNP mutagenicity. Substitution of S9 with a similar concentration of bovine serum albumin did not affect DNP activity. Thus non-specific binding to microsomal protein is not involved. However, when lipids, derived from uninduced microsomes, were added to incubations of DNP and S. typhimurium TA98, mutagenicity was decreased. Furthermore, substitution of microsomal lipids with a suspension of phosphatidylcholine (PC), a major lipid constituent of microsomes, affected DNP mutagenicity similarly. An increase in PC concentration resulted in a greater inhibitory effect. The reduction in DNP mutagenicity observed with microsomal lipids or with PC was less than that detected with uninduced S9, whilst the mutagenicity of 2-nitrofluorene was reduced to an approximately equal extent by lipids and S9. This phenomenon may be responsible for the S9-mediated detoxification of other mutagenic nitroaromatic compounds and may have important implications for mutagenicity testing.

Air Pollutants↗