Effect of carcinogen dose fractionation, diet and source of F344 rat on the induction of colonic aberrant crypts by 2-amino-3-methylimidazo[4,5-f]quinoline

Meirong Xu1, Rongliang Chen1 and Roderick H. Dashwood1,2,3

1 Linus Pauling Institute and
2 Department of Environmental and Molecular Toxicology, Oregon State University, Corvallis, OR 97331-6512, USA


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Carcinogen dose fractionation, diet and source of laboratory animal were examined as variables in the induction of colonic aberrant crypt foci (ACF) by the heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline (IQ). In the first experiment, male F344 rats from the National Cancer Institute (NCI rats) were fed AIN-93G diet and, starting in the third week, IQ was given by gavage on alternating days, the total carcinogen dose of 105 mg being fractionated proportionally over 2, 4, 8 or 14 weeks. Only the high dose (2 week) treatment with IQ was effective for the induction of ACF at 16 weeks, producing on average 3.8 ACF/colon versus 0.5 ACF/colon in all other groups (P < 0.05). The 2 week IQ dosing protocol was used in a second experiment in which male F344 rats from Simonsen Laboratories (SN) or NCI were fed AIN-93G, AIN-76A or chow diet. On average, SN rats on chow diet had twice the number of aberrant crypts compared with NCI rats given the same diet and three to four times as many aberrant crypts as NCI rats fed AIN diets. Hepatic cytochrome P4501A1 (CYP1A1) levels were essentially unaffected by diet, but methoxyresorufin O-demethylase activities and CYP1A2 protein levels were increased 2- to 3-fold in animals fed chow versus AIN diets. During the 2 week period of carcinogen administration, IQ markedly induced CYP1A proteins and negated the differences among groups related to diet. No consistent diet-related changes were detected in the activities of aryl sulfotransferase or N-acetyltransferase, but UDP-glucuronosyltransferase activities were elevated 2- to 3-fold in rats given chow versus AIN diets. In summary, high dose treatment with IQ was required for the induction of ACF, rats on the chow diet had more aberrant crypts than those given AIN diets and male F344 rats purchased from different vendors and fed chow diet differed with respect to their sensitivity to induction of ACF.

Abbreviations: ACF, aberrant crypt foci; AST, arylsulfotransferase; EROD, 7-ethoxyresorufin O-deethylase; IQ, 2-amino-3-methylimidazo[4,5-f]quinoline; MROD, methoxyresorufin O-demethylase; NAT, N-acetyltransferase; NCI, National Cancer Institute; PABA, p-aminobenzoic acid; PC, Purina chow; PhIP, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine; SN, Simonsen Laboratories; UDPGT, UDP-glucuronosyltransferase.


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Heterocyclic amine mutagens, such as 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), are generated during the cooking of meat and fish (1,2). In the male F344 rat, chronic administration of IQ or PhIP induces tumors at multiple sites (2), including adenocarcinomas of the colon, and short-term treatment leads to the formation of colonic aberrant crypt foci (ACF). The ACF represents a putative preneoplastic lesion (35) and has been used as an intermediate biomarker to screen for chemopreventive agents that might be effective against heterocyclic amines and other colon carcinogens (68).

We have reported that the induction of ACF by IQ and PhIP can be inhibited by several chemopreventive agents, including conjugated linoleic acids, chlorophyllin, indole-3-carbinol, green tea and black tea (913). However, in the course of conducting these and other studies we observed that ACF induction by heterocyclic amines can be influenced significantly by the dose and frequency of carcinogen administration, the type of diet and the source of experimental animal. To investigate these variables in greater detail, we examined the effect of carcinogen dose fractionation on ACF induction by IQ and compared the sensitivity of F344 rats from two different vendors after feeding AIN-76A, AIN-93G or chow diet. To provide some insight into the changes that might occur in the metabolism of IQ in the various treatment groups, several phase I and phase II enzymes were examined prior to and after carcinogen dosing.


    Materials and methods
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Chemicals
IQ was purchased from Toronto Research Chemicals (Toronto, Canada). Chemicals and reagents for western analysis and enzyme assays were from sources described elsewhere (911,14).

Animals and treatments
In Experiment 1, 28-day-old weanling male F344 rats were purchased from the National Cancer Institute (NCI). Immediately upon arrival, the NCI rats were divided randomly into groups of 10 animals and given water and AIN-93G diet (ICN Biomedicals, Aurora, OH) ad libitum. After 2 weeks, dosing with IQ or vehicle alone (controls) was started. One group of animals received 100 mg/kg body wt IQ for 2 weeks, given on alternating days by oral gavage, according to a previous study (9). Alternatively, the total cumulative carcinogen dose (105 mg) was fractionated proportionally over periods of 4, 8 or 14 weeks, administered by oral gavage on alternating days. Rats given IQ for periods <14 weeks received vehicle alone from the time of the last carcinogen dose until the end of the study. Rats were killed in week 16 and the colons were removed, fixed in buffered formalin, stained with 0.2% methylene blue and the ACF were scored as described previously (15).

The protocol used in Experiment 2 is shown in Figure 1aGo and a key to the diets and animals is given in Figure 1bGo. Each of the four groups comprised 20 weanling male F344 rats of the same age (28 days) and average body weight (100 ± 10 g). One group of rats from Simonsen Laboratories (Gilroy, CA), referred to hereafter as SN rats, and a group of NCI rats were fed Purina chow (PC) diet (PMI Feeds, St Louis, MO). Because the primary goal of this experiment was to study the influence of diet (rather than vendor) on ACF induction, two additional groups of NCI rats were included and fed either AIN-76A or AIN-93G diet (ICN Biomedicals). Diets and water were given ad libitum throughout the study. After 2 weeks on the assigned diet, three rats in each group were killed before dosing with IQ and the livers were frozen in liquid nitrogen and stored at –80°C. During the next 2 weeks (experiment weeks 3 and 4), each rat was given the carcinogen on alternating days by oral gavage (100 mg/kg body wt IQ at each injection). Twelve hours after the final injection of IQ, three rats in each group were killed and the livers were frozen and stored at –80°C. Upon thawing, portions of each liver were chopped into small pieces in ice-cold saline prior to homogenization and differential centrifugation. Protein concentrations of the cytosolic and microsomal fractions were determined using bovine serum albumin as standard (16). At the end of experiment week 10, the 14 animals that remained in each group were killed and the colons were scored with respect to size and number of ACF. No differences were detected among the various groups with respect to food and water intake or body weight gain throughout the study (not shown).



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Fig. 1. (a) Protocol used to study the effect of diet and source of male F344 rat on IQ-induced ACF and hepatic enzyme activities and (b) key showing the shading patterns used in Figures 1–4GoGoGoGo to distinguish the diets and animals. Numbers in parentheses indicate number of rats per group. Average IQ dose 7.5 mg/day, by oral gavage.

 
Alkoxyresorufin O-dealkylase assays
Liver microsomes were compared in assays for 7-ethoxyresorufin O-deethylase (EROD) and methoxyresorufin O-demethylase (MROD), which are mainly indicative of CYP1A1 and CYP1A2, respectively. A fluorometric assay was used to determine the activities of EROD and MROD (17). The product in both assays, resorufin, was quantified by its fluorescence using 540 nm excitation and 580 nm emission filters.

Western blotting
Details of the protocol used for western analysis were reported elsewhere (11,17). Briefly, liver microsomal proteins were separated by SDS–PAGE, transferred onto PVDF-plus membrane and incubated with 0.01 mg/ml anti-CYP1A1/1A2 antibody for 1 h followed by anti-mouse IgG–horseradish peroxidase conjugate. Detection was according to the manufacturer's guidelines using ECL reagents 1 and 2 (Amersham Corp., Arlington Heights, IL). The film was scanned with a densitometer to determine relative band density (arbitrary units).

NADPH-cytochrome P450 reductase activity
This assay was conducted with liver microsomes using cytochrome c as substrate, as described previously (14,18). Kinetic parameters were determined by computerized linear regression analysis of the Lineweaver–Burk plot (SigmaPlot 4.0; Jandel Scientific, San Rafael, CA).

Phase II enzyme activities
UDP-glucuronosyltransferase (UDPGT) activities of the liver microsomes were measured according to the method of Bock et al. (19). Aryl sulfotransferase (AST) activities of the liver cytosols were quantified as described by Matsui and Watanabe (20). In both the UDPGT and AST assays, enzyme activities were calculated from the loss of p-nitrophenol substrate, monitored spectrophometrically at 400 nm. Hepatic N-acetyltransferase (NAT) activities were measured using p-aminobenzoic acid (PABA) as substrate, according to the method of Andres et al. (21); the final concentrations of acetyl-CoA and PABA were 0.44 and 0.2 mM, respectively.

Statistical analysis
Results are expressed as means ± SD within a group. Data were analyzed by ANOVA using the SAS statistical package and the Waller–Duncan K-ratio was used to determine significant differences among means.


    Results
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Table IGo summarizes the results from Experiment 1, in which a cumulative dose of 105 mg IQ was fractionated over 2, 4, 8 or 14 weeks. None of the vehicle controls had ACF (not shown). In the 2 week carcinogen treatment group (7.5 mg/day IQ average dose administered) each rat had one or more ACF, whereas less than half the animals had ACF in the remaining groups given IQ. After correcting for rats bearing ACF, the total number of aberrant crypts/colon and the number of ACF/colon were significantly higher in the 2 week IQ group versus all other groups (P < 0.05). Thus, only the 2 week, high dose treatment was effective for induction of ACF by IQ, and this exposure protocol was used in Experiment 2.


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Table I. Effect of carcinogen dose fractionation on IQ-induced ACF
 
Figure 2a–cGo summarizes the results from Experiment 2 in terms of total aberrant crypts/rat, ACF/rat and aberrant crypts/ACF, respectively. No difference was observed between NCI rats fed AIN-76A diet versus AIN-93G diet (course hatching versus open bars, respectively). Rats given PC diet had a significantly greater number of aberrant crypts/colon than animals fed AIN diets (Figure 2aGo) and the average size of ACF was significantly greater in SN rats given PC diet compared with NCI rats given the AIN diets (Figure 2cGo). Moreover, SN rats on the PC diets had significantly more aberrant crypts, ACF and aberrant crypts/ACF than NCI rats on PC diet (compare solid versus fine hatched bars, Figure 2a–cGo). Rats given the PC diet had increased numbers of intermediate sized foci with 2–3 and 4–6 aberrant crypts/focus, i.e. categories ACF-2 and ACF-3, respectively (Figure 2dGo).




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Fig. 2. Effect of diet and source of male F344 rat on IQ-induced ACF formation. In (d) ACF are categorized as follows: ACF-1, 1 aberrant crypt/focus; ACF-2, 2–3 aberrant crypts/focus; ACF-3, 4–6 aberrant crypts/focus; ACF-4, >6 aberrant crypts/focus (see ref. 15 for further details). Data are given as means ± SD, n = 14; groups with different superscripts differ significantly, P < 0.05, by ANOVA.

 
Hepatic EROD and MROD activities were examined before and after IQ treatment (Figure 3Go) and the corresponding CYP1A1 and CYP1A2 proteins were analyzed by western blotting (Figure 4Go). As reported previously (11,17), high dose IQ treatment increased hepatic EROD and MROD activities (Figure 3Go) and CYP1A1 and 1A2 levels were induced in all groups (Figure 4Go). During this period, no consistent diet-related changes were detected among the various groups with respect to EROD/MROD activities and the levels of CYP1A proteins. In contrast, before IQ dosing MROD activities were significantly higher in rats given PC diet versus AIN diets (Figure 3bGo) and a corresponding 2- to 3-fold increase in CYP1A2 protein levels was detected (Figure 4bGo).



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Fig. 3. Effect of diet and source of male F344 rat on hepatic (a) EROD and (b) MROD activities prior to and after IQ dosing. Data represent means ± SD, n = 3; different superscripts indicate statistically different, P < 0.05, by ANOVA. For a key to the various groups see Figure 1bGo.

 


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Fig. 4. Effect of diet and source of male F344 rat on hepatic (a) CYP1A1 and (b) CYP1A2 prior to and after 2 week carcinogen dosing. Data shown were taken from a single western blot (upper) and are representative of findings from three or more independent determinations. For details of the conditions used for SDS–PAGE and for immunodetection using anti-CYP1A1/1A2 antibody see Materials and methods. For a key to the various groups see Figure 1bGo.

 
Differences in the activity of hepatic microsomal NADPH-cytochrome P450 reductase were also observed among the various treatment groups (Figure 5Go). Vmax values were higher in rats fed chow diet (solid symbols) compared with rats given AIN diets (open symbols); by computerized regression analysis, the results for PC groups were 183.9 and 152.1 versus 103.3 and 102.7 nmol/min/mg for the AIN-76A and AIN-93G groups, respectively (P < 0.05). The Km was lower in NCI rats fed AIN-76A diet (33 µM) compared with SN rats given chow (106 µM), but other differences in Km were not significant. Changes in NADPH-cytochrome P450 reductase affect cytochromes P450 indirectly, as distinct from direct alterations in the levels or activities of CYP1A1 and CYP1A2.



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Fig. 5. Lineweaver–Burk plot of hepatic NADPH-cytochrome P450 reductase activities. Data points and bars, means ± SD (n = 3). Kinetic parameters (Vmax and Km) were obtained by regression analysis using SigmaPlot v.4.0 (see text).

 
To determine whether the relative sensitivity of SN rats compared with NCI rats might be due, in part, to changes in the activities of phase II enzymes, assays were conducted for UDPGT, AST and NAT (Table IIGo). No consistent diet-related changes in AST or NAT were detected among the various treatment groups, but UDPGT activities were significantly higher in rats given the PC diet compared with those fed the AIN diets. Thus, mean activities for UDPGT in the two groups given the PC diet were 29.64 and 39.46 versus 15.95 and 12.64 nmol/min/mg in the AIN-fed rats (P < 0.05). Similar trends were observed for AST, NAT and UDPGT after IQ dosing (data not shown). Collectively, the results indicated that UDPGT activities were 2- to 3-fold higher in rats fed the PC diet compared with those given the AIN diets.


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Table II. Effect of diet and source of rat on phase II enzyme activities (nmol/min/mg)
 

    Discussion
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
In the present investigation, we examined certain parameters associated with the induction of colonic ACF by IQ in the male F344 rat. In Experiment 1, short-term, high dose treatment with IQ for 2 weeks was significantly more effective in inducing ACF than the same amount of carcinogen fractionated over longer periods of time. In general, therefore, one might predict that human exposure to heterocyclic amines at chronic low levels will pose less risk than acute, high level ingestion, as might occur for example during frequent `all-you-can-eat' consumption of well-done meat. Epidemiological studies generally find no association between modest intake of cooked meat and cancers at various sites, but increased risk has been reported for colorectal and other cancers in individuals consuming higher quantities of well-done meat (2224).

In Experiment 2, rats fed a PC diet were significantly more sensitive to the induction of colonic aberrant crypts than animals given AIN diets, but no differences were detected between groups given AIN-76A versus AIN-93G diet. The latter observation is important in that the American Institute of Nutrition recommended substituting AIN-93G diet for AIN-76A diet in rodent studies (25). Although AIN-76A was used extensively for 16 years, nutritional and technical problems were encountered which prompted the following principal changes: the use of soybean oil in place of corn oil; substitution of corn starch for sucrose; reduced phosphorus and manganese levels; use of L-cystine as an amino acid supplement for casein; increased amounts of vitamins E, K and B12 (25). From Figure 2Go, the suggested substitution of AIN-93G for AIN-76A (25) is unlikely to significantly affect the anticipated results from chronic feeding studies using IQ as an initiating agent, but rats given the PC diet are predicted to be more sensitive to heterocyclic amine-induced carcinogenesis. However, in a previous study, F344 rats given chow or AIN-76A diet for 6 weeks prior to a single oral injection of carcinogen had no differences in hepatic IQ–DNA adduct levels at 24 h (26). One reason for this discrepancy might be related to the chow diet. Laboratory rodent chow is composed of such ingredients as ground yellow corn, soybean meal, beet pulp, brewers yeast, alfalfa, molasses, wheatgerm, whey, fish meal and meat meal, and as a consequence the PC diet is less well defined than AIN diets. For example, the phytochemical composition of alfalfa most likely varies from batch to batch and this could influence the `basal' levels of phase I and phase II enzymes.

In the present study, no changes in NAT or AST activities were observed in response to the various dietary treatments, but the activities of NADPH-cytochrome P450 reductase, CYP1A2 and UDPGT were augmented in rats given the PC diet versus the AIN diets, presumably resulting in enhanced activation of IQ and increased elimination of IQ glucuronides. It should be pointed out that the present investigation used pseudo-substrates for the enzyme assays and specific isoforms of the phase II enzymes such as NAT1, NAT2 and UDPGT1A were not examined (27,28). Thus, it will be important to study the effects of the PC and AIN diets on activation and detoxification pathways using heterocyclic amines as substrates, coupled with thorough examination of the carcinogen metabolite profiles in vivo.

An additional, unexpected finding from this work was that male F344 rats from NCI and from Simonsen Laboratories differed significantly with respect to their sensitivity to induction of colonic aberrant crypts by IQ. To our knowledge, this is the first carcinogenicity study to compare the sensitivity of male F344 rats from different suppliers and showing lower numbers of aberrant crypts in rats from NCI. Although the PC diet alone was tested in rats from different vendors, we anticipate fewer IQ-induced aberrant crypts in NCI versus SN rats after feeding either of the AIN diets. Based on the results for different vendors (Figure 2Go), an investigation of other commonly used suppliers of laboratory animals appears to be warranted. Indeed, these findings have potentially important implications for inter-laboratory comparisons and for efforts to maintain a database on carcinogenic potencies of various chemicals tested in rodent carcinogenicity bioassays (29). The results provide one possible explanation for inconsistencies among studies that appear identical in all other respects, i.e. diet, duration of experiment, dose and type of carcinogen and species and sex of experimental animal.

In summary, the present investigation has shown that the induction of ACF by IQ can be significantly influenced by a multiplicity of factors, including carcinogen dose fractionation, diet and source of laboratory animal. Short duration treatment with IQ at the highest dose was required for induction of ACF, rats given the PC diet had more aberrant crypts than those fed the AIN diets and animals purchased from two different vendors and fed the PC diet differed with respect to their sensitivity to induction of ACF.


    Acknowledgments
 
This work was supported in part by Public Health Service grant CA65525 from the National Institutes of Health.


    Notes
 
3 To whom correspondence should be addressed Email: rod.dashwood{at}orst.edu Back


    References
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 

  1. Skog,K.I., Johansson M.A.E. and Jagerstad,M.I. (1998) Carcinogenic heterocyclic amines in model systems and cooked foods: a review on formation, occurrence and intake. Food Chem. Toxicol., 36, 879–896.[ISI][Medline]
  2. Sugimura,T. (1997) Overview of carcinogenic heterocyclic amines. Mutat. Res., 376, 211–219.[ISI][Medline]
  3. Bird,R.P. (1987) Observation and quantification of aberrant crypts in the murine colon treated with a colon carcinogen—preliminary findings. Cancer Lett., 37, 147–151.[ISI][Medline]
  4. Pretlow,T., Barrow,B., Aston,W., O'Riordan,M., Jurcisek,J.A. and Stellato,T. (1991) Aberrant crypts: putative preneoplastic foci in human colonic mucosa. Cancer Res., 51, 1564–1567.[Abstract]
  5. Vivona,A.A., Shpitz,B., Medline,A., Bruce,W.R., Hay,K., Ward,M.A., Stern,H.S. and Gallinger,S. (1993) K-ras mutations in aberrant crypt foci, adenomas and adenocarcinomas during azoxymethane-induced colon carcinogenesis. Carcinogenesis, 14, 1777–1781.[Abstract]
  6. Lam,L.K.T. and Zhang,J. (1991) Reduction of aberrant crypt formation in the colon of CF1 mice by potential chemopreventive agents. Carcinogenesis, 12, 2311–2315.[Abstract]
  7. Pereira,M.A., Barnes,L.H., Rassman,V., Kelloff,G. and Steele,V.E. (1994) Use of azoxymethane-induced foci of aberrant crypts in rat colon to identify potential cancer chemopreventive agents. Carcinogenesis, 15, 1049–1054.[Abstract]
  8. Dashwood,R.H. (1999) Early detection and prevention of colorectal cancer. Oncol. Rep., 6, 277–281.[ISI][Medline]
  9. Liew,C., Schut,H.A.J., Chin,S.F., Pariza,M.W. and Dashwood,R.H. (1995) Protection of conjugated linoleic acids against 2-amino-3-methylimidazo[4,5-f]quinoline-induced colon carcinogenesis in the F344 rat: a study of inhibitory mechanisms. Carcinogenesis, 16, 3037–3043.[Abstract]
  10. Guo,D., Schut,H.A.J., Davis,C.D., Snyderwine,E.G., Bailey,G.S. and Dashwood,R.H. (1995) Protection by chlorophyllin and indole-3-carbinol against 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced DNA adducts and colonic aberrant crypts in the F344 rat. Carcinogenesis, 16, 2931–2937.[Abstract]
  11. Xu,M., Bailey,A.C., Hernaez,J.F., Taoka,C.R., Schut,H.A.J. and Dashwood,R.H. (1996) Protection by green tea, black tea and indole-3-carbinol against 2-amino-3-methylimidazo[4,5-f]quinoline-induced DNA adducts and colonic aberrant crypts in the F344 rat. Carcinogenesis, 17, 1429–1434.[Abstract]
  12. Dashwood,R.H. (1997) Chlorophylls as anticarcinogens. Int. J. Oncol., 10, 721–727.[ISI]
  13. Dashwood,R.H., Xu,M., Hernaez,J.F., Hasaniya,N., Youn,K. and Razzuk,A. (1999) Cancer chemopreventive mechanisms of tea against heterocyclic amine mutagens from cooked meat. Proc. Soc. Exp. Biol. Med., 220, 239–243.[Abstract]
  14. Tachino,N., Guo,D., Dashwood,W.M., Yamane,S., Larsen,R. and Dashwood,R.H. (1994) Mechanisms of the in vitro antimutagenic action of chlorophyllin against benzo[a]pyrene: studies of enzyme inhibition, molecular complex formation and degradation of the ultimate carcinogen. Mutat. Res., 308, 191–203.[ISI][Medline]
  15. Tachino,N., Hayashi,R., Liew,C., Bailey,G. and Dashwood,R.H. (1995) Evidence for ras gene mutation in 2-amino-3-methylimidazo[4,5-f]quinoline-induced colonic aberrant crypts in the rat. Mol. Carcinog., 12, 187–192.[ISI][Medline]
  16. Lowry,O.H., Rosebrough,N.J., Farr,A.L. and Randall,R.J. (1951) Protein measurement with the Folin phenol reagent. J. Biol. Chem., 193, 265–275.[Free Full Text]
  17. Xu,M., Schut,H.A.J., Bjeldanes,L.F., Williams,D.E., Bailey,G.S. and Dashwood,R.H. (1997) Inhibition of 2-amino-3-methylimidazo[4,5-f]quinoline–DNA adducts by indole-3-carbinol: dose–response studies in the rat colon. Carcinogenesis, 18, 2149–2153.[Abstract]
  18. Hasaniya,N., Youn,K., Xu,M., Hernaez,J. and Dashwood,R.H. (1997) Inhibitory activity of green and black tea in a free radical-generating system using 2-amino-3-methylimidazo[4,5-f]quinoline as substrate. Jpn J. Cancer Res., 88, 553–558.[ISI][Medline]
  19. Bock,K.W., Burchell,B., Dutton,G.J., Hanninen,O., Mulder,G.J., Owens,I.S., Siest,G. and Tephly,T.R. (1983) UDP-glucuronosyltransferase activities: guidelines for consistent interim terminology and assay conditions. Biochem. Pharmacol., 32, 953–955.[ISI][Medline]
  20. Matsui,M. and Watanabe,H.K. (1982) Developmental alteration of hepatic UDP-glucuronosyltransferase and sulphotransferase towards androsterone and 4-nitrophenol in Wistar rats. Biochem. J., 204, 441–447.[ISI][Medline]
  21. Andres,H.H., Klem,A.J., Szabo,S.M. and Weber,W.W. (1985) New spectrophotometric and radiochemical assays for acetyl-CoA:arylamine N-acetyltransferase applicable to a variety of arylamines. Anal. Biochem., 145, 367–375.[ISI][Medline]
  22. Felton,J.S., Malfatti,M.A., Knize,M.G., Salmon,C.P., Hopmans,E.C. and Wu,R.W. (1997) Health risks of heterocyclic amines. Mutat. Res., 376, 37–41.[ISI][Medline]
  23. Augustsson,K., Skog,K., Jagerstad,M., Dickman,P.W. and Steineck,G. (1999) Dietary heterocyclic amines and cancer of the colon, rectum, bladder and kidney: a population-based study. Lancet, 353, 703–707.[ISI][Medline]
  24. Zheng,W., Gustafson,D.R., Sinha,R., Cerhan,J.R., Moore,D., Hong,C.P., Anderson,K.E., Kushi,L.H., Sellers,T.A. and Folsom,A.R. (1998) Well-done meat intake and the risk of breast cancer. J. Natl Cancer Inst., 90, 1724–1729.[Abstract/Free Full Text]
  25. Reeves,P.G. (1997) Components of the AIN-93 diets as improvements in the AIN-76A diet. J. Nutr., 127 (suppl.), 838S–841S.[Medline]
  26. Schut,H.A.J. (1994) Dietary modulation of DNA adduct formation of the food mutagen 2-amino-3-methylimidazo[4,5-f]quinoline in the male Fischer 344 rat. Environ. Health Perspect., 102 (suppl. 6), 57–60.
  27. Oda,Y., Yamazaki,H. and Shimada,T. (1999) Role of human N-acetyltransferases, NAT1 or NAT2, in genotoxicity of nitroarenes and aromatic amines in Salmonella typhimurium NM6001 and NM6002. Carcinogenesis, 20, 1079–1083.[Abstract/Free Full Text]
  28. Nowell,S.A., Massengill,J.S., Williams,S., Radominska-Pandya,A., Tephly,T.R., Cheng,Z., Strassberg,C.P., Tukey,R.H., MacLeod,S.L., Lang,N.P. and Kadlubar,F.F. (1999) Glucuronidation of 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine by human microsomal UDP-glucuronosyltransferases: identification of specific UTG1A family isoforms involved. Carcinogenesis, 20, 1107–1114.[Abstract/Free Full Text]
  29. Gold,L.S., Slone,T.H. and Ames,B.N. (1998) What do animal cancer tests tell us about human cancer risk?: overview of analyses of the carcinogenic potency database. Drug Metab. Rev., 30, 359–404.[ISI][Medline]
Received April 2, 1999; revised July 8, 1999; accepted August 19, 1999.