Chemoprevention of azoxymethane-induced rat colon carcinogenesis by dietary capsaicin and rotenone

The modifying effects of dietary administration of capsaicin, which is the principal pungent capsicum fruit, and rotenone, which is a naturally occurring pesticide derived from Derris and Lonchorcarpus species, on azoxymethane (AOM)-induced colon tumorigenesis were investigated in male F344 rats. Gavage with capsaicin and rotenone significantly elevated phase II enzymes, glutathione S-transferase (GST) and quinone reductase (QR), in the liver and colon. In an aberrant crypt foci (ACF) bioassay, feeding of capsaicin and rotenone at a dose of 500 ppm for 4 weeks significantly inhibited ACF formation induced by AOM (20 mg/kg body weight, once a week for 2 weeks). In a subsequent long-term study designed to confirm the protective effects of both compounds on ACF development, one group was treated with AOM alone and four other groups received the carcinogen treatment plus diets containing 500 ppm test compounds for 4 weeks (initiation phase) and for 34 weeks (post-initiation phase). Two groups were treated with capsaicin or rotenone alone (500 ppm in diet) and one group was maintained on the basal diet. At the termination of the study, dietary exposure of capsaicin during the initiation phase was found to significantly reduce the incidence of colonic adenocarcinoma (60% vs. 24%, 60% reduction, P=0.0407). Rotenone feeding during the post-initiation phase also reduced the frequency of colonic adenocarcinoma (60% vs. 19%, 68% reduction, P=0.0226). Our results suggest that two natural compounds, capsaicin and rotenone, might be useful for the prevention of human colon cancers.     

Dietary supplementation of the citrus antioxidant auraptene inhibits N,N-diethylnitrosamine-induced rat hepatocarcinogenesis

OBJECTIVES: We have previously reported that an antioxidant, auraptene (AUR), isolated from citrus fruit effectively inhibits chemically induced carcinogenesis in digestive tracts, such as the oral cavity, esophagus and large bowel. In this study, we investigated the modifying effects of dietary supplementation with AUR on N,N-diethylnitrosamine (DEN)-initiated hepatocarcinogenesis in male F344 rats in two different experiments to determine whether the compound exerts a cancer-chemopreventive action in other organs. METHODS: In the first experiment, animals were fed diets containing AUR at dose levels of 100 and 500 ppm for 7 weeks 1 week before, during, and 1 week after the start of liver carcinogenesis induced by DEN (40 ppm in drinking water for 5 weeks) to predict the modulatory effect on hepatocarcinogenesis. After 7 weeks, the numbers of hepatocellular enzyme-altered foci (EAF; cm(2)) which stained positive for the placental form of glutathione S-transferase (GST-P) and transforming growth factor (TGF)-alpha were determined on immunohistochemically stained sections. In the second experiment conducted to confirm the findings, animals subjected to DEN treatment were fed AUR-containing diets (100 and 500 ppm) during either the initiation stage ('initiation' feeding for 7 weeks) or post-initiation phase ('post-initiation' feeding for 25 weeks) of DEN-induced hepatocarcinogenesis. RESULTS: In the first experiment, feeding with AUR at both doses during DEN exposure decreased the mean numbers of GST-P-positive and TGF-alpha-positive EAF/cm(2), and the reduction in the number of TGF-alpha-positive EAF by feeding 500 ppm AUR was statistically significant (p < 0.005). In the second experiment, the 'initiation' feeding with 500 ppm AUR significantly inhibited the incidence (33 vs. 83%, p = 0.000511) and multiplicity (0.67 +/- 1.09 vs. 1.96 +/- 1.85, p < 0.005) of liver cell carcinoma. Also, the 'post-initiation' feeding with AUR at both doses significantly reduced the development of hepatocellular carcinoma (100 ppm: incidence, 15%, p = 0.000006; multiplicity: 0.25 +/- 0.64, p < 0.001; 500 ppm: incidence, 11%, p = 0.000002; multiplicity, 0.26 +/- 0.81, p < 0.001). In addition, AUR feeding reduced cell proliferation and the apoptotic index in liver cell neoplasms. CONCLUSIONS: The results suggest that the citrus antioxidant AUR is a potential chemopreventive agent against DEN-induced hepatocarcinogenesis in rats.     

Citrus auraptene inhibits chemically induced colonic aberrant crypt foci in male F344 rats

The modifying effect of dietary administration of auraptene isolated from the peel of citrus fruit (Citrus natsudaidai Hayata) on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) was investigated in rats. Male F344 rats were given s.c. injections of AOM (15 mg/kg body wt) once a week for 3 weeks to induce ACF. They also received diets containing 100 or 500 p.p.m. auraptene for 5 weeks, starting 1 week before the first dose of AOM. At termination of the study (week 5) dietary administration of auraptene caused a significant reduction in the frequency of ACF in a dose- dependent manner (P < 0.05). Feeding of auraptene suppressed expression of cell proliferation biomarkers (5-bromo-2'-deoxyuridine labeling- index, ornithine decarboxylase activity, polyamine content and number of silver stained nucleolar organizer region protein particles) in the colonic mucosa and the occurrence of micronuclei caused by AOM. Also, auraptene increased the activities of phase II enzymes (glutathione S- transferase and quinone reductase) in the liver and colon. These findings might suggest that inhibition of AOM-induced ACF may be associated, in part, with increased activity of phase II enzymes in the liver and colon and suppression of cell proliferation in the colonic mucosa.      

Dietary prevention of azoxymethane-induced colon carcinogenesis with rice-germ in F344 rats.

The modifying effect of dietary administration of defatted rice-germ and gamma-aminobutyric acid (GABA)-enriched defatted rice-germ on azoxymethane (AOM)-induced colon carcinogenesis was investigated in two experiments with male F344 rats. In the first experiment (the pilot study), the effects of the defatted rice-germ, the GABA-enriched defatted rice-germ and rice-germ on AOM-induced (15 mg/kg body wt once a week for 3 weeks) formation of aberrant crypt foci (ACF) were examined. The latter two preparations (2.5% in the diet) significantly inhibited ACF formation (P < 0.005). In the second experiment, a long-term study of the effects of rice-germ was done. One group was treated with AOM alone, four groups received the carcinogen and were fed the diets containing 2.5% rice-germ or 2.5% GABA-enriched defatted rice-germ for 5 (initiation phase) or 30 weeks (post-initiation phase), two groups were treated with rice-germ or GABA-enriched defatted rice-germ alone and one group was kept on the basal diet. At the termination of the study, dietary exposure to rice-germ during the initiation phase significantly reduced the incidence of colonic adenocarcinoma (71 versus 29%, P < 0.01). GABA-enriched defatted rice-germ or rice-germ during the post-initiation phase also decreased the frequency of colonic adenocarcinoma (71 versus 20%, GABA-enriched defatted rice-germ feeding, P < 0.01; 27%, rice-germ feeding, P < 0.01). These data suggest that constituents of rice-germ are possible dietary preventatives for human colon cancers.     

Chemoprevention of Azoxymethane-induced Rat Colon Carcinogenesis by a Xanthine Oxidase Inhibitor, 1'-Acetoxychavicol Acetate

In our studies to find natural compounds with chemopreventive efficacy in foods, using azoxymethane (AOM)-induced colonic aberrant crypt foci and colonic mucosal cell proliferation as biomarkers, a x an thine oxidase inhibitor, 1′-acetoxychavicol acetate (ACA), present in the edible plant Languas galanga from Thailand was found to be effective. This study was conducted to test the ability of ACA to inhibit AOM-induced colon tumorigenesis when it was fed to rats during the initiation or post-initiation phase. Male F344 rats were given three weekly s.c. injections of AOM (15mg/kg body weight) to induce colonic neoplasms. They were fed diet containing 100 or 500 ppm ACA for 4 weeks, starting one week before the first dosing of AOM (the initiation feeding). The other groups were fed the ACA diet for 34 weeks, starting one week after the last AOM injection (the post-initiation feeding). At the termination of the study (week 38), AOM had induced 71% incidence of colonic adenocarcinoma (12/17 rats). The initiation feeding with ACA caused significant reduction in the incidence of colon carcinoma (54% inhibition by 100 ppm ACA feeding and 77% inhibition by 500 ppm ACA feeding, P=0.03 and P=0.001, respectively). The post-initiation feeding with ACA also suppressed the incidence of colonic carcinoma (45% inhibition by 100 ppm ACA feeding and 93% inhibition by 500 ppm ACA feeding, P=0.06 and P=0.00003, respectively). Such inhibition was dose-dependent and was associated with suppression of proliferation biomarkers, such as ornithine decarboxylase activity in the colonic mucosa, and blood and colonic mucosal polyamine contents. ACA also elevated the activities of phase II enzymes, glutathione S-transferase (GST) and quinone reductase (QR), in the liver and colon. These results indicate that ACA could inhibit the development of AOM-induced colon tumorigenesis through its suppression of cell proliferation in the colonic mucosa and its induction of GST and QR. The results confirm our previous finding that ACA feeding effectively suppressed the development of colonic aberrant crypt foci. These findings suggest possible chemopreventive ability of ACA against colon tumorigenesis.     

Silymarin,a naturally occurring polyphenolic antioxidant flavonoid,inhibits azoxymethane-induced colon carcinogenesis in male F344 rats

The modifying effect of dietary administration of the polyphenolic antioxidant flavonoid silymarin, isolated from milk thistle [Silybum marianum (L.) Gaertneri], on AOM-induced colon carcinogenesis was investigated in male F344 rats. In the short-term study, the effects of silymarin on the development of AOM-induced colonic ACF, being putative precursor lesions for colonic adenocarcinoma, were assayed to predict the modifying effects of dietary silymarin on colon tumorigenesis. Also, the activity of detoxifying enzymes (GST and QR) in liver and colonic mucosa was determined in rats gavaged with silymarin. Subsequently, the possible inhibitory effects of dietary feeding of silymarin on AOM-induced colon carcinogenesis were evaluated using a long-term animal experiment. In the short-term study, dietary administration of silymarin (100, 500 and 1,000 ppm in diet), either during or after carcinogen exposure, for 4 weeks caused significant reduction in the frequency of colonic ACF in a dose-dependent manner. Silymarin given by gavage elevated the activity of detoxifying enzymes in both organs. In the long-term experiment, dietary feeding of silymarin (100 and 500 ppm) during the initiation or postinitiation phase of AOM-induced colon carcinogenesis reduced the incidence and multiplicity of colonic adenocarcinoma. The inhibition by feeding with 500 ppm silymarin was significant (p < 0.05 by initiation feeding and p < 0.01 by postinitiation feeding). Also, silymarin administration in the diet lowered the PCNA labeling index and increased the number of apoptotic cells in adenocarcinoma. beta-Glucuronidase activity, PGE(2) level and polyamine content were decreased in colonic mucosa. These results clearly indicate a chemopreventive ability of dietary silymarin against chemically induced colon tumorigenesis and will provide a scientific basis for progression to clinical trials of the chemoprevention of human colon cancer.     

Dietary Conjugated Linolenic Acid Inhibits Azoxymethane-induced Colonic Aberrant Crypt Foci in Rats

The modifying effects of dietary feeding of conjugated linolenic acid (CLN) isolated from the seeds of bitter gourd ( Momordica charantia ) on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) were investigated in male F344 rats to predict its possible cancer chemopreventive efficacy. The effect of CLN on the proliferating cell nuclear antigen (PCNA) index in colonic ACF was also examined. Rats were given subcutaneous injections of AOM (20 mg/ kg body weight) once a week for 2 weeks to induce ACF. They also received the experimental diet containing 0.01%, 0.1% or 1% CLN for 5 weeks, starting one week before the first dosing of AOM. AOM exposure produced a substantial number of ACF (108±21/rat) at the end of the study (week 4). Dietary administration of CLN caused a significant reduction in the frequency of ACF: 87±14 (19.4% reduction, P <0.05) at a dose of 0.01%, 69±28 (36.1% reduction, P <0.01) at a dose of 0.1% and 40±d6 (63.0% reduction, P <0.001) at a dose of 1%. Also, CLN administration lowered the PCNA index and induced apoptosis in ACF. These findings might suggest possible chemopreventive activity of CLN in the early phase of colon tumorigenesis through modulation of cryptal cell proliferation activity and/or apoptosis.     

Suppression of Azoxymethane-induced Rat Colon Aberrant Crypt Foci by Dietary Protocatechuic Acid

The modifying effect of dietary exposure to protocatechuic acid (PCA) on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) was investigated in male F344 rats. The effects of PCA feeding on the silver-stained nucleolar organizer regions protein (AgNORs) count in the colonic epithelial cells and on the ornithine decarboxylase (ODC) activity in the colonic mucosa were also estimated. Animals were given weekly s.c. injections of AOM (15 mg/kg body weight) for 3 weeks to induce ACF. These rats were fed diet containing 1000 or 2000 ppm PCA for 5 weeks, starting one week before the first dosing of AOM. All rats were killed 2 weeks after the last AOM injection, to measure the number of ACF, ODC activity, and AgNORs count per nucleus in the colon. In rats given AOM and PCA, the frequency of ACF/colon was significantly decreased compared with that in rats given AOM alone ( P < 0.005 at 1000 and P < 0.05 at 2000 ppm). ODC activity in the colon of rats given AOM and PCA at both doses was also significantly lower than that of rats treated with AOM alone ( P < 0.05). Similarly, the mean AgNORs count in rats fed PCA was significantly smaller than that of rats treated with AOM alone ( P < 0.0001). Treatment with PCA alone did not affect these three biomarkers. These results provide further evidence that PCA could be a chemopreventive agent against rat colon carcinogenesis.     

Modifying effects of a flavonoid morin on azoxymethane-induced large bowel tumorigenesis in rats.

The modifying effect of dietary exposure to a flavonoid morin during the initiation and post-initiation phases of azoxymethane (AOM)-initiated colorectal carcinogenesis was investigated in male F344 rats. A total of 55 animals were initiated with AOM by weekly s. c. injections of 15 mg/kg body wt for 3 weeks to induce colorectal neoplasms. Rats were fed a diet containing 500 p.p.m. morin for 5 ('initiation feeding') or 28 ('post-initiation feeding') weeks. Other groups contained rats treated with morin alone (500 p.p.m. in diet) and untreated rats. At the end of the study (32 weeks), the incidence of adenocarcinoma in the large intestine of rats initiated with AOM together with (43%) or followed by (29%) a diet containing morin was smaller than that of rats given AOM alone (75%). A significant difference was found between 'post-initiation feeding' and untreated groups (P = 0.023). Although both 'initiation feeding' and 'post-initiation feeding' of morin reduced polyamine levels in colorectal mucosa and blood, 'post-initiation feeding' of morin significantly decreased the proliferating cell nuclear antigen-positive index in aberrant crypt foci. 'Post-initiation feeding' of morin significantly elevated glutathione S-transferase and quinone reductase activities in the liver and large bowel, but 'initiation feeding' caused a significant elevation of these enzymes activities only in the large bowel. These results indicate that morin could exert a weak chemopreventive effect on large bowel tumorigenesis induced by AOM when fed during the post-initiation phase.     

Intake of beer inhibits azoxymethane-induced colonic carcinogenesis in male Fischer 344 rats

Modulatory effects of beer consumption on azoxymethane (AOM)-induced rat colonic carcinogenesis in male Fischer 344 rats were investigated. Single cell gel electrophoresis assay indicated that DNA damage of colonocytes, induced by a single AOM injection (15 mg/kg body weight), was significantly reduced in rats fed beer or malt extract for 2 weeks. Examination of aberrant crypt foci (ACF) formation in colonic mucosa, induced by AOM (15 mg/kg body weight; twice weekly), revealed that feeding of beer during the whole experimental period of 5 weeks significantly reduced the number of ACF by 35%. In the post-initiation protocol, a reduction in ACF formation by 26% was not significant. The efficacy in inhibition of ACF formation varied with the brand of beer. ACF formation was significantly reduced in rats treated with freeze-dried beer (FD Beer), but not with ethanol, suggesting that nonvolatile components of beer are responsible for the reduction. Significant suppression of ACF formation was observed in groups treated with hot water extract of malt, especially with extracts of colored malts, although no reduction was observed by feeding with hops extract. A long-term experiment of 42 weeks indicated that intake of beer decreased tumor incidence by 22% and decreased the number of neoplastic lesions, including adenocarcinomas and adenomas, by 44%. These results suggest that components of beer have chemopreventive effects on colonic carcinogenesis induced by AOM and that intake of beer may contribute to a reduction in the risk of cancer susceptibility.     

Dietary conjugated linolenic acid inhibits azoxymethane-induced colonic aberrant crypt foci in rats.

The modifying effects of dietary feeding of conjugated linolenic acid (CLN) isolated from the seeds of bitter gourd (Momordica charantia) on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) were investigated in male F344 rats to predict its possible cancer chemopreventive efficacy. The effect of CLN on the proliferating cell nuclear antigen (PCNA) index in colonic ACF was also examined. Rats were given subcutaneous injections of AOM (20 mg/kg body weight) once a week for 2 weeks to induce ACF. They also received the experimental diet containing 0.01%, 0.1% or 1% CLN for 5 weeks, starting one week before the first dosing of AOM. AOM exposure produced a substantial number of ACF (108 +/- 21/rat) at the end of the study (week 4). Dietary administration of CLN caused a significant reduction in the frequency of ACF: 87 +/- 14 (19.4% reduction, P < 0.05) at a dose of 0.01%, 69 +/- 28 (36.1% reduction, P < 0.01) at a dose of 0.1% and 40 +/- 6 (63.0% reduction, P < 0.001) at a dose of 1%. Also, CLN administration lowered the PCNA index and induced apoptosis in ACF. These findings might suggest possible chemopreventive activity of CLN in the early phase of colon tumorigenesis through modulation of cryptal cell proliferation activity and/or apoptosis.     

Azoxymethane-induced beta-catenin-accumulated crypts in colonic mucosa of rodents as an intermediate biomarker for colon carcinogenesis

It is now well established that bile acids act as colon tumor promoters. However, a previous study provided conflicting data showing that dietary exposure of cholic acid (CHA), a primary bile acid, inhibits the carcinogen-induced formation of aberrant crypt foci (ACF), possible preneoplastic lesions, in colonic mucosa of rodents. Recently we found beta-catenin-accumulated crypts (BCAC) in colonic mucosa of rats initiated with azoxymethane (AOM) and provided evidence that BCAC might be preneoplastic lesions independent from ACF. In the present study, we investigated the modifying effects of dietary CHA on the formation of BCAC as well as ACF in male F344 rats after exposure to AOM to determine if the differences in the effect of CHA on these lesions could account for this discrepancy. The results indicate that administration of CHA (0.5%) in the diet during the post-initiation phase significantly reduced the total number, multiplicity and size of ACF (P < 0.00001) in AOM-exposed colonic mucosa as reported previously. The number of ACF even with >4 aberrant crypts/focus was also decreased significantly (P < 0.0002), suggesting that the large ACF are little resistant to continuous feeding of 0.5% CHA diet. Interestingly, the dietary CHA significantly enhanced both the multiplicity (P < 0.002) and size (P < 0.00001), but not the incidence, of AOM-induced BCAC when compared with the control diet group. Importantly, the number of large BCAC with >6 crypts/lesion was increased significantly by the dietary CHA (P < 0.003). Our results support the concept that BCAC are precursors of colon tumors and indicate the usefulness of BCAC as intermediate biomarkers for colon carcinogenesis, although the methodology for their detection requires further improvement.     

A xanthine oxidase inhibitor 1'-acetoxychavicol acetate inhibits azoxymethane-induced colonic aberrant crypt foci in rats

The modifying effect of dietary administration of a xanthine oxidase inhibitor 1'-acetoxychavicol acetate (ACA) present in an edible plant Languas galanga in Thailand on the development of azoxymethane (AOM)- induced colonic aberrant crypt foci (ACF) was investigated in rats. Male F344 rats were given s.c. injections of AOM (15 mg/kg body wt) once a week for 3 weeks to induce colonic ACF. They were fed the diets containing 100 or 200 ppm ACA for 5 weeks, starting 1 week before the first dosing of AOM. At the termination of the study (week 5), AOM induced 118 +/- 28 ACF/colon. Dietary administration of ACA caused significant reduction in the frequency of ACF (41% inhibition by 100 ppm ACA feeding and 37% inhibition by 200 ppm ACA feeding, P<0.01). Such inhibition might be associated with suppression of the proliferation biomarkers' expression such as ornithine decarboxylase activity in the colonic mucosa, number of silver-stained nucleolar organizer regions' protein in the colonic mucosal cell nuclei and blood polyamine content. These results indicate that ACA could inhibit the development of AOM-induced ACF through its suppression of cell proliferation in the colonic mucosa and ACA might be a possible chemopreventive agent against colon tumourigenesis.      

The influence of simple sugars and starch given during pre- or post-initiation on aberrant crypt foci in rat colon

The aim of the present study was to investigate the enhancing effect of dietary sugar on the development of aberrant crypt foci (ACF) in male F344 rats initiated with azoxymethane (AOM). The potential role of sugar as either a co-initiator or a promoter was investigated by giving diets high in sucrose and dextrin (61%) during either the pre-initiation, the initiation, and/or the post-initiation stage of the ACF development. The colonic cell proliferation, activity of colonic phase II enzymes, and a biomarker of lipid peroxidation were additionally examined in order to obtain information on the specific mechanisms involved in the suggested effect of sucrose and dextrin on ACF development. The number of large sized and the total number of ACF were significantly increased by feeding sucrose and dextrin in the post-initiation period. No positive association between colonic cell proliferation and ACF was seen. The level of oxidative stress in the cytosol from the proximal colon and colonic glutathione transferase and quinone reductase was not affected by the sugar treatments. The overall results from this study show that sucrose and dextrin enhance the number of preneoplastic lesions in AOM-initiated rats, and act primarily as promoters in the development of ACF.     

Preventive effect of fermented brown rice and rice bran against colon carcinogenesis in male F344 rats

Epidemiological and preclinical studies demonstrate that nutrition plays an important role in the etiology of cancer. It has been reported that rice components, especially rice germ plays a key role in prevention of cancer. The experiments described here examined the potential anticancer properties of brown rice fermented by Aspergillus Oryzae (FBRA) in male F344 rats using inhibition of the formation of azoxymethene (AOM) induced aberrant crypt foci (ACF) and tumors in the colon as the measure of preventive efficacy. The agent was administered at 2.5 and 5% levels in the diet during the initiation phase (during and until 1 week after carcinogen treatment) and/or post-initiation phase (beginning 1 week after carcinogen treatment) of carcinogenesis. In the ACF and tumor studies, rats were sacrificed 5 or 40 weeks after the initiation of AOM treatment (15 mg/kg body weight, once weekly for 3 weeks), respectively. Colonic ACF and tumors were evaluated histopathologically. Administration of 2.5 and 5% FBRA in the diet continuously during initiation and post-initiation period significantly inhibited the ACF formation in rats treated with AOM, compared with rats treated with AOM alone (99+/-24.1 and 79+/-18.4 vs. 139.5+/-27.7, respectively). In addition, administration of 5% FBRA in the diet during the post-initiation phase significantly suppressed the incidence (44 vs.18%) and multiplicity (0.93+/-0.96 vs. 0.18+/-0.40) of colon adenocarcinomas as compared to those given the control diet. In addition, 5% FBRA in the diet during post-initiation phase caused significant inhibition of cell proliferation in the colonic mucosa as compared to the group fed the control diet (81% reduction, p<0.05). These observations demonstrated for the first time that FBRA inhibits colon tumor development in rats, and suggest that it is a promising dietary supplement for prevention of human colon cancer.     

Inhibition of azoxymethane-induced aberrant crypt foci in rats by natural compounds, caffeine, quercetin and morin.

The modifying effects of dietary administration of natural compounds, caffeine, quercetin and morin, which are present in our daily food, on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) were investigated in rats and compared to that of a metabolic inhibitor of AOM, disulfiram. Male F344 rats were given s. c. injections of AOM (15 mg/kg body weight) once a week for 3 weeks to induce ACF. They also received the experimental diets containing one of test compounds (500 ppm) for 5 weeks, starting one week before the first dosing of AOM. At the termination of the study (week 5), AOM exposure produced 101.0+/-10.2 ACF/rat. Disulfiram almost completely inhibited ACF development (0.60+/-0.90, 99% reduction). Dietary administration of test compounds caused significant reduction in the frequency of ACF: caffeine (70.4+/-16.6, 30% reduction), quercetin (53.0+/-8.4, 48% reduction) and morin (37. 6+/-18.1, 63% reduction). Numbers of cells positive for proliferative cell nuclear antigen in ACF and surrounding crypts were lowered by feeding of test compounds. Feeding of these test compounds also suppressed polyamine content in the colonic mucosa and blood as did disulfiram. These findings might indicate possible chemopreventive effects of caffeine, quercetin and morin, through their modulation of cell proliferation activity in crypt cells, on colon tumorigenesis.     

Chemoprevention of azoxymethane-induced rat colon carcinogenesis by the naturally occurring flavonoids, diosmin and hesperidin

The modulating effects of dietary feeding of two flavonoids, diosmin and hesperidin, both alone and in combination, during the initiation and post-initiation phases on colon carcinogenesis initiated with azoxymethane (AOM), were investigated in male F344 rats. Animals were initiated with AOM by weekly s.c. injections of 15 mg/kg body wt for 3 weeks to induced colon neoplasms. Rats were fed the diets containing diosmin (1000 ppm), hesperidin (1000 ppm) or diosmin (900 ppm) + hesperidin (100 ppm) for 5 weeks (initiation treatment) or 28 weeks (post-initiation treatment). The others contained the groups of rats treated with diosmin, hesperidin alone or in combination, and untreated. At the end of the study (32 weeks), the incidence and multiplicity of neoplasms (adenoma and adenocarcinoma) in the large intestine of rats initiated with AOM together with, or followed by, a diet containing diosmin or hesperidin were significantly smaller than those of rats given AOM alone (P <0.001). The combination regimen during the initiation and post-initiation stages also inhibited the development of colonic neoplasms, but the tumor data did not indicate any beneficial effect of diosmin and hesperidin administered together as compared with when these agents were given individually. In addition, feeding of diosmin and hesperidin, both alone and in combination, significantly inhibited the development of aberrant crypt foci. As for cell proliferation biomarkers, dietary exposure of diosmin and hesperidin significantly decreased the 5'-bromodeoxyuridine- labeling index and argyrophilic nuclear organizer region's number in crypt cells, colonic mucosal ornithine decarboxylase activity, and polyamine levels in the blood. These results indicate that diosmin and hesperidin, both alone and in combination, act as a chemopreventive agent against colon carcinogenesis, and such effects may be partly due to suppression of cell proliferation in the colonic crypts, although precise mechanisms should be clarified.      

Extract of vinegar "Kurosu" from unpolished rice inhibits the development of colonic aberrant crypt foci induced by azoxymethane

The modifying effects of administrating an ethyl acetate extract of "Kurosu" (EK), a vinegar made from unpolished rice, on development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) were investigated in male F344 rats. We also assessed the effects of EK on proliferating cell nuclear antigen (PCNA) index in ACF, prostaglandin (PG) E2 expression in the colonic mucosa and activities of detoxifying enzymes of glutathione S-transferase (GST) and quinone reductase (QR) in the liver. To induce ACF, rats were given two weekly subcutaneous injections of AOM (20 mg/kg body wt). They also received drinking water containing 0, 0.05, 0.1 or 0.2% EK for 4 weeks, starting 1 week before the first dosing of AOM. AOM exposure produced 140 +/- 23 ACF/rat at the end of the study (week 4). EK administration dose-dependently inhibited ACF formation and inhibition by 0.2% EK was statistically significant (P < 0.002). Treatment with EK significantly lowered PCNA index in ACF and reduced PGE2 content in the colonic mucosa, while EK elevated liver GST and QR activities. These findings suggest that EK may be effective for inhibiting colonic ACF, through induction of liver GST and QR and possibly alteration of PGE2 production.     

Chemoprevention of colonic aberrant crypt foci by an inducible nitric oxide synthase-selective inhibitor

Inducible nitric oxide synthase (iNOS) is overexpressed in colonic tumors of humans and also in rats treated with a colon carcinogen. iNOS appear to regulate cyclooxygenase-2 (COX-2) expression and production of proinflammatory prostaglandins, which are known to play a key role in colon tumor development. Experiments were designed to study the inhibitory effects of S,S'-1,4-phenylene-bis(1,2-ethanediyl)bis-isothiourea (PBIT) a selective iNOS-specific inhibitor, measured against formation of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF). Beginning at 5 weeks of age, male F344 rats were fed experimental diets containing 0 or 50 p.p.m. of PBIT, or 2000 p.p.m. of curcumin (non-specific iNOS inhibitor). One week later, rats were injected s.c. with AOM (15 mg/kg body wt, once weekly for 2 weeks). At 17 weeks of age, all rats were killed, colons were evaluated for ACF formation and colonic mucosa was assayed for isoforms of COX and NOS activities. Both COX and iNOS activities in colonic mucosa of the AOM-treated rats were significantly induced. Importantly, 50 p.p.m. PBIT suppressed AOM-induced colonic ACF formation to 58% (P < 0.0001) and crypt multiplicity containing four or more crypts per focus to 78% (P < 0.0001); it also suppressed AOM-induced iNOS activity. Curcumin inhibited colonic ACF formation by 45% (P < 0.001). These observations suggest that iNOS may play a key regulatory role in colon carcinogenesis. Developing iNOS-specific inhibitors may provide a selective and safe chemopreventive strategy for colon cancer treatment.     

Prevention of colonic aberrant crypt foci and modulation of large bowel microbial activity by dietary coffee fiber, inulin and pectin

The present experiments were aimed at developing novel dietary fibers to aid in reduction of colon cancer risk. We assessed the effects of coffee (non-fiber fraction), coffee fiber (arabino-galactose polymer) and inulin (oligo-fructose) in male F344 rats using formation of azoxymethane (AOM)-induced aberrant crypt foci (ACF) in the colon as the measure of preventive efficacy (or lack of such). At 5 weeks of age, groups of rats were fed the AIN-76A (control) and experimental diets that contained 1% coffee, 10% coffee fiber, 10% inulin, 10% pectin (positive control for fiber) or 200 p.p.m. piroxicam (a known ACF inhibitor). At 7 weeks of age, all animals were s.c injected with AOM (15 mg/kg body wt) once weekly for 2 weeks. All rats were killed 8 weeks after the last AOM injection and ACF were counted. The contents of the cecum were analyzed for bacterial beta-glucuronidase activity and short-chain fatty acids (SCFAs). Dietary administration of coffee fiber significantly suppressed AOM-induction of colonic ACF, in terms of total number, as well as crypt multiplicity and number of ACF/cm2 colon (P < 0.01-0.001). Inulin diet had no significant effect on total ACF, but had reduced the number of ACF/cm2 (P < 0.05). Whereas coffee had no effect on ACF formation, 10% pectin diet and 200 p.p.m. piroxicam significantly suppressed colonic ACF (P < 0.001) as had been expected. A significant reduction of cecal beta-glucuronidase activity was observed in the rats fed coffee, coffee fiber and pectin diets. Further, coffee fiber, inulin and pectin increased cecal SCFA levels 3- to 5-fold. These results suggest that coffee fiber can prevent colon cancer risk. Further studies are warranted to determine the full potential of this fiber in pre-clinical efficacy studies.