Inhibition by dietary ethanol of experimental colonic carcinogenesis induced by high-dose azoxymethane in F344 rats

Epidemiological studies have shown an association between consumption of alcoholic beverages and increased occurrence of large bowel carcinoma, but studies in experimental models of colonic carcinogenesis have produced conflicting results. We assessed the effects of chronic dietary ethanol consumption during the preinduction and induction phase (period of acclimatization and carcinogen administration) in a high-dose azoxymethane-treated rat model (14 mg/kg/wk for 10 wk). Ten-wk-old male Fischer 344 rats were given 33% of calories as ethanol or no ethanol (controls). Pair-feeding with Lieber-DeCarli-type liquid diets provided comparable total carbohydrates, proteins, fats, and calories. After 3 wk of dietary acclimatization, injections of azoxymethane (AOM) were given s.c. to all rats in Wk 1 to 10. At necropsy in Wk 25, dramatic suppression of gastrointestinal tumorigenesis was evident in the ethanol-fed group: the prevalence of colonic tumors was 5% as compared with 91% in controls; and the prevalence of small bowel tumors was 0% versus 74% (P less than 0.0001). In an analogous study of [14C]AOM metabolism, exhaled 14CO2 was decreased in the ethanol-fed rats, indicating suppression of AOM metabolism. Similarly, in the ethanol-fed rats the levels of the DNA adducts O6-methylguanine and 7-methylguanine 24 h after AOM injection were reduced in the colonic mucosa to 14 +/- 7% and 61 +/- 11% of controls and in the liver to 80 +/- 9% and 86 +/- 6 of controls. By contrast, rats changed from the ethanol diet to no-ethanol diet for 12 h prior to the dose of [14C]AOM metabolized the carcinogen at a faster rate than controls, indicating loss of suppression with cessation of ethanol intake along with induction of metabolizing enzymes; DNA adduct levels were reduced in the colonic mucosa to 90 +/- 13% and 76 +/- 9% of controls and in the liver to 81 +/- 6% and 85 +/- 3% of controls. Our findings indicate that dietary ethanol during the preinduction and induction phase of the AOM model dramatically inhibits tumorigenesis, even with high dosage of carcinogen, and suggest that: (a) inhibition of tumorigenesis may result from suppression of metabolic activation of AOM and the consequent reduced formation of DNA adducts during the induction (initiation) phase of the model; (b) these anti-initiation effects of ethanol are unrelated to the epidemiological association between consumption of alcoholic beverages and large bowel cancer; and (c) mechanisms of action of agents found to modulate carcinogenesis in experimental models should be determined before the results can be generalized to human beings.     

Effects of chronic dietary beer and ethanol consumption on experimental colonic carcinogenesis by azoxymethane in rats

Epidemiological studies have shown an association between consumption of alcoholic beverages, particularly beer, and carcinoma of the large bowel, especially the rectum. We studied the effects of chronic dietary beer and ethanol consumption on experimental colonic carcinogenesis, fecal bile acid and neutral sterol levels, fecal bacterial flora, and colonic epithelial DNA synthesis. Ten-week-old male Fischer 344 rats were pair fed throughout the study with Lieber-DeCarli-type liquid diets providing comparable total carbohydrates, proteins, fats, and calories. The diets provided 23 or 12% of calories as alcohol in beer (Hi-Beer and Lo-Beer groups), 18 or 9% of calories as reagent ethanol (Hi-EtOH and Lo-EtOH groups), or no alcohol (control group). After 3 weeks of dietary acclimatization, 10 weekly s.c. injections of the bowel carcinogen azoxymethane, 7 mg/kg, were given (weeks 1-10). At necropsy in week 26, the high alcohol groups (Hi-Beer and Hi-EtOH) showed a significantly reduced incidence of tumors in the right colon (42 and 46% versus 81% in control, P less than 0.01 and P = 0.02) but no effect on left colonic tumorigenesis. By contrast, the low alcohol groups (Lo-Beer and Lo-EtOH) showed a trend toward increased incidence and proportion of tumors in the left colon (incidence of 42 and 35% versus 15% in control, P = 0.06 for Lo-Beer; 28 and 30% of tumors in left colon versus 11%, P = 0.08 and P = 0.07) but no effect on right colonic tumorigenesis. Numbers of right colonic tumors were inversely correlated with alcohol consumption of all rats (r = -0.350, P less than 0.001), but left colonic tumors were not correlated. Fecal bile acid and neutral sterol levels, fecal bacterial counts, and colonic epithelial DNA synthesis did not correlate with the effects of alcohol consumption on colonic tumorigenesis. Our findings suggest that: modulation of experimental colonic tumorigenesis by chronic dietary beer and ethanol consumption was due to alcohol rather than other beverage constituents; tumorigenesis in the right and left colon was affected differentially by the levels of alcohol consumption, reflecting complex interactions among the potential mechanisms for alcohol effects in the model used.     

Inhibition of chemically induced mammary and colon tumor promotion by caloric restriction in rats fed increased dietary fat

Tumor promotion associated with increased dietary fat may be inhibited by reduction in total caloric intake. This hypothesis was tested in rats given either 7,12-dimethylbenz(a)anthracene to induce mammary tumors or 1,2-dimethylhydrazine to induce colon tumors. One week after dosage with either carcinogen, the rats were fed semipurified diets that provided 4% fat with ad libitum calories or 13.1% fat with a reduction of calories by 40% from ad libitum intake. Rats treated with 7,12-dimethylbenz(a)anthracene and subjected to caloric restriction weighed 40% less than those fed ad libitum; rats treated with 1,2-dimethylhydrazine were heavier at the onset of caloric restriction and lost weight and weighed approximately 40% less than animals fed ad libitum. At 20 weeks after 7,12-dimethylbenz(a)anthracene administration, rats fed ad libitum had 80% tumor incidence while in those fed restricted calories, 20% had tumors (P less than 0.001). All other measures of mammary tumor growth were significantly reduced in rats given restricted calories. Six months after 1,2-dimethylhydrazine administration, colon tumor incidence was 100% in rats fed ad libitum and 53% in those fed the calorie-restricted diet (P less than 0.001). This reduction of colonic carcinogenesis was seen despite a significant increase in mucosal labeling index following [3H]thymidine autoradiography. This paradoxical finding may be due to the increased fat content of the calorie-restricted diet. These data demonstrate that the tumor-promoting effects of dietary fat can be more than offset by a reduction in total caloric intake and that the promoting effect of fat may be due, at least in part, to its greater caloric density.     

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.     

Pomegranate seed oil rich in conjugated linolenic acid suppresses chemically induced colon carcinogenesis in rats

Pomegranate (Punica granatum L.) seed oil (PGO) contains more than 70% cis(c)9,trans(t)11,c13-18:3 as conjugated linolenic acids (CLN). Our previous short-term experiment demonstrated that seed oil from bitter melon (Momordica charantia) (BMO), which is rich in c9,t11,t13-CLN, inhibited the occurrence of colonic aberrant crypt foci (ACF) induced by azoxymethane (AOM). In this study, we investigated the effect of dietary PGO on the development of AOM-induced colonic malignancies and compared it with that of conjugated linoleic acid (CLA). To induce colonic tumors, 6-week old male F344 rats were given subcutaneous injections of AOM (20 mg/kg body weight) once a week for 2 weeks. One week before the AOM treatment they were started on diet containing 0.01%, 0.1%, or 1% PGO or 1% CLA for 32 weeks. Upon termination of the bioassay (32 weeks) colon tumors were evaluated histopathologically. AOM exposure produced colonic adenocarcinoma with an incidence of 81% and multiplicity of 1.88 +/- 1.54 at week 32. Administration of PGO in the diet significantly inhibited the incidence (AOM + 0.01% PGO, 44%, P < 0.05; AOM + 0.1% PGO, 38%, P < 0.01; AOM + 1% PGO, 56%) and the multiplicity (AOM + 0.01% PGO, 0.56 +/- 0.73, P < 0.01; AOM + 0.1% PGO, 0.50 +/- 0.73, P < 0.005; AOM + 1% PGO, 0.88 +/- 0.96, P < 0.05) of colonic adenocarcinomas, although a clear dose-response relationship was not observed at these dose levels. CLA feeding also slightly, but not significantly, reduced the incidence and multiplicity of colonic adenocarcinomas. The inhibition of colonic tumors by PGO was associated with an increased content of CLA (c9,t11-18:2) in the lipid fraction of colonic mucosa and liver. Also, administration of PGO in the diet elevated expression of peroxisome proliferator-activated receptor (PPAR) gamma protein in the non-tumor mucosa. These results suggest that PGO rich in c9,t11,c13-CLN can suppress AOM-induced colon carcinogenesis, and the inhibition is associated in part with the increased content of CLA in the colon and liver and/or increased expression of PPARgamma protein in the colon mucosa.     

A rapid dual organ rat carcinogenesis bioassay for evaluating the chemoprevention of breast and colon cancer

In this study we evaluated the effect of dietary administration of a high fat, low fiber diet (HRD) with or without 2% phytic acid (PA) on the development of mammary cancer and/or colon cancer in rats exposed to methylnitrosourea (MNU), azoxymethane (AOM) or MNU + AOM. The rats were fed a HRD alone or a HRD + 2% PA. At the end of week 2, the rats were given either a s.c. injection of MNU (50 mg/kg body wt) or one of normal saline (vehicle). At the end of weeks 3 and 4, the rats were given either a s.c. injection of AOM (15 mg/kg body wt per week) or one of normal saline (vehicle). Nine weeks after the injection of MNU or saline, 10 rats from each group were sacrificed and the mammary tumor incidence and the number of colonic aberrant crypt foci (ACF) were compared between different groups. The administration of different diets was continued for an additional 21 weeks and the mammary tumor and colon tumor incidence between different groups were compared. Results showed that rats injected with MNU alone did not develop ACF or colon tumors while those injected with AOM alone did not develop mammary tumors. Linear regression analysis of the number of ACF at 11 weeks versus colonic tumor incidence at 32 weeks, and the linear regression analysis of mammary tumor incidence at 11 weeks versus mammary tumor incidence at 32 weeks, both showed good linear correlation. These results demonstrate the potential value of the short term dual organ carcinogenesis bioassay for screening chemopreventive agents for their relative ability to inhibit the development of mammary cancer and/or colon cancer while on high risk diet.     

Effect of different levels of calorie restriction on azoxymethane-induced colon carcinogenesis in male F344 rats

Epidemiological and animal model studies indicate that increased calorie intake increases the risk for colon cancer development. Previous studies in animal models restricted the calorie intake severely, and none of these studies have investigated a dose-response effect of different levels of calorie restriction on colon carcinogenesis. The present study was designed to investigate the effect of various levels of calorie restriction on colon carcinogenesis in male F344 rats fed the low and high fat diets and the effect of these diets on the activities of colonic mucosal and tumor ornithine decarboxylase (ODC) and protein tyrosine kinase. Starting at 5 weeks of age, groups of male F344 rats were fed the low fat or high fat diets ad libitum. At 7 weeks of age, all animals except the vehicle-treated groups were given s.c. injections of azoxymethane (AOM) (15 mg/kg body weight, once weekly for 2 weeks). Four days after the second injection, groups of animals were restricted to 90, 80, or 70% of total calories consumed by the high fat ad libitum group (i.e., 10, 20, and 30% calorie restriction, respectively). In the low fat groups, animals were restricted to 80% of total calories consumed by the low fat ad libitum group (i.e., 20% restriction). Thirty-six weeks after AOM injections, all animals were necropsied and colon tumors were used for histopathology and ODC and protein tyrosine kinase analysis. In the second experiment, the protocol was the same as above except that the animals were sacrificed 5 days after the second AOM injection and colonic mucosal ODC and protein tyrosine kinase activities were assayed. The incidence and multiplicity of colon tumors were significantly inhibited in animals fed the high fat 20% calorie-restricted and high fat 30% calorie-restricted diets, as compared to those fed the high fat ad libitum diet. The regression coefficient representing the dose-response effect of different levels of calorie restriction in both high fat groups is significant. Results also indicate that AOM treatment significantly increased the colonic mucosal ODC and protein tyrosine kinase activities. This stimulation was inhibited by feeding the calorie-restricted diets. ODC and protein tyrosine kinase activities were lower in the colon tumors of animals fed the calorie-restricted diets.     

Effect of different levels of dietary corn oil and lard during the initiation phase of colon carcinogenesis in F344 rats

The effect of various levels of polyunsaturated fat (corn oil) and saturated fat (lard) fed during the initiation stage of colon carcinogenesis was studied in male F344 rats. The animals were fed the diets containing 5, 13.6, and 23.5% corn oil or lard 2 weeks before, during, and until 1 week after sc injection of 15 mg azoxymethane [(AOM) CAS: 25843-45-2]/kg body weight, once weekly for 2 weeks (designated as initiation). One week after AOM treatment, groups of animals were transferred to their respective 5% corn oil or lard diets. Additional groups consuming 5% corn oil or lard were transferred to 23.5% corn oil or lard, respectively (post-initiation stage). All animals were fed these diets until the termination of the experiment. Fecal bile acids and colonic mucosal ornithine decarboxylase activity were measured in vehicle-treated animals fed the experimental diets for 14 weeks. Body weights and intakes of total calories, protein, nonnutritive fiber, and micronutrients were comparable among the various dietary groups. The animals fed the 23.5% corn oil diet during the postinitiation stage had a higher incidence of colon tumors than did those fed the 5% corn oil diet, whereas feeding of 23.5 and 13.6% corn oil diets during the initiation stage had no effect. In contrast, animals fed the 23.5 and 13.6% lard diet during the initiation stage and 23.5% lard diet during the postinitiation stage developed more colon adenocarcinomas than did those fed the 5% lard diet. The excretion of fecal deoxycholic acid, lithocholic acid, and 12-ketolithocholic acid and the activity of colonic mucosal ornithine decarboxylase activity were higher in animals fed the 23.5% corn oil or lard diet during the postinitiation compared to the levels in animals fed the 5% corn oil or lard diet.     

Effect of restricted caloric intake on azoxymethane-induced colon tumor incidence in male F344 rats

The effect of 30% caloric restriction on azoxymethane (AOM)-induced colon carcinogenesis was investigated in male F344 rats. Starting at 5 weeks of age, groups of animals were fed ad libitum a high-fat (23.5%) semipurified diet. At 7 weeks of age, all animals except the vehicle-treated groups were s.c. injected with AOM (15 mg/kg body wt, once weekly for 2 weeks). Four days after the second AOM injection, groups of animals were continued on high-fat diet and fed ad libitum (ad libitum group) whereas other groups were restricted to 70% of total calories (calorie-restricted group) consumed by the ad libitum group, but received same amounts of fiber, vitamins, and minerals. Thirty-two weeks after AOM injections, all animals were necropsied. The animals in the calorie-restricted group developed significantly fewer colon tumors and had a lower colon tumor incidence than did the rats in the ad libitum group. The size of colon tumors was also reduced in the calorie-restricted group.     

Mechanisms in the chemoprevention of colon cancer: modulation of protein kinase C, tyrosine protein kinase and diacylglycerol kinase activities by 1,4-phenylenebis-(methylene)selenocyanate and impact of low-fat diet

Epidemiological and experimental studies suggest an inverse relationship between the intake of dietary selenium and/or low fat-intake and colon cancer risk. Efficacy studies in rodents suggest that the organoselenium compound 1, 4-phenylenebis(methylene)selenocyanate (p-XSC), is a more effective and less toxic chemopreventive agent than other organic or inorganic selenium compounds such as selenomethionine and Na2SeO3. The efficacy of p-XSC against colon cancer is significantly augmented by a low-fat diet. To explore the mechanisms by which this combined inhibiting effect against colon carcinogenesis comes about, we have investigated protein kinase C (PKC), tyrosine protein kinase (TPK), diacylglycerol kinase (DGK) activities and 8-isoprostane levels in colonic mucosa and tumor tissues in an azoxymethane (AOM)-induced rat colon cancer model. Weanling male F344 rats were fed the semipurified AIN-76A diet until seven weeks of age. Then various experimental groups were fed the low- or high-fat diets containing 0 or 20 ppm p-XSC (10 ppm as selenium). At seven weeks of age, groups of rats were injected s.c. with azoxymethane (AOM; 15 mg/kg body wt., once weekly for 2 weeks) and continued on their respective experimental diets until 38 weeks after the second AOM treatment. They were then sacrificed and colonic mucosal and tumor samples were evaluated for PKC, TPK, DGK and 8-isoprostane levels. Administration of p-XSC along with a low-fat diet significantly inhibited Ca+2-dependent and -independent PKC (P<0.05-0.01) activities in colonic mucosa and tumors. Administration of p-XSC either low-fat or high-fat diet significantly suppressed both colonic mucosal and tumor TPK activity (P<0.05-0.01). Suppression of TPK activity was more pronounced in rats maintained on a low-fat diet containing p-XSC. In contrast, rats receiving p-XSC with either low- or high fat diet showed significantly increased DGK activity (P<0.01-0.0001). Rats fed low-fat or high-fat plus p-XSC had lower-levels of 8-isoprostane in the colonic tumors than animals who had been given low- or high-fat diets without the organoselenium compound. Interestingly, 8-isoprostane levels were lower in the colon tumors of the rats fed the low-fat diet than those fed the high-fat diet. Our findings suggest that p-XSC induced down-regulation of PKC and TPK activities and up-regulation of DGK activity. These events may in part be responsible for the chemopreventive activity against colon carcinogenesis. Further, this study implies that p-XSC with a low-fat dietary regimen will augment regulation of PKC, TPK and DGK activities in the colon.     

Modifying effects of Terminalia catappa on azoxymethane-induced colon carcinogenesis in male F344 rats.

The modifying effects of dietary administration of an herb, Terminalia catappa (TC), were investigated on rat colon carcinogenesis induced by a carcinogen azoxymethane (AOM). The number of aberrant crypt foci (ACF) and beta-catenin accumulated crypts (BCACs) in the colon, and proliferating cell nuclear antigen (PCNA) labelling index in the colonic epithelium were examined in a total of 36 male F344 rats. All animals were randomly divided into five experimental groups (4-10 rats in each group). At 6 weeks of age, rats in groups 1, 2 and 3 were given s.c. injections of AOM once a week for 2 weeks at a concentration of 20 mg/kg body weight. One week before the first injection of AOM, rats in groups 2 and 3 were fed a diet containing 0.02 and 0.1% TC, respectively, throughout the experiment. Rats in group 4 were fed a diet containing 0.1% TC. Rats in group 5 were served as untreated controls. All animals were sacrificed at the experimental week 5 after the start of the experiment. Oral administration of TC at both doses significantly decreased the numbers of both ACF/colon/rat (P<0.05 for 0.02% TC, P<0.005 for 0.1% TC) and BCAC/cm/rat (P<0.05 for both 0.02 and 0.1% TC), when compared with the control group (group 1). Colonic PCNA labelling index in groups 2 and 3 was also significantly lower than that in group 1 (P<0.001 for 0.02% TC, P<0.005 for 0.1% TC). These results suggest that TC has a potent short-term chemopreventive effect on biomarkers of colon carcinogenesis and this effect may be associated with the inhibition of the development of ACF and BCACs.     

Catalpa seed oil rich in 9t,11t,13c-conjugated linolenic acid suppresses the development of colonic aberrant crypt foci induced by azoxymethane in rats

Catalpa (Catalpa ovata) seed oil (CPO) is a unique oil that contains a high amount of 9trans,11trans,13cis-conjugated linolenic acid. In the present study, we investigated whether dietary administration with CPO affects the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) in male F344 rats to elucidate its possible cancer chemopreventive efficiency. Also, the effect of CPO on the fatty acid composition of liver tissue and colonic mucosa, the serum levels of total cholesterol and triglyceride, and the mRNA expression of cyclooxygenase (COX)-2 in the colonic mucosa were measured. In addition, the cell proliferation activity and apoptotic index in the colonic mucosa were estimated immunohistochemically. Animals were given two weekly subcutaneous injections of AOM (20 mg/kg body weight). They also received the experimental diet containing 0.01%, 0.1% or 1% CPO for 4 weeks, starting one week before the first dosing of AOM. AOM exposure produced a substantial number of ACF (99+/-28) at the end of the study (week 4). Dietary administration of CPO reduced the number of ACF (AOM + 0.01% CPO, 32+/-11, P<0.001; AOM + 0.1% CPO, 35+/-18, P<0.001; AOM + 1% CPO, 18+/-10, P<0.001). 9t,11t-conjugated linoleic acid was detected in the liver tissue and colonic mucosa of rats fed the CPO-containing diet. Additionally, dietary administration with CPO decreased the serum triglyceride level and the expression of COX-2 mRNA in the colonic mucosa. The indices of cell proliferation and apoptosis in the colonic mucosa of rats treated with AOM and 1% CPO have significant differences when compared with the AOM alone group. These findings suggest the possible chemopreventive activity of CPO in the early phase of colon carcinogenesis.     

Modifying effect of tuna orbital oil rich in docosahexaenoic acid and vitamin D3 on azoxymethane-induced colonic aberrant crypt foci in rats

The modifying effect of dietary tuna (Thunnus thynnus orientalis) orbital oil rich in docosahexaenoic acid (DHA) and vitamin D3 (VD3) on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) was investigated in male F344 rats. Animals were given three weekly subcutaneous injections of AOM (15 mg/kg body weight) to induce ACF. The rats were fed the experimental diet containing 5% tuna orbital oil (low fish oil), 23.5% tuna orbital oil (high fish oil), 5% corn oil (low corn oil) or 23.5% corn oil (high corn oil) for 5 weeks, starting 1 week before the first dose of AOM. Animals were sacrificed 2 weeks after the last AOM injection to count colonic ACF and assay the expression of cyclooxygenase (COX)-1 and -2. High corn oil diet significantly increased the development of ACF, when compared with low corn oil diet (P<0.005). High fish oil diet also increased ACF formation compared with low fish oil diet (P<0.01), but the increase was smaller than high corn oil diet. The frequency of ACF was significantly lower in the rats fed high fish oil diet than high corn oil diet (P<0.02). Moreover, frequency of ACF consisted of 4 or more crypts in rats fed the high fish oil diet was significantly lower than that of rats given high corn oil diet. COX-1 and COX-2 expression did not significantly differ among the groups. These results suggest that fish oil derived from tuna, which contains high amounts of DHA and VD3, suppresses the formation and growth of ACF without affecting COX-1 and COX-2 expression, and may have a preventive effect on colon carcinogenesis.     

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 seed oil rich in conjugated linolenic acid from bitter melon inhibits azoxymethane-induced rat colon carcinogenesis through elevation of colonic PPARgamma expression and alteration of lipid composition

Our previous short-term experiment demonstrated that seed oil from bitter melon (Momordica charantia) (BMO), which is rich in cis(c)9, trans(t)11, t13-conjugated linolenic acid (CLN), inhibited the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF). In our study, the possible inhibitory effect of dietary administration of BMO on the development of colonic neoplasms was investigated using an animal colon carcinogenesis model initiated with a colon carcinogen AOM. Male F344 rats were given subcutaneous injections of AOM (20 mg/kg body weight) once a week for 2 weeks to induce colon neoplasms. They also received diets containing 0.01%, 0.1% or 1% BMO for 32 weeks, starting 1 week before the first dosing of AOM. At the termination of the study (32 weeks), AOM induced 83% incidence (15/18 rats) of colonic adenocarcinoma. Dietary supplementation with 0.01% and 0.1% BMO caused significant reduction in the incidence (47% inhibition by 0.01% BMO, p<0.02; 40% inhibition by 0.1% BMO, p<0.05; and 17% inhibition by 1% BMO) and the multiplicity (64% inhibition by 0.01% BMO, p<0.005; 58% inhibition by 0.1% BMO, p<0.02; and 48% inhibition by 1% BMO, p<0.05) of colonic adenocarcinoma, though a clear dose response was not observed. Such inhibition was associated with the increased content of CLA (c9,t11-18:2) in the lipid composition in colonic mucosa and liver. Also, BMO administration in diet enhanced expression of peroxisome proliferator-activated receptor (PPAR) gamma protein in the nonlesional colonic mucosa. These findings suggest that BMO rich in CLN can suppress AOM-induced colon carcinogenesis and the inhibition might be caused, in part, by modification of lipid composition in the colon and liver and/or increased expression of PPARgamma protein level in the colon mucosa.     

Chemopreventive efficacy of sulindac sulfone against colon cancer depends on time of administration during carcinogenic process.

Epidemiological and model studies with laboratory animals have provided evidence that nonsteroidal anti-inflammatory drugs reduce the risk of colon cancer. Sulindac, a nonsteroidal anti-inflammatory drug, has been shown to inhibit azoxymethane (AOM)-induced colon carcinogenesis in rats when administered continuously before, during, and after carcinogen treatment (initiation and postinitiation periods) or when given continuously beginning 14 weeks after carcinogen administration (promotion/ progression stage). The present study was designed to investigate the chemopreventive efficacy of sulindac sulfone (exisulind), the sulfone metabolite of sulindac, when administered during the promotion/progression stage of colon carcinogenesis in comparison to the effect during the initiation and postinitiation periods. We have also studied the modulating effect of exisulind on colonic tumor apoptosis. At 5 weeks of age, groups of male F344 rats were fed diets containing 0%, 0.06%, and 0.12% exisulind. At 7 weeks of age, groups of animals were injected s.c. with AOM (15 mg/kg body weight, once weekly for 2 weeks). Animals intended for the promotion/progression study and receiving 0% exisulind were switched to an experimental diet containing 0.12% exisulind at 14 weeks after the second AOM treatment. All rats remained on their respective dietary regimens until the termination of the study, 50 weeks after the second AOM injection. Colon tumors were evaluated histopathologically for tumor type. Administration of 0.06% and 0.12% exisulind during the initiation and postinitiation periods significantly inhibited the incidence and multiplicity of invasive and/or noninvasive adenocarcinomas of the colon. The inhibition of colon tumorigenesis by exisulind was associated with a significant retardation of body weight gain shortly after sulfone administration and increased apoptosis in the colon tumors. In contrast, administration of the higher dose (0.12%) of exisulind during the promotion/progression stage had only minimal effects on colon tumorigenesis and apoptosis in the colon tumors, suggesting that early administration, but not late administration, may be required for chemopreventive efficacy of this drug.     

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.     

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.      

Effects of androgen manipulations on chemically induced colonic tumours and on macroscopically normal colonic mucosa in male Sprague-Dawley rats

Epidemiological and experimental studies suggest that androgens influence colonic carcinogenesis. We investigated the effects of hormonal manipulations (surgical and chemical castration, hormone substitution) on colonic tumour development, tumour and mucosal histopathology, and epithelial proliferation in macroscopically normal colonic mucosa in male rats, after induction of chemical colon carcinogenesis by subcutaneous injections of azoxymethane (AOM). Chemical castration with cyproterone acetate, but not surgical castration, resulted in increased colonic tumorigenesis, which was accompanied by decreased crypt length, decreased number of cells per crypt, and increased crypt epithelial mitotic index in the right colon. Chemically castrated rats also had crypt hyperplasia and increased numbers of dysplastic foci in the left colon which were not seen with surgical castration. By contrast, rats given testosterone after surgical castration showed decreased colonic tumorigenesis with an increased proportion of tumours in the left colon and lower percentage of tumours with invasion. The grossly normal mucosa of the testosterone-substituted castrated rats showed decreased crypt length in the right colon similar to the other groups of castrated rats, but no significant increase in mitotic index. Our results suggest that the anti-androgenic progestin cyproterone is a potent enhancer of colonic tumorigenesis and epithelial proliferative abnormalities after AOM administration. Exogenous testosterone after castration alters tumour distribution and characteristics and suppresses epithelial proliferative abnormalities. Finally, androgen effects on the colonic mucosa are more prominent in the right than in the left colon, suggesting different influences of hormones on the epithelium of these anatomical sites.     

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.