Inhibitory effect of peptide Epitalon on colon carcinogenesis induced by 1,2-dimethylhydrazine in rats

The effect of synthetic pineal peptide Epitalon (Ala-Glu-Asp-Gly) on colon carcinogenesis was firstly studied in rats. Eighty 2-month-old outbred male LIO rats were subdivided into four groups and were weekly exposed to five subcutaneous injections of 1,2-dimethylhydrazine (DMH) at a single dose of 21 mg/kg body weight. Additionally, 5 days a week, some of the rats were given subcutaneous injections of saline at a dose of 0.1 ml during the whole experiment (group 1, control) or Epitalon at a single dose of 1 microg during the whole experiment (group 2), Epitalon after termination of carcinogen injections (group 3) or during the period of DMH exposure (group 4). Colon carcinomas developed in 90-100% of DMH-treated rats. The number of total colon tumors per rat was 4.1; 2.7; 3.7; 2.9 in groups 1, 2, 3, 4, respectively (the difference in groups 2 and 4 compared with group 1 is significant). In rats from group 2, colon tumors were smaller than in control animals. In group 2, the incidence, as well the multiplicity of tumors in ascending and descending colon, were significantly decreased in comparison with group 1. In group 4, the mean number of tumors per rat was significantly decreased, too. A trend to decrease the number of tumors in the rectum in rats from groups 2, 3 and 4, treated with Epitalon was found. Epitalon inhibited also the development of tumors in jejunum and ileum. Thus, our results demonstrated an inhibitory effect of Epitalon on chemically induced bowel carcinogenesis in rats.     

Inhibitory effect of synthetic interferon inductor Cycloferone on 1,2-dimethylhydrazine-induced colon carcinogenesis in rats

The effect of a synthetic interferon inductor Cycloferone on colon carcinogenesis was firstly studied in rats. Seventy-five 2-month-old outbred female LIO rats were subdivided into three groups and were weekly exposed to 15 s.c. injections of 1,2-dimethylhydrazine (DMH) at a single dose of 7 mg/kg body wt. From the day of the fist injection of DMH rats from group 2 were given weekly i.p. injections of Cycloferone (62.5 mg/kg) until the end of the experiment. DMH-treated rats (group 3) were exposed to weekly i.p. injections of Cycloferone (62.5 mg/kg) starting in the week after the last injection of the carcinogen. Rats from group 1 were exposed to DMH and treated weekly with 0.2 ml i.p. of normal saline. Additional groups of rats were treated weekly with Cycloferone (62.5 mg/kg) or with 0.2 ml of saline. The experiment was ended 6 months after the first injection of DMH. In DMH-treated rats (groups 1, 2 and 3) colon adenocarcinomas developed in 87, 61 and 59%, respectively. The number of colon tumors per tumor-bearing rat was 2.5, 1.9 and 1.3 in groups 1, 2 and 3, respectively. Treatment with Cycloferone significantly inhibits carcinogenesis in ascending and descending colon. The incidence of tumors of the rectum was decreased in the group 2 as compared with the group 1. There were no cases of tumors of rectum in rats from group 3. The treatment with Cycloferone alone as well as with normal saline failed to induce any tumors in rats. Thus, our results demonstrated inhibitory effect of Cycloferone on colon carcinogenesis induced by DMH in rats.     

Effect of Konjac mannan on 1,2-dimethylhydrazine-induced intestinal carcinogenesis in Fischer 344 rats

The effect of Konjac mannan (KM) on 1,2-dimethylhydrazine (DMH-induced intestinal carcinogenesis was studied in male F344 rats. Rats were fed a diet containing 5% KM at 5 weeks of age. At 6 weeks of age, all animals were given a weekly intraperitoneal injection of 20 mg DMH/kg body wt for 13 weeks and autopsied 13 weeks after the last injection of DMH. The weight gain was lower in rats fed the KM diet than in rats fed the control diet throughout the experiment (P less than 0.05). The incidence of DMH-induced colon tumors was lower in animals fed the KM diet compared to animals fed the control diet (P less than 0.05). The number of colon adenocarcinoma per animal was also lower in animals fed the KM than in animals fed the control diet (P less than 0.05). However, the incidence of tumors of the small intestine did not significantly differ between the groups fed the KM and control diets. The present study demonstrated that colon tumorigenesis induced by DMH in F344 rat was inhibited by maintaining the KM diet.     

Promoting and synergistic effects of chrysazin on 1,2-dimethylhydrazine-induced carcinogenesis in male ICR/CD-1 mice

The modifying effects of chrysazin on 1,2-dimethylhydrazine(DMH)-indueed colon and liver carcinogenesis were examined inmate ICR/CD-1 mice. Starting at 6 weeks of age, mice were dividedinto four groups, two of which were treated with s.c. injectionsof DMH (20 mg/kg body wt) once a week for 12 weeks. A week afterthe final injection of DMH, one group was kept on the basaldiet throughout the study (group I), and the other group wasfed the diet containing chrysazin (mixed in basal diet at 0.2%concentration) alone for 42 weeks (group II). The other twogroups were injected with normal saline and given the diet containing0.2% chrysazin for 42 weeks (group III), or the basal diet duringthe experiment (group TV). The incidence and multiplicity ofcolon tumors of group II were significantly greater than thoseof group I (P<0.05, P<0.01). The incidence and multiplicityof the hepatocellular neoplasms of group II were larger thanthose of group I (P< 0.002, P< 0.02 respectively). Ingroup III, colon tumors were not found, though a few liver neoplasmsand severe inflammatory lesions of the colon were observed.The activity of ornithine decarboxylase of the colonk mucosain mice exposed to chrysazin was stronger than that of animalswithout chrysazin. The results suggest that the promoting effectof chrysazin is probably related to an increase of cell proliferationin the target organ. A synergistic effect of DMH with chrysazinwas also observed in liver tumorigenesis.     

Carcinogenicity of 1,2-DimethyIhydrazine in Colorectal Tissue Heterotopically Transplanted into the Glandular Stomach of Rats

The present study was designed to examine the effect of the intestinal carcinogen 1,2-dimethyl-hydrazine (DMH) on grafted colorectal mucosa implanted into the glandular stomach of rats. Four groups were studied: Group 1 received the operation and DMH, Group 2 received the operation alone, Group 3 received DMH alone and Group 4 (controls) received only a sham operation. For Groups 1 and 2, about 8-mm diameter segments of colorectal tissue obtained from various sites in the large intestine of 8-week-old male F344 rats were isologously implanted into the fundic region of the stomachs of age-matched rats. DMH was injected at a dose of 20 mg/kg body weight i.m. per week for 20 weeks beginning 4 weeks after the operation. The animals were then observed for 8 months after the initial DMH treatment. In Group 1, adenocarcinomas developed in 41 of 60 successful implants (68%). Furthermore, poorly differentiated type tumors were observed in the grafts obtained from the rectum. This finding was contrary to that for intrinsic rectal tumors, all of which were well differentiated. The histochemical staining of mucin in the tissues from different sites of the large intestine revealed that sulfomucin, which normally exists essentially only in the intrinsic descending colon or rectum, was present in the grafts from the proximal ascending or ascending colon. No gastric tumors were observed in the control rats, which received either DMH or sham operations alone. Tumors in the intrinsic large intestine were observed only in rats that received DMH. These results indicate that colorectal mucosa implanted into the glandular stomach, like the intrinsic large intestine, is still sensitive to tumorigenesis caused by DMH.     

Carcinogenicity of 1,2-dimethylhydrazine in colorectal tissue heterotopically transplanted into the glandular stomach of rats.

The present study was designed to examine the effect of the intestinal carcinogen 1,2-dimethylhydrazine (DMH) on grafted colorectal mucosa implanted into the glandular stomach of rats. Four groups were studied: Group 1 received the operation and DMH, Group 2 received the operation alone, Group 3 received DMH alone and Group 4 (controls) received only a sham operation. For Groups 1 and 2, about 8-mm diameter segments of colorectal tissue obtained from various sites in the large intestine of 8-week-old male F344 rats were isologously implanted into the fundic region of the stomachs of age-matched rats. DMH was injected at a dose of 20 mg/kg body weight i.m. per week for 20 weeks beginning 4 weeks after the operation. The animals were then observed for 8 months after the initial DMH treatment. In Group 1, adenocarcinomas developed in 41 of 60 successful implants (68%). Furthermore, poorly differentiated type tumors were observed in the grafts obtained from the rectum. This finding was contrary to that for intrinsic rectal tumors, all of which were well differentiated. The histochemical staining of mucin in the tissues from different sites of the large intestine revealed that sulfomucin, which normally exists essentially only in the intrinsic descending colon or rectum, was present in the grafts from the proximal ascending or ascending colon. No gastric tumors were observed in the control rats, which received either DMH or sham operations alone. Tumors in the intrinsic large intestine were observed only in rats that received DMH. These results indicate that colorectal mucosa implanted into the glandular stomach, like the intrinsic large intestine, is still sensitive to tumorigenesis caused by DMH.     

Calcium inhibits colon carcinogenesis in an experimental model in the rat

Different dietary factors can affect colorectal cancer incidence. However, the effect of increased levels of dietary calcium on neoplasms is unclear. The present study was designed to examine the effect of a low calcium supplement on experimental colon carcinogenesis induced by parenteral administration of dimethylhydrazine (DMH). One hundred and twenty 10-week-old Sprague-Dawley rats were divided into five groups of equal sex distribution. The 10 rats in group A (control group) received no treatment; the 30 rats in group B (DMH group) were injected subcutaneously with 18 weekly doses of 21 mg/kg DMH; the 20 rats in group C (EDTA control group) received EDTA solution only; the 30 rats in group D (calcium group) received calcium at 3.2 g/l by adding calcium lactate to the drinking water from the start until the conclusion of the experiment; and the 30 rats in group E (DMH + calcium group) received oral calcium supplements at the same dose as the rats in group D (calcium group) and the same DMH injections as the rats in group B (DMH group). The rats were sacrificed at 25-34 weeks. In group E, we observed a significant diminution in the number of tumours (P = 0.01); an increase in the number of tumour-free animals (P = 0.006); a change in tumour location towards the distal colon (P < 0.025); more adenomas (P = 0.02); and a diminution of adenocarcinomas and mucinous carcinomas, although this was not significant. We conclude that a low dietary calcium supplement in rats inhibits colon cancer carcinogenesis induced by DMH, and changes tumour location towards the distal colon.     

Long term administration of granulocyte-macrophage colony stimulating factor decreases development of 1-2 dimethylhydrazine-induced colon cancer in rats

BACKGROUND AND OBJECTIVES: The antitumoral activities of granulocyte-macrophage colony stimulating factor (GM-CSF) were shown earlier. In this study, the effects of GM-CSF were investigated on colon cancer induced by 18 weeks of 1-2 dimethylhydrazine (DMH) administration in rats. METHODS: Four groups received subcutaneous saline (n = 20), 15 mg/kg DMH (n = 30), DMH +6 microg/kg GM-CSF (n = 30), and DMH +12 microg/kg (n = 30) GM-CSF. RESULTS: The average number of tumors (2.8 vs. 1.5) and mean tumor volume (179 +/- 36 vs. 27 +/- 9 mm(3); means +/- SEM) were reduced in DMH + GM-CSF groups as compared to the DMH group (n = 30, P < 0.01). DMH-induced enhancement of free radicals and lipid peroxidation were decreased in DMH + GM-CSF group (n = 8-12, P < 0.05). The magnitude of DMH-induced alterations in superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities was lowered in the DMH + GM-CSF group (n = 12-16, P < 0.05). DMH-induced increases in the total nitrite/nitrate levels and the nitric oxide synthase (NOS) activity (n = 10-12, P < 0.05) were also reduced in the DMH + GM-CSF group (n = 8-9, P < 0.05). CONCLUSIONS: The results indicate that GM-CSF inhibits the development of DMH-induced colon cancer in rats and suggest that inhibition of oxidative stress and NO pathway are involved in the observed antitumoral effects.     

Modification by five antioxidants of 1,2-dimethylhydrazine-initiated colon carcinogenesis in F344 rats

The effects of antioxidants given in the post initiation phaseof colon tumor development were investigated in male F344 ratstreated with 1 ,2-dimethylhydrazine (DMH). Animals (20/group)were given s.c. injections of DMH at a dose of 20 mg/kg oncea week for four consecutive weeks. One week after the last injection,rats were fed diet containing 5% sodium L-asorbate (SA), 0.5%butylated hydroxyanisole (BHA), 0.8% ethoxyquin (EQ), 1.0% propylgallate or 0.5% butylated hydroxytoluene (BHT) for 36 weeks.A control group was fed the basal diet not containing antioxidants.The experiment was terminated 40 weeks after the first injectionof DMH and all intestinal tumors were confirmed histologically.SA significantly increased the incidence of adenomas and thenumber of tumors per rat of the colon (especially of the distalcolon). Although EQ and BHT did not affect the number of ratswith colon tumors, the number of tumors per rat occurring inthe distal colon was significantly increased by EQ while beingdecreased by BHT. No modification of tumor development was observedwith BHA or PG. Thus, modification of tumor development by SA,EQ and BHT was apparent, mainly in the distal colon.     

The effect of endotoxin on 1,2-dimethylhydrazine-induced colonic tumors in rats

The effect of endotoxin on colon tumors was studied in male Sprague-Dawley rats. Colon tumors were induced in weanling rats by the administration of 20 weekly subcutaneous injections of 1,2-dimethylhydrazine (DMH). When colon tumors were detected by colonoscopy in 80% of the rats around week 24 after DMH injection, the animals were divided randomly into two groups. One group served as the control. The other group received six endotoxin (Escherichia coli) treatments every fifth day. The first dose was 50 micrograms/100 g (intraperitoneally); the remaining doses were 100 micrograms/100 g (subcutaneously). Rats were killed 2 weeks after the last endotoxin injection. Endotoxin treatments resulted in larger colon tumors. The median tumor size was 71 mm2 for endotoxin-treated and 31 mm2 for untreated rats (P less than 0.02). Endotoxin treatments also resulted in a significantly higher incidence (P less than 0.05) of ulcer development in the small intestine, that is 47% in the endotoxin-treated versus 23% in the untreated rats. After a single subcutaneous injection of endotoxin (100 micrograms/100 g), the colon mucosal reduced glutathione (GSH) level was raised by 21% at 16 hours, reached a peak on day 2, then decreased to baseline by day 4. The increased GSH level in the colon mucosa was maintained up to the third endotoxin injection. By the fifth injection, no increase in the GSH level was observed. These results suggest that the growth of colon tumors in rats induced by DMH could be enhanced by endotoxin treatments. The enhanced tumor growth may be due to an increase in the colon GSH level and/or other mediators released by macrophages as a result of endotoxin treatments.     

Chronic Trypanosoma cruzi infection associated with low incidence of 1,2-dimethylhydrazine-induced colon cancer in rats

Experimental data have demonstrated that chronic infection with intracellular parasites may enhance resistance against some types of tumour. This phenomenon has not yet been demonstrated for experimental Trypanosoma cruzi chronic infection. This study investigated the effect of a specific colon cancer inducing drug, 1,2-dimethylhydrazine (DMH), on chronically T.cruzi infected Wistar rats. Infection was obtained by inoculation of 10(5) tripomastigote forms by subcutaneous (s.c.) route. Acute phase of the infection was monitored every other day by examination of a blood smear from each animal until negativation. In the early chronic phase of the infection, colon adenocarcinoma was induced by weekly s.c. injections of DMH at a dose of 20 mg/kg body weight for 12 weeks. 102 animals were divided in four test groups: 39 infected rats received DMH (group 1); 32 non-infected rats received DMH (group 2); 16 infected rats and 15 non-infected animals were used as control groups. Animals were killed 6 months after the first dose of DMH. The whole colon was removed and prepared for light microscopic examination. Twelve animals from group 1 and 22 from group 2 had colon adenocarcinomas, the proportion of cancer being 30.7 and 68.7%, respectively (chi(2) = 10.16; P < 0.05). The relative risk of having a colon tumor in infected animals (group 1) was 0.45 (IC 95% 0.26-0.76), which is a protective risk compared with non-infected animals. These findings show that chronic infection with T.cruzi is associated with a lower incidence of DMH-induced colon cancer in rats.     

Bombesin enhances experimental carcinogenesis induced in rat colon by 1,2-dimethylhydrazine

The effects of bombesin on the colonic mucosa and on the incidence,number, size and histology of colon cancers induced by 1,2-dimethylhydrazine(DMH) were studied in Fischer 344 rats. In experiment 1, ratswere randomized into three groups to receive either saline orbombesin (10 or 30 µg/kg body wt) to determine the labelingindex of normal colonic mucosa. In experiment 2, rats were given20 weekly injections of DMH (20 µg/kg body wt) and receivedeither saline or bombesin (10 or 30 µg/kg body wt) everyother day for 24 weeks. Administration of bombesin significantlyincreased the labeling indices of colonic mucosa in a dose-dependentmanner. Chronic administration of bombesin at both dosages withDMH caused significant increases in the incidence, number anddepth of involvement of colon cancers; however, it did not affectthe size and histological type of colon cancers. In addition,bombesin at the dose of 30 µg/kg significantly increasedthe labeling index of colon cancer. These results suggest thatbombesin stimulates the cell proliferation of colonic mucosaand colon cancer and enhances colon carcinogenesis in rats.     

Site specific reduction of colon cancer incidence, without a concomitant reduction in cryptal cell proliferation, in 1,2-dimethylhydrazine treated rats by diets containing 10% pectin with 5% or 20% corn oil

The effects of specific dietary interventions on incidence ofcarcinogen-induced cancer and on cryptal cell proliferationin areas of the colon located either over aggregates of lymphoidnodules (ALN) or away from ALN was investigated. Groups of dimethylhydrazine(DMH) treated rats or non-DMH-treated rats were fed a basalAIN-76 diet less fiber of any type, or the basal fiber freediet supplemented with 10% pectin and with 5%, 10%, or 20% cornoil. The adenocarcinoma (AC) incidence was determined in regionsof the colon, i.e. ascending, descending, descending over theALN and descending away from the ALN. The results indicate that:(i) factors associated with ALN promote AC formation, (ii) dietarymodifications (addition of pectin and of 20% corn oil to thediet) each cause significant site specific suppression of ACincidence, (iii) DMH-treatment rendered crypts non-responsiveto the suppression of cryptal cell proliferation which occurredin the rats not treated with DMH (suggestive of a DMH-inducedloss in the regulation of cell proliferation) and (iv) reductionof AC incidence was not always accompanied by reduction in cryptcell proliferation. Studies of intervention procedures designedto prevent colon cancer should take into account the colon sitespecific tumorigenic response to the preventive agent and shouldnot rely on a single biomarker to predict the efficacy of theintervention.     

Chemopreventive effect of trans-resveratrol--a phytoalexin against colonic aberrant crypt foci and cell proliferation in 1,2-dimethylhydrazine induced colon carcinogenesis

Prevention of cancer remains a primary need and new chemopreventive agents must be developed for this purpose. Towards this goal, a chemoprevention study was conducted to evaluate the activity of resveratrol (Res), a phytoalexin, as an inhibitor of colon carcinogenesis. Wistar male rats were divided into six groups, group 1 were control rats, group 2 were control rats that received Res (8 mg/kg body wt p.o. everyday), rats in groups 3-6 were treated weekly with 1,2-dimethylhydrazine (DMH, 20 mg/kg body wt, s.c. x 15 times). In addition, groups 4, 5 and 6 received Res as in group 2. Modifying effects were assessed using aberrant crypt foci (ACF) and the extent of histopathological lesions as end point markers. At the end of 30 weeks, Res markedly reduced tumor incidence, the degree of histological lesions and also the size of tumors significantly (P < 0.05) as compared with the rats treated with unsupplemented DMH. The number of ACF consisting of more than six aberrant crypts per rat was observed in group 6 (6.2 +/- 1.4), group 5 (7.7 +/- 1.0) and group 4 (8.2 +/- 1.4) which were significantly lower than that of group 3 (22.3 +/- 2.4) (P < 0.05). The most pronounced inhibition of ACF development was noted in rats fed Res for the entire period and also during the post-initiation period. Also, Res administration lowered the number of argyrophilic nucleolar organizing region-associated proteins (AgNORs) per nucleus in non-lesional colonic crypts, which reflects the cell proliferation activity. Oxidative imbalance in DMH-treatment was significantly (P < 0.01) modulated on Res supplementation as indicated by optimal concentration of thiobarbituric acid reactive substances (TBARS), superoxide dismutase (SOD), catalase (CAT) and reduced glutathione (GSH). The results of our study suggest Res to be an effective chemopreventive agent, which suppresses DMH-induced colon carcinogenesis at various stages.     

Dietary whey protein protects against azoxymethane-induced colon tumors in male rats.

Epidemiological studies have suggested a relationship between diet and colon cancer incidence. Results from animal studies suggest that whey protein, but not casein protein, may provide protective effects against experimentally induced breast cancer in animals. In the current study, we investigated the effects of casein and whey diets on chemically induced colon cancer in male rats. Pregnant female Sprague Dawley rats (days 3-4 of gestation) were maintained on modified AIN-93G diets formulated with a single protein source of either casein or whey. Life-time exposure to these diets was studied in the F1 generation (experiment A) or the F2 generation (experiment B). Male offspring were weaned to the same diets as the dams and were maintained on these diets throughout the study. At age 90 days, all rats received azoxymethane once a week for 2 weeks (s.c., 15 mg/kg). Forty weeks after the last azoxymethane injection, all rats were euthanized, the colon was examined visually for tumors, and each tumor was histologically evaluated. The weights and distribution of all of the tumors were recorded. In experiment A, rats fed the casein diet had a 56% incidence of colon tumors compared with 30% of the rats on whey-based diets (P < 0.05). In experiment B, rats fed the casein diet had 50% incidence of colon tumors compared with 29% in the whey group (P < 0.05). There were no significant effects of diet on tumor multiplicity or mass. These results suggest that consumption of whey protein-containing diets may reduce the risk of developing colon tumors.     

Effects of a 15% orange-pulp diet on tumorigenesis and immune response in rats with colon tumors

This study evaluated whether the feeding of rats with a 15% orange-pulp diet affects the lymphatic system and the tumorigenic response in rats exposed to a high dose of carcinogen. Five-week-old Sprague Dawley rats were divided into 2 groups fed a control chow diet or the same diet with 15% orange pulp. All rats were injected with 1,2-dimethylhydrazine (DMH) (20 mg/kg) weekly for 6 weeks. At 8 months, tumors, spleens and descending colon were taken from each group for analyses. Feeding rats the 15% orange-pulp diet did not reduce the tumor number but modified the number of adenocarcinomas found in the orange-pulp group compared to controls: 66.7% vs. 93.7%. The number of endophytic tumors was also significantly lower in the experimental group: 6.3% vs. 32.3% in controls. DMH affected the size of the splenic structures. The size of follicles and germinal centers decreased significantly in tumor-bearing rats compared to tumor-free rats. This effect was changed in rats fed the orange-pulp diet. In tumor-bearing rats from this group, only the area of the marginal zone decreased and the red pulp increased compared to tumor-free rats. The size of germinal centers significantly increased compared to tumor-bearing rats in controls. The total number of lymphoid cells decreased in germinal centers of spleens obtained from control tumor-bearing rats compared to tumor-free rats. DMH alone significantly increased the total number of cells in the colon mucosa of the rats fed the control diet. In tumor-bearing rats exposed to the carcinogen and fed the 15% orange-pulp diet, the total number of cells and the number of Ki-67+ cells increased in the depth of tumors whereas the number of CD8+ T cells increased in the colon mucosa, at the border of tumors and its depth. The caspase-3 protein a cysteine protease was elevated in tumors from rats fed the orange-pulp diet. Although the 15% orange-pulp diet did not change the number of tumors in the tumor-bearing rats, feeding rats orange pulp significantly decreased the number of endophytic tumors and increased the number of exophytic tumors. Increased activity of T cell killers in tumors and higher level of proteins involved with apoptosis following consumption of the orange pulp indicate a clear tumor suppressor effect of these dietary fibers.     

Chemoprevention of 2-amino-1-methyl-6-phenylimidazo- [4,5-b]pyridine-induced colon carcinogenesis by 1-O-hexyl-2,3,5-trimethylhydroquinone after initiation with 1,2-dimethylhydrazine in F344 rats

The aim of this study is to investigate the chemopreventive effects of the synthetic phenolic antioxidant 1-O-hexyl-2,3, 5-trimethylhydroquinone (HTHQ) on 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-associated colon carcinogenesis in rats after initiation with 1,2-dimethylhydrazine (DMH) in male F344 rats. Groups of 20-22, 6-week-old male F344 rats were given four subcutaneous injections of 40 mg/kg body wt of DMH during the initial 4 weeks. They were then maintained on powdered basal diet containing 0.03% PhIP alone, PhIP together with 0.5 or 0.125% HTHQ, 0.5 or 0.125% HTHQ alone or basal diet for 32 weeks. A small number (1.1 +/- 1.1/rat) of colon tumors were induced by DMH treatment alone. After initiation with DMH, the number of colon tumors was greatly increased to 8.3 +/- 5.6 by the administration of PhIP. Additional treatment with HTHQ dose-dependently decreased the multiplicity of colon adenocarcinomas to 4.9 +/- 2.8 and 2.6 +/- 1.4 with 0.125 and 0.5%, respectively. This treatment similarly reduced atypical hyperplasias of the ventral prostate. Furthermore, HTHQ significantly reduced the multiplicity of duodenal adenocarcinomas induced by DMH + PhIP or DMH alone. Immunohistochemically, HTHQ was revealed to suppress PhIP-DNA adduct formation in the epithelial cells of the colon and prostate in a separate 2 weeks experiment. The present results clearly showed that HTHQ has chemopreventive potential for PhIP-associated colon and prostate carcinogenesis. The observed inhibition may largely be due to interference with PhIP-DNA adduct formation. In addition, HTHQ has been demonstrated to inhibit duodenal carcinogenesis in the post-initiation stage.     

Early detection of cancer-associated gene alterations in DNA isolated from rat feces during intestinal tumor-induction with 1,2-dimethylhydrazine

A highly sensitive mutation detection method was applied to reveal tarry K-ras alterations in exfoliated intestinal epithelium of Fischer-344 rats during the course of 1,2-dimethylhydrazine (DMH)-induced carcinogenesis. Ten weekly s.c. injections of DMH (50 mg/kg) in combination with consumption of a low-fiber diet resulted in 100% incidence of intestinal tumors at 20 weeks after initial DMH injection. Analysis of DNA extracted from fresh fecal samples obtained individually showed that proportion of codon 12 K-ras oncogene mutant alleles (G-->A transition at the second position of codon 12) was increased in some rats at 4 weeks and clearly in all rats at 8 weeks after initial DMH injection, i.e. much earlier than the first tumors appeared (14 weeks). A gradual increase of mutant K-ras fraction in DNA samples extracted from feces led to an extremely high level of the mutant reaching 10% of the oncogene alleles at the end of the experiment (20 weeks). K- and H-ras oncogene and p53 tumor suppressor gene mutations were analyzed in the resulting colon and duodenal tumors. 14 of 17 colon tumors had K-P as mutations (11 - G-->A transition at codon 12 second base; 3 - G-->A transition at codon 13 second base). G-->A transitions at codon 12 first base of H-ras were detected in 3 colon tumors. All 5 duodenal tumors induced in the experiment had G-->A transition at codon 12 second base of K-ras. 3 of these tumors also had H-ras mutations. No mutation was detected within exons 4-7 of p53 gene indicating that p53 alterations may not be involved in the rapid development of tumors induced by high doses of DMH. Our observations suggest that detection of K-ras mutations in stool samples are predictive of later tumor development from a very early stage.     

Dietary calcium and vitamin D modulate 1,2-dimethylhydrazine-induced colonic carcinogenesis in the rat

To determine whether supplemental dietary calcium and/or vitamin D deficiency are involved in modulating colon cancer induced by 1,2-dimethylhydrazine (DMH), Sprague-Dawley rats were fed diets containing either: (a) a normal content of calcium (0.87%) and phosphorus (0.60%) with 2.2 IU of vitamin D3 per g of feed (group A); (b) the same diet as group A, but with calcium and phosphorus increased to 1.80 and 0.80%, respectively (group B); or (c) a vitamin D-deficient diet with supplemental calcium (1.80%) and phosphorus (0.80%) (group C). After 6 weeks on their respective diets, one-half the animals in each group were given s.c. injections of either vehicle or DMH (20 mg/kg body weight/week) for 26 weeks. Animals were then sacrificed and the incidence of tumors as well as the number of tumors per tumor-bearing rat were determined. Colonic mucosal polyamine levels were measured after 15 weeks of exposure to vehicle or DMH, before development of histologically recognizable neoplasms. The results of these experiments demonstrated that neither calcium supplementation alone nor supplemental calcium in conjunction with vitamin D deficiency altered the incidence of colonic cancer induced by this carcinogen. Supplemental calcium, however, significantly decreased the number of rats with multiple tumors and reduced tumor size. Moreover, vitamin D deficiency abolished these protective effects of calcium on colon cancer in this experimental model. DMH treatment increased polyamine levels in the premalignant colonic mucosa in group A rats. This carcinogen-induced effect was blunted by high dietary calcium. Vitamin D-deficient, calcium-supplemented rats (group C) showed an increase in N1-acetylspermidine, but not the other polyamines, with DMH treatment.     

Interaction of dietary fat and route of carcinogen administration on 1,2-dimethylhydrazine-induced colon tumorigenesis in rats

Since the results of an earlier study indicating no effect ofdietary fat on dimethylhydrazine (DMH)-induced colon cancerin rats differed from those of other investigators, the presentstudy was initiated to determine if the modulating effect offat intake on colon tumorigenesis was dependent on the routeof DMH administration. Male weanling Sprague-Dawley rats (160)were fed one of two nutritionally balanced diets containing5% or 24% corn oil (CO). Following 3 weeks adaptation to theirrespective diets, 40 rats from each diet group were treatedwith five doses of DMH (30 mg/kg) by intragastric i.g.) gavageor subcutaneous (s.c.) injection, over a 3 week period. Ratswere sacrificed when they showed clinical signs of colon tumorand surviving animals were killed 51 weeks after the initialDMH treatment. The cumulative probability of death with coloncarcinoma did not differ between the dietary or treatment groups.There was no effect of route of administration or dietary faton total intestinal tumor incidence. The number of rats withcolon carcinoma was: 5%CO.IG=25; 24%CO.IG=27; 5%CO.SC=23; 24%CO.SC=19.Polypoid tumor incidence was significantly higher in the 24%CO.SCgroup (12/40) compared to the 5%CO.SC group (3/40) (Chi-squared= 5.25; p <0.03) while sessile tumor incidence was the inverse.Marginally significant differences in tumor morphology werenoted between the IG groups.