Triamterene Suppresses Bombesin-enhanced Peritoneal Metastasis of Intestinal Adenocarcinomas Induced by Azoxymethane

The effects of combined administration of bombesin and the diuretic triamterene on the incidence of peritoneal metastasis of intestinal cancers induced by azoxymethane (AOM) and the labeling index of intestinal cancers were investigated in male inbred Wistar rats. From the start of the experiment, rats were given weekly s.c. injections of AOM (7.4 mg/kg body weight) for 10 weeks and s.c. injections of bombesin (40 μg/kg body weight) every other day, and from week 16, s.c. injections of triamterene (10 and 20 mg/kg body weight) every other day until the end of the experiment in week 45. Bombesin significantly increased the incidence of intestinal tumors and cancer metastasis to the peritoneum in week 45. It also significantly increased the labeling index of intestinal cancers. Although administration of both doses of triamterene with bombesin had little or no influence on the enhancement of intestinal carcinogenesis by bombesin, or the location, histologic type, depth of invasion, or labeling index of intestinal cancers, it significantly reduced the incidence of cancer metastasis. These findings indicate that triamterene suppresses cancer metastasis through a mechanism that does not affect the proliferation of intestinal cancers.     

Enhancement by methionine enkephalin of colon carcinogenesis induced by azoxymethane

The effect of the opioid receptor agonist methionine enkephalin (Met-enkephalin) and the opioid receptor antagonist naloxone on colonic carcinogenesis induced by azoxymethane was investigated in Wistar rats. Rats received ten weekly injections of 7.4 mg/kg of body weight of azoxymethane and injections of Met-enkephalin (50 micrograms/kg of body weight), naloxone (2 mg/kg of body weight), or Met-enkephalin (50 micrograms/kg of body weight) plus naloxone (2 mg/kg of body weight) once every 2 days. In wk 40, the group treated with Met-enkephalin had a significantly increased incidence of colonic tumors. A combination of Met-enkephalin and naloxone attenuated the enhancing effect by Met-enkephalin on the development of colonic tumors. Administration of naloxone alone had no influence on colonic tumorigenesis. During and after administration of the carcinogen, the bromodeoxyuridine-labeling indices of the colon mucosa and/or cancers were significantly increased in rats treated with Met-enkephalin. However, a combination of Met-enkephalin and naloxone significantly decreased the labeling indices of the colon mucosa and/or cancers. These findings indicate that Met-enkephalin enhanced colon carcinogenesis and that naloxone attenuated this enhancement. Because naloxone is an opioid receptor antagonist, these findings also indicate that the enhancing effect of Met-enkephalin on colon carcinogenesis may be mediated through opioid receptors.     

Chemoprevention by amiloride of experimental carcinogenesis in rat colon induced by azoxymethane

The effects of amiloride on the incidence and histology of colontumors induced by azoxymethane, on the labeling index of thecolon mucosa and on the activity of ornithine decarboxylasein the colon wall were investigated in Wistar rats. Rats received10 weekly injections of 7.4 mg/kg body wt azoxymethane and s.c.injections of 5 or 7.5 mg/kg body wt amiloride in depot formevery other day for 35 weeks. Prolonged administration of amilorideat a dose of 7.5 mg/kg, but not 5 mg/kg, significantly reducedthe incidence of colon tumors at week 35. However, administrationof amiloride had little or no significant influence on the histologicaltypes of colon tumors and cancers. Administration of amilorideat 7.5 mg/kg significantly decreased the labeling index of thecolon mucosa and ornithine decarboxylase activity in the colonwall during and after administration of azoxymethane. Thesefindings suggest that amiloride inhibits development of colontumors. A possible mechanism of inhibition of colon carcinogenesisby amiloride is its suppression of proliferation of colon tumorcells.     

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.     

Dietary n-3 PUFA increases the apoptotic response to 1,2-dimethylhydrazine, reduces mitosis and suppresses the induction of carcinogenesis in the rat colon

The effect of dietary fish oil on colonic crypt cell apoptosis and proliferation was examined in male Wistar rats, 24 and 48 h after administration of 1,2-dimethylhydrazine (DMH), and its influence on the induction of aberrant crypt foci (ACF) in the distal colon was assessed. Rats (125-150 g) fed a high-fat semi-synthetic diet containing corn oil (CO) were given DMH (30 mg/kg body wt) or a sham injection of EDTA/NaCl. Animals were then fed either the CO diet or a diet in which fish oil (EPA 18.7%; DHA 8%) was substituted for corn oil. Subgroups of rats (n = 5) were killed after 24 and 48 h, and crypt cell apoptosis and proliferation were quantified by morphological criteria in isolated intact crypts from the mid and distal colon. Consumption of the fish oil diet (FO) was associated with increased apoptotic cell death (P < 0.001) and suppression of proliferation (P < 0.05) in colonic crypts both 24 and 48 h after DMH. In a second experiment, animals were given three injections of DMH or sham injections of carrier at weekly intervals. For 48 h after each injection animals were fed either the CO or FO diet, but otherwise maintained on the CO throughout. The number and crypt multiplicity of ACF in the distal colon were determined after 18 weeks, and animals given the FO diet for the 48 h period following carcinogen administration were found to have significantly fewer ACF than rats fed the CO diet (P < 0.05). The data demonstrate that the fatty acid composition of the diet is an important determinant in the induction of carcinogenesis by DMH. The proliferative and apoptotic response of the colonic crypt to carcinogen and fish oil, coupled with the reduced incidence of ACF, suggest n-3 PUFA can protect against the carcinogenic effects of DMH by mediating changes in the balance proliferation and cell death.     

Promotion by bombesin of gastric carcinogenesis induced by N-methyl-N'-nitro-N-nitrosoguanidine in Wistar rats

The effects of bombesin on the incidence, number, histological type, and depth of involvement of gastric cancers induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) were investigated in male Wistar rats. Rats received alternate-day s.c. administration of 20 or 40 micrograms/kg body weight of bombesin in depot form after p.o. treatment with the carcinogen for 25 weeks. Prolonged administration of bombesin at 40 micrograms/kg led to a significant increase in the incidence and number per rat of gastric cancers of the glandular stomach at Week 52. In rats that had received alternate-day injections of 20 micrograms/kg of bombesin, the number of gastric cancers per rat, but not the incidence of cancer, was significantly more than in untreated rats. However, bombesin at both dosages did not affect the histological appearance of the lesions or their depth of involvement. At Weeks 30 and 52, norepinephrine concentrations in the fundic and antral portion of the gastric walls and labeling indices in the antral and fundic mucosae were significantly higher in rats treated with bombesin at both dosages than in untreated rats. These findings indicate that bombesin enhances gastric carcinogenesis after administration of N-methyl-N'-nitro-N-nitrosoguanidine is stopped and that this effect may be related to its effects in increasing tissue norepinephrine concentrations in the stomach wall and increasing cell proliferation in the gastric mucosa.     

Increased induction of aberrant crypt foci by 1,2-dimethylhydrazine in rats fed diets containing purified genistein or genistein-rich soya protein

The isoflavonoid genistein inhibits mitosis and increases apoptosisin a variety of tumour cell lines in vitro, and may exert anticarcinogeniceffects in vivo. To assess its effects on the colon, rats werefed a semi-synthetic control diet, or similar diets enrichedwith genistein (0.25 g/kg), either as the pure isoflavone oras part of a soya protein isolate, for 7 days before receivingsubcutaneous injections of saline or 1,2-dimethylhydrazine (DMH).After 48 h, rats given saline were killed and samples of theirsmall and large intestinal mucosa were obtained for assessmentof crypt cell mitosis and apoptosis by visual analysis of isolatedintact crypts. Rats given DMH were fed control diet and killedafter 48 h for assessment of crypt cytokinetics or maintainedfor 42 days then killed and their colonic mucosa analysed foraberrant crypt foci (ACF). Two further groups were given controldiet before DMH, followed by the genistein or soya-based dietfor 42 days before assessment of ACF. Neither genistein norsoya protein isolate had a significant effect on crypt cellmitosis or apoptosis in untreated rats, or on the proliferativeresponse to treatment with DMH. However, consumption of puregenistein or the soya protein isolate before treatment withDMH was associated with a 3-fold (P < 0.001) or 2-fold (P< 0.05) increase, respectively, in ACF in the distal colon.There was no significant effect of genistein or soya proteinisolate given after DMH treatment. We conclude that genisteinhas no detectable effect on colonic crypt mitosis or apoptosisin the rat in vivo, but that it promotes induction of ACF byan as yet undefined mechanism when fed immediately before treatmentwith DMH.     

Ornithine decarboxylase inhibitor attenuates bombesin enhancement of intestinal carcinogenesis and metastasis induced by azoxymethane

The effects of combined administration of bombesin (40 μ/kg body weight) and the ornithine decarboxylase (ODC) inhibitor, 1,3-diaminopropane (DAP), on the development of large and small intestinal tumors and the incidence of their metastasis to the peritoneum induced by azoxymethane (AOM, 7.4 mg/kg body weight), the ODC activity of the intestinal wall, and the labeling index of the intestinal mucosa and tumor were investigated in inbred Wistar rats. Rats received weekly s.c. injections of AOM for 10 weeks, s.c. injections of bombesin every other day, and drinking water containing DAP (2.5 g/l) until the end of the experiment at week 40. Administration of bombesin significantly increased the incidence of intestinal tumors at week 40. It had no influence on the location, size, histological features or depth of involvement of intestinal adenocarcinomas, but significantly increased the incidence of their metastasis to the peritoneum. It also resulted in a significant increase in the intestinal ODC activity and labeling index. Administration of DAP with bombesin significantly reduced the enhancement of intestinal carcinogenesis by bombesin. Although the combined use of DAP with bombesin had little or no influence on the location, size, histological features, or depth of involvement of intestinal cancers, the incidence of their metastasis was significantly reduced. DAP significantly attenuated bombesin enhancement of the intestinal ODC activity and labeling index. These findings indicate that ODC inhibition attenuated the enhancement of intestinal carcinogenesis and metastasis to the peritoneum.     

Effect of dietary sodium ascorbate on 1,2-dimethylhydrazine- or methylnitrosourea-induced colon carcinogenesis in rats

The effect of dietary sodium ascorbate (SA) on colon carcinogenesisevoked by 1,2-dimethylhydrazine (DMH) or N-methyl-N-nitrosourea(MNU) was studied in female F344 rats. Animals were fed dietscontaining 0, 0.25 and 1% of SA and given s.c. a single doseof 150 mg DMH/kg body wt., 10 weekly s.c. injections of 20 mgDMH/kg body wt. or intrarectal administration of 2 mg MNU, twicea week for 2 weeks. The incidence of colon and kidney tumorswas lower in rats fed the 0.25 or 1% SA and treated with a singledose of DMH than in the animals fed the diet without SA; however,the tumor incidences did not differ between the SA- and controldiet-fed animals and treated with multiple doses of DMH or MNU.     

Inhibition by the dopamine antagonist haloperidol of experimental carcinogenesis induced by azoxymethane in rat colon.

The effects of the dopamine agonist bromocriptine and antagonist haloperidol on the incidence and histology of colon tumors induced by azoxymethane and on the labeling index of colon mucosa were investigated in Wistar rats. Rats received weekly s.c. injections of 7.4 mg/kg of body weight azoxymethane for 10 weeks and s.c. injections of 2 mg/kg of body weight bromocriptine or 2 mg/kg of body weight haloperidol, in depot form, every other day until the end of the experiment in week 30. Administration of haloperidol resulted in a significant decrease in the incidence of colon tumors. It also caused a significant decrease in the incidence of adenocarcinomas, with 75% of the tumors being adenomas, and in the labeling index of the colon epithelial cells. In contrast, bromocriptine had no influence on the incidence or histology of colon tumors or the labeling index of the colon mucosa. These findings indicate that the dopamine antagonist haloperidol inhibits colon carcinogenesis and that this effect may be related to its effect in decreasing the proliferation of colon epithelial cells.     

Presence of well‐differentiated distal,but not poorly differentiated proximal,rat colon carcinomas is correlated with increased cell proliferation in and lengthening of colon crypts

To determine whether colon crypt proliferative parameters were significantly altered by the stage of colon carcinogenesis or the type or location of colon tumors in rats, male Sprague‐Dawley rats received an injection of the carcinogen 1,2‐dimethylhydrazine (12 mg DMH base/kg body weight) or DMH vehicle once a week for 8 weeks, then were killed 24 weeks later. Three hours before sacrifice, rats were injected with 1 mg/kg body weight colchicine to arrest mitotic cells at metaphase. Transverse sections of the colon mucosa were taken 6 cm from the anus and at least 3 cm from any tumor, fixed in formalin, then stained with hematoxylin & eosin (H&E) for analyses of proliferative parameters. Only complete, mid‐axial crypts were scored for mitotic count (MC), crypt proliferative zone (PZ) height and crypt height (CH). Serial tumor sections were stained with H&E for histological evaluation or used in immunohistochemical detection of transforming growth factor α (TGFα). DMH treatment significantly increased MC, PZ and CH regardless of tumor status. The PZ and CH of rats with a carcinoma located in the distal colon were significantly increased compared with DMH‐treated rats without an adenocarcinoma (AC) or with rats which had a tumor located in the proximal colon. Distal colon ACs were found to be well differentiated and to have greater TGFα immunoreactivity than the generally less differentiated proximal colon carcinomas. Distal colon AC production and systemic circulation of a soluble colon crypt stimulating factor such as TGFα may explain the significant increase in PZ and CH in histologically normal colonic mucosa located away from the tumor. Int. J. Cancer 80:68–71, 1999. © 1999 Wiley‐Liss, Inc.     

Inhibition of ornithine decarboxylase with 2-difluoromethylornithine: reduced incidence of dimethylhydrazine-induced colon tumors in mice

2-Difluoromethylornithine (DFMO) was administered to 1,2-dimethylhydrazine (DMH)-treated mice to reduce colonic polyamine levels and mucosal hyperplasia. Mice received 1% DFMO in drinking water throughout the experiment and were given injections of DMH (20 mg/kg) weekly for 28 weeks. DFMO inactivated 93% of colonic ornithine decarboxylase activity. Although DMH treatment did not induce colonic ornithine decarboxylase activity by Week 28, the putrescine content was increased 31% in DMH-treated mice (p less than 0.01). Concurrent treatment with DFMO depressed putrescine content (42 to 63%) and spermidine content (27 to 38%), but it increased spermine content (18 to 22%). At Week 28 of treatment with DMH alone, RNA content was increased 8.6% (p less than 0.01), DNA content 10% (p less than 0.01), DNA specific activity 24% (p less than 0.01), and crypt depth 20% (p less than 0.01), but not in mice receiving DMH and DFMO. At 28 weeks, 13 of 17 mice (76%) treated with DMH alone had histologically confirmed colon cancers; of mice treated with DMH and DFMO, two of 18 (11%) had colonic tumors. Throughout the experiment, 50 colon cancers developed in 16 DMH-treated mice (mean, 3.12 tumors/mouse); three mice treated with DMH and DFMO developed three colon cancers total (p less than 0.001). Reduction of colonic polyamine levels after DFMO treatment prevents proliferative changes induced by DMH and reduces the incidence of tumors.     

Effect of polyamine oxidase inhibition on the colonic malignant transformation process induced by 1,2-dimethylhydrazine

Recent studies of colon adenocarcinomas in humans and experimentally induced colonic tumors in rodents have demonstrated selective elevations in the level of N1-acetylspermidine in these malignant tissues. The exact relationship of these alterations in acetylated polyamine levels to the malignant transformation process, however, remains unclear. In order to clarify this issue, rats were given s.c. injections of 1,2-dimethylhydrazine (DMH; 20 mg/kg body wt/week) or diluent for up to 26 weeks. After 10 weeks of carcinogen treatment, one-half of the animals in each group were also concomitantly given i.p. injections of MDL 72527 (20 mg/kg body wt/week), a specific inhibitor of polyamine oxidase, until they were killed. Animals were killed after 15 weeks of DMH treatment and polyamine levels as well as the activities of polyamine oxidase, ornithine decarboxylase and spermidine-N1-acetyltransferase were measured and compared in rat proximal and distal colonic mucosa of each group. Polyamine levels were also assessed in each of these groups after 26 weeks of treatment with this carcinogen +/- MDL 72527. In addition, in view of recent studies that have indicated that polyamines may influence certain oncogenes in human colonic carcinoma cells, tumors from DMH +/- MDL 72527 were analyzed for K-ras mutations. The results of these experiments demonstrated for the first time that: (i) MDL 72527 was a specific inhibitor of polyamine oxidase in normal and malignant colonic tissue; (ii) concomitant administration of this agent with DMH enhanced the elevation of colonic N1-acetylspermidine and significantly reduced the mean colonic tumor burden, as assessed by total tumor area per rat, produced by this carcinogen alone; (iii) analysis of K-ras mutations revealed a similar incidence (62-69%) in adenocarcinomas for both groups (+/- MDL 72527); (iv) however, analysis of the K-ras-mutated and non-mutated tumors revealed that in both carcinogen-treated groups (+/- MDL 72527), tumors with such mutations were smaller than their counterparts without such genetic alterations. Moreover, MDL 72527 reduced the average size of tumors, with and without such mutations, to a similar extent.     

Induction by bombesin of peritoneal metastasis of gastric cancers induced by N-methyl-N′-nitro-N-nitrosoguanidine in Wistar rats

Background. We investigated the effect of the gastrointestinal regulatory peptide, bombesin, on the development of peritoneal metastasis from gastric cancers induced in rats by N-methyl-N′-nitro-N-nitrosoguanidine (MNNG), and on Rho activity in the gastric cancers. Methods. Rats were allocated to three groups. All groups received MNNG (100 μg/ml) solution for 25 weeks from the start of the experiment. Group 1 (controls) received olive oil injections from the start of MNNG treatment; group 2 animals received alternate-day s.c. injections of bombesin (40 μg/kg body weight) in olive oil from the start of the experiment until the end of the experiment at week 52; and group 3 received the s.c. bombesin injection on alternate days from week 26 until week 52. The effect of bombesin on Rho activity in gastric cancer was examined by Western blotting. Results. Bombesin given from the start of the experiment (group 2) and after the MNNG treatment (group 3) both significantly increased the incidence of gastric cancer metastasis, compared with controls, at week 52: The incidence of metastasis was significantly higher in group 2 than in group 3. Bombesin from the start of the experiment (group 2) significantly increased the incidence of tumors with deeper invasion or more infiltrative growth pattern, or lymphatic vessel tumor invasion, while bombesin after MNNG treatment (group 3) significantly increased the incidence of lymphatic vessel invasion. Bombesin also increased the activity of Rho protein in the tumors. Conclusion. Bombesin significantly increased the incidence of peritoneal metastasis from gastric cancers through the activation of Rho protein. Received: October 19, 2000 / Accepted: January 9, 2001     

Genistein attenuates peritoneal metastasis of azoxymethane-induced intestinal adenocarcinomas in Wistar rats

The effects of the soybean isoflavonoid genistein on the development of bombesin-enhanced peritoneal metastasis from intestinal adenocarcinomas induced by azoxymethane (AOM) were investigated in male inbred Wistar rats. From the beginning of the experiment, rats were given 10 weekly s.c. injections of AOM (7.4 mg/kg body weight) and s.c. injections of bombesin (40 microg/kg body weight) every other day, and from week 16, s.c. injections of genistein (5 or 10 mg/kg body weight) every other day until the end of the experiment in week 45. Bombesin significantly increased the incidence of intestinal tumors and of cancer metastasis to the peritoneum. Although genistein administered at either dose had little or no effect on the enhancement of intestinal carcinogenesis by bombesin or on the location, histologic type, depth of involvement, labeling index, or growth pattern of intestinal cancers, it significantly decreased the incidence of cancer metastasis. Genistein also significantly decreased the incidence of lymphatic vessel invasion of adenocarcinomas, which was enhanced by bombesin. Our findings indicate that genistein attenuates cancer metastasis by inhibiting cancer cell invasion into lymphatic vessels through activities that do not affect the growth of intestinal cancers.     

Enhanced levels of oleate-dependent and Arf-dependent phospholipase D isoforms in experimental colon cancer

Phospholipase D (PLD) (EC 3.1.4.4) is one of the intracellar signal transduction enzymes and plays an important role in a variety of cellular functions. In order to clarify the role of PLD in proliferation and tumorigenesis of colon cancer, we investigated the activities of oleate-dependent and ADP-ribosylation factor (Arf)-dependent types of PLD in experimental colon cancer of the rat. We produced colon cancer in Wistar rats by injecting the carcinogen, dimethylhydrazine dihydrochloride (DMH). The control rats were injected with physiological saline. Mucosal scrapings from the colon were homogenized and centrifuged to obtain the microsomal or membrane fraction. We measured the two types of PLD activities in these fractions using the transphosphatidylation reaction. Both oleate-dependent and Arf-dependent PLD activities were significantly higher in the colon cancer tissue than normal colonic mucosa. The mean specific activity of oleate-dependent PLD) in colon cancers was 1.66 +/- 0.75 (SD) nmol/min/mg whereas the value for normal colonic mucosa was 0.18 +/- 0.09 nmol/min/mg (P < 0.01; Mann-Whitney U-test). On the other hand, the mean specific activity of Arf-dependent PLD in colon cancers was 76.36 +/- 29.37 pmol/min/mg whereas the value for normal colonic mucosa was 19.90 +/- 11.97 pmol/min/mg (P < 0.01; Mann-Whitney U-test). These results suggest that PLD is implicated in the proliferation and tumorigenesis of colon cancer. The present study provides the first evidence for the enhanced levels of two types of PLD in colon cancer and raises the possibility that these PLDs can be used as the potential target for the treatment of colon cancer.     

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.     

Chemopreventive Effect of Amorphophallus campanulatus (Roxb.) Blume Tuber Against Aberrant Crypt Foci and Cell Proliferation in 1, 2-Dimethylhydrazine Induced Colon Carcinogenesis

Colorectal cancer is one of the leading causes of cancer death, both in men and women. This study investigatedthe effects of Amorphophallus campanulatus tuber methanolic extract (ACME) on aberrant crypt foci (ACF)formation, colonic cell proliferation, lipid peroxidative damage and the antioxidant status in a long term preclinicalmodel of 1, 2-dimethylhydrazine (DMH) induced colon carcinogenesis in rats. Male Wistar rats were dividedinto six groups, viz., group I rats served as controls; group II rats treated as drug controls receiving 250 mg/kg body weight of ACME orally; group III rats received DMH (20 mg/kg body weight) subcutaneously once aweek for the first 15 weeks; groups IV, V and VI rats received ACME along with DMH during the initiation,post- initiation stages and the entire period of the study, respectively. All the rats were sacrificed at the end of 30weeks and the intestinal and colonic tissues from different groups were subjected to biochemical and histologicalstudies. Administration of DMH resulted in significant (p≤0.05) intestinal and colonic lipid peroxidation (MDA)and reduction of antioxidants such as catalase, glutathione peroxidase, glutathione reductase, glutathione-Stransferaseand reduced glutathione. Whereas the supplementation of ACME significantly (p≤0.05) improvedthe intestinal and colonic MDA and reduced glutathione levels and the activities of antioxidant enzymes inDMH intoxicated rats. ACME administration also significantly suppressed the formation and multiplicity ofACF. In addition, the DMH administered rats showed amplified expression of PCNA in the colon and decreasedexpression of this proliferative marker was clearly noted with initiation, post-initiation and entire period ofACME treatment regimens. These results indicate that ACME could exert a significant chemopreventive effecton colon carcinogenesis induced by DMH.     

Melatonin and colon carcinogenesis: I. Inhibitory effect of melatonin on development of intestinal tumors induced by 1,2-dimethylhydrazine in rats

The effect of pineal indole hormone melatonin on colon carcinogenesis was firstly studied in rats. Two-month-old outbred female LIO rats were weekly exposed to 15 (experiment 1, groups 1 and 2) or to five (experiment 2, groups 1 and 2) s.c. injections of 1,2-dimethylhydrazine (DMH) at a single dose of 21 mg/kg of body weight. From the day of the first injection of the carcinogen DMH, the rats from groups 2 (experiments 1 and 2) were given melatonin five days a week during the night-time (from 18:00 h to 8:00 h), dissolved in tap water at 20 mg/l. The experiment was finalized in 6 months after the first injection of DMH. In both experiments the majority of tumors were localized in the descending colon. Tumors of the small intestines developed only in rats from experiment 1. Total incidence of colon tumors as well as tumors in different parts of the colon and the mean number of tumors per rat were much higher in rats from both groups in experiment 1 than that in rats from experiment 2. In experiment 1 melatonin failed to influence the total incidence of colon tumors. However, incidence of carcinomas in the ascending colon was significantly reduced (P < 0.01). The multiplicity of total colon tumors per rat, as well as the mean number of tumors, ascending and descending colon per rat, was also decreased under the influence of melatonin (group 2 vs group 1, P < 0.01). In the same experiment, melatonin slightly decreased the depth of tumor invasion and increased number of highly differentiated colon carcinomas induced by DMH. The percentage of small tumours in the descending colon among rats from group 2 was higher than that of group 1. Treatment with melatonin was also followed by a decrease in the multiplicity of DMH- induced tumors of the duodenum (group 2 vs group 1, P < 0.05) and by a decrease in the incidence of jejunum and ileum tumors (group 2 vs group 1, P < 0.05). In experiment 2, the inhibitory effect of melatonin on DMH-induced colon carcinogenesis was much more expressed than that in experiment 1. Thus, in group 1 the incidence of total colon tumors, ascending and descending colon tumors, was significantly decreased in comparison with group 2; also melatonin reduced the number of tumors per rat in the ascending and descending colon. The number of colon tumors that invaded only mucosa was significantly higher in group 2 than in group 1, P < 0.05. The ratio of highly differentiated tumors was increased (P < 0.05) and the ratio of low-differentiated tumors was decreased (P < 0.05) in rats exposed to melatonin (group 4) as compared with group 3. The number of large size tumors in the ascending and descending colon was decreased whereas the number of small size tumors (<10 mm2) was increased in those parts of the colon that were under the influence of melatonin in experiment 2. Thus, our results demonstrate the inhibitory effect of melatonin on intestinal carcinogenesis induced by DMH in rats.      

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.