Selenium as a chemopreventive agent in experimentally induced colon carcinogenesis

AIM: To elucidate the chemopreventive efficacy of selenium during experimentally induced colon carcinogenesis.METHODS: Thirty-two male wistar rats were divided into four groups: group I (normal control); group II [1,2-dimethylhydrazine (DMH) treated]; group III (selenium treated); and group IV (DMH + selenium treated). Groups II and IV were given subcutaneous injections of DMH (30 mg/kg body weight) every week for 20 wk. Selenium, in the form of sodium selenite, was given to groups III and IV at 1 ppm in drinking water ad libitum for 20 wk. At the end of the study, rats were sacrificed and their colons were analyzed for the development of tumors, antioxidant enzyme levels and histological changes.RESULTS: 100% of the DMH treated rats developed tumors, which was reduced to 60% upon simultaneous selenium supplementation. Similarly, tumor multiplicity decreased to 1.1 following selenium supplementation to DMH treated rats. Levels of lipid peroxidation, glutathione-S-transferase, superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPx) decreased following DMH treatment, whereas levels of glutathione (GSH) and glutathione reductase (GR) significantly increased in DMH treated rats. Selenium administration to DMH treated rats led to an increase in the levels of lipid peroxidation, SOD, catalase, glutathione-S-transferase and GPx, but decreased the levels of GSH and GR. Histopathological studies on DMH treated rats revealed dysplasia of the colonic histoarchitecture, which showed signs of improvement following selenium treatment.CONCLUSION: The study suggests the antioxidative potential of selenium is a major factor in providing protection from development of experimentally induced colon carcinogenesis.     

Effect of selenium on azoxymethane-induced intestinal cancer in rats fed high fat diet

The effects of selenium supplementation on azoxymethane-induced intestinal cancer were studied in male Sprague-Dawley rats given 8 weekly injections of azoxymethane (8 mg/kg body wt), and fed a 30% beef fat diet. Selenium-supplemented groups received 8 ppm H₂SeO₃ in drinking water. Blood selenium levels of supplemented rats increased rapidly the first 9 weeks of the experiment, followed by a plateau significantly higher than that for non-selenium controls. There was a significant increase in liver and intestinal selenium levels in supplemented groups.The average number of intestinal tumors was 6.5 in the control group, and 3.1 in the selenium-supplemented group. There was a significant reduction in tumor incidence in the proximal half of the colon of selenium-treated rats. There was also increased concentration of tissue selenium in the proximal half of the colon of these rats.     

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.     

Effects of dietary fish oil and corn oil on protein kinase C distribution in the rat colon with and without 1,2 dimethylhydrazine treatment and in rat colonic adenocarcinoma

The activity and subcellular distribution of Protein Kinase C (PKC) was determined in the colons of Sprague-Dawley rats that were fed either a low fat rat chow or rat chow supplemented with 17% corn oil (40% ingested calories as fat). Rats given the high fat diets were either given no carcinogen or treated prior to or subsequent to the initiation of the test diets with 1,2 dimethylhydrazine (DMH). Rats were sacrificed and PKC activity determined in the soluble and particulate fractions of the colonic tissue 13 weeks after the initiation of the diets or DMH treatment, which was before tumor induction. In addition several rats were maintained on their diets until colon tumor formation occurred and PKC activity determined in the colonic tumor and compared to age matched control colonic tissue. In the absence of DMH, fish oil and corn oil equally augmented PKC activity and decreased the ratio of soluble/particulate PKC. With DMH treatment, corn oil augmented PKC as above, but fish oil supplementation resulted in a pattern of PKC activity and distribution more typical of a low fat diet, particularly when fish oil supplementation preceded DMH treatment. PKC activity in DMH induced colonic carcinomas was markedly depressed regardless of the fat source in the diet, when compared to colonic tissue from a non-DMH treated age matched low fat control.     

Ornithine decarboxylase activity in the rat and human colon

We have investigated the effect of age, a high-fat diet, sodium deoxycholate, and the ornithine analogue alpha-difluoromethylornithine on ornithine decarboxylase (ODC) activity in the rat colon. The relative levels of ODC activity were also determined in normal mucosa and tumor tissue from rat and human colon. The colonic ODC activity induced by intrarectal instillation of sodium deoxycholate in male Sprague-Dawley rats was highest in young animals, and it decreased with increasing age. A high level of dietary fat caused both an increased in basal colonic ODC activity and enhanced ODC induction by deoxycholate. alpha-Difluoromethylornithine given in drinking water inhibited, in a dose-dependent fashion, deoxycholate-induced ODC activity. The frequency of azoxymethane-induced intestinal tumors was also significantly reduced by alpha-difluoromethylornithine. Since colonic ODC activity is increased in carcinogenesis by known promoting agents and decreased by tumor inhibitors, this short-term assay may provide a useful system for identifying colon tumor promoters and inhibitors. The ODC activity in colon tumors of Sprague-Dawley rats was found to be significantly higher than in normal-appearing mucosa in the same animals. Similarly, ODC activity in human colon cancer was found to be higher than that of the normal-appearing mucosa in the same specimen. These results strengthen the utilization of the rat model for studies, the results of which may apply to the human situation.     

Effects of D,L-2-difluoromethylornithine and indomethacin on mammary tumor promotion in rats fed high n-3 and/or n-6 fat diets

Virgin female Sprague-Dawley rats (50 days of age) were administered a single intragastric 10-mg dose of 7,12-dimethylbenz(a)anthracene (DMBA). Twenty-one days later they were placed on diets containing either 20% corn oil (CO), 15% menhaden oil plus 5% corn oil (MO + CO), 20% CO plus 0.5% w/w of the irreversible ornithine decarboxylase inhibitor, D,L-2-difluoromethylornithine (CO + DFMO), 20% CO plus 0.004% w/w of the cyclooxygenase inhibitor indomethacin (CO + INDO), 20% CO + 0.004% INDO + 0.5% DFMO (CO + INDO + DFMO), or 15% MO + 5% CO + 0.5% DFMO (MO + CO + DFMO). The incidence of DMBA-induced mammary tumors was significantly reduced in rats fed diets containing DFMO but not in rats fed the diet containing indomethacin. Incidences of mammary tumors at 16 weeks post-DMBA were 86% in rats fed the CO diet, 83% in rats ingesting the diet containing CO + INDO, 28% in rats fed CO + DFMO, 32% in rats fed diet containing CO + INDO + DFMO, 59% in rats fed the MO + CO diet, and 24% in rats fed the MO + CO + DFMO diet. The average number of tumors and tumor burden per tumor-bearing rat were reduced and tumor latency was increased in all rats fed diets containing DFMO. Body weight gain, but not food intake, of rats fed the 20% fat + 0.5% DFMO diets was significantly less than in rats fed the 20% fat diets. Prostaglandin E and leukotriene (LTB4) syntheses, ODC activity and mammary tumorigenesis were significantly inhibited by feeding the diet containing menhaden oil or by adding 0.5% DFMO to any of the high fat diets. Feeding a 20% CO diet containing 0.004% INDO significantly reduced prostaglandin synthesis and ODC activity and increased LTB4 synthesis of mammary tumors but did not inhibit mammary tumorigenesis. This study suggests that the 5-lipoxygenase product LTB4 may be involved in mammary tumor production. Whereas a decrease in LTB4 appears to be associated with a decrease in tumorigenesis, an increase (as seen in the indomethacin group) was not associated with any change in the tumorigenic response.     

Follow-up of tumor development in the colons of living rats and implications for chemoprevention trials: assessment of aspirin-difluoromethylornithine combination

We aimed to establish a reliable procedure allowing the follow-up of tumor development by computed tomographic (CT) colonography in an animal model of colon carcinogenesis in order to assess the chemopreventive efficacy of aspirin and difluoromethylornithine (DFMO) given in combination. Fischer rats received an intraperitoneal injection (25 mg/kg) of dimethylhydrazine (DMH) once a week for two weeks in order to initiate colon carcinogenesis. Five months after the last injection of DMH, a first CT colonography was performed and rats were then randomly separated into two groups (control and experimental). The experimental group received a 0.1% mixture of aspirin and DFMO in drinking water. CT colonography was performed at 6, 7 and 8 months. Data showed a precise correlation between location and size of tumors found at autopsy and those detected by CT colonography at 8 months. All tumors were also detected on the CT views obtained previously. Animals of the aspirin/DFMO group exhibited an inactivation of ornithine decarboxylase, a key enzyme in polyamine biosynthesis, and a two-fold reduction in the prostaglandin E2 content of the colonic mucosa (p<0.01). In rats with tumors at the start of the aspirin/DFMO treatment, a significant slow-down of tumor development was observed. In contrast, in rats where no tumors were detected at the start of the treatment, tumor formation was inhibited. Our data show that CT colonography represents a reliable method to assess in a living animal the efficacy of chemopreventive agents.     

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.     

Induction of colon tumors in 1,2-dimethylhydrazine-resistant Lobund Wistar rats by methylazoxymethanol acetate.

Sprague-Dawley and Lobund Wistar rats, which were sensitive and resistant to induction of colon tumors by 1,2-dimethylhydrazine (DMH), respectively, were treated with methylazoxymethanol (MAM), the product of DMH metabolism by the microsomal mixed-function oxidase system. Although the colon tissue in both stocks of rats had similar NAD+-dependent dehydrogenase activities that are considered necessary to activate MAM to an ultimate carcinogen, still a sevenfold greater incidence of colon tumors was found in the Sprague-Dawley rats, and their tumors were more extensive. The results indicated that the difference in susceptibility to colon tumor induction between the rat stocks was partially related to metabolic activation of the DMH and to other, as yet undetermined, endogenous factors.     

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.     

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.     

Reduced growth rate of dimethylhydrazine-induced colon tumors in rats.

alpha-Difluoromethylornithine (DFMO) treatment has been shown to modify carcinogenesis in many experimental tumor models, including breast, urinary bladder, and colon. This study was designed to determine whether DFMO treatment can inhibit tumor growth on chemical-induced colon cancer in rats. Effectiveness of DFMO in combination with mitomycin C (MMC) was also evaluated. Forty-two Sprague-Dawley rats received dimethylhydrazine (20 mg/kg) s.c. once weekly for 20 wk to induce colon cancer. Then a double-contrast barium enema was performed, and colon tumors were detected. The animals were divided into four groups that were subjected to the following treatment: none; DFMO alone; MMC alone; and a combination of DFMO plus MMC. After 5 wk of treatment, the barium enema was repeated. For the evaluation of treatment efficacy, tumor doubling time was adopted. The mean tumor doubling time in the control group was 20.7 +/- 9.1 days (SD). "Response" was judged as effective when tumor doubling time in treatment groups was more than 38.9 days, calculated from the mean + 2 SDs in the control group. Response rates in the DFMO, MMC, and DFMO plus MMC groups were 40.0%, 10.0%, and 82.3%, respectively. DFMO was a more effective inhibitor of tumor growth than MMC, and DFMO in combination with MMC resulted in a synergic diminution of tumor growth. The double-contrast barium enema is useful to observe sequential tumor growth and may be appropriate for the evaluation of new treatment on experimental colon cancer in rats.     

Celecoxib and difluoromethylornithine in combination have strong therapeutic activity against UV-induced skin tumors in mice

The cyclooxygenase-2 (COX-2) inhibitor celecoxib and the ornithine decarboxylase (ODC) inhibitor difluoromethylornithine (DFMO) were each previously shown to prevent skin tumor development when administered throughout the course of UV irradiation. This raised the question of whether maintenance or continued growth of existing tumors required prostaglandins, the product of COX, or polyamines, the product of ODC. To address this question, SKH hairless mice were irradiated 3 times/week with 90 mJ/cm(2); this dose was increased 10% weekly to a maximum of 175 mJ/cm(2). UV was stopped at 27 weeks, at which time there were an average of 5 papillomas/mouse. The mice were then placed in one of four treatment groups: group 1, no treatment; group 2, 0.4% DFMO in the drinking water; group 3, 500 p.p.m. celecoxib in the diet (AIN76); group 4, both DFMO and celecoxib. The control group continued to produce new tumors in a nearly linear manner such that by week 31 the tumor number had nearly doubled, i.e. approximately 10 tumors/mouse. The group receiving DFMO showed significant tumor regression, losing an average of 1 tumor/mouse/week, such that 50% of the tumors remained at week 31. The celecoxib group showed a 25% reduction in tumor number. The group receiving the combination of celecoxib and DFMO showed the greatest regression, with an 89% reduction in tumor number compared with the control group. There was also a corresponding reduction in the size of the tumors. To determine whether tumor regression was permanent or required continued treatment, all treatments were stopped at 31 weeks. Over the next 4 weeks, tumors reappeared at the same rate in all treatment groups. It is concluded that the combination of celecoxib and DFMO are potent therapeutic agents for skin cancer, although the benefits are lost with the cessation of treatment.     

Modulation of mouse skin tumor promotion by dietary 13-cis-retinoic acid and {alpha}-difluoromethylornithine

The effects of dietary supplementation of 13-cis-retinoic acid(13-cis-RA) and -difluoromethylornithine (DFMO) in the drinkingwater on 12-O-tetradecanoylphorbol-13-acetate (TPA)-promotedskin tumor formation was determined. Administration of 13-cis-RAin the diet and DFMO in the drinking water was started 1 weekand 2 days before the first TPA application to the dimethylbenz[a]anthracene-initiatedskin of either female CD-I or SENCAR mice, respectively. Dietary13-cis-RA failed to inhibit both the tumor yield and the incidence;papillomas per mouse at 0, 5, 50, 100 and 200 mg/kg diet 13-cis-RAdoses were 25, 30, 22, 28 and 25 respectively at 18 weeks ofpromotion treatment and at all doses 100% of the mice bore papillomas.However, dietary 13-cis-RA dramatically reduced the size ofskin tumor promoted with TPA. 13-cis-RA at doses of 5, 50, 100and 200 mg/kg diet inhibited skin papillomas (> 4 mm diameter)per mouse by 28, 55, 76 and 93%, respectively. Retinoid treatmentdid not affect body weight gains and the survival was more than80% in all groups. In accord with our previous findings, DFMOwhen given in drinking water, was a very effective inhibitorof mouse skin tumor promotion by TPA; DFMO at 0.25% concentrationinhibited the number of papillomas by 50%. Inhibition of skintumor promotion by combined treatments with dietary 13-cis-RA(100 mg/kg) and DFMO (0.25%) in the drinking water was possiblyadditive. The retinoid and DFMO preclude TPA-increased ornithinedecarboxylase (ODC) activity and the accumulation of putrescineby differential effects on ODC, an enzyme associated with skintumor promotion by TPA.     

The effects of chronic ethanol administration on polyamine content during dimethylhydrazine-induced colorectal carcinogenesis in the rat

Chronic ethanol (EtOH) consumption has been implicated as aco-carcinogen, selectively promoting rectal tumor formation.We studied the effects of EtOH consumption on tumor formationand polyamine content (putrescine, spermidine and spermine)in proximal and distal colon and rectum of Sprague-Dawley ratstreated with the procarcinogen 1,2-dimethylhydrazine (DMH).Sixty-four adult male rats were pair fed nutritionally completeliquid diets with 36% of calories supplied as EtOH or isocaloriccarbohydrates. Both groups received 4 weeks of the liquid dietfollowed by 4 weeks of standard laboratory chow during which50% of the rats in each group received DMH (30 mg/kg) or vehicles.c. weekly. This cycle was repeated four times (32 weeks).Animals were sacrificed at the end of each 8 week cycle andnormal appearing and available tumor bearing tissue from proximaland distal colon and rectum was obtained for polyamine contentand histology. Five animals, unexposed to DMH or EtOH servedas baseline controls. There were no consistent regional differencesin putrescine, spermidine or spermine of baseline controls.A progressive decrease in tissue putrescine was seen in allthree regions of the control group and was significant at 24and 32 weeks versus baseline controls. In all three regions,chronic EtOH consumption prevented the decrease in tissue putrescine.Spermidine content was also significantly increased in the distalcolon of EtOH-treated animals compared to baseline values. Consistentchanges in spermine content were seen in any treatment groupor region. A significant increase in putrescine content of normalappearing and tumor-bearing tissue of DMH treated animals at32 weeks was noted. Chronic EtOH administration did not augmentrectal or colonic polyamine content in DMH-treated animals.Likewise, chronic EtOH consumption did not alter the number,size or distribution of large bowel tumors of DMH treated animals.     

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.     

Evaluation of chemopreventive effect of dietary selenium-rich egg on mouse skin tumor induced by 2'-(4-nitrophenoxy)oxirane and 12-O-tetradecanoylphorbol-13-acetate

Chemopreventive effect of dietary selenium (as sodium seleniteor as Se-rich egg) on mouse skin tumor induced by topical applicationof 2'-(4-nitrophenoxy)oxirane (NPO) as tumor initiator and 12-O-tetradecanoylphorbol-13-acet-ate(TPA) as tumor promoter was evaluated in relation to the dietarysource and levels of selenium. Selenium supplementation (0.3p.p.m.) to the basal diet (0.07 p.p.m. Se) as sodium seleniteor as Se-rich egg brought about a 40 or 37% reduction respectively,in the incidence of papilloma formation at 12 weeks after NPOtreatment. Tumor yield (number of papillomas per mouse) at 14weeks after NPO treatment in the basal diet group, basal dietsupplemented with 0.3 p.p.m. Se as sodium selenite group andbasal diet supplemented with 0.3 p.p.m. Se as Se-rich egg groupwere 7.5 ± 2.1, 2.7 ± 2.3 and 4.1 ± 3.5respectively. Dietary supplementation of 1.0 p.p.m. Se as Se-richegg to the basal diet reduced the incidence and the multiplicityof papillomas during the early phase of promotion (11 weeks)but its antitumor activity decreased thereafter, indicatingthat the accumulation of tissue selenium above the saturatedlevel may not be beneficial. Selenium concentrations in blood,liver and skin tissue of mice in basal diet group (0.33 ±0.02, 0.54 ± 0.10 and 0.21 ± 0.03 p.p.m. respectively)increased significantly (P<0.05) by the supplementation of0.3 p.p.m. Se as selenite (0.58 ± 0.02, 1.17 ±0.10 and 0.31 ± 0.05 p.p.m. respectively), 0.3 p.p.m.Se as Se-rich egg (0.59 ± 0.02, 1.25 ± 0.11 and0.33 ± 0.06 p.p.m. respectively) and 1.0 p.p.m. Se asSe-rich egg (1.20 ± 0.05, 2.32 ± 0.28 and 0.51± 0.01 p.p.m. respectively). Glutathione peroxidase activityin blood of mice of the basal diet group (2.4 ± 0.4 EU/mg) increased significantly (P<0.05) by dietary seleniumsupplementation (3.8 ± 0.6 EU/mg in 0.3 p.p.m. Selenite;3.7 ± 0.8 EU/mg in 0.3 p.p.m. Se as Se-rich egg; 4.9±0.9 EU/mg in 1.0 p.p.m. Se as Se-rich egg) and the enzymeactivities in liver and skin tissue also increased by 0.3 p.p.m.Se (as selenite or Se-rich egg) supplementation, but no furtherincreases in their activities were obtained by 1.0 p.p.m. Se(as Se-rich egg). It is, therefore, concluded that a moderatelevel of dietary selenium (0.3 p.p.m.) has an efficient chemopreventiveactivity at the promotional stage of carcinogenesis and thatdietary selenium rich egg as well as dietary selenite exertedantitumor activity.     

Polyamine-directed preferential nutritional repletion of normal tissues in tumor-bearing hosts

Our previous study demonstrated that total parenteral nutrition (TPN) increases erythrocyte (RBC) polyamines selectively in cancer patients but not in non-cancer patients, suggesting that these changes may relate to tumor presence. We therefore studied the possible relationship between RBC polyamine levels and potential tumor growth induced by TPN. In addition, we attempted to control tumor growth during TPN with alpha-difluoromethylornithine (DFMO), a suicide inhibitor of ornithine decarboxylase (ODC), in a fibrosarcoma-bearing rat model. Rats in a control group received chow ad libitum for 3 weeks and their tumors grew exponentially. A restricted intake (RI) tumor-bearing group received 8 g chow per day per rat; this produced malnourished rats in which tumor growth significantly slowed. The third group (RI----TPN) received the RI diet for 2 weeks and TPN for 1 week, which resulted in an increase in tumor growth. The last group (RI----TPN + DFMO) received a 2-week RI regimen and 1-week TPN plus DFMO regimen, which inhibited tumor growth to the level of the RI group due to the addition of DFMO (1 g/kg/day). Although the addition of TPN to the RI regimen markedly increased tumor ODC activity, as well as tumor putrescine and RBC putrescine levels, the addition of DFMO to the RI----TPN regimen entirely abolished these increases. Despite DFMO treatment, liver ODC activity in RI----TPN + DFMO group was unaffected. The plasma albumin level, an indicator of host nutrition, of the RI----TPN + DFMO group was significantly higher than that of the RI group. Liver, spleen, kidney and total body weights of the RI----TPN or RI----TPN + DFMO group were significantly higher than those of the RI group. These results indicate that TPN can enhance tumor growth in malnourished hosts and that the addition of DFMO to TPN can effectively control this TPN-enhanced tumor growth, leading to preferential nutritional repletion of the host.     

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.     

Supplemental dietary calcium fails to alter the acute effects of 1,2-dimethylhydrazine on O6-methylguanine, O6-alkylguanine-DNA alkyltransferase and cellular proliferation in the rat colon

Prior studies from our laboratory have demonstrated that K-rasG to A mutations were detectable in a high percentage of carcinomaswhich developed in the colons of animals treated with the knowncolonic procarcinogen, 1,2-dimethyl-hydrazine (DMH). Moreover,in this model, the incidence of these mutations was decreasedby a supplemental dietary calcium regimen which concomitantlydecreased the frequency of rats with multiple tumors as wellas tumor size. In an attempt to clarify the possible mechanism(s)involved in this antimutagenic effect of supplemental calcium,two groups of Sprague—Dawley rats were fed semisyntheticdiets containing either 0.87 or 1.80% cakium by weight for 3weeks, s.c. injected with 100 mg/kg of DMH and killed priorto and at various time periods (16-144 h) after injection. Thecolons of animals were analyzed and compared with respect toO⁶-methylguanine content in DNA, O⁶-alkylguanine-DNA alkyltransferaselevels as well as cellular proliferation, as assessed by immunohistochemicalstaining of colonic crypts by bromodeoxyundine. In certain experiments,these parameters were also analyzed in the proximal and distalcolon before and at various times after administration of DMH.The results of these experiments demonstrated that supplementaldietary calcium was not found to influence significantly O⁶-methylguaninelevels, alkyltransferase levels or cellular proliferation inthe entire colon or in either colonic segment before or afterthe acute administration of DMH. DMH did, however, differentiallyalter all three of these biochemical parameters in the colonicsegments (distal > proximal), possibly due to a greater degreeof metabolic activation in the distal colon.