Supplementation with Brazil nuts and green tea extract regulates targeted biomarkers related to colorectal cancer risk in humans.

Se and green tea have been shown in epidemiological, observational and preclinical studies to be inversely related to the risk of developing colorectal cancer (CRC). However, there are limited studies to evaluate their regulatory effects on genes/proteins that relate to CRC oncogenesis in human subjects, such as selenoproteins, WNT signalling pathway, inflammation and methylation. This study examined the effects of supplementation of Se using Brazil nuts and green tea extract (GTE) capsules, alone and in combination, on targeted biomarkers. In total, thirty-two volunteers (>50 years of age) with plasma Se≤1·36 µmol/l were randomised to one of three treatment groups: nine to Se (approximately 48 µg/d) as six Brazil nuts, eleven to four GTE capsules (800 mg (-)-epigallocatechin-3-gallate) and twelve to a combination of Brazil nuts and GTE. Blood and rectal biopsies were obtained before and after each intervention. Plasma Se levels, rectal selenoprotein P (SePP) and ß-catenin mRNA increased significantly in subjects consuming Brazil nuts alone or in combination, whereas rectal DNA methyltransferase (DNMT1) and NF-κB mRNA were reduced significantly in subjects consuming GTE alone or in combination. None of the interventions significantly affected rectal acetylated histone H3 or Ki-67 expression at the protein level or plasma C-reactive protein. Effects of the combination of Brazil nuts and GTE did not differ from what would be expected from either agent alone. In conclusion, supplementation of Brazil nuts and/or GTE regulates targeted biomarkers related to CRC oncogenesis, specifically genes associated with selenoproteins (SePP), WNT signalling (ß-catenin), inflammation (NF-κB) and methylation (DNMT1). Their combination does not appear to provide additional effects compared with either agent alone.

Combination of selenium and green tea improves the efficacy of chemoprevention in a rat colorectal cancer model by modulating genetic and epigenetic biomarkers.

Dietary supplementation of selenium and green tea holds promise in cancer prevention. In this study, we evaluated the efficacies of selenium and green tea administered individually and in combination against colorectal cancer in an azoxymethane (AOM)-induced rat colonic carcinogenesis model and determined the underlying mechanisms of the protection. Four-week old Sprague-Dawley male rats were fed with diets containing 0.5% green tea extract, 1 ppm selenium as selenium-enriched milk protein, or combination of 1 ppm selenium and 0.5% green tea extract. Animals received 2 AOM (15 mg/kg) treatments to induce colonic oncogenesis. Rats were killed 8 or 30 wk later after the last AOM to examine the effect of dietary intervention on aberrant crypt foci (ACF) formation or tumor development. On sacrifice, colons were examined for ACF and tumors, the mRNA levels of SFRP5 and Cyclin D1, and the proteins levels of ß-catenin, COX-2, Ki-67, DNMT1 and acetyl histone H3. The combination of selenium and green tea resulted in a significant additive inhibition of large ACF formation, this effect was greater than either selenium or green tea alone, P<0.01; the combination also had a significant additive inhibition effect on all tumor endpoints, the effect of the combination diet on tumor incidence, multiplicity and size was greater than selenium or green tea alone, P<0.01. Rats fed the combination diet showed marked reduction of DNMT1 expression and induction of histone H3 acetylation, which were accompanied by restoration of SFRP5 mRNA in normal-appearing colonic crypts. The combination diet also significantly reduced ß-catenin nuclear translocation, Cyclin D1 expression and cell proliferation. These data show, for the first time, that combination of selenium and green tea is more effective in suppressing colorectal oncogenesis than either agent alone. The preventive effect is associated with regulation of genetic and epigenetic biomarkers implicated in colonic carcinogenesis.

Selenium-rich foods: a promising approach to colorectal cancer prevention.

Selenium (Se), an essential trace element, has also been identified as an anticarcinogenic agent, with supporting evidence from epidemiological studies, clinical intervention trials, preclinical intervention studies (animal cancer models) and cell culture studies. Natural organic and inorganic sources of Se as well as synthetic organoselenium compounds have been shown to be effective; safety and efficacy factors favour the organic forms. Intakes that are several fold that purported to meet nutritional requirements (adult recommend dietary allowance--55 µg Se/day) are associated with reductions in cancer risk, but are not currently met by most diets, unless Se-rich foods are included. Further clinical studies and development of tools for speciating Se in foods will enable progress to be made in determining desirable Se forms and foods with respect to providing safe and effective ways of reducing cancer risk.

The influence of selenium-enriched milk proteins and selenium yeast on plasma selenium levels and rectal selenoprotein gene expression in human subjects.

Certain forms of dietary Se may have advantages for improving human Se status and regulating the risk for disease, such as cancers, including colorectal cancer (CRC). The present study compared the effects of a Se-enriched milk protein (dairy-Se) with a Se-rich yeast (yeast-Se) on plasma Se levels and rectal selenoprotein gene expression since we reasoned that if these genes were not regulated, there was little potential for regulating the risk for CRC in this organ. A total of twenty-three healthy volunteers with plasma Se in the lower half of the population range were supplemented with dairy-Se (150 µg/d) or yeast-Se (150 µg/d) for 6 weeks, followed by 6 weeks of washout period. Blood was sampled every 2 weeks, and rectal biopsies were obtained before and after Se supplementation and after the washout period. Plasma Se levels and glutathione peroxidase (GPx) activity, and rectal mRNA of selenoprotein P (SeP), cytosolic GPx-1 (GPx-1), gastrointestinal GPx-2 (GPx-2) and thioredoxin reductase-1 (TrxR-1) were measured. Plasma Se levels increased rapidly in both Se groups (P < 0·001); plasma GPx activity was not significantly changed. Rectal SeP mRNA increased at 6 weeks compared with baseline in both Se groups (P < 0·05); only dairy-Se resulted in a sustained elevation of SeP after the washout period (P < 0·05). Rectal GPx-1 and GPx-2 mRNA were higher with dairy-Se (P < 0·05) than with yeast-Se at 6 weeks. In conclusion, three rectal selenoprotein mRNA were differentially regulated by dairy-Se and yeast-Se. Changes in rectal selenoproteins are not predicted by changes in plasma Se; dairy-Se effectively regulates the expression of several rectal selenoproteins of relevance to the risk for CRC.

Selenium-enriched milk proteins and selenium yeast affect selenoprotein activity and expression differently in mouse colon.

Certain forms of dietary Se may have an advantage in improving Se status and reducing cancer risk. The present study compared the effects of an Se-enriched milk protein product (dairy-Se) with an Se yeast (yeast-Se) on selenoprotein activity and expression in the mouse colon. Mice were fed four diets for 4 weeks: a control milk protein diet (Se at 0.068 parts per million (ppm)), dairy-Se diets with Se at 0.5 and 1 ppm, and a yeast-Se diet with Se at 1 ppm. Cytosolic glutathione peroxidase-1 (GPx-1) activity, mRNA of selenoprotein P (SeP), GPx-1, gastrointestinal glutathione peroxidase-2 (GPx-2) and thioredoxin reductase-1 (TrxR-1) were examined in the mouse colon. Dairy-Se diets did not significantly affect GPx-1 mRNA and GPx-1 activity but produced a dose-dependent increase in SeP and GPx-2 mRNA, with a significantly higher level achieved at 1 ppm Se (P < 0.05). Yeast-Se at 1 ppm significantly increased GPx-1 mRNA and GPx-1 activity (P < 0.01) but not GPx-2 mRNA. Neither Se supplement had any effect on TrxR-1. The present study indicates that selenoprotein levels in the mouse colon are regulated differently depending on the Se supplement. As we have previously shown that dairy-Se at 1 ppm was protective against colorectal cancer (CRC) in an azoxymethane-induced CRC mouse model, this up-regulation of colonic GPx-2 and SeP with Se supplementation may be crucial to its chemopreventive action.

Suppression of colorectal oncogenesis by selenium-enriched milk proteins: apoptosis and K-ras mutations.

The chemical form and bioavailability of dietary selenium may influence its protectiveness against colorectal cancer. Selenium is readily incorporated into milk proteins by feeding cows with selenized-yeast. This study examined whether a dairy source of organic selenium (as milk proteins) is more effective than a yeast source at inhibiting oncogenesis in carcinogen-treated mice and whether it regulates the homeostatic response to carcinogen-induced DNA damage. Dietary interventions are as follows: selenium-enriched milk protein isolate (Tatura-Bio Se; 0.5 or 1 ppm selenium) or milk protein control and selenized-yeast (Sel-Plex; 1 or 4 ppm selenium) with casein or casein alone as control. After 4 weeks on diet, mice received a single azoxymethane (10 mg/kg) injection to induce mutations and were killed 6 hours later. Measures were as follows: plasma selenium, cell proliferation, and acute apoptotic response to azoxymethane (AARGC). Separate groups of mice on the same diets were given 4 weekly azoxymethane (15 mg/kg) injections to induce oncogenesis. Mice were killed 6 or 30 weeks after the last azoxymethane injection. Measures were as follows: aberrant crypt foci (ACF), cancers, and K-ras mutations. Dairy-selenium at 1 ppm significantly suppressed ACF and cancers, whereas yeast-selenium at an equivalent selenium intake had no effect. Dairy-selenium significantly increased plasma selenium levels and AARGC, and reduced cell proliferation and frequency of K-ras mutations in ACF relative to an equivalent dose of selenium from yeast. Selenium-enriched milk protein isolate is superior to selenized-yeast in terms of its bioavailability and capacity to suppress oncogenesis. Suppression may be a consequence of enhanced apoptotic deletion of azoxymethane-induced DNA lesions and the subsequent reduction in frequency of K-ras mutations.

Whey protein isolate and glycomacropeptide decrease weight gain and alter body composition in male Wistar rats.

The effect of feed protein type on body composition and growth has been examined. Evidence exists that whey protein concentrate is effective at limiting body fat expansion. The presence of caseinomacropeptide, a mixture of glycosylated and non-glycosylated carbohydrate residues, in particular glycomacropeptide (GMP) in whey protein concentrate may be important for this effect. The influence of whey protein isolate (WPI) and GMP on weight gain and body composition was examined by feeding Wistar rats ad libitum for 7 weeks with five semi-purified American Institute of Nutrition-based diets differing in protein type: (1) casein; (2) barbequed beef; (3) control WPI (no GMP); (4) WPI+GMP at 100 g/kg; (5) WPI+GMP at 200 g/kg. Body composition was assessed, and plasma samples were assayed for TAG, insulin and glucose. Body-weight gain was lower (- 21 %) on the control WPI diet relative to casein, with a non-significant influence associated with GMP inclusion (- 30 %), the effect being equivalent at both levels of GMP addition. Renal and carcass fat mass were reduced in the highest GMP diet when compared with WPI (P < 0.05). Plasma insulin was lowered by GMP at the highest addition compared with WPI alone (- 53 %; P < 0.01). Plasma TAG in the WPI+GMP (200 g/kg) group were lower (- 27 %; P < 0.05) than the casein and beef groups. In conclusion, GMP appears to have a significant additional influence when combined with WPI on fat accumulation. WPI alone appears to have the predominant influence accounting for 70 % of the overall effect on body-weight gain. Mechanisms for this effect have not been identified but food intake was not responsible.

Selenised casein protects against AOM-induced colon tumors in Sprague Dawley rats.

Selenium (Se) has been shown to be protective against cancers in animal models at concentrations exceeding those considered essential for normal nutritional requirements. Organic forms of Se provided as dairy proteins were obtained from cows fed diets supplemented with yeast Se. The casein extracted from milk was found to contain approximately half the Se of the Se-enriched milk. This casein was included in a semi-purified AIN rodent diet so as to provide 1 ppm Se and 25% protein and was compared with AIN diets containing no added Se (control, 0.05 ppm), 1 ppm and 4 ppm Se as selenised yeast (Sel-Plex) Their influence on colon tumor expression was examined in rats induced with azoxymethane, the diets being introduced post-induction. The selenised casein diet at this concentration was effective in reducing colon tumor incidence (by 29%) and burden (decreased 52%, P < 0.05) relative to the control in rats 26 wk post-induction. Selenised yeast, when added at similar (1 ppm) and increased Se concentration (4 ppm), did not influence significantly colon tumor expression. However, in a second study, with Se yeast providing Se at 1 ppm, 4 ppm, and 8 ppm throughout the experiment, a significant reduction in tumors was observed with 8 ppm Se (colon tumor incidence was 15% lower and colon tumor burden was 35% lower, P < 0.05). However this was associated with a significantly lower body weight in the rats (down 10.5%, P < 0.05) indicating a possible disturbance with normal energy intake or metabolism. The form in which Se is presented in the diet may influence significantly its bioavailability and/or anticancer potential at given concentrations within a safe range. The efficacy of selenised casein and indeed other potential dietary sources deserve further investigation with regard to their ability to prevent colon tumors at concentrations considered safe in the diet.

A high-whey-protein diet reduces body weight gain and alters insulin sensitivity relative to red meat in wistar rats.

A high-protein diet can reduce body weight and increase insulin sensitivity, but whether the type of dietary protein affects these outcomes is unknown. We hypothesized that feeding insulin-resistant rats a high-protein diet (32%) containing whey protein concentrate (WPC) would reduce body weight and tissue lipid levels and increase insulin sensitivity more than a diet containing red meat (RM). Rats were fed a high-fat diet (300 g fat/kg diet) for 9 wk, then switched to a diet containing either 80 or 320 g protein/kg diet, provided by either WPC or RM, for 6 wk (n = 8). The rats were then killed after overnight food deprivation. High dietary protein reduced energy intake (P < 0.001) and visceral (P < 0.001), subcutaneous (P < 0.001), and carcass fat (P < 0.05). Increasing the dietary density of WPC, but not of RM, reduced body weight gain by 4% (P < 0.001). Dietary WPC also reduced plasma insulin concentration by 40% (P < 0.05) and increased insulin sensitivity, compared to RM (P < 0.05). These findings support the conclusions that a high-protein diet reduces energy intake and adiposity and that whey protein is more effective than red meat in reducing body weight gain and increasing insulin sensitivity.

Antitumor activity of extract of Zingiber aromaticum and its bioactive sesquiterpenoid zerumbone.

The anticancer properties of zerumbone (2,6,9 humulatriene-8-one, a sesquiterpenoid) from Zingiber aromaticum were compared with those of curcumin from Curcuma longa in an in vitro MTT tetrazolium salt assay using HT-29, CaCo-2, and MCF-7 cancer cells and in an azoxymethane (AOM)-induced animal model of colon cancer using aberrant crypt foci (ACFs) as a preneoplastic marker. The IC50 of zerumbone was approximately 10 mM and that of curcumin was 25 mM. Cell cycle arrest in HT-29 cells was observed at G0/G1 for 10 and 12.5 mM and G2/M for 25 mM after 24 h at concentrations of 10-25 mM of zerumbone, and a concentration-dependent increase in apoptosis (2-6% of viable cells) was observed after 48 h using the same concentration range. Male Sprague-Dawley rats were fed extracts in an AIN diet prepared from the equivalent of 4% by weight of dried rhizomes of Z. aromaticum and C. longa. ACFs were induced by two doses (15 mg/kg body weight) subcutaneously of AOM 1 wk apart, the rats were killed 10 wk later, and the ACFs were assessed in the colon. Total ACFs were significantly reduced by Z. aromaticum extract (down 21%, P < 0.05) relative to control, the effect being most evident with large ACFs (>3 aberrant crypts per focus). Similar reductions were observed with 4% C. longa extract in the diet (down 24%, P < 0.01) and with 2,000 ppm curcumin, the effect being particularly evident with large ACFs. The concentration of zerumbone in the Z. aromaticum extract diet was assayed at 300 ppm and of curcumin in the C. longa extract diet was also 300 ppm, i.e., the extract of C. longa was as effective at one-seventh the concentration of curcumin as the positive control. Zerumbone is effective as an anticancer agent, possibly by its apoptosis-inducing and antiproliferative influences. This latter possibility is currently being investigated.

Whole-grain rye and wheat foods and markers of bowel health in overweight middle-aged men.

BACKGROUND: Whole-grain cereal foods including rye have been identified as providing significant health benefits that do not occur when refined-cereal foods are ingested. OBJECTIVES: Foods (90 g) containing whole-grain rye flour and whole-grain wheat flour were compared with low-fiber refined-cereal foods for their effects on markers of bowel health and the metabolic markers insulin and glucose. DESIGN: Three 4-wk interventions were undertaken in a randomized crossover design with 28 overweight men aged 40-65 y who had no history of bowel disease. Against a background intake of 14 g dietary fiber (DF), the men were fed low-fiber cereal grain foods providing 5 g DF for a total of 19 g DF/d. High-fiber wheat foods provided 18 g DF, and high-fiber rye foods provided 18 g DF, both giving a total of 32 g DF/d. Fecal samples (48-h) and fasting and postprandial blood samples were collected at the end of each period and assayed. RESULTS: Both high-fiber rye and wheat foods increased fecal output by 33-36% (P = 0.004) and reduced fecal beta-glucuronidase activity by 29% (P = 0.027). Postprandial plasma insulin was decreased by 46-49% (P = 0.0001) and postprandial plasma glucose by 16-19% (P = 0.0005). Rye foods were associated with significantly (P = 0.0001) increased plasma enterolactone (47% and 71%) and fecal butyrate (26% and 36%), relative to wheat and low-fiber options, respectively. CONCLUSIONS: High-fiber rye and wheat food consumption improved several markers of bowel and metabolic health relative to that of low-fiber food. Fiber from rye appears more effective than that from wheat in overall improvement of biomarkers of bowel health.

A diet containing alpha-cellulose and fish oil reduces aberrant crypt foci formation and modulates other possible markers for colon cancer risk in azoxymethane-treated rats.

There is a need for better understanding of the roles of dietary fats and fibers in colon cancer risk. We examined the effect of different dietary fiber and fat sources on an azoxymethane (AOM)-induced colon cancer in rats. In a 2 x 3 factorial design, rats were fed a semipurified diet containing soy-derived fiber (Fibrim), alpha-cellulose (Solkafloc) or resistant starch (RS; Hi-maize) at 10 g dietary fiber/100 g diet, combined with fish oil (FO) or sunflower seed oil (SSO) at 10 g/100 g diet, and lard added to all diets at 10 g/100 g, to provide a total of 20 g mixed fat/100 g diet. Sprague-Dawley rats (28 d of age) consumed diets for 4 wk and then two doses of AOM (15 mg/kg body) were administered 1 wk apart by subcutaneous injection. Rats were killed after 13 wk of consuming experimental diets. Colons were fixed in formalin and aberrant crypt foci (ACF) were quantified after staining. ACF counts were higher (+66%, P < 0.01) in rats fed SSO and RS, than in those fed alpha-cellulose and FO. Rats fed FO had 19% fewer ACF than those fed SSO (P < 0.05). alpha-Cellulose was associated with the highest cecal butyrate concentration (P < 0.001), the highest beta-glucuronidase specific activity (P < 0.001) and the lowest cecal water cytotoxicity (P < 0.001) relative to soy fiber- and RS-fed rats. There were inverse correlations between the number of ACF and cecal butyrate concentration (r = -0.33, P < 0.05) and between cecal water cytotoxicity and beta-glucuronidase activity (r = -0.70, P < 0.001). The greatest protection was associated with alpha-cellulose as the fiber source and FO as the fat source as measured by colon ACF numbers in rats.