Effects of partial replacement of red and processed meat with non-soya legumes on bone and mineral metabolism and amino acid intakes in BeanMan randomised clinical trial.

The transition towards more plant-based diets may pose risks for bone health such as low vitamin D and Ca intakes. Findings for the contribution of animal and plant proteins and their amino acids (AA) to bone health are contradictory. This 6-week clinical trial aimed to investigate whether partial replacement of red and processed meat (RPM) with non-soya legumes affects AA intakes and bone turnover and mineral metabolism in 102 healthy 20-65-year-old men. Participants were randomly assigned to diet groups controlled for RPM and legume intake (designed total protein intake (TPI) 18 E%): the meat group consumed 760 g RPM per week (25 % TPI) and the legume group consumed non-soya legume-based products (20 % TPI) and 200 g RPM per week, the upper limit of the Planetary Health Diet (5 % TPI). No differences in bone (bone-specific alkaline phosphatase; tartrate-resistant acid phosphatase 5b) or mineral metabolism (25-hydroxyvitamin D; parathyroid hormone; fibroblast growth factor 23; phosphate and Ca) markers or Ca and vitamin D intakes were observed between the groups (P > 0·05). Methionine and histidine intakes were higher in the meat group (P ≤ 0·042), whereas the legume group had higher intakes of arginine, asparagine and phenylalanine (P ≤ 0·013). Mean essential AA intakes in both groups met the requirements. Increasing the proportion of non-soya legumes by reducing the amount of RPM in the diet for 6 weeks did not compromise bone turnover and provided on average adequate amounts of AA in healthy men, indicating that this ecologically sustainable dietary change is safe and relatively easy to implement.

Partial substitution of red or processed meat with plant-based foods and the risk of type 2 diabetes.

High consumption of red and processed meat has been associated with increased type 2 diabetes (T2D) risk. These kinds of diets are also environmentally unsustainable. We examined a modeled association between a partial substitution of red meat or processed meat with plant-based foods (legumes, vegetables, fruit, cereals, or a combination of these) and T2D risk among Finnish adults. We used pooled data from five Finnish cohorts (n = 41,662, 22% women, aged ≥ 25 years, 10.9 years median follow-up with 1750 incident T2D cases). Diet was assessed by a validated food frequency questionnaire. In the substitution models, 100 g/week of red meat or 50 g/week of processed meat were substituted with similar amounts of plant-based substitutes. Cohort-specific hazard ratios (HRs) were estimated by Cox proportional hazards multivariable model and pooled using a two-staged random-effects model. We observed small, but statistically significant, reductions in T2D risk in men when red or processed meat were partially substituted with fruits (red meat: HR 0.98, 95% CI 0.97-1.00, P = 0.049, processed meat: 0.99, 0.98-1.00, P = 0.005), cereals (red meat: 0.97, 0.95-0.99, P = 0.005, processed meat: 0.99, 0.98-1.00, P = 0.004) or combination of plant-based foods (only processed meat: 0.99, 0.98-1.00, P = 0.004) but not with legumes or vegetables. The findings of women were similar but not statistically significant. Our findings suggest that even small, easily implemented, shifts towards more sustainable diets may reduce T2D risk particularly in men.

Partial replacement of red and processed meat with legumes: a modelling study of the impact on nutrient intakes and nutrient adequacy on the population level.

OBJECTIVE: The shift towards plant-based diets with less meat and more legumes is a global target and requires an understanding of the consequences of dietary adequacy on the population level. Our aim was to model the impact of partial replacement of red and processed meat with legumes on nutrient intakes and population shares below dietary reference intakes. DESIGN: Modelling study with three scenarios anchored in meat cut-offs: ≤ 70 g/d (Finnish dietary guideline); ≤ 50 g/d (Danish dietary guideline); ≤ 30 g/d (EAT-Lancet recommendation). In all subjects, the amount of meat in grams over the cut-off was replaced with the same amount of legumes. The SPADE method was used to model usual intake distributions. Meaningful differences in average intakes and in population shares below dietary reference intakes compared to the reference (FinDiet) were evaluated based on non-overlapping 95 % CI. SETTING: Finnish national food consumption survey (FinDiet 2017). SUBJECTS: Finnish adults (n 1655) aged18-74 years (47 % men). RESULTS: The scenarios introduced increases in the average intakes of fibre, folate, K, Mg, Cu and Fe, and decreases in intakes of saturated fat, niacin, vitamin B12, Se and Zn. Meaningful shifts of the usual intake distributions of fibre and folate towards improvement in intakes emerged already in 'scenario 70 g'. Overall, distribution shifts towards a higher probability of inadequate intakes of the studied nutrients were not observed. CONCLUSIONS: These results support the public health message to partly replace meat with legumes and may benefit nutrition policy actions towards sustainable diets in the Nordic countries and beyond.

Replacing dietary animal-source proteins with plant-source proteins changes dietary intake and status of vitamins and minerals in healthy adults: a 12-week randomized controlled trial.

PURPOSE: A shift towards more plant-based diets promotes both health and sustainability. However, controlled trials addressing the nutritional effects of replacing animal proteins with plant proteins are lacking. We examined the effects of partly replacing animal proteins with plant proteins on critical vitamin and mineral intake and statuses in healthy adults using a whole-diet approach. METHODS: Volunteers aged 20-69 years (107 female, 29 male) were randomly allocated into one of three 12-week intervention groups with different dietary protein compositions: ANIMAL: 70% animal-source protein/30% plant-source protein; 50/50: 50% animal/50% plant; PLANT: 30% animal/70% plant; all with designed protein intake of 17 E%. We analysed vitamin B-12, iodine, iron, folate, and zinc intakes from 4-day food records, haemoglobin, ferritin, transferrin receptor, folate, and holotranscobalamin II from fasting blood samples, and iodine from 24-h urine. RESULTS: At the end point, vitamin B-12 intake and status were lower in PLANT than in 50/50 or ANIMAL groups (P ≤ 0.007 for all). Vitamin B-12 intake was also lower in 50/50 than in ANIMAL (P < 0.001). Iodine intake and status were lower in both 50/50 and PLANT than in ANIMAL (P ≤ 0.002 for all). Iron and folate intakes were higher in PLANT than in ANIMAL (P < 0.001, P = 0.047), but no significant differences emerged in the respective biomarkers. CONCLUSIONS: Partial replacement of animal protein foods with plant protein foods led to marked decreases in the intake and status of vitamin B-12 and iodine. No changes in iron status were seen. More attention needs to be paid to adequate micronutrient intakes when following flexitarian diets. CLINICAL TRIAL REGISTRY: NCT03206827; registration date: 2017-06-30.

Partial Replacement of Animal Proteins with Plant Proteins for 12 Weeks Accelerates Bone Turnover Among Healthy Adults: A Randomized Clinical Trial.

BACKGROUND: Plant-based diets may reduce the risk of chronic diseases, but can also lead to low calcium and vitamin D intakes, posing a risk for bone health. OBJECTIVES: We investigated whether partial replacement of animal proteins with plant-based proteins using a whole-diet approach affects bone and mineral metabolism in healthy adults in 3 groups fed diets differing in protein composition. METHODS: This 12-week clinical trial was comprised of 107 women and 29 men (20-69 years old; BMI mean ± SD, 24.8 ± 3.9) randomly assigned to consume 1 of 3 diets designed to provide 17 energy percent (E%) protein: "animal" (70% animal protein, 30% plant protein of total protein intake), "50/50" (50% animal, 50% plant), and "plant" (30% animal, 70% plant) diets. We examined differences in bone formation [serum intact procollagen type I amino-terminal propeptide (S-iPINP)], bone resorption [serum collagen type 1 cross-linked C-terminal telopeptide (S-CTX)], mineral metabolism markers (primary outcomes), and nutrient intakes (secondary outcomes) by ANOVA/ANCOVA. RESULTS: S-CTX was significantly higher in the plant group (mean ± SEM, 0.44 ± 0.02 ng/mL) than in the other groups (P values < 0.001 for both), and differed also between the animal (mean ± SEM, 0.29 ± 0.02 ng/mL) and 50/50 groups (mean ± SEM, 0.34 ± 0.02 ng/mL; P = 0.018). S-iPINP was significantly higher in the plant group (mean ± SEM, 63.9 ± 1.91 ng/mL) than in the animal group (mean ± SEM, 55.0 ± 1.82 ng/mL; P = 0.006). In a subgroup without a history of vitamin D supplement use, plasma parathyroid hormone was significantly higher in the plant than in the animal group (P = 0.018). Vitamin D and calcium intakes were below recommended levels in the plant group (mean ± SEM, 6.2 ± 3.7 µg/d and 733 ± 164 mg/d, respectively). CONCLUSIONS: Partial replacement of animal proteins with plant-based proteins for 12 weeks increased the markers of bone resorption and formation among healthy adults, indicating a possible risk for bone health. This is probably caused by lower vitamin D and calcium intakes from diets containing more plant-based proteins, but it is unclear whether differences in protein intake or quality play a major role. This trial was registered at clinicaltrials.gov as NCT03206827.

Replacing Animal-Based Proteins with Plant-Based Proteins Changes the Composition of a Whole Nordic Diet-A Randomised Clinical Trial in Healthy Finnish Adults.

Increased consumption of plant-based foods and decreased consumption of animal-based foods is recommended for healthy diets and sustainable food production. We investigated the effects of partial replacement of dietary animal proteins with plant-based ones on intake of energy-yielding nutrients, fibre, and plasma lipoproteins. This 12-week randomised clinical intervention comprised 107 women and 29 men (20-69 years) in three diet groups with different dietary protein compositions ("ANIMAL": Animal 70%/plant 30%; "50/50": Animal 50%/plant 50%; "PLANT": Animal 30%/plant 70%; all: Protein intake 17 E%). Nutrient intakes were assessed by 4-day food records. Saturated fat intake (E%) was lower and polyunsaturated fatty acid intake (E%) higher in the PLANT and 50/50 groups compared to the ANIMAL group (p < 0.001 for all). Fibre intake was higher in the PLANT (p ˂ 0.001) and 50/50 (p = 0.012) groups. Total and LDL cholesterol were lower in the PLANT than in the ANIMAL group (p = 0.003 for both) but no differences in HDL cholesterol or triglycerides were observed (p > 0.05). Replacing animal protein with plant protein sources in the diet led to an increased fibre intake and improved dietary fat quality as well as blood lipoprotein profile. Flexitarian diets could provide healthy and more sustainable alternatives for the current, predominantly animal-based diets.

Western diet enhances intestinal tumorigenesis in Min/+ mice, associating with mucosal metabolic and inflammatory stress and loss of Apc heterozygosity.

Western-type diet (WD) is a risk factor for colorectal cancer, but the underlying mechanisms are poorly understood. We investigated the interaction of WD and heterozygous mutation in the Apc gene on adenoma formation and metabolic and immunological changes in the histologically normal intestinal mucosa of ApcMin/+ (Min/+) mice. The diet used was high in saturated fat and low in calcium, vitamin D, fiber and folate. The number of adenomas was twofold higher in the WD mice compared to controls, but adenoma size, proliferation or apoptosis did not differ. The ratio of the Min to wild-type allele was higher in the WD mice, indicating accelerated loss of Apc heterozygosity (LOH). Densities of intraepithelial CD3ε+ T lymphocytes and of mucosal FoxP3+ regulatory T cells were higher in the WD mice, implying inflammatory changes. Western blot analyses from the mucosa of the WD mice showed suppressed activation of the ERK and AKT pathways and a tendency for reduced activation of the mTOR pathway as measured in phosphoS6/S6 levels. The expression of pyruvate dehydrogenase kinase 4 was up-regulated in both mRNA and protein levels. Gene expression analyses showed changes in oxidation/reduction, fatty acid and monosaccharide metabolic pathways, tissue organization, cell fate and regulation of apoptosis. Together, our results suggest that the high-risk Western diet primes the intestine to tumorigenesis through synergistic effects in energy metabolism, inflammation and oxidative stress, which culminate in the acceleration of LOH of the Apc gene.

Changes in intestinal immunity, gut microbiota, and expression of energy metabolism-related genes explain adenoma growth in bilberry and cloudberry-fed ApcMin mice.

We showed previously that ellagitannin-rich cloudberries and anthocyanin-rich bilberries reduce the number of intestinal adenomas in multiple intestinal neoplasia/+ (ApcMin) mice. We also found that cloudberries decreased the size of adenomas, whereas bilberries increased it. Here we hypothesized that the difference in adenoma growth could be explained by dissimilar effects of the berries on intestinal immune responses and gut microbiota, potentially driven by the distinct polyphenol compositions of the 2 berries. Our objectives were to investigate lymphocyte subtypes and the predominant cecal bacterial diversity in mice fed with bilberries and cloudberries, and to analyze global gene expression profiles in the intestinal mucosa. Immunostainings of CD3+ T lymphocytes, FoxP3+ regulatory T lymphocytes, and CD45R+ B lymphocytes revealed a smaller ratio of intraepithelial to all mucosal CD3+ T lymphocytes in the cloudberry-fed mice compared with controls, suggesting an attenuation of inflammation. Bilberry feeding induced no changes in the density of any of the lymphocyte subtypes. The predominant bacterial diversity in cecal contents, analyzed using polymerase chain reaction-denaturating gradient gel electrophoresis, was higher in the bilberry group than in the control or cloudberry groups. The microbial profiles of cloudberry-fed mice clustered together and were associated with small adenoma size. Pathway analyses of gene expression data showed that cloudberry down-regulated and bilberry up-regulated the expression of energy metabolism-related genes in the intestinal mucosa. In conclusion, attenuation of intestinal inflammation, changes in microbial profiles, and down-regulation of mucosal energy metabolism may account for the smaller adenoma size in cloudberry-fed mice in comparison to bilberry-fed mice.

Ellagitannin-rich cloudberry inhibits hepatocyte growth factor induced cell migration and phosphatidylinositol 3-kinase/AKT activation in colon carcinoma cells and tumors in Min mice.

Berries have been found to inhibit colon carcinogenesis in animal models, and thus represent a potential source of compounds for prevention and treatment of colorectal cancer. The mechanistic basis for their effects is not well understood. We used human colon carcinoma cells and Min mice to investigate the effects of ellagitannin-rich cloudberry (Rubus chamaemorus) extract on cancer cell migration and underlying cell signaling. Intrinsic and hepatocyte growth factor (HGF) -induced cell motility in human HT29 and HCA7 colon carcinoma cells was assessed carrying out cell scattering and scratch wound healing assays using time-lapse microscopy. Activation of Met, AKT, and ERK in cell lines and tumors of cloudberry-fed Min mice were determined using immunoprecipitation, Western blot and immunohistochemical analyses. Cloudberry extract significantly inhibited particularly HGF-induced cancer cell migration in both cell lines. Cloudberry extract inhibited the Met receptor tyrosine phosphorylation by HGF and strongly suppressed HGF-induced AKT and ERK activation in both HT29 and HCA7 cells. Consistently, cloudberry feeding (10% w/w freeze-dried berries in diet for 10 weeks) reduced the level of active AKT and prevented phosphoMet localization at the edges in tumors of Min mice. These results indicate that cloudberry reduces tumor growth and cancer cell motility by inhibiting Met signaling and consequent activation of phosphatidylinositol 3-kinase/AKT in vitro and in tumors in vivo. As the Met receptor is recognized to be a major target in cancer treatment, our results suggest that dietary phytochemicals may have therapeutic value in reducing cancer progression and metastasis.

Plant sterol feeding induces tumor formation and alters sterol metabolism in the intestine of Apc(Min) mice.

Dietary plant sterols reduce the absorption of cholesterol and therefore increase intraluminal cholesterol concentration. We examined how plant sterol esters from functional foods affect intestinal tumorigenesis in tumor-prone adenomatous polyposis coli (Apc)(Min) mice. Feeding plant sterols at 0.8% increased the number of intestinal adenomas, and the effect was significant in female mice. The concentration of mucosal free sitosterol increased by eightfold in plant sterol males and by threefold in plant sterol females when compared with respective controls. The concentration of mucosal free cholesterol was significantly lower in plant sterol males than in control males, and the decrease in free cholesterol was accompanied with a significant increase in nuclear sterol regulatory element binding protein-2. No difference was found in the levels of ß-catenin, cyclin D1, epidermal growth factor receptor, extracellular signal-regulated kinase 1/2, or caveolin-1 in either gender after plant sterol feeding. Among all measured parameters, higher levels of estrogen receptor ß and free cholesterol in the mucosa were among the strongest predictors of increased intestinal tumorigenesis. In addition, gene expression data showed significant enrichment of up-regulated genes of cell cycle control and cholesterol biosynthesis in plant sterol females. The results indicate that high intake of plant sterols accelerates intestinal tumorigenesis in female Apc (Min)mice; however, the mechanism behind the adverse effect remains to be discovered.

Cancer-predicting gene expression changes in colonic mucosa of Western diet fed Mlh1+/- mice.

Colorectal cancer (CRC) is the second most common cause of cancer-related deaths in the Western world and interactions between genetic and environmental factors, including diet, are suggested to play a critical role in its etiology. We conducted a long-term feeding experiment in the mouse to address gene expression and methylation changes arising in histologically normal colonic mucosa as putative cancer-predisposing events available for early detection. The expression of 94 growth-regulatory genes previously linked to human CRC was studied at two time points (5 weeks and 12 months of age) in the heterozygote Mlh1(+/-) mice, an animal model for human Lynch syndrome (LS), and wild type Mlh1(+/+) littermates, fed by either Western-style (WD) or AIN-93G control diet. In mice fed with WD, proximal colon mucosa, the predominant site of cancer formation in LS, exhibited a significant expression decrease in tumor suppressor genes, Dkk1, Hoxd1, Slc5a8, and Socs1, the latter two only in the Mlh1(+/-) mice. Reduced mRNA expression was accompanied by increased promoter methylation of the respective genes. The strongest expression decrease (7.3 fold) together with a significant increase in its promoter methylation was seen in Dkk1, an antagonist of the canonical Wnt signaling pathway. Furthermore, the inactivation of Dkk1 seems to predispose to neoplasias in the proximal colon. This and the fact that Mlh1 which showed only modest methylation was still expressed in both Mlh1(+/-) and Mlh1(+/+) mice indicate that the expression decreases and the inactivation of Dkk1 in particular is a prominent early marker for colon oncogenesis.

Plant stanols induce intestinal tumor formation by up-regulating Wnt and EGFR signaling in Apc Min mice.

The rate of APC mutations in the intestine increases in middle-age. At the same period of life, plant sterol and stanol enriched functional foods are introduced to diet to lower blood cholesterol. This study examined the effect of plant stanol enriched diet on intestinal adenoma formation in the Apc(Min) mouse. Apc(Min) mice were fed 0.8% plant stanol diet or control diet for nine weeks. Cholesterol, plant sterols and plant stanols were analyzed from the caecum content and the intestinal mucosa. Levels of ß-catenin, cyclin D1, epidermal growth factor receptor (EGFR) and extracellular signal-regulated kinase 1/2 (ERK1/2) were measured from the intestinal mucosa by Western blotting. Gene expression was determined from the intestinal mucosa using Affymetrix and the data were analyzed for enriched categories and pathways. Plant stanols induced adenoma formation in the small intestine, however, the adenoma size was not affected. We saw increased levels of nuclear ß-catenin, phosphorylated ß-catenin (Ser675 and Ser552), nuclear cyclin D1, total and phosphorylated EGFR and phosphorylated ERK1/2 in the intestinal mucosa after plant stanol feeding. The Affymetrix data demonstrate that several enzymes of cholesterol synthesis pathway were up-regulated, although the cholesterol level in the intestinal mucosa was not altered. We show that plant stanols induce adenoma formation by activating Wnt and EGFR signaling. EGFR signaling seems to have promoted ß-catenin phosphorylation and its translocation into the nucleus, where the expression of cyclin D1 was increased. Up-regulated cholesterol synthesis may partly explain the increased EGFR signaling in the plant stanol-fed mice.

Berries as chemopreventive dietary constituents--a mechanistic approach with the ApcMin/+ mouse.

Berries contain a number of compounds that are proposed to have anticarcinogenic properties. We wanted to see if pure ellagic acid, natural ellagitannins and three wild berries have any effect on the adenoma formation in Apc- mutated Min/+ mice. Min/+ mice were fed high-fat AIN93-G diets containing 10% (w/w) freeze-dried bilberry (Vaccinium myrtillus), lingonberry (Vaccinium vitis-idaea), cloudberry (Rubus chamaemorus), cloudberry seeds or cloudberry pulp or pure ellagic acid at 1564 mg/kg for 10 weeks. beta-Catenin and cyclin D1 protein levels in the adenomas and in the normal-appearing mucosa were determined by Western blotting and immunohistochemistry. Early changes in gene expression in the normal-appearing mucosa were analyzed by Affymetrix microarrays. Three wild berries significantly reduced tumour number (15-30%, p < 0.05), and cloudberry and lingonberry also reduced tumour size by over 60% (p < 0.01). Cloudberry resulted in decreased levels of nuclear beta-catenin and cyclin D1 and lingonberry in the level of cyclin D1 in the large adenomas (p < 0.05). Affymetrix microarrays revealed changes in genes implicated in colon carcinogenesis, including the decreased expression of the adenosine deaminase, ecto-5f-nucleotidase and PGE2 receptor subtype EP4. Ellagic acid had no effect on the number or size of adenomas in the distal or total small intestine but it increased adenoma size in the duodenum when compared with the control diet (p < 0.05). Neither cloudberry seed nor pulp had any effect on the adenoma formation. Berries seem to have great potential as a source of chemopreventive components.

Chemoprevention by white currant is mediated by the reduction of nuclear beta-catenin and NF-kappaB levels in Min mice adenomas.

BACKGROUND: Berries are a good natural source of phenolic compounds and many berries or their compounds have been shown to be chemopreventive. White currant is an interesting berry, as it contains low levels of dominant berry phenolics such as ellagic acid, anthocyanins and other flavonoids. AIMS OF THE STUDY: To study if white currant is chemopreventive in an experimental model for intestinal tumorigenesis and further study the effects on beta-catenin and NF-kappaB signaling pathways. METHODS: Multiple intestinal neoplasia (Min) mice were fed an AIN-93G based control diet or a diet containing 10% freeze dried white currant (Ribes x pallidum) for 10 weeks. Cell signaling parameters were analysed from intestinal adenomas and surrounding mucosa by Western blotting and immunohistochemistry. RESULTS: The white currant diet reduced the number of adenomas from 81 (min-max 47-114) to 51 (36-84) in the total small intestine of Min mice (P<0.02). Most of the adenomas develop in the distal part of the small intestine, and in this area white currant reduced the number from 49 to 29.5 (P<0.01) and also the size of the adenomas from 0.88 mm to 0.70 mm (P<0.02). In the colon white currant increased the number of adenomas (0.3+/-0.6 vs. 0.8+/-0.6, mean +/- SD, P<0.05), but did not affect the size. White currant reduced nuclear beta-catenin and NF-kappaB protein levels in the adenomas (P<0.05 and P<0.02, respectively). They were correlated with the size of adenomas (P<0.01). CONCLUSIONS: This study shows that white currant is effective in preventing cancer initiation and progression in the Min mouse. Whether the positive effects are due to its special phenolic composition needs to be studied in more detail.

Three Nordic berries inhibit intestinal tumorigenesis in multiple intestinal neoplasia/+ mice by modulating beta-catenin signaling in the tumor and transcription in the mucosa.

Berries contain a number of compounds that are proposed to have anticarcinogenic properties. We studied the effects and molecular mechanisms of wild berries with different phenolic profiles on intestinal tumorigenesis in multiple intestinal neoplasia/+ mice. The mice were fed a high-fat AIN93-G diet (Con) or AIN93-G diets containing 10% (w:w) freeze-dried bilberry, lingonberry (LB), or cloudberry (CB) for 10 wk. All 3 berries significantly inhibited the formation of intestinal adenomas as indicated by a 15-30% reduction in tumor number (P < 0.05). CB and LB also reduced tumor burden by over 60% (P < 0.05). Compared to Con, CB and LB resulted in a larger (P < 0.05) proportion of small adenomas (43, 69, and 64%, respectively) and a smaller proportion of large adenomas (56, 29, and 33%, respectively). Beta-catenin and cyclin D1 in the small and large adenomas and in the normal-appearing mucosa were measured by Western blotting and immunohistochemistry. CB resulted in decreased levels of nuclear beta-catenin and cyclin D1 and LB in the level of cyclin D1 in the large adenomas (P < 0.05). Early changes in gene expression in the normal-appearing mucosa were analyzed by Affymetrix microarrays, which revealed changes in genes implicated in colon carcinogenesis, including the decreased expression of the adenosine deaminase, ecto-5'-nucleotidase, and prostaglandin E2 receptor subtype EP4. Our results indicate that berries are potentially a rich source of chemopreventive components.

Ellagic acid and natural sources of ellagitannins as possible chemopreventive agents against intestinal tumorigenesis in the Min mouse.

Ellagic acid has been shown to have chemopreventive effects in various experimental cancer models. We wanted to see whether pure ellagic acid and natural ellagitannins from cloudberry (Rubus chamaemorus) seed and pulp have any effect on adenoma formation in Apc-mutated Min mice. From the age of 5 wk, the mice were fed either a control diet, a diet containing pure ellagic acid at 1,564 mg/kg, or diets containing 4.7% (wt/wt) cloudberry seeds or 5.3% cloudberry pulp. The concentrations of ellagitannins and free ellagic acid in the seed diet were 807 and 42 mg/kg and in the pulp diet 820 and 34 mg/kg, respectively. After the 10-wk feeding period, ellagic acid had no effect on the number or size of adenomas in the distal or total small intestine, but it increased adenoma size in the duodenum when compared with the control diet (1.50+/-0.29 vs. 1.16+/-0.31 mm; P=0.029). Neither cloudberry seed nor pulp diets had any effect on the adenoma formation. Chemopreventive effects and mechanisms of whole cloudberry and other similar sources of phenolic compounds should, however, be studied, further taking into account food matrix and interactions with other dietary constituents that may be involved in the bioavailability and metabolism of ellagitannins.

Promotion of adenoma growth by dietary inulin is associated with increase in cyclin D1 and decrease in adhesion proteins in Min/+ mice mucosa.

We have earlier shown that dietary fructo-oligosaccharide inulin enhances adenoma growth in multiple intestinal neoplasia (Min/+) mice. To further explore inulin-induced early biochemical changes in the normal-appearing mucosa, Min/+ mice were fed from the age of 5 weeks to the ages of 8 and 15 weeks a control diet or an inulin-enriched diet (10% w/w). In addition, the wild-type littermates were fed with the same diets until the age of 8 weeks, in order to determine whether similar changes happen both in the wild-type and Min/+ mice. The mucosa without adenomas was collected and fractionated to nuclear, cytosolic and membrane pools. The protein levels of beta-catenin, cyclin D1 and E-cadherin were determined by Western blotting at both time points, and immunohistochemical stainings were done for 8-week-old mice. The promotion of adenoma growth by inulin (week 15, 1.3-fold increase, P=.0004) was associated with accumulation of cytosolic and nuclear beta-catenin, and increased amount of cytosolic cyclin D1 (1.5-fold increase, P=.003) in the normal-appearing mucosa of the Min/+ mice. Furthermore, inulin feeding reduced the membranous pools of beta-catenin and E-cadherin. Also in the wild-type mice the drop in membranous beta-catenin was clear (P=.015), and, moreover, a subset of crypts had enhanced nuclear beta-catenin staining. These data indicate that dietary inulin can already activate in the normal-appearing mucosa beta-catenin signaling, which in the presence of Apc mutation induces adenoma growth and even in the wild-type mice direction of the changes is similar.

Promotion of intestinal tumor formation by inulin is associated with an accumulation of cytosolic beta-catenin in Min mice.

Inulin, polydisperse beta (2-1) fructan, has been suggested to protect against colon carcinogenesis and is currently used in a number of food applications. However, the data regarding the role of inulin in intestinal carcinogenesis remains controversial since the results of our previous study suggested that inulin promotes intestinal tumor formation in Min mice, an animal model for intestinal cancer with a mutation in the Apc tumor suppressor gene (Carcinogenesis 2000;21:1167-73). In our present study, we further examined the effects of inulin on intestinal tumor formation in Min mice by carefully analyzing beta-catenin expression and cellular localization at 3 different time points during the tumorigenic process. Min mice were fed a high-fat inulin-enriched (10% w/w) diet or the high-fat diet without any added fiber from the age of 6 weeks to the ages of 9, 12 or 15 weeks. The results showed that inulin significantly increased the number (by 20%) and especially the size (by 44%) of adenomas in the small intestine. At week 15, the promotion of tumor development was accompanied by an accumulation of cytosolic beta-catenin in the adenoma tissue. In the normal appearing mucosa, levels of membrane beta-catenin and PCNA were reduced in the inulin-fed mice, possibly indicating impaired enterocyte migration. These data do not support the earlier suggestions on the cancer preventive effects of inulin and emphasize the need for further research and evaluation where health claims for inulin are concerned.