Effects of the whole seed and a protein isolate of faba bean (Vicia faba) on the cholesterol metabolism of hypercholesterolaemic rats

The aim of the present work was to analyse the hypocholesterolaemic efficiency of a Vicia fabaprotein isolate in relation to the intact legume. In addition, the mechanisms underlying the effects of this isolate were investigated. Hypercholesterolaemic rats were divided into three groups n10 3 and fed high-fat diets rich in cholesterol-containing casein, whole seeds of Vicia faba or the protein isolate of faba beans as protein source, for 2 weeks ad libitum. The protein isolate was prepared by isoelectric precipitation and spray dried. Analyses of serum, liver and faeces, as well as of the activity of hepatic 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase, were assessed by enzymatic methods. The rats fed on Vicia faba diets showed significantly lower body weights and energy intakes than rats fed on casein diets. The wholeseed diet induced a significant reduction in plasma triacylglycerol. Feeding rats on diets containing faba bean seeds, or the protein isolate, induced a significant decrease in plasma (LDL+VLDL)-cholesterol but not in HDL-cholesterol. Hepatic cholesterol and triacylglycerol were also reduced. The hypocholesterolaemic effects of Vicia faba were not the result of a reduction in cholesterol synthesis as assessed from HMG-CoA reductase activity, but the result of an increase in steroid faecal excretion. The faba bean-protein isolate obtained under our experimental conditions was useful in improving the metabolic alterations induced by feeding with a hypercholesterolaemic diet compared with casein. The effectiveness of the whole seeds was higher than that of the protein isolate

Shifts in microbiota species and fermentation products in a dietary model enriched in fat and sucrose

The gastrointestinal tract harbours a “superorganism” called the gut microbiota, which is known to play a crucial role in the onset and development of diverse diseases. This internal ecosystem, far from being a static environment, could be willingly manipulated by diet and dietary components. Feeding animals with high-fat sucrose diets entails diet-induced obesity, a model which is usually used in research to mimic the obese phenotype of Western societies. The aim of the present study was to identify gut microbiota dysbiosis and associated metabolic changes produced in 5 male Wistar rats fed a high-fat sucrose (HFS) diet for six weeks and to compare it with the basal microbial composition. For this purpose, DNA extracted from faeces at baseline and after the treatment was analysed by amplification of the V4-V6 region of the 16S ribosomal DNA (rDNA) gene using 454 pyrosequencing. Short-chain fatty acids (SCFA), acetate, propionate and butyrate, were also evaluated by gas chromatography-mass spectrometry (GC-MS). At the end of the treatment, gut microbiota composition significantly differed at phylum level (Firmicutes, Bacteroidetes and Proteobacteria) and class level (Erisypelotrichi, Deltaproteobacteria, Bacteroidia and Bacilli). Interestingly, Clostridia class showed a significant decrease after the HFS-diet treatment, which correlated with visceral adipose tissue, and is likely mediated by dietary carbohydrates. Of particular interest, Clostridium cluster XIVa species were significantly reduced and changes were identified in the relative abundance of other specific bacterial species (Mitsuokella jalaludinii, Eubacterium ventriosum, Clostridium sp. FCB90-3, Prevotella nanceiensis, Clostridium fusiformis, Clostridium sp. BNL1100 and Eubacterium cylindroides) that, in some cases, showed opposite trends to their relative families. These results highlight the relevance of characterizing gut microbial population differences at species level and contribute to understand the plausible link between the 1 diet and specific gut bacterial species that are able to influence the inflammatory status, intestinal barrier function and obesity development. Keywords: gut microbiota, pyrosequencing, high-fat sucrose diet, short chain fatty acids, Erysipelotrich

Reshaping faecal gut microbiota composition by the intake of trans-resveratrol and quercetin in high-fat sucrose diet-fed rats

Diet‐induced obesity is associated to an imbalance in the normal gut microbiota composition. Resveratrol and quercetin, widely known for their health beneficial properties, have low bioavailability and, when reach the colon, they are targets of the gut microbial ecosystem. Hence, the use of these molecules in obesity might be considered as a potential strategy to modulate intestinal bacterial composition. The purpose of this study was to determine whether trans‐resveratrol and quercetin administration could counteract gut microbiota dysbiosis produced by high‐fat sucrose diet (HFS) and in turn, improve gut health. Wistar rats were randomized into four groups fed a HFS diet supplemented or not with trans‐resveratrol (15 mg/kg BW/day), quercetin (30 mg/kg BW/day) or a combination of both polyphenols at those doses. Administration of both polyphenols together prevented body‐weight gain and reduced serum insulin levels. Moreover, individual supplementation of trans‐resveratrol and quercetin effectively reduced serum insulin levels and insulin resistance. Quercetin supplementation generated a great impact on gut microbiota composition at different taxonomic levels, attenuating Firmicutes/Bacteroidetes ratio and inhibiting the growth of bacterial species previously associated to diet‐induced obesity (Erysipelotrichaceae, Bacillus, Eubacterium 1 cylindroides). Overall, the administration of quercetin was found to be effective in lessening HFS diet‐induced gut microbiota dysbiosis. In contrast, trans‐resveratrol supplementation alone or in combination with quercetin, scarcely modified the profile of gut bacteria, but acted at intestinal level altering the mRNA expression of tight‐junction proteins (TJPs) and inflammation associated genes

Metabolic faecal fingerprinting of trans-resveratrol and quercetin following a high-fat sucrose dietary model using liquid chromatography coupled to high-resolution mass spectrometry

Faecal non‐targeted metabolomics deciphers metabolic end‐products resulting from the interactions among food, host genetics, and gut microbiota. Faeces from Wistar rats fed a high‐fat sucrose (HFS) diet supplemented with trans‐resveratrol and quercetin (separately or combined) were analysed by liquid chromatography coupled to high‐resolution mass spectrometry (LC‐HRMS). Metabolomics in faeces are categorised into four clusters based on the type of treatment. Tentative identification of significantly differing metabolites highlighted the presence of carbohydrate derivatives or conjugates (3‐phenylpropyl glucosinolate and dTDP‐D‐mycaminose) in quercetin group. The trans‐resveratrol group was differentiated by compounds related to nucleotides (uridine monophosphate and 2,4‐dioxotetrahydropyrimidine D‐ribonucleotide). Marked associations between bacterial species (Clostridium genus) and the amount of some metabolites were identified. Moreover, trans‐resveratrol and resveratrol‐derived microbial metabolites (dihydroresveratrol and lunularin) were also identified. Accordingly, this study confirms the usefulness of omics‐based techniques to discriminate individuals depending on the physiological effect of food constituents and represents an interesting tool to assess the impact of future personalized therapies

Reshaping faecal gut microbiota composition by the intake of trans-resveratrol and quercetin in high-fat sucrose diet-fed rats

Diet‐induced obesity is associated to an imbalance in the normal gut microbiota composition. Resveratrol and quercetin, widely known for their health beneficial properties, have low bioavailability and, when reach the colon, they are targets of the gut microbial ecosystem. Hence, the use of these molecules in obesity might be considered as a potential strategy to modulate intestinal bacterial composition. The purpose of this study was to determine whether trans‐resveratrol and quercetin administration could counteract gut microbiota dysbiosis produced by high‐fat sucrose diet (HFS) and in turn, improve gut health. Wistar rats were randomized into four groups fed a HFS diet supplemented or not with trans‐resveratrol (15 mg/kg BW/day), quercetin (30 mg/kg BW/day) or a combination of both polyphenols at those doses. Administration of both polyphenols together prevented body‐weight gain and reduced serum insulin levels. Moreover, individual supplementation of trans‐resveratrol and quercetin effectively reduced serum insulin levels and insulin resistance. Quercetin supplementation generated a great impact on gut microbiota composition at different taxonomic levels, attenuating Firmicutes/Bacteroidetes ratio and inhibiting the growth of bacterial species previously associated to diet‐induced obesity (Erysipelotrichaceae, Bacillus, Eubacterium 1 cylindroides). Overall, the administration of quercetin was found to be effective in lessening HFS diet‐induced gut microbiota dysbiosis. In contrast, trans‐resveratrol supplementation alone or in combination with quercetin, scarcely modified the profile of gut bacteria, but acted at intestinal level altering the mRNA expression of tight‐junction proteins (TJPs) and inflammation associated genes

Effects of the whole seed and a protein isolate of faba bean (Vicia faba) on the cholesterol metabolism of hypercholesterolaemic rats

The aim of the present work was to analyse the hypocholesterolaemic efficiency of a Vicia fabaprotein isolate in relation to the intact legume. In addition, the mechanisms underlying the effects of this isolate were investigated. Hypercholesterolaemic rats were divided into three groups n10 3 and fed high-fat diets rich in cholesterol-containing casein, whole seeds of Vicia faba or the protein isolate of faba beans as protein source, for 2 weeks ad libitum. The protein isolate was prepared by isoelectric precipitation and spray dried. Analyses of serum, liver and faeces, as well as of the activity of hepatic 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase, were assessed by enzymatic methods. The rats fed on Vicia faba diets showed significantly lower body weights and energy intakes than rats fed on casein diets. The wholeseed diet induced a significant reduction in plasma triacylglycerol. Feeding rats on diets containing faba bean seeds, or the protein isolate, induced a significant decrease in plasma (LDL+VLDL)-cholesterol but not in HDL-cholesterol. Hepatic cholesterol and triacylglycerol were also reduced. The hypocholesterolaemic effects of Vicia faba were not the result of a reduction in cholesterol synthesis as assessed from HMG-CoA reductase activity, but the result of an increase in steroid faecal excretion. The faba bean-protein isolate obtained under our experimental conditions was useful in improving the metabolic alterations induced by feeding with a hypercholesterolaemic diet compared with casein. The effectiveness of the whole seeds was higher than that of the protein isolate

Analysis of mutant allele fractions in driver genes in colorectal cancer – biological and clinical insights

Sequencing of tumors is now routine and guides personalized cancer therapy. Mutant allele fractions (MAFs, or the 'mutation dose') of a driver gene may reveal the genomic structure of tumors and influence response to targeted therapies. We performed a comprehensive analysis of MAFs of driver alterations in unpaired primary and metastatic colorectal cancer (CRC) at our institution from 2010 to 2015 and studied their potential clinical relevance. Of 763 CRC samples, 622 had detailed annotation on overall survival in the metastatic setting (OSmet) and 89 received targeted agents matched to KRAS (MEK inhibitors), BRAF (BRAF inhibitors), or PIK3CA mutations (PI3K pathway inhibitors). MAFs of each variant were normalized for tumor purity in the sample (adjMAFs). We found lower adjMAFs for BRAFV600E and PIK3CA than for KRAS, NRAS, and BRAF non-V600 variants. TP53 and BRAFV600E adjMAFs were higher in metastases as compared to primary tumors, and high KRAS adjMAFs were found in CRC metastases of patients with KRAS wild-type primary tumors previously exposed to EGFR antibodies. Patients with RAS- or BRAFV600E -mutated tumors, irrespective of adjMAFs, had worse OSmet. There was no significant association between adjMAFs and time to progression on targeted therapies matched to KRAS, BRAF, or PIK3CA mutations, potentially related to the limited antitumor activity of the employed drugs (overall response rate of 4.5%). In conclusion, the lower BRAFV600E and PIK3CA adjMAFs in subsets of primary CRC tumors indicate subclonality of these driver genes. Differences in adjMAFs between metastases and primary tumors suggest that approved therapies may result in selection of BRAFV600E - and KRAS-resistant clones and an increase in genomic heterogeneity with acquired TP53 alterations. Despite significant differences in prognosis according to mutations in driver oncogenes, adjMAFs levels did not impact on survival and did not help predict benefit with matched targeted agents in the metastatic setting