Introduction: Physiological microflora is characterised by wide diversity. The microbial community is mostly composed of bacteria, but also includes fungi, archaea and viruses. Anaerobic commensal bacteria (Firmicutes and Bacteroidetes) dominate 90% of the colon. The composition and products of the gut microbiota have a significant effect on an individual's immune system, and their interactions may ultimately promote immune tolerance or inflammatory immune response. Blood cell transplantation (HSCT) and its associated standard procedures of conditioning, antibiotic exposure and dietary prophylaxis represent modification and disruption of the gut microbiota leading to the development of some serious post-transplant complications affecting the OS (overall survival) and TRM (treatment related mortality) of patients. Objectives: The aim of this work was to investigate the representation of individual bacterial strains in patients undergoing allogeneic HSCT, as well as the effect of transplantation on the composition and diversity of their gut microbiota. Methods: Stool samples were obtained from 52 patients who underwent an allogeneic hematopoietic cell transplant at the Institute of Hematology and Blood Transfusion in Prague. A cut-off date for the first sample was set for the start of...

Plant-derived protein has gained popularity in recent years due to its health and environmental impact. Studies comparing the health benefits of animal and plant protein have mostly focused on soy as a plant protein representative due to its popularity. Demand for other protein sources such as peas and fava beans has been increasing; thus, the health impact of plant protein sources other than soy must be assessed. Evidence has shown that diet influences the gut microbiota and the metabolites in the body. Metabolites associated with amino acids are strong contributors to the metabolite distinction between dietary animal and plant foods. Certain gut bacteria are also known to be able to metabolize amino acids, thus influencing their survival in the gut. To discern the impact of plant and animal-based proteins, namely soy, pea, faba bean, beef, chicken, and pork, on the host gut microbiota as well as the metabolic profile, male C57BL/6J mice were fed with the proteins for 8 weeks. Results showed that each protein source influenced the gut microbiota and metabolic profile differently. However, these impacts were not caused by the amino acid profile alone. Other factors, such as myoglobin in meats and phenolic compounds in plant proteins, also play a role.

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No / Inflammatory bowel diseases are associated with increased risk of developing colitis-associated colorectal cancer (CAC). Epidemiological data show that the consumption of ω-3 polyunsaturated fatty acids (ω-3 PUFAs) decreases the risk of sporadic colorectal cancer (CRC). Importantly, recent data have shown that eicosapentaenoic acid-free fatty acid (EPA-FFA) reduces polyp formation and growth in models of familial adenomatous polyposis. However, the effects of dietary EPA-FFA are unknown in CAC. We tested the effectiveness of substituting EPA-FFA, for other dietary fats, in preventing inflammation and cancer in the AOM-DSS model of CAC. The AOM-DSS protocols were designed to evaluate the effect of EPA-FFA on both initiation and promotion of carcinogenesis. We found that EPA-FFA diet strongly decreased tumor multiplicity, incidence and maximum tumor size in the promotion and initiation arms. Moreover EPA–FFA, in particular in the initiation arm, led to reduced cell proliferation and nuclear β-catenin expression, whilst it increased apoptosis. In both arms, EPA-FFA treatment led to increased membrane switch from ω-6 to ω-3 PUFAs and a concomitant reduction in PGE2 production. We observed no significant changes in intestinal inflammation between EPA-FFA treated arms and AOM-DSS controls. Importantly, we found that EPA-FFA treatment restored the loss of Notch signaling found in the AOM-DSS control and resulted in the enrichment of Lactobacillus species in the gut microbiota. Taken together, our data suggest that EPA-FFA is an excellent candidate for CRC chemoprevention in CAC.

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Type II Diabetes Mellitus (T2D) and cardiovascular diseases (CVD) are non-communicable chronic diseases that involves impairments in glucose metabolism and vascular function. Multiple factors may increase the risk for T2D, including but not limited to genetics, obesity and lifestyle, such as physical inactivity and diet. The gut microbiota, the human's largest population of microorganisms, plays an essential role in health and disease. The physiology and function of the gastrointestinal tract can be influenced by the diet. Phosphatidylcholine (PC), a source of choline in the diet, is rich in Western-type diets. Gut microbiota metabolize choline to trimethylamine (TMA) which circulates and is oxidized in the liver to form trimethylamine N-oxide (TMAO). As a result, ingestion of PC or choline could increase levels of TMAO. Preclinical studies indicate a role of TMAO in the development of atherosclerosis. Likewise, multiple observations support a potential role of TMAO in the development of insulin resistance and T2D. Much of the research has been conducted on rodent models, while others are observational human studies. Whether acute and short-term increases in TMAO contribute to impairments in insulin sensitivity in humans remains unknown. To address this, we performed two studies utilizing a double-blind, placebo controlled, crossover design. Eligible participants consumed a 1000mg/day dose of choline bitartrate and placebo (maltodextrin) the night before each testing session (for the acute choline study) or for 4 weeks (for the short-term choline ingestion study). Oral glucose tolerance test, continuous glucose monitoring, flow-mediated dilation, and applanation tonometry was performed the day after the acute choline load and before and after the short-term choline ingestion period. We hypothesized that gut microbiota-generated increase in TMAO will impair insulin sensitivity, glucose tolerance, endothelial function and arterial stiffness in healthy sedentary humans. Following acute choline ingestion, significant increases in plasma TMAO (p = 0.013) and choline (p = 0.003) were evident. There was no statistically significant difference in insulin sensitivity, glucose tolerance or in any of the endothelial function and arterial stiffness measurements. Four weeks of 1000mg choline ingestion per day, significantly increased plasma (p = 0.042) and urine (p = 0.008) TMAO concentrations compared to the placebo. However, no significant differences were observed for any other measurements of insulin sensitivity, glucose tolerance, glycemic variability, endothelial function, and arterial stiffness. More research is needed to elucidate the mechanisms behind the mechanistic observations between elevated TMAO concentrations and T2D and CVD. / Doctor of Philosophy / Type 2 diabetes mellitus (T2D) and cardiovascular diseases (CVD) increase the risk of all-cause mortality. Choline is a nutrient that can be found in foods such as red meat, dairy, fish, and eggs. Choline is metabolized from bacteria in our gut and a metabolite called trimethylamine (TMA) is formed. TMA is then oxidized in the liver and trimethylamine-N-oxide (TMAO) is produced. A Western-type diet is rich in red meat, dairy, fish, and eggs and has been shown to increase production of the compound TMAO. Preclinical studies have suggested a causal role of TMAO in atherosclerosis and T2D and elevated plasma TMAO concentrations have been associated with an increased risk for CVD and T2D in observational studies. However, the causal nature of this relationship in humans is unknown. The studies described herein aimed to investigate the effects of increases in TMAO on insulin sensitivity and vascular function in healthy adults. The first study tested the effect of increasing TMAO on insulin sensitivity, glucose tolerance, and vascular function following an acute choline load (1000mg) and placebo (carbohydrate) the night before each testing session. In the second study, we examined the effect of increasing TMAO on insulin sensitivity, glucose tolerance, and vascular function in healthy adults, following a short-term choline load (1000mg/day) and placebo (carbohydrate) for 4 weeks. Acute and short-term choline ingestion significantly increased plasma TMAO concentrations. No significant differences were observed following acute or short-term choline ingestion for any measurement of insulin sensitivity, glucose tolerance 24-hout glycemic variability, vascular function., and arterial stiffness.

The global pandemic of diet-related non-communicable diseases and the fact that global food production represents one of the largest contributors to greenhouse gas emissions, have identified unhealthy and unsustainably food chains as a major societal health challenge and a risk to ecosystem stability. This thesis aimed to investigate if digestion of nutritious, less highly processed foods could lead to health-promoting changes in the gut microbiota. Our modern Western-style diet (MWD) is characterized by high intake of extremely processed foods, which contain significant concentrations of inflammatory advanced glycation end-products (AGE) implicated in metabolic disease development. Novel observations in this thesis showed that chronic exposure to dietary AGE modulated gut microbiota (GM) community structure rendering it more similar to the GM previously observed in diabetic/obese mice. Further, I demonstrated that elevated systemic inflammatory markers could be mediated by AGE induced changes in GM composition. Measuring the potential of whole plant foods to improve gut health, a local broccoli ecotype (Broccolo of Torbole, BR) and Moringa oleifera were investigated using in vitro models of the human GM and intestinal epithelium. BR significantly reduced bacterial richness and evenness, increased Escherichia-Shigella relative abundance and decreased Alistipes and Ruminococcus 1. The GM extensively metabolized BR polyphenols and increased concentrations of short chain fatty acids. However, BR did not impact on intestinal permeability, using a Caco-2 monolayer model and trans-epithelial electrical resistance (TEER). This thesis provided novel insights on the fate of Moringa glucosinolates and polyphenols during faecal fermentation and on their potential beneficial activity on gut health, with glucomoringin significantly increasing TEER. Microbial communities are also involved in healthy and sustainable food production. Characterizing the successional development of local organic sauerkraut production, this thesis established a culture collection of sauerkraut lactic acid bacteria of potential future biotechnological evaluation and measured metabolite production during sauerkraut fermentation. Sauerkraut water improved immune response of a Caco-2-peripheral blood mononuclear cell (PBMC) in vitro model of the gut associated immune system upon inflammatory LPS challenge. Finally, since sustainable diets rely on sustainable and nutritious foods, I analyzed the role of the GM in improving the sustainability of farmed trout. Novel sustainable feeds containing poultry by-products (P) or insect protein (Hermetia illucens (H) meal), were investigated for their potential impact on fish growth performance, GM composition and inflammatory biomarkers. P increased the relative abundance of protein-degraders Paeniclostridium and Bacteroidales, while H increased chitin-degraders Actinomyces and Bacillus. This study also provided evidence of feed-chain microbiome transmission of Actinomyces from insect H feed to trout GM. The analysis of gut microbiomes therefore represents an innovative strategy to define healthy reference diets, to characterize the potential health effects of local and traditionally produced foods, to identify new sustainable and nutritious crops, and to drive the urgently needed transformation of the global food system. In order to obtain more sustainable, healthy and nutritious food production systems a better understanding and management of microbiomes along the food chain has never been more important.

Tannins represent a heterogeneous group of high-molecular-weight polyphenols that are ubiquitous among plant families, especially in cereals, as well as in many fruits and vegetables. Hydrolysable and condensed tannins, in addition to phlorotannins from marine algae, are the main classes of these bioactive compounds. Despite their low bioavailability, tannins have many beneficial pharmacological effects, such as anti-inflammatory, antioxidant, antidiabetic, anticancer, and cardioprotective effects. Microbiota-mediated hydrolysis of tannins produces highly bioaccessible metabolites, which have been extensively studied and account for most of the health effects attributed to tannins. This review article summarises the effect of the human microbiota on the metabolism of different tannin groups and the expected health benefits that may be induced by such mutual interactions. Microbial metabolism of tannins yields highly bioaccessible microbial metabolites that account for most of the systemic effects of tannins. This article also uses explainable artificial intelligence to define the molecular signatures of gut-biotransformed tannin metabolites that are correlated with chemical and biological activity. An understanding of microbiota–tannin interactions, tannin metabolism-related phenotypes (metabotypes) and chemical tannin-metabolites motifs is of great importance for harnessing the biological effects of tannins for drug discovery and other health benefits.

京都大学 / 新制・課程博士 / 博士(農学) / 甲第24678号 / 農博第2561号 / 新制||農||1100(附属図書館) / 学位論文||R5||N5459(農学部図書室) / 京都大学大学院農学研究科応用生物科学専攻 / (主査)教授 佐藤 健司, 教授 菅原 達也, 教授 舟場 正幸 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DGAM

The CpG Island Methylator Phenotype (CIMP) is a distinct form of aberrant DNA methylation in cancer, and it is seen in 20-40% of colorectal cancers (CRC) where its causes remain elusive. Intestinal microbiota represents an important environmental component implicated in CRC development. Interestingly, microbiota have been shown to modulate DNA methylation in preclinical models but the relationship between tumor-infiltrating bacteria and CIMP status in currently unknown. Our hypothesis is that the gut microbiota affects colonic neoplasia through modulating aberrant DNA methylation in host epigenome. To test this hypothesis, we analyzed CIMP status in CRC patient tumor samples. We used a genome-wide approach to determine the CIMP status by filtering cancer-related sites. A total of 1317 CpG sites were filtered and used to determine distinct CIMP classifications that aligned with well-known characteristics of CIMP cases, including localization in the proximal colon, a higher prevalence in female patients, and a higher frequency of MLH1 hypermethylation. To study the association between CIMP and the gut microbiota, we analyzed the enrichment of four bacterial species associated with CRC, including Bacteroides fragilis, Escherichia coli, Fusobacterium nucleatum, and Klebsiella pneumoniae. Notably, they exhibited higher enrichment in CIMP-Positive tumor samples, except for E. coli. This analysis also identified a group of samples referred to as bacterial "Superhigh," characterized by remarkably high abundances of these three bacterial species. The bacterial Superhigh cases displayed a significant association with CIMP status and MLH1 methylation. We validated the association between the CRC-associated bacteria and CIMP by analyzing the Cancer Genome Atlas (TCGA) 450K methylation array data and whole exome sequencing data. The analysis demonstrated that bacterial Superhigh cases in the TCGA datasets also had significantly higher odds of being CIMP-Positive and having MLH1 methylation. To expand our investigation, we conducted 16S rRNA gene sequencing to identify additional bacterial taxa linked to CIMP. Numerous bacterial genera and species were found to be enriched in CRC tumor tissues, with specific enrichments in CIMP-Positive and CIMP-High groups. Notably, Bergeyella, Campylobacter concisus, and Fusobacterium canifelinum were significantly enriched in CIMP-Positive tumors. Additionally, I studied the causal relationship between gut microbiota and CpG island methylation by colonizing germ-free mice ApcMinΔ850/+;Il10–/– with E. coli NC101 & K. pneumoniae, specific pathogen free bacteria, and the mouse bacterial Consortium. Differential methylation analyses of adjacent normal colon tissue revealed a pronounced tissue side-specific difference, particularly in non-CpG island regions. The tissue specificity diminished with the increasing tumorigenic potential of the microbiota group. Comparisons between microbiota groups and germ-free mice indicated a more significant increase in methylation within CpG islands when gut microbiota with higher tumorigenic potential was present. In conclusion, our study underscores the association between CIMP in CRC and the gut microbiota and the causal relationship between the cut microbiota and CpG island methylation. It highlights specific bacterial taxa that may impact DNA methylation especially in CpG islands and contribute to the development ang progression of CIMP in colorectal cancer. / Biomedical Sciences