Our ancestral diet consisted of much more nondigestible fiber than that of many societies today. Thus, from an evolutionary perspective the human genome and its physiological and nutritional requirements are not well aligned to modern dietary habits. Fiber reaching the colon is anaerobically fermented by the gut bacteria, which produce short-chain fatty acids (SCFAs) as metabolic byproducts. SCFAs play a role in intestinal homeostasis, helping to explain why changes in the microbiota can contribute to the pathophysiology of human diseases. Recent research has shown that SCFAs can also have effects on tissues and organs beyond the gut, through their circulation in the blood. SCFAs not only signal through binding to cognate G-protein-coupled receptors on endocrine and immune cells in the body but also induce epigenetic changes in the genome through effects on the activity of histone acetylase and histone deacetylase enzymes. Furthermore, epigenetic imprinting likely occurs in utero, highlighting the importance of the maternal diet in early life. Here we review current understanding of how SCFAs impact on human and animal physiology and discuss the potential applications of SCFAs in the prevention and treatment of human diseases. SCFAs Are the Main Players in the Interplay between Diet, Microbiota, and HealthSCFAs have fewer than six carbons in the aliphatic tail, and the most abundant in the intestine are acetate (C2), propionate (C3), and butyrate (C4). SCFAs are a metabolic by-product of microbial fermentation of complex polysaccharides not digested, or only partly digested, in the human small intestine. These nondigestible polysaccharides (NDPs) are found in plant cell walls and are further classified into soluble and nonsoluble dietary fibers. The soluble NDPs are highly fermentable and typically generate greater quantities of SCFAs in the colon than do soluble fibers.Currently there are three well characterized human SCFA-sensing G-protein-coupled receptors (GPCRs) which are differentially expressed on different sets of immune cells as well as epithelium and endocrine cells central to the regulation of metabolism (Table 1). Studies in GPCR-genedeficient mice have established the importance of SCFA signaling through these metabolite receptors in the control of inflammation and intestinal homeostasis but the functional roles of these receptors on different cell types is still not fully understood. HighlightsShort chain fatty acids (SCFAs) contribute to intestinal homeostasis and the regulation of energy metabolism.SCFAs circulating in the blood influence tissue-specific acetylation of histones 3 and 4 in a tissue-specific fashion.Delivery of SCFAs to the colon, using specialized diets, prevents onset of diabetes in nonobese diabetic (NOD) mice.During gestation, SCFAs can cause epigenetic imprinting in utero and protect against allergic airway disease.SCFAs regulate the blood-brain barrier and neuroimmunoendocrine functions.

An important practical limitation of the three-dimensional geometry of stem-cell derived intestinal organoids is that it prevents easy access to the apical epithelium for testing food components, microorganisms, bioactive and toxic compounds. To this end, we here report on a new robust method for generating confluent intestinal cell monolayers from single-cell suspensions of enzymatically-dissociated porcine organoids using modified culture conditions. With this method, cell seeding densities can be standardised, overcoming problems with methods based on mechanical dissociation of organoids. Confluent monolayers formed tight junctions with high transepithelial electrical resistance in three days and could be used in experiments for up to two weeks. Multilineage differentiation of ileal stem cells was demonstrated by immunohistochemistry and RT-qPCR of cell-specific transcripts, also unequivocally confirming the controversial existence of Paneth-like cells in the porcine small intestine. The method described here is useful to standardize primary epithelial monolayer formation from intestinal organoids and allows rapid and robust studies of intestinal physiology.

ScopeMetabolic flexibility is the ability to switch metabolism between carbohydrate oxidation (CHO) and fatty acid oxidation (FAO) and is a biomarker for metabolic health. The effect on metabolic health of nicotinamide riboside (NR) as an exclusive source of vitamin B3 is unknown and is examined here for a wide range of NR.Design and methodsNine‐week‐old male C57BL/6JRcc mice received a semi‐purified mildly obesogenic (40 en% fat) diet containing 0.14% L‐tryptophan and either 5, 15, 30, 180, or 900 mg NR per kg diet for 15 weeks. Body composition and metabolic parameters were analyzed. Metabolic flexibility was measured using indirect calorimetry. Gene expression in epididymal white adipose tissue (eWAT) was measured using qRT‐PCR .ResultsThe maximum delta respiratory exchange ratio when switching from CHO to FAO (maxΔRERCHO1→FAO) and when switching from FAO to CHO (maxΔRERFAO→CHO2) were largest in 30 mg NR per kg diet (30NR). In eWAT, the gene expression of Pparγ, a master regulator of adipogenesis, and of Sod2 and Prdx3, two antioxidant genes, were significantly upregulated in 30NR compared to 5NR.Conclusion30NR is most beneficial for metabolic health, in terms of metabolic flexibility and eWAT gene expression, of mice on an obesogenic diet.

The emergence of intestinal organoids, as a stem cell-based self-renewable model system, has led to many studies on intestinal development and cell-cell signaling. However, potential issues regarding the phenotypic stability and reproducibility of the methodology during culture still needs to be addressed for different organoids. Here we investigated the transcriptomes of jejunum organoids derived from the same pig as well as batch-to-batch variation of organoids derived from different pigs over longterm passage. The set of genes expressed in organoids closely resembled that of the tissue of origin, including small intestine specific genes, for at least 17 passages. Minor differences in gene expression were observed between individual organoid cultures. In contrast, most small intestine-specific genes were not expressed in the jejunum cell line IPEC-J2, which also showed gene expression consistent with cancer phenotypes. We conclude that intestinal organoids provide a robust and stable model for translational research with clear advantages over transformed cells.

In pigs, high protein diets have been related to post-weaning diarrhoea, which may be due to the production of protein fermentation metabolites that were shown to have harmful effects on the intestinal epithelium in vitro. In this review, we discussed in vivo effects of protein fermentation on the microbial composition and their protein catabolic activity as well as gut and overall health. The reviewed studies applied different dietary protein levels, which was assumed to result in contrasting fermentable protein levels. A general shift to N-utilisation microbial community including potential pathogens was observed, although microbial richness and diversity were not altered in the majority of the studies. Increasing dietary protein levels resulted in higher protein catabolic activity as evidenced by increased concentration of several protein fermentation metabolites like biogenic amines in the digesta of pigs. Moreover, changes in intestinal morphology, permeability and pro-inflammatory cytokine concentrations were observed and diarrhoea incidence was increased. Nevertheless, higher body weight and average daily gain were observed upon increasing dietary protein level. In conclusion, increasing dietary protein resulted in higher proteolytic fermentation, altered microbial community and intestinal physiology. Supplementing diets with fermentable carbohydrates could be a promising strategy to counteract these effects and should be further investigated.