BackgroundMammalian small intestinal tight junctions (TJ) link epithelial cells to one another and function as a permselective barrier, strictly modulating the passage of ions and macromolecules through the pore and leak pathways, respectively, thereby preventing the absorption of harmful compounds and microbes while allowing regulated transport of nutrients and electrolytes. Small intestinal epithelial permeability is ascribed primarily to the properties of TJs between adjoining enterocytes (ENTs), because there is almost no information on TJ composition and the paracellular permeability of nonenterocyte cell types that constitute a small but significant fraction of the intestinal epithelia.ResultsHere we directed murine intestinal crypts to form specialized organoids highly enriched in intestinal stem cells (ISCs), absorptive ENTs, secretory goblet cells, or Paneth cells. The morphological and morphometric characteristics of these cells in organoids were similar to those in vivo. The expression of certain TJ proteins varied with cell type: occludin and tricellulin levels were high in both ISCs and Paneth cells, while claudin-1, -2, and -7 expression was greatest in Paneth cells, ISCs, and ENTs, respectively. In contrast, the distribution of claudin-15, zonula occludens 1 (ZO-1), and E-cadherin was relatively homogeneous. E-cadherin and claudin-7 marked mainly the basolateral membrane, while claudin-2, ZO-1, and occludin resided in the apical membrane. Remarkably, organoids enriched in ENTs or goblet cells were over threefold more permeable to 4 and 10 kDa dextran compared to those containing stem and Paneth cells. The TJ-regulator larazotide prevented the approximately tenfold increases in dextran flux induced by the TJ-disrupter AT1002 into organoids of different cell types, indicating that this ZO toxin nonselectively increases permeability. Forced dedifferentiation of mature ENTs results in the reacquisition of ISC-like characteristics in TJ composition and dextran permeability, suggesting that the post-differentiation properties of TJs are not hardwired.ConclusionsDifferentiation of adult intestinal stem cells into mature secretory and absorptive cell types causes marked, but potentially reversible, changes in TJ composition, resulting in enhanced macromolecular permeability of the TJ leak pathway between ENTs and between goblet cells. This work advances our understanding of how cell differentiation affects the paracellular pathway of epithelia.Electronic supplementary materialThe online version of this article (10.1186/s12915-018-0481-z) contains supplementary material, which is available to authorized users.

Nutrient sensing triggers responses by the gut-brain axis modulating hormone release, feeding behavior and metabolism that become dysregulated in metabolic syndrome and some cancers. Except for absorptive enterocytes and secretory enteroendocrine cells, the ability of many intestinal cell types to sense nutrients is still unknown; hence we hypothesized that progenitor stem cells (intestinal stem cells, ISC) possess nutrient sensing ability inherited by progenies during differentiation. We directed via modulators of Wnt and Notch signaling differentiation of precursor mouse intestinal crypts into specialized organoids each containing ISC, enterocyte, goblet, or Paneth cells at relative proportions much higher than in situ as determined by mRNA expression and immunocytochemistry of cell type biomarkers. We identified nutrient sensing cell type(s) by increased expression of fructolytic genes in response to a fructose challenge. Organoids comprised primarily of enterocytes, Paneth, or goblet, but not ISC, cells responded specifically to fructose without affecting nonfructolytic genes. Sensing was independent of Wnt and Notch modulators and of glucose concentrations in the medium but required fructose absorption and metabolism. More mature enterocyte- and goblet-enriched organoids exhibited stronger fructose responses. Remarkably, enterocyte organoids, upon forced dedifferentiation to reacquire ISC characteristics, exhibited a markedly extended lifespan and retained fructose sensing ability, mimicking responses of some dedifferentiated cancer cells. Using an innovative approach, we discovered that nutrient sensing is likely repressed in progenitor ISCs then irreversibly derepressed during specification into sensing-competent absorptive or secretory lineages, the surprising capacity of Paneth and goblet cells to detect fructose, and the important role of differentiation in modulating nutrient sensing. Small intestinal stem cells differentiate into several cell types transiently populating the villi. We used specialized organoid cultures each comprised of a single cell type to demonstrate that ) differentiation seems required for nutrient sensing,) secretory goblet and Paneth cells along with enterocytes sense fructose, suggesting that sensing is acquired after differentiation is triggered but before divergence between absorptive and secretory lineages, and ) forcibly dedifferentiated enterocytes exhibit fructose sensing and lifespan extension.

The fruit of mume, Japanese apricot (Prunus mume Sieb. et Zucc.), was evaluated for its phenolics content, high performance liquid chromatography (HPLC) profile and antioxidative activities. The phenolics content of mume fruit was relatively high, the flesh of fully matured fruit containing up to 1% of phenolics on a dry weight basis. Reflecting such a high content of phenolics, the ORAC (oxygen radical absorbance capacity) value for mume fruit flesh showed high values, ranging from 150 to 320 µmol/g Trolox equivalent, depending upon the stage of maturation. 5-O-Caffeoylqunic acid (chlorogenic acid), 3-O-caffeoylquinic acid and tetra-O-acylated sucrose-related compounds were isolated from the flesh of mume fruit, although many unknown peaks were also apparent in the HPLC chromatogram. An alkali hydrolysate comprised four main phenolic acids, caffeic acid, cis/trans-p-coumaric acid and ferulic acid. No flavonoids were observed in the analysis. These results suggest that the majority of phenolics in mume fruit were hydroxycinnamic acid derivatives.

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Mume fruit, the Japanese apricot (Prunus mume SIEB. et ZUCC.), is popular in Japan and is mostly consumed in the pickled form called umeboshi. This fruit is known to have anti-microbial properties, but the principal constituents responsible for the antimicrobial properties have not yet been elucidated. We investigated the antimicrobial activities of the phenolic compounds in P. mume against enterobacteria. In this study, growth inhibitory activities were measured as an index of the antibacterial activities. The phenolic compounds were prepared from a byproduct of umeboshi called umesu or umezu (often translated as "mume vinegar"). Umesu or umezu phenolics (UP) contain approximately 20% phenolic compounds with p-coumaric acid as a standard and do not contain citric acid. We observed the inhibitory effects of UP against the growth of some enterobacteria, at a relatively high concentration (1250-5000 µg/mL). Alkali hydrolysates of UP (AHUP) exhibited similar antibacterial activities, but at much lower concentrations of 37.5-300 µg/mL. Since AHUP comprises hydroxycinnamic acids such as caffeic acid, p-coumaric acid, and ferulic acid, the antibacterial activities of each of these acids were examined. Our study shows that the phenolic compounds in P. mume other than citric acid contribute to its antimicrobial activity against enterobacteria in the digestive tract.

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Attempts to isolate the molt-inhibiting hormone (MIH) of crustaceans from crab eyestalks (ES) resulted in the characterization of xanthurenic acid as an inhibitor of ecdysone biosynthesis in the cultured Y-organ-complex (YOC) homogenate. It was also found that 3-hydroxy-L-kynurenine present in the ES is transformed into xanthurenic acid in the YOC and body fluid. Its mode of inhibitory action in ecdysone biosynthesis is probably inactivation of cytochrome P-450.