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Since many nutrients, including the three major ones of glucose, dipeptides, and cholesterol, are mainly absorbed in the small intestine, the assessment of their effects on intestinal tissue is important for the study of food absorption. However, cultured intestinal cell lines, such as Caco-2 cells, or animal models, which differ from normal human physiological conditions, are generally used for the evaluation of intestinal absorption and digestion. Therefore, it is necessary to develop an alternative in vitro method for more accurate analyses. In this study, we demonstrate inhibitory effects on nutrient absorption through nutrient transporters using three-dimensional xenogeneic-free human intestinal organoids (XF-HIOs), with characteristics of the human intestine, as we previously reported. We first show that the organoids absorbed glucose, dipeptide, and cholesterol in a transporter-dependent manner. Next, we examine the inhibitory effect of natural ingredients on the absorption of glucose and cholesterol. We reveal that glucose absorption was suppressed by epicatechin gallate or nobiletin, normally found in green tea catechin or citrus fruits, respectively. In comparison, cholesterol absorption was not inhibited by luteolin and quercetin, contained in some vegetables. Our findings highlight the usefulness of screening for the absorption of functional food substances using XF-HIOs.

Patient-derived organoids (PDOs) established from tissues from various tumor types gave the foundation of ex vivo models to screen and/or validate the activity of many cancer drug candidates. Due to their phenotypic and genotypic similarity to the tumor of which they were derived, PDOs offer results that effectively complement those obtained from more complex models. Yet, their potential for predicting sensitivity to combination therapy remains underexplored. In this review, we discuss the use of PDOs in both validation and optimization of multi-drug combinations for personalized treatment strategies in CRC. Moreover, we present recent advancements in enriching PDOs with diverse cell types, enhancing their ability to mimic the complexity of in vivo environments. Finally, we debate how such sophisticated models are narrowing the gap in personalized medicine, particularly through immunotherapy strategies and discuss the challenges and future direction in this promising field.

In this study, we propose a microfluidic organoid-trapping device used to immobilize human intestinal organoids and apply fluidic stimuli to them. The proposed device has a microchannel with a trapping region with wall gaps between the channel walls and the bottom surface, and a constriction to clog the organoids in the channel. Since the introduced culture medium escapes from the gap, organoids can be cultured without excessive deformation by hydrostatic pressure. Owing to the characteristics of the organoid-trapping device, we succeeded in trapping human intestinal organoids in the channel. Furthermore, to demonstrate the applicability of the device for culturing intestinal organoids, we induced organoid fusion to form large organoids by aligning the organoids in the channel and applying fluidic shear stress to the organoids to regulate their surface structures. Therefore, we believe that organoid-trapping devices will be useful for investigating organoids aligned or loaded with fluidic stimulation.

Recognition of cell-surface glycans is an important step in the attachment of several viruses to susceptible host cells. The molecular basis of glycan interactions and their functional consequences are well studied for human norovirus (HuNoV), an important gastrointestinal pathogen. Histo-blood group antigens (HBGAs), a family of fucosylated carbohydrate structures that are present on the cell surface, are utilized by HuNoVs to initially bind to cells. In this review, we describe the discovery of HBGAs as genetic susceptibility factors for HuNoV infection and review biochemical and structural studies investigating HuNoV binding to different HBGA glycans. Recently, human intestinal enteroids (HIEs) were developed as a laboratory cultivation system for HuNoV. We review how the use of this novel culture system has confirmed that fucosylated HBGAs are necessary and sufficient for infection by several HuNoV strains, describe mechanisms of antibody-mediated neutralization of infection that involve blocking of HuNoV binding to HBGAs, and discuss the potential for using the HIE model to answer unresolved questions on viral interactions with HBGAs and other glycans.

Predicting the absorption of orally administered drugs is crucial to drug development. Current in vitro models lack physiological relevance, robustness, and reproducibility, thus hindering reliable predictions. In this study, we developed a reproducible and robust culture method to generate a human intestinal organoid-derived monolayer model that can be applied to study drug absorption through a step-by-step approach. Our model showed similarity to primary enterocytes in terms of the drug absorption-related gene expression profile, tight barrier function, tolerability toward artificial bile juice, drug transporter and metabolizing enzyme function, and nuclear receptor activity. This method can be applied to organoids derived from multiple donors. The permeability of launched 19 drugs in our model demonstrated a correlation with human Fa values, with an R 2 value of 0.88. Additionally, by combining the modeling and simulation approaches, the estimated FaFg values for seven out of nine drugs, including CYP3A substrates, fell within 1.5 times the range of the human FaFg values. Applying this method to the drug discovery process might bridge the gap between preclinical and clinical research and increase the success rates of drug development.

BackgroundIntestinal stem cells (ISCs) play indispensable roles in the maintenance of homeostasis, and also in the regeneration of the damaged intestinal epithelia. However, whether the inflammatory environment of Crohn’s disease (CD) affects properties of resident small intestinal stem cells remain uncertain.MethodsCD patient-derived small intestinal organoids were established from enteroscopic biopsy specimens taken from active lesions (aCD-SIO), or from mucosa under remission (rCD-SIO). Expression of ISC-marker genes in those organoids was examined by immunohistochemistry, and also by microfluid-based single-cell multiplex gene expression analysis. The ISC-specific function of organoid cells was evaluated using a single-cell organoid reformation assay.ResultsISC-marker genes, OLFM4 and SLC12A2, were expressed by an increased number of small intestinal epithelial cells in the active lesion of CD. aCD-SIOs, rCD-SIOs or those of non-IBD controls (NI-SIOs) were successfully established from 9 patients. Immunohistochemistry showed a comparable level of OLFM4 and SLC12A2 expression in all organoids. Single-cell gene expression data of 12 ISC-markers were acquired from a total of 1215 cells. t-distributed stochastic neighbor embedding analysis identified clusters of candidate ISCs, and also revealed a distinct expression pattern of SMOC2 and LGR5 in ISC-cluster classified cells derived from aCD-SIOs. Single-cell organoid reformation assays showed significantly higher reformation efficiency by the cells of the aCD-SIOs compared with that of cells from NI-SIOs.ConclusionsaCD-SIOs harbor ISCs with modified marker expression profiles, and also with high organoid reformation ability. Results suggest modification of small intestinal stem cell properties by unidentified factors in the inflammatory environment of CD.

Intestinal organoids have emerged as the new paradigm for modelling the healthy and diseased intestine with patient-relevant properties. In this study, we show directed differentiation of induced pluripotent stem cells towards intestinal-like phenotype within a microfluidic device. iPSCs are cultured against a gel in microfluidic chips of the OrganoPlate, in which they undergo stepwise differentiation. Cells form a tubular structure, lose their stem cell markers and start expressing mature intestinal markers, including markers for Paneth cells, enterocytes and neuroendocrine cells. Tubes develop barrier properties as confirmed by transepithelial electrical resistance (TEER). Lastly, we show that tubules respond to pro-inflammatory cytokine triggers. The whole procedure for differentiation lasts 14 days, making it an efficient process to make patient-specific organoid tubules. We anticipate the usage of the platform for disease modelling and drug candidate screening.

Human intestinal Organoids (HIOs) may be set up from primary human tissue following dissociation, release, and culture of intestinal LGR5 + stem cells. Here, we characterise the use of a semi-automated mechanical tissue dissociation technique, which demonstrates an expedited workflow and streamlined protocol, standardising key aspects of the process while consistently maintaining a high level of cultured organoid viability.

Sheep are an important livestock species whose gastrointestinal tract is essential for overall health. Feed contaminants such as bacterial toxins and mycotoxins severely damage the sheep intestine, yet the mechanisms remain mostly elusive partially due to the lack of physiologically relevant in vitro models. Here, we investigated molecular mechanisms underlying deoxynivalenol (DON)-induced toxicity by developing intestinal organoids from isolated intestinal crypts of Hu sheep. The organoids had a central lumen and monolayer epithelium, and could be continuously passaged, cryopreserved, and resuscitated. Histological and transcriptomic analysis showed that the intestinal organoids recapitulate the cell lineages and gene expression characteristics of the original intestinal tissues. Statistical analysis indicated that DON exposure significantly inhibited organoid formation efficiency, as well as the proliferation and activity of intestinal organoid cells. RNA-seq and Western blotting analysis further revealed that DON exposure induces intestinal toxicity by inhibiting the PI3K/AKT/GSK3β/β-catenin signaling pathway. Our study provides a novel example of organoid application in toxicity studies and reveals the signaling pathway involved in DON-induced toxicity in sheep, which is of great significance for improving mitigation strategies for DON.

The intestinal flora has attracted much attention in recent years. An imbalance in the intestinal flora can cause not only intestinal diseases but also cause a variety of parenteral diseases, such as endocrine diseases, nervous system diseases and cardiovascular diseases. Research on the mechanism of disease is likely to be hampered by sample accessibility, ethical issues, and differences between cellular animal and physiological studies. However, advances in stem cell culture have made it possible to reproduce 3D human tissues in vitro that mimic the cellular, anatomical and functional characteristics of real organs. Recent studies have shown that organoids can be used to simulate the development and disease of the gut and intestinal flora and have a wide range of applications in intestinal flora physiology and disease. Intestinal organoids provide a preeminent in vitro model system for cultivating microbiota that influence GI physiology, as well as for understanding how they encounter intestinal epithelial cells and cause disease. The mechanistic details obtained from such modelling may provide new avenues for the prevention and treatment of many gastrointestinal (GI) disorders. Researchers are now starting to take inspiration from other fields, such as bioengineering, and the rise of interdisciplinary approaches, including organoid chip technology and microfluidics, has greatly accelerated the development of organoids to generate intestinal organoids that are more physiologically relevant and suitable for gut microbiota studies. Here, we describe the development of organoid models of gut biology and the application of organoids to study the pathophysiology of diseases caused by intestinal dysbiosis.