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.

Postprandial hyperglycemia is an important causative factor of type 2 diabetes mellitus, and permanent localization of intestinal GLUT2 in the brush border membrane is an important reason of postprandial hyperglycemia. Berberine, a small molecule derived from Coptidis rhizome, has been found to be potent at lowering blood glucose, but how berberine lowers postprandial blood glucose is still elusive. Here, we investigated the effect of berberine on intestinal glucose transporter 2 (GLUT2) translocation and intestinal glucose absorption in type 2 diabetes mouse model. Type 2 diabetes was induced by feeding of a high-fat diet and injection of streptozotocin and diabetic mice were treated with berberine for 6 weeks. The effects of berberine on intestinal glucose transport and GLUT2 translocation were accessed in isolated intestines and intestinal epithelial cells (IEC-6), respectively. We found that berberine treatment improved glucose tolerance and systemic insulin sensitivity in diabetic mice. Furthermore, berberine decreased intestinal glucose transport and inhibited GLUT2 translocation from cytoplasm to brush border membrane in intestinal epithelial cells. Mechanistically, berberine inhibited intestinal insulin-like growth factor 1 (IGF-1R) phosphorylation and thus reduced localization of PLC-β2 in the membrane, leading to decreased GLUT2 translocation. These results suggest that berberine reduces intestinal glucose absorption through inhibiting IGF-1R-PLC-β2-GLUT2 signal pathway.

Comparative Gene Identification-58 (CGI-58), a lipid droplet (LD)-associated protein, promotes intracellular triglyceride (TG) hydrolysis in vitro. Mutations in human CGI-58 cause TG accumulation in numerous tissues including intestine. Enterocytes are thought not to store TG-rich LDs, but a fatty meal does induce temporary cytosolic accumulation of LDs. Accumulated LDs are eventually cleared out, implying existence of TG hydrolytic machinery in enterocytes. However, identities of proteins responsible for LD-TG hydrolysis remain unknown. Here we report that intestine-specific inactivation of CGI-58 in mice significantly reduces postprandial plasma TG concentrations and intestinal TG hydrolase activity, which is associated with a 4-fold increase in intestinal TG content and large cytosolic LD accumulation in absorptive enterocytes during the fasting state. Intestine-specific CGI-58 knockout mice also display mild yet significant decreases in intestinal fatty acid absorption and oxidation. Surprisingly, inactivation of CGI-58 in intestine significantly raises plasma and intestinal cholesterol, and reduces hepatic cholesterol, without altering intestinal cholesterol absorption and fecal neutral sterol excretion. In conclusion, intestinal CGI-58 is required for efficient postprandial lipoprotein-TG secretion and for maintaining hepatic and plasma lipid homeostasis. Our animal model will serve as a valuable tool to further define how intestinal fat metabolism influences the pathogenesis of metabolic disorders, such as obesity and type 2 diabetes.

Regulation of lipid absorption by enterocytes can influence metabolic status in humans and contribute to obesity and related complications. The intracellular steps of chylomicron biogenesis and transport from the Endoplasmic Reticulum (ER) to the Golgi complex have been described, but the mechanisms for post-Golgi transport and secretion of chylomicrons have not been identified. Using a newly generated Dennd5b-/- mouse, we demonstrate an essential role for this gene in Golgi to plasma membrane transport of chylomicron secretory vesicles. In mice, loss of Dennd5b results in resistance to western diet induced obesity, changes in plasma lipids, and reduced aortic atherosclerosis. In humans, two independent exome sequencing studies reveal that a common DENND5B variant, p.(R52K), is correlated with body mass index. These studies establish an important role for DENND5B in post-Golgi chylomicron secretion and a subsequent influence on body composition and peripheral lipoprotein metabolism.

Oleocanthal (OLC), a phenolic compound of extra virgin olive oil (EVOO), has emerged as a potential therapeutic agent against a variety of diseases due to its anti-inflammatory activity. The aim of the present study is to explore its in vivo intestinal absorption and metabolism. An in situ perfusion technique in rats was used, involving simultaneous sampling from the luminal perfusate and mesenteric blood. Samples were analysed by UHPLC-MS-MS for the presence of oleocanthal (OLC) and its metabolites. OLC was mostly metabolized by phase I metabolism, undergoing hydration, hydrogenation and hydroxylation. Phase II reactions (glucuronidation of hydrogenated OLC and hydrated metabolites) were observed in plasma samples. OLC was poorly absorbed in the intestine, as indicated by the low effective permeability coefficient (2.23 ± 3.16 × 10-5 cm/s) and apparent permeability coefficient (4.12 ± 2.33 × 10-6 cm/s) obtained relative to the values of the highly permeable reference compound levofloxacin (LEV). The extent of OLC absorption reflected by the area under the mesenteric blood-time curve normalized by the inlet concentration (AUC) was also lower than that of LEV (0.25 ± 0.04 vs. 0.64 ± 0.03, respectively). These results, together with the observed intestinal metabolism, suggest that OLC has a moderate-to-low oral absorption; but higher levels of OLC are expected to reach human plasma vs. rat plasma.

Reduced activity of nutrient-sensing signaling networks can extend organismal lifespan, yet the underlying biology remains unclear. We show that the anti-aging effects of rapamycin and reduced intestinal insulin/insulin growth factor (IGF) signaling (IIS) require the Drosophila FoxA transcription factor homolog Fork Head (FKH). Intestinal FKH induction extends lifespan, highlighting a role for the gut. FKH binds to and is phosphorylated by AKT and Target of Rapamycin. Gut-specific FKH upregulation improves gut barrier function in aged flies. Additionally, it increases the expression of nutrient transporters, as does lowered IIS. Evolutionary conservation of this effect of lowered IIS is suggested by the upregulation of related nutrient transporters in insulin receptor substrate 1 knockout mouse intestine. Our study highlights a critical role played by FKH in the gut in mediating anti-aging effects of reduced IIS. Malnutrition caused by poor intestinal absorption is a major problem in the elderly, and a better understanding of the mechanisms involved will have important therapeutic implications for human aging.

The absorption and antioxidant activity of polyphenols from grape pomace (GP) are important aspects of its valorization as a feed additive in the diet of weaned piglets. This study aimed to evaluate the presence of polyphenols from GP both in vitro in IPEC cells and in vivo in the duodenum and colon of piglets fed with diets containing or not 5% GP and also to compare and correlate the aspects of their in vitro and in vivo absorption. Total polyphenolic content (TPC) and antioxidant status (TAS, CAT, SOD and GPx enzyme activity, and lipid peroxidation-TBARS level) were assessed in duodenum and colon of piglets fed or not a diet with 5% GP. The results of UV-Vis spectroscopy demonstrated that in cellular and extracellular medium the GP polyphenols were oxidized (between λmax = 276 nm and λmax = 627.0 nm) with the formation of o-quinones and dimers. LC-MS analysis indicated a procyanidin trimer possibly C2, and a procyanidin dimer as the major polyphenols identified in GP, 12.8% of the procyanidin trimer and 23% of the procyanidin dimer respectively being also found in the compound feed. Procyanidin trimer C2 is the compound accumulated in duodenum, 73% of it being found in the colon of control piglets, and 62.5% in the colon of GP piglets. Correlations exist between the in vitro and in vivo investigations regarding the qualitative evaluation of GP polyphenols in the cells (λmax at 287.1 nm) and in the gut (λmax at 287.5 nm), as oxidated metabolic products. Beside the presence of polyphenols metabolites this study shows also the presence of the unmetabolized procyanidin trimers in duodenum and colon tissue, an important point in evaluating the benefic actions of these molecules at intestinal level. Moreover the in vivo study shows that a 5% GP in piglet’s diet increased the total antioxidant status (TAS) and decreased lipid peroxidantion (TBARS) in both duodenum and colon, and increased SOD activity in duodenum and CAT and GPx activity in colon. These parameters are modulated by the different polyphenols absorbed, mainly by the procyanidin trimers and catechin on one side and the polyphenols metabolites on the other side.

Iron is a trace metal, key for the development of living organisms. Its absorption process is complex and highly regulated at the transcriptional, translational and systemic levels. Recently, the internalization of the DMT1 transporter has been proposed as an additional regulatory mechanism at the intestinal level, associated to the mucosal block phenomenon. The short-term effect of iron exposure in apical uptake and initial absorption rates was studied in Caco-2 cells at different apical iron concentrations, using both an experimental approach and a mathematical modeling framework. This is the first report of short-term studies for this system. A non-linear behavior in the apical uptake dynamics was observed, which does not follow the classic saturation dynamics of traditional biochemical models. We propose a method for developing mathematical models for complex systems, based on a genetic programming algorithm. The algorithm is aimed at obtaining models with a high predictive capacity, and considers an additional parameter fitting stage and an additional Jackknife stage for estimating the generalization error. We developed a model for the iron uptake system with a higher predictive capacity than classic biochemical models. This was observed both with the apical uptake dataset used for generating the model and with an independent initial rates dataset used to test the predictive capacity of the model. The model obtained is a function of time and the initial apical iron concentration, with a linear component that captures the global tendency of the system, and a non-linear component that can be associated to the movement of DMT1 transporters. The model presented in this paper allows the detailed analysis, interpretation of experimental data, and identification of key relevant components for this complex biological process. This general method holds great potential for application to the elucidation of biological mechanisms and their key components in other complex systems.

Iron deficiency is one of the most prevalent nutritional disorders worldwide. The standard treatment involves iron supplementation, but this task is challenging because of poor solubility and organoleptic issues. Moreover, the need to increase iron bioavailability represents a challenge for treating iron-related disorders. In this study, gastroresistance and iron intestinal absorption of an innovative granular formulation composed of ferric pyrophosphate, modified starch and phospholipids branded as Ferro Fosfosoma® was investigated. Gastroresistant properties were studied using standard protocols, and a bioaccessible fraction was obtained by exposing a food supplement to in vitro digestion. This fraction was used for investigating iron absorption in Caco-2 and human follicle-associated intestinal epithelium (FAE) models. Ferro Fosfosoma® showed an improved resistance to gastric digestion and higher intestinal absorption than ferric pyrophosphate salt used as a control in both models. In the FAE model, Ferro Fosfosoma® induces larger iron absorption than in the Caco-2 monolayer, most likely due to the transcytosis ability of M cells. The larger iron absorption in the Ferro Fosfosoma®-treated FAE model corresponds to higher ferritin level, proving physiological iron handling that was once delivered by granular formulation. Finally, the formulation did not induce any alterations in viability and barrier integrity. To conclude, Ferro Fosfosoma® favors iron absorption and ferritin expression, while preserving any adverse effects.

Aloe products are one of the top selling health-functional foods in Korea, however the adequate level of intake to achieve desirable effects are not well understood. The objective of this study was to determine the intestinal uptake and metabolism of physiologically active aloe components using in vitro intestinal absorption model. The Caco-2 cell monolayer and the everted gut sac were incubated with 5-50 microM of aloin, aloe-emodin, and aloesin. The basolateral appearance of test compounds and their glucuronosyl or sulfated forms were quantified using HPLC. The % absorption of aloin, aloe-emodin, and aloesin was ranged from 5.51% to 6.60%, 6.60% to 11.32%, and 7.61% to 13.64%, respectively. Up to 18.15%, 18.18%, and 38.86% of aloin, aloe-emodin, and aloesin, respectively, was absorbed as glucuronidated or sulfated form. These results suggest that a significant amount is transformed during absorption. The absorption rate of test compounds except aloesin was similar in two models; more aloesin was absorbed in the everted gut sac than in the Caco-2 monolayer. These results provide information to establish adequate intake level of aloe supplements to maintain effective plasma level.

In this work, we studied the intestinal absorption of a peptide with a molecular weight of 4353 Da (MEDI7219) and a protein having a molecular weight of 11 740 Da (PEP12210) in the rat intestinal instillation model and compared their absorption to fluorescein isothiocyanate (FITC)-labeled dextrans of similar molecular weights (4 and 10 kDa). To increase the absorption of the compounds, the permeation enhancer sodium caprate (C10) was included in the liquid formulations at concentrations of 50 and 300 mM. All studied compounds displayed an increased absorption rate and extent when delivered together with 50 mM C10 as compared to control formulations not containing C10. The time period during which the macromolecules maintained an increased permeability through the intestinal epithelium was approximately 20 min for all studied compounds at 50 mM C10. For the formulations containing 300 mM C10, it was noted that the dextrans displayed an increased absorption rate (compared to 50 mM C10), and their absorption continued for at least 60 min. The absorption rate of MEDI7219, on the other hand, was similar at both studied C10 concentrations, but the duration of absorption was extended at the higher enhancer concentration, leading to an increase in the overall extent of absorption. The absorption of PEP12210 was similar in terms of the rate and duration at both studied C10 concentrations. This is likely caused by the instability of this molecule in the intestinal lumen. The degradation decreases the luminal concentrations over time, which in turn limits absorption at time points beyond 20 min. The results from this study show that permeation enhancement effects cannot be extrapolated between different types of macromolecules. Furthermore, to maximize the absorption of a macromolecule delivered together with C10, prolonging the duration of absorption appears to be important. In addition, the macromolecule needs to be stable enough in the intestinal lumen to take advantage of the prolonged absorption time window enabled by the permeation enhancer.

Gastric inhibitory polypeptide (GIP) is released from the small intestine upon meal ingestion and increases insulin secretion from pancreatic β cells. Although the GIP receptor is known to be expressed in small intestine, the effects of GIP in small intestine are not fully understood. This study was designed to clarify the effect of GIP on intestinal glucose absorption and intestinal motility. Intestinal glucose absorption in vivo was measured by single-pass perfusion method. Incorporation of [(14)C]-glucose into everted jejunal rings in vitro was used to evaluate the effect of GIP on sodium-glucose co-transporter (SGLT). Motility of small intestine was measured by intestinal transit after oral administration of a non-absorbed marker. Intraperitoneal administration of GIP inhibited glucose absorption in wild-type mice in a concentration-dependent manner, showing maximum decrease at the dosage of 50 nmol/kg body weight. In glucagon-like-peptide-1 (GLP-1) receptor-deficient mice, GIP inhibited glucose absorption as in wild-type mice. In vitro examination of [(14)C]-glucose uptake revealed that 100 nM GIP did not change SGLT-dependent glucose uptake in wild-type mice. After intraperitoneal administration of GIP (50 nmol/kg body weight), small intestinal transit was inhibited to 40% in both wild-type and GLP-1 receptor-deficient mice. Furthermore, a somatostatin receptor antagonist, cyclosomatostatin, reduced the inhibitory effect of GIP on both intestinal transit and glucose absorption in wild-type mice. These results demonstrate that exogenous GIP inhibits intestinal glucose absorption by reducing intestinal motility through a somatostatin-mediated pathway rather than through a GLP-1-mediated pathway.

Hephaestin is a vertebrate multicopper ferroxidase important for the transfer of dietary iron from intestinal cells to the blood. Hephaestin is mutated in the sex-linked anemia mouse, resulting in iron deficiency. However, sex-linked anemia mice still retain some hephaestin ferroxidase activity. They survive, breed, and their anemia improves with age. To gain a better understanding of the role of hephaestin in iron homeostasis, we used the Cre-lox system to generate knockout mouse models with whole body or intestine-specific (Villin promoter) ablation of hephaestin. Both types of mice were viable, indicating that hephaestin is not essential and that other mechanisms, multicopper ferroxidase-dependent or not, must compensate for hephaestin deficiency. The knockout strains, however, both developed a microcytic, hypochromic anemia, suggesting severe iron deficiency and confirming that hephaestin plays an important role in body iron acquisition. Consistent with this, the knockout mice accumulated iron in duodenal enterocytes and had reduced intestinal iron absorption. In addition, the similarities of the phenotypes of the whole body and intestine-specific hephaestin knockout mice clarify the important role of hephaestin specifically in intestinal enterocytes in maintaining whole body iron homeostasis. These mouse models will serve as valuable tools to study the role of hephaestin and associated proteins in iron transport in the small intestine and other tissues.

Non-alcoholic fatty liver disease (NAFLD), in which abnormal lipid metabolism plays an important role in disease progression, has become a pandemic. Abnormal lipid metabolism, for example an increased fat intake, has been thought to be an initial factor leading to NAFLD. The small intestine is the main site of dietary lipid absorption. A number of clinical trials have shown that acupuncture has positive effects in the regulation of lipid metabolism, which is closely associated with the progression of NAFLD. We therefore hypothesized that, acupuncture can improve the conditions of NAFLD by regulating intestinal absorption of lipid.

Flying mammals present unique intestinal adaptations, such as lower intestinal surface area than nonflying mammals, and they compensate for this with higher paracellular absorption of glucose. There is no consensus about the mechanistic bases for this physiological phenomenon. The surface area of the small intestine is a key determinant of the absorptive capacity by both the transcellular and the paracellular pathways; thus, information about intestinal surface area and micro-anatomical structure can help explain differences among species in absorptive capacity. In order to elucidate a possible mechanism for the high paracellular nutrient absorption in bats, we performed a comparative analysis of intestinal villi architecture and enterocyte size and number in microchiropterans and rodents. We collected data from intestines of six bat species and five rodent species using hematoxylin and eosin staining and histological measurements. For the analysis we added measurements from published studies employing similar methodology, making in total a comparison of nine species each of rodents and bats. Bats presented shorter intestines than rodents. After correction for body size differences, bats had ~41% less nominal surface area (NSA) than rodents. Villous enhancement of surface area (SEF) was ~64% greater in bats than in rodents, mainly because of longer villi and a greater density of villi in bat intestines. Both taxa exhibited similar enterocyte diameter. Bats exceeded rodents by ~103% in enterocyte density per cm2 NSA, but they do not significantly differ in total number of enterocytes per whole animal. In addition, there is a correlation between SEF and clearance per cm2 NSA of L-arabinose, a nonactively transported paracellular probe. We infer that an increased enterocyte density per cm2 NSA corresponds to increased density of tight junctions per cm2 NSA, which provides a partial mechanistic explanation for understanding the high paracellular absorption observed in bats compared to nonflying mammals.

Loganin, iridoid glycosides, is the main bioactive ingredients in the plant Strychnos nux-vomica L. and demonstrates various pharmacological effects, though poor oral bioavailability in rats. In this study, the intestinal absorption mechanism of loganin was investigated using the human intestinal Caco-2 cell monolayer model in both the apical-to-basolateral (A-B) and the basolateral-to-apical (B-A) direction; additionally, transport characteristics were systematically investigated at different concentrations, pHs, temperatures, and potential transporters. The absorption permeability (PappAB) of loganin, which ranged from 12.17 to 14.78 × 10-6cm/s, was high at four tested concentrations (5, 20, 40, and 80μM), while the major permeation mechanism of loganin was found to be passive diffusion with active efflux mediated by multidrug resistance-associated protein (MRP) and breast cancer resistance protein (BCRP). In addition, it was found that loganin was not the substrate of efflux transporter P-glycoprotein (P-gp) since the selective inhibitor (verapamil) of the efflux transporter exhibited little effects on the transport of loganin in the human intestinal Caco-2 cells. Meanwhile, transport from the apical to the basolateral side increased 2.09-fold after addition of a MRP inhibitor and 2.32-fold after addition of a BCRP inhibitor. In summary, our results clearly demonstrate, for the first time, a good permeability of loganin in the human intestinal Caco-2 cell model and elucidate, in detail, the intestinal absorption mechanism and the effects of transporters on iridoid glycosides compounds.

Labrasol® is a self-emulsifying excipient that contains saturated polyglycolysed C6-C14 glycerides and this additive is known to improve the intestinal absorption of poorly absorbed drugs after oral administration. However, the effects of formulations similar to Labrasol® on the intestinal absorption of poorly absorbed drugs have not been characterized. In this study, we used insulin as a model peptide drug and examined the absorption-enhancing effects of Labrasol® and its related formulations for insulin absorption in rats. The co-administration of Labrasol-related formulations with insulin reduced the blood glucose levels. Among these formulations, Capryol 90 was the most effective additive. Notably, the effect of Capryol 90 was greater at pH 3.0 than at pH 7.0. Additionally, almost no mucosal damage was observed in the presence of these formulations, as these formulations did not affect the activity of lactate dehydrogenase (LDH) and the amount of protein released from the small intestine. In mechanistic studies, Capryol 90 improved the stability of insulin and suppressed the association with insulin under acidic conditions. The loosening of the tight junctions (TJs) could be the underlying mechanism by which Capryol 90 improved intestinal insulin absorption via a paracellular route. These findings suggest that Capryol 90 is an effective absorption enhancer for improving the intestinal absorption of insulin, without inducing serious damage to the intestinal epithelium.

Intestinal cell proliferation and cell production is best quantified by measuring the rate of accumulation of vincristine arrested metaphases in microdissected intestinal crypts to determine the crypt cell production rate (CCPR). Studies of intestinal adaptation could be much more informative if a valid measure of intestinal function could also be included. One such method is the water absorption capacity. The CCPR of the jejunum and intestinal water absorption were measured in 19 groups of hypo and hyperproliferative rats which were in a 'steady state' of cell production and turnover. The minimum values were obtained after hypophysectomy and the maximum values were observed in lactation. Crypt cell production rate and absorption were significantly correlated (p less than 0.001) to each other. There was a significant (p less than 0.001) correlation between both CCPR and absorption and dry weight of the intestinal segment studies and food intake. Body weight was a poor predictor of either CCPR or absorption. The combined study of CCPR and water absorption is thus a practical and convenient approach to the study of intestinal cell proliferation and intestinal adaptation.

Mineralocorticoid receptor (MR) has pathological roles in various cell types, including renal tubule cells, myocytes, and smooth muscle cells; however, the role of MR in intestinal epithelial cells (IECs) has not been sufficiently evaluated. The intestine is the sensing organ of ingested sodium; accordingly, intestinal MR is expected to have essential roles in blood pressure (BP) regulation.

In this paper, two novel liver-targeting nanoparticles, norcantharidin-loaded chitosan nanoparticles (NCTD-CS-NPs) and norcantharidin-associated galactosylated chitosan nanoparticles (NCTD-GC-NPs), were prepared using ionic cross-linkage. The physical properties, particle size, encapsulation efficiency, and drug release characteristics of the nanoparticles were investigated in vitro. To investigate the intestinal absorption mechanisms of the two preparations, a series of experiments was carried out, including in situ circulation method, in vitro everted gut sacs, and Ussing chamber perfusion technique. The absorption rate constants (Ka) of NCTD at different segments were found to be duodenum > jejunum > ileum > colon. The concentration had no distinctive effect on absorption kinetics, suggesting that drug absorption is not dose-dependent. The transport of NCTD was found to be inhibited by P-glycoprotein (P-gp) inhibitor, indicating that NCTD might be the substrate of P-gp. The order of the absorption enhancer effects were as follows: low molecular weight chitosan (CS-8kDa) > high molecular weight chitosan (CS-30kDa) > Poloxamer > sodium dodecyl sulfate (SDS) > sodium deoxycholate (SDCh). The results indicate that the chitosan nanoparticles can improve intestinal absorption of NCTD.