Intestinal Caco-2 Cell Monolayers Research Articles

Transcellular transport of domoic acid across intestinal Caco-2 cell monolayers

The intestinal absorption mechanism of domoic acid (DA) was investigated using Caco-2 cells. DA is a tricarboxylic amino acid that contains a glutamic acid moiety, and causes deficits in short-term memory by binding to glutamate receptors as an agonist of glutamic acid. Caco-2 cell monolayers cultured on permeable membranes were incubated with 100μM DA on either the apical or basolateral side, and the transcellular transport of DA was measured. The transcellular transport of DA from the apical to basolateral side was about twofold that in the opposite direction. The transcellular transport of DA from the apical side was optimal at a neutral pH, and was temperature- and Cl−-dependent, but was Na+-independent. Coincubation of DA with 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS), an anion exchange inhibitor, significantly decreased the apical-to-basolateral transport of DA by 48%, and coincubation with probenecid (a non-specific anion transport inhibitor) significantly decreased the transport of DA by 31%. In contrast, coincubation with glutamic acid, succinic acid (a dicarboxylic acid), or citric acid (a tricarboxylic acid) did not decrease the transport of DA. These results suggest that the apical-to-basolateral transport of DA across the Caco-2 cell monolayers is mediated by DIDS-sensitive anion transporters.

Intestinal safety of water-soluble beta-cyclodextrins in paediatric oral solutions of spironolactone: effects on human intestinal epithelial Caco-2 cells.

Effects of water-soluble beta-cyclodextrins (beta CDs) on intestinal epithelial integrity were investigated, to establish the safe use of these beta CDs as solubilizers of spironolactone in paediatric enteral solutions. Mannitol permeability and transepithelial resistance (TER) of human intestinal epithelial Caco-2 cell monolayers during exposure to dimethyl-beta-cyclodextrin (DM beta CD), hydroxypropyl-beta-cyclodextrin (HP beta CD) and sulphobutyl ether beta-cyclodextrin (SBE beta CD) were followed. Staining methods were used to discern cells with damaged membranes and to study the integrity of cytoskeletal actin and tight junctions. Cytotoxicity of the beta CDs was tested by effects on intracellular dehydrogenase activity. Exposure to HP beta CD and SBE beta CD solutions had only minor effects on the integrity of Caco-2 cell monolayers. In contrast, DM beta CD clearly increased the epithelial permeability for the hydrophilic marker [14C]mannitol across Caco-2 monolayers, decreased TER and showed a dose-dependent cytotoxicity. According to staining, DM beta CD increased the permeability of the apical cell membrane without discernable effects on cytoskeletal actin. HP beta CD and SBE beta CD appear to be safe additives for use in enteral spironolactone preparations with respect to their acute local effects on epithelial integrity.

Effect of Bioactive Dietary Polyphenols on Zinc Transport across the Intestinal Caco-2 Cell Monolayers

Polyphenolic compounds are known to possess many beneficial health effects, including the antioxidative activities of scavenging reactive oxygen species and chelating metals, such as iron and zinc. Tea and red wine are thought to be important sources of these compounds. However, some polyphenolic compounds can also reduce the absorption of iron, and possibly other trace metals, when included in a diet. There is very little information on the effect of dietary polyphenolic compounds on the status of trace elements other than iron. The effects of epigallocatechin-3-gallate (EGCG), green tea extract (GT), and grape seed extract (GSE) on the absorption of (65)Zn were examined and compared with their effects on (55)Fe absorption in human intestinal Caco-2 cells grown on microporous membrane inserts. The levels of EGCG, GT, and GSE used in this study were within physiological ranges and did not affect the integrity of the Caco-2 cell monolayers. GSE significantly (P < 0.05) reduced zinc transport across the cell monolayer, and the decreased zinc transport was associated with a reduction in apical zinc uptake. However, EGCG and GT did not alter zinc absorption. In contrast, the polyphenolic compounds in EGCG, GT, and GSE almost completely blocked transepithelial iron transport across the cell monolayer. The effect of GSE on zinc absorption was very different from that on iron absorption. Whereas GSE decreased zinc absorption by reducing apical zinc uptake, the polyphenolic compounds inhibited iron absorption by enhancing apical iron uptake. GSE inhibited zinc absorption similarly to that observed for phytate. Phytate significantly (P < 0.05) decreased transepithelial zinc transport by reducing apical zinc uptake. The inhibition of zinc absorption may be due to the presence of procyanidins in GSE, which bind zinc with high affinity and block the transport of zinc across the apical membrane of enterocytes. Further research on the absorption of zinc as zinc-polyphenol complexes and free zinc should provide further insight into the process of dietary zinc absorption in the presence of GSE and other bioactive dietary polyphenols. The present study suggests that some individuals should consider their zinc status if they regularly consume procyanidin-containing foods in their diet. However, further studies, especially in vivo studies, are needed to confirm these results.

Intestinal transport of bis(12)‐hupyridone in Caco‐2 cells and its improved permeability by the surfactant Brij‐35

The objective of the present study was to elucidate the mechanisms of intestinal transport of bis(12)-hupyridone (B12H) to predict its oral bioavailability. The effect of the B12H concentration and the contribution of the drug efflux transporters, P-glycoprotein (P-gp or ABCB1) and multidrug resistance-associated proteins (MRPs or ABCC) on B12H absorption were measured and evaluated using the human intestinal epithelial Caco-2 cell monolayer in the presence of transporter inhibitors. The results indicated that B12H was absorbed in a dose-dependent manner at concentrations ranging from 132 to 264 µM. However, only apical efflux was observed in the directional transport studies for B12H below 88 µM (P(app) (AP-to-BL): virtually zero; P(app) (BL-to-AP): 1.591 ± 0.071 × 10(-5) cm s(-1) ). P-gp and mixed P-gp/MRP inhibitors significantly increased the absorptive transport (P(app) (AP-to-BL)) to 0.619 ± 0.018 × 10(-5) and 0.608 ± 0.025 × 10(-5) cm s(-1) , respectively, while decreasing secretory transport (P(app) (BL-to-AP)) by >75%. A multiple-MRP inhibitor, probenecid, increased the P(app) (AP-to-BL) to 0.329 ± 0.015 × 10(-5) cm s(-1) while decreasing the P(app) (BL-to-AP) by 50%. Another multiple-MRP inhibitor, indomethacin, only modestly decreased the P(app) (BL-to-AP) by ∼30% and had no effect on the absorptive transport (P(app) (AP-to-BL): virtually zero). In addition, the effect of various pharmaceutical excipients (e.g. Pluronic F-68, Tween-80 and Brij-35) on B12H transport was determined and compared. Among them, Brij-35 effectively enhanced B12H absorption at a concentration lower than its critical micelle concentration (CMC, 60 µM). Therefore, Brij-35 can be used as a potential enhancer to improve intestinal absorption of B12H for oral administration.

Kaempferol Enhances Intestinal Barrier Function through the Cytoskeletal Association and Expression of Tight Junction Proteins in Caco-2 Cells

Kaempferol, a natural flavonoid present in fruits, vegetables, and teas, provides beneficial effects for human health. We investigated the promotive effect of kaempferol on tight junction (TJ) barrier integrity in human intestinal Caco-2 cell monolayers. Transepithelial electrical resistance (TER; a TJ integrity marker) across the monolayers rapidly and markedly increased during the first 6 h after kaempferol administration and remained elevated until 48 h without any changes in the lucifer yellow or dextran fluxes. Immunoblot analysis demonstrated that kaempferol promoted the actin cytoskeletal association of the TJ proteins, zonula occludens (ZO)-1, ZO-2, occludin, claudin-1, claudin-3, and claudin-4, which was associated with the increase in TER. Kaempferol-mediated ZO-2 and claudin-4 expression was relatively smaller or occurred later than the kaempferol-promoted cytoskeletal association. Confocal microscopy showed that kaempferol-induced assembly of occludin and claudin-3 occurred at the TJ at 6 h postadministration. Extraction of cholesterol with methyl-β-cyclodextrin suppressed the kaempferol-mediated increase in TER. Sucrose density gradient centrifugation showed that the kaempferol treatment increased the TJ protein distributions in the cholesterol-rich lipid microdomain fraction. Taken together, these results indicate that the membrane lipid microdomain is involved in the kaempferol-mediated promotion of TJ protein assembly and intestinal TJ integrity.

BPR0C261 is a novel orally active antitumor agent with antimitotic and anti‐angiogenic activities

BPR0C261 is a synthetic small molecule compound cytotoxic against human cancer cells and active prolonging the lifespan of leukemia mice. In the present study, we further investigated the mechanisms of its anticancer action and found that BPR0C261 inhibited microtubule polymerization through interacting with the colchicine binding sites on tubulins, disrupted microtubule arrangement and caused cell cycle arrest at G(2)/M phase in cancer cells. BPR0C261 also inhibited the clonogenic growths of cancer cells and showed cytotoxicity against human cervical cancer cells of multidrug-resistant phenotype. In addition, BPR0C261 concentration-dependently inhibited the proliferation and migration of HUVECs and disrupted the endothelial capillary-like tube formations in HUVEC and rat aorta ring cultures. Given orally, BPR0C261 inhibited angiogenesis in s.c. implanted Matrigel plugs in mice. Notably, its IC(50) values against the endothelial cell growths were approximately 10-fold lower than those against the cancer cells. It was found orally absorbable in mice and showed a good oral bioavailability (43%) in dogs. BPR0C261 permeated through the human intestinal Caco-2 cell monolayer, suggesting oral availability in humans. Orally absorbed BPR0C261 distributed readily into the s.c. xenografted tumors in nude mice in which the tumor tissue levels of BPR0C261 were found oral dose-dependent. BPR0C261 showed in vivo activities against human colorectal, gastric, and nasopharyngeal tumors in nude mice. Most interestingly, the combination of BPR0C261 plus cisplatin synergistically prolonged the lifespans of mice inoculated with murine leukemia cells. Thus, BPR0C261 is a novel orally active tubulin-binding antitumor agent with antimitotic, apoptosis-inducing, and vasculature disrupting activities.

Erythropoietin regulates intestinal iron absorption in a rat model of chronic renal failure

Erythropoietin is produced by the kidney and stimulates erythropoiesis; however, in chronic renal disease its levels are reduced and patients develop anemia that is treatable with iron and recombinant hormone. The mechanism by which erythropoietin improves iron homeostasis is still unclear, but it may involve suppression of the iron regulatory peptide hepcidin and/or a direct effect on intestinal iron absorption. To investigate these possibilities, we used the well-established 5/6th nephrectomy rat model of chronic renal failure with or without human recombinant erythropoietin treatment. Monolayers of human intestinal Caco-2 cells were also treated with erythropoietin to measure any direct effects of this hormone on intestinal iron transport. Nephrectomy increased hepatic hepcidin expression and decreased intestinal iron absorption; these effects were restored to levels found in sham-operated rats on erythropoietin treatment of the rats with renal failure. In Caco-2 cells, the addition of erythropoietin significantly increased the expression of apical divalent metal transporter 1 (DMT1) and basolateral ferroportin and, consequently, iron transport across the monolayer. Taken together, our results show that erythropoietin not only exerts a powerful inhibitory action on the expression of hepcidin, thus permitting the release of iron from reticuloendothelial macrophages and intestinal enterocytes, but also acts directly on enterocytes to increase iron absorption.

Permeation of Disaccharides Derived from Chondroitin Sulfate through Human Intestinal Caco-2 Cell Monolayersviathe Paracellular Pathway

The intestinal absorption of chondroitin sulfate (CS) and its hydrolysate, particularly the disaccharides (Di-CS), was investigated by using human intestinal epithelial Caco-2 cell monolayers. Although the transepithelial transport of CS was not detectable, dose- and time-dependent transport of Di-CS was observed. The transport of Di-CS was proved to be energy-independent and its permeability increased inversely with the transepithelial electrical resistance (TER) value of the Caco-2 cell monolayers. The transport rate for Di-CS was also increased by treating the cell monolayers with such tight junction-opening agents as interferon-gamma. These results suggest that Di-CS permeated across the Caco-2 cell monolayers mainly via the paracellular pathway. The permeability of Caco-2 cell monolayers to authentic chondroitin 4-sulfate disaccharides (Di-4S), chondroitin 6-sulfate disaccharides (Di-6S) and chondroitin 0-sulfate disaccharides (Di-0S) was almost same, suggesting that sulfation did not affect the transport rate of Di-CS.

Metabolites of Galangin by 2,3,7,8-Tetrachlorodibenzo-p-dioxin-Inducible Cytochrome P450 1A1 in Human Intestinal Epithelial Caco-2 Cells and Their Antagonistic Activity toward Aryl Hydrocarbon Receptor

Galangin, a dietary flavonoid, inhibited cytochrome P450 1A1 (CYP1A1) expression induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). This inhibitory activity remained after permeating human intestinal epithelial Caco-2 cell monolayers, but was reduced when galangin permeated TCDD-pretreated Caco-2 cells. The present study tested whether TCDD affected the intestinal metabolism of flavonoids. LC-MS/MS analyses showed that galangin and two galangin glucuronoconjugates were reduced 0.7-fold, whereas kaempferol (a galangin oxidate) and kaempferol glucuronoconjugate were increased 1.5-fold by permeating TCDD-pretreated Caco-2 cells, as compared to untreated Caco-2 cells. An assay using recombinant human CYP1A1 and the CYP1A1 inhibitor alpha-naphthoflavone revealed that CYP1A1 oxidized galangin to kaempferol. These results indicated that galangin was metabolized to kaempferol by TCDD-inducible CYP1A1 in Caco-2 cells. A previous study revealed that kaempferol had much weaker inhibitory activity than galangin toward TCDD-induced CYP1A1 expression. Therefore, the oxidative metabolism of galangin to kaempferol in TCDD-pretreated Caco-2 cells implicated reduction in the inhibitory activity of galangin.

Transport of Hop Bitter Acids across Intestinal Caco-2 Cell Monolayers

Several health-beneficial properties of hop bitter acids have been reported (inhibition of bone resorption and anticarcinogenic and anti-inflammatory activities); however, scientific data on the bioavailability of these compounds are lacking. As a first approach to study the bioavailability, the epithelial transport of hop alpha- and beta-acids across Caco-2 monolayers was investigated. Hop acids were added either to the apical or to the basolateral chamber and, at various time points, amounts transported to the receiving compartment were determined. The monolayer integrity control was performed by using marker compounds (atenolol and propranolol), transepithelial electrical resistance (TEER) measurement, and determination of the fluorescein efflux. The TEER and fluorescein efflux confirmed the preservation of the monolayer integrity. The membrane permeability of the alpha-acids (apparent permeability coefficients for apical to basolateral transport (P(appAB)) ranged from 14 x 10(-6) to 41 x 10(-6) cm/s) was determined to be substantially higher than that of the beta-acids (P(appAB) values ranging from 0.9 x 10(-6) to 2.1 x 10(-6) cm/s). Notably, the beta-acids exhibited significantly different bidirectional P(app) values with efflux ratios around 10. The involvement of carrier-mediated transport for beta-acids (active efflux pathway by P-gp, BCRP, and/or MRP-2 type efflux pumps) could be confirmed by transport experiments with specific inhibitors (verapamil and indomethacin). It appears that alpha-acids are efficiently absorbed, whereas the permeability of beta-acids is low. Limiting factors in the absorption of beta-acids could involve P-gp and MRP-2 type efflux transporters and phase II metabolism.

Role of n−6 PUFA derived lipid mediators on epithelial barrier function in intestinal Caco-2 cell monolayers

An abstract is not available for this content. As you have access to this content, full HTML content is provided on this page. A PDF of this content is also available in through the ‘Save PDF’ action button.

Synthesis, characterization and evaluation of thiolated quaternary ammonium-chitosan conjugates for enhanced intestinal drug permeation

In a previous report quaternary ammonium-chitosan conjugates (N +-Chs) endowed with intestinal drug permeability-enhancing properties were described. They are characterized by short pendant chains of n adjacent diethyl-dimethylene-ammonium groups substituted onto the primary amino group of the chitosan (Ch) repeating units. In the present work two N +-Chs, one having DS (degree of substitution) = 59.2 ± 4.5%, n = 1.7 ± 0.1 (N +(60)-Ch), the other one having DS = 40.6 ± 1.3%, n = 3.0 ± 0.2 (N +(40)-Ch) were used to synthesize novel multifunctional non-cytotoxic Ch derivatives, each carrying thiol along with quaternary ammonium groups (N +-Ch–SH), with increased potential to enhance transepithelial drug transport. They have been obtained by transforming the residual free amino groups of N +(60)-Ch and N +(40)-Ch into 3-mercaptopropionamide moieties. The former yielded 4.5 ± 0.7% thiol-bearing groups, the latter, 5.2 ± 1.1% of such groups, on a Ch repeating unit basis. The multifunctional derivatives have improved the ability of the parent N +-Chs to enhance the permeability of the water-soluble macromolecular fluorescein isothiocyanate dextran, MW 4400 Da (FD4) and that of the lipophilic dexamethasone (DMS) across the excised rat intestinal mucosa and Caco-2 cell monolayer, respectively. The data from the present work altogether point to a synergism of quaternary ammonium and thiol groups to improve the intestinal drug absorption enhancing properties of the multifunctional Ch derivatives.

Stereoselective transport and uptake of propranolol across human intestinal Caco‐2 cell monolayers

The transport and uptake of individual propranolol (PPL) enantiomers were studied in human intestinal Caco-2 cell monolayers, and a reversed-phase HPLC-UV assay was used for quantitative analysis. S-PPL and R-PPL across Caco-2 cell monolayers was determined in the concentrations range of 10-500 microM in both apical (AP) to basolateral (BL) and BL to AP directions. S-PPL exhibited greater permeability than R-PPL in the AP to BL direction, whereas in the BL to AP direction S-enantiomer transported less than R-enantiomer. Uptake of R-PPL was significantly higher than that of S-PPL either from AP side or from BL side. The statistically significant differences in uptake were observed at the concentrations range from 10 to 50 microM. Furthermore, the apparent Michaelis constant (K(m)) and maximal velocity (V(max)) also showed significant difference between the two enantiomers. Moreover, the AP to BL transport of PPL enantiomer was markedly decreased by lowering the pH of the apical side but it did not affect the stereoselectivity of PPL across Caco-2 cell monolayers. The transport and uptake of PPL in the BL to AP direction was not influenced by several protein inhibitors. The results suggest that PPL enantiomers showed stereoselective transport and uptake across the Caco-2 cell monolayers. A special transport mechanism capable of directing the PPL enantiomers might be present in the Caco-2 monolayers.

Quercetin Enhances Intestinal Barrier Function through the Assembly of Zonnula Occludens-2, Occludin, and Claudin-1 and the Expression of Claudin-4 in Caco-2 Cells

Dietary flavonoids provide various beneficial effects for our health. We investigated the promotive effects of quercetin and myricetin on the intestinal barrier function in human intestinal Caco-2 cell monolayers. Transepithelial electrical resistance (TER) across the monolayers increased rapidly during incubation with quercetin, peaking at 6 h. Lucifer yellow flux, a paracellular marker, was dose-dependently lower after quercetin and myricetin treatments, although quercetin exhibited a more potent effect. Immunoblot analysis of tight junction (TJ) proteins revealed that zonnula occludens (ZO)-2, occludin, and claudin-1 were distributed to the actin cytoskeleton fraction by quercetin without increasing their respective whole-cell levels and this distribution was correlated with the increases in TER. The claudin-4 level was elevated by quercetin in both the cytoskeleton fraction and whole cells after 12 h. Confocal microscopy showed the assembly of claudin-1 and -4 at the TJ by quercetin. An inhibitor of protein kinase Cδ (PKCδ), rottlerin, enhanced the barrier function with changes in the distribution and expression of TJ proteins in a manner very similar to that of quercetin. Phosphorylation of PKCδ indicating the enzymatic activity in the cells was decreased by quercetin after 1 h. In the kinase assay, quercetin exhibits direct inhibition of the PKCδ isoform. This study demonstrates that quercetin enhances the intestinal barrier function through the assembly of ZO-2, occludin, and claudin-1 by inhibiting PKCδ and the increase in claudin-4 expression has an additional role after 12 h.

Transepithelial transport of ambroxol hydrochloride across human intestinal Caco-2 cell monolayers

This study aimed i) to characterize the transepithelial transport of the mucolytic agent ambroxol hydrochloride across the intestinal barrier, ii) to classify the ambroxol according to Biopharmaceutics Classification System (BCS) and iii) to predict ambroxol absorption in humans. Transport of ambroxol (100, 300 and 1000 micromol/l) was studied in a human colon carcinoma cell line Caco-2 in apical to basolateral and basolateral to apical direction, under iso-pH 7.4 and pH-gradient (6 vs. 7.4) conditions. The relative contribution of the paracellular route was estimated using Ca2+-free transport medium. Ambroxol samples from receiver compartments were analysed by HPLC with UV detection (242 nm). Results showed that ambroxol transport is linear with time, pH-dependent and direction-independent, displays non-saturable (first-order) kinetics. Thus, the transport seems to be transcellular mediated by passive diffusion. Estimated high solubility and high permeability (P(app) = 45 x 10(-6) cm/s) of ambroxol rank it among well absorbed compounds and class I of BCS. It can be expected that the oral dose fraction of ambroxol absorbed in human intestine is high.

A study of intestinal absorption of bicyclol in rats: active efflux transport and metabolism as causes of its poor bioavailability

To determine the possible mechanism of poor bioavailability of bicyclol, and clarify the respective contribution of P- glycoprotein (P-gp) and Cytochrome 3A (CYP3A). Rat in situ single-pass intestinal perfusion and Caco-2 cell monolayer model with selective inhibitors of CYP3A and P-gp were employed. In rat intestinal perfusion, bicyclol (50 microM) appearance in mesenteric blood (Pblood) was increased 3, 12 and 16-fold after addition of inhibitors of P-gp (LSN335984), CYP3A (troleandomycin, TAO) or P-gp and CYP3A (Cyclosporin A, CsA), respectively, whereas permeability of midazolam (CYP3A substrate only) was unchanged after addition of LSN335984 and increased 5 fold after addition of TAO. Moreover, the cumulative amount of bicyclol in mesenteric blood was increased at concentration range 10-100 microM of bicyclol in perfusate. The basolateral to apical permeability value of bicyclol in Caco-2 monolayer was significantly deceased by LSN335984 and CsA. The poor bioavailability of bicyclol is mostly due to P-gp mediated efflux and metabolism by CYP3A in intestine, with CYP3A making more contribution than P-gp.

Transport and metabolism of flavonoids from Chinese herbal remedy Xiaochaihu-tang across human intestinal Caco-2 cell monolayers1

To investigate the limiting factors for oral bioavailability of flavonoids derived from the Chinese herbal remedy Xiaochaihu-tang. The investigational flavonoids included baicalin, wogonoside, oroxylin-A 7-O-beta-Dglucopyranosiduronide, liquiritin, liquiritin apioside, isoliquiritin, and isoliquiritin apioside, as well as their aglycones baicalein, wogonin, oroxylin-A, liquiritigenin, and isoliquiritigenin. Caco-2 cell monolayers were used and the apparent permeability both in apical to basolateral and basolateral to apical directions was measured for each investigational compound. Meanwhile, chemoinformatics was carried out to provide insight into the mechanism governing the permeability. In addition, carrier-mediated transport with or without inhibitors, as well as the metabolism by conjugation, was also examined with Caco- 2 cell monolayers for the flavonoids. The investigational flavonoid aglycones exhibited favorable membrane permeability, but efficient glucuronidation and/or sulfation by the enterocytes may limit their bioavailability. For the flavonoid glycosides, their poor membrane permeability was found to be caused by high hydrogen-bonding potential. Among the glycosides, oroxylin-A 7-O-beta- D-glucopyranosiduronide, isoliquiritin, and isoliquiritin apioside were transported under the mediation of the efflux transporters multidrug resistance-associated protein and/or P-glycoprotein. The limiting factors of oral bioavailability for the flavonoids derived from Xiaochaihu-tang appeared to include poor membrane permeability, significant efflux, and efficient intestinal metabolism by conjugation.

Intestinal Transport of Free Anthraquinones in Caco-2 Cell Model

AIM: To investigate the transepithelial transport of the six free anthraquinones 1,8-dihydroxyanthraquinone (DQ),chrysophanol (CP),emodin (EN),rhein (RN),aloe-emodin (AE) and citreorosein (CN) in the human intestinal Caco-2 cell monolayer model. METHODS: The monolayers of the human intestinal epithelial cancer cell line Caco-2 were incubated with DQ,CP,EN,RN,AE and CN to model its intestinal absorption and transport,respectively. The determination of the compounds was performed by high-performance liquid chromatography. RESULTS: The apparent permeability coefficients (Papp) in the Caco-2 cell monolayers for DQ,CP,EN,RN,AE and CN were (2.16±0.38)×10-6,(3.00±0.09)×10-7,(1.99±0.94)×10-6,(4.58±0.20)×10-6,(6.24±0.19)×10-6 and (7.34±0.61)×10-6 cm ·s-1 from the apical-to-basolateral direction,respectively,and (3.20±0.10)×10-6,(4.50 ± 0.02)×10-7,(2.22 ± 0.62)×10-6,(4.94 ± 0.39)×10-6,(2.87 ± 0.18)×10-6 and (6.80 ± 0.37)×10-6 cm ·s-1 from the basolateral-to-apical direction,respectively. The Papp values of DQ,EN,RN,AE and CN were about 10 times less than those (6.30 ± 0.26)×10-5 cm ·s-1 of propranolol as a reference standard compound of high permeability and the main mechanism of intestinal absorption by passive diffusion,and about 10 times higher than those ( 1×10-7 cm ·s-1) of atenolol as a reference standard compound of poor permeability and the main mechanism of intestinal absorption by passive diffusion. Whereas,the Papp value of CP was at nearly the same magnitude with those of atenolol. In the presence of iodoacetamide,the Papp of AE were not affected in both apical-to-basolateral and basolateral-to-apical directions. CONCLUSION: The intestinal absorption of all test compounds was passive diffusion as the dominating process in Caco-2 cell monolayer model. DQ,EN,RN,AE and CN will be moderately absorbed compounds and CP was estimated to be a poorly absorbed compound.

Absorption of Black Currant Anthocyanins by Monolayers of Human Intestinal Epithelial Caco-2 Cells Mounted in Ussing Type Chambers

Anthocyanins (ACNs) have been reported to have multiple biological properties imparting benefits to human health. Their role in human nutrition, however, needs to be related to biokinetic data, such as bioavailability. The purpose of the present study was to focus on the potential absorption of black currant ( Ribes nigrum L.) ACNs. Caco-2 monolayers were used as an in vitro model of the absorptive intestinal epithelium. For absorption studies, Caco-2 cells grown on permeable filters were mounted into Ussing type chambers. The monolayer integrity was monitored by measuring the transepithelial electrical resistance (TEER). Luminal to serosal transport of ACNs was examined by comparing ACN disappearance from the luminal solution of Ussing chambers not containing any inserts (control chambers) with that of Ussing chambers containing inserts. ACNs (C total ACN approximately 180 microM) were not detected in any serosal solution. However, it was shown that ACNs disappeared from the luminal side, not due to ACN degradation processes but rather--at least in part--due to physiological actions of the cells. The luminal net disappearance of ACNs was calculated (max(t20 min) approximately 11% for total ACNs) and labeled as "absorption efficiency". This apical transport might occur to a much larger extent than the further translocation across the basolateral membrane. Thus, cell metabolism and translocation across the basolateral membrane may be the key determinants of ACN absorption and bioavailability.

High‐ and low‐capacity taurine transport across the luminal membrane of the small intestinal epithelium

Taurine is a “semi‐essential” beta‐amino acid; yet it is not clear which transporter(s) mediates taurine uptake across the small intestinal luminal membrane. The three intestinal beta‐amino acid transporters [taurine transporter TauT (SLC6A6), proton‐coupled amino acid transporter PAT1 (SLC36A1), and ATB0,+ transporter (SLC6A14)] were expressed in Xenopus laevis oocytes, and radiolabeled β‐alanine or taurine uptake characterized. TauT and PAT1 both transport taurine and β‐alanine whereas ATB0,+ transports only β‐alanine. To determine the relative contribution of each carrier to small intestinal transport, β‐alanine and taurine uptakes were measured across the apical membrane of human intestinal Caco‐2 cell monolayers. Use of selective inhibitors and ion substitutions identified roles for PAT1 and TauT (but not ATB0,+) in small intestinal uptake of these beta‐amino acids (ATB0,+ function may be more important in the colon). Under physiological conditions (apical pH 6.5, presence of Na+), the relative contributions of PAT1 and TauT to taurine uptake were dependent upon the taurine concentration. The high‐capacity, low‐affinity PAT1 may provide the major route for small intestinal taurine absorption from taurine‐rich foods whereas the low‐capacity, high‐affinity TauT may scavenge small quantities of taurine from the lumen. Supported by The Wellcome Trust (grant number 078640/Z/05/Z).