Lucifer Yellow Flux Research Articles

Dietary limonene promotes gastrointestinal barrier function via upregulating tight/adherens junction proteins through cannabinoid receptor type-1 antagonistic mechanism and alters cellular metabolism in intestinal epithelial cells.

Limonene, a dietary monocyclic monoterpene commonly found in citrus fruits and various aromatic plants, has garnered increasing interest as a gastrointestinal protectant. This study aimed to assess the effects of limonene on intestinal epithelial barrier function and investigate the involvement of cannabinoid receptor type-1 (CB1R) in vitro. Additionally, the study focused on examining the metabolomic changes induced by limonene in the intestinal epithelial cells (Caco-2). Initial analysis of transepithelial electrical resistance (TEER) revealed that both l-limonene and d-limonene, isomers of limonene, led to a dose- and time-dependent increase in TEER in normal cells and those inflamed by pro-inflammatory cytokines mixture (CytoMix). Furthermore, both types of limonene reduced CytoMix-induced paracellular permeability, as demonstrated by a decrease in Lucifer yellow flux. Moreover, d-limonene and l-limonene treatment increased the expression of tight junction molecules (TJs) such as occludin, claudin-1, and ZO-1, at both the transcriptional and translational levels. d-Limonene upregulates E-cadherin, a molecule involved in adherens junctions (AJs). Mechanistic investigations demonstrated that d-limonene and l-limonene treatment significantly inhibited CB1R at the protein, while the mRNA level remained unchanged. Notably, the inhibitory effect of d-limonene on CB1R was remarkably similar to that of pharmacological CB1R antagonists, such as rimonabant and ORG27569. d-limonene also alters Caco-2 cell metabolites. A substantial reduction in β-glucose and 2-succinamate was detected, suggesting limonene may impact intestinal epithelial cells' glucose uptake and glutamate metabolism. These findings suggest that d-limonene's CB1R antagonistic property could effectively aid in the recovery of intestinal barrier damage, marking it a promising gastrointestinal protectant.

Influences of Yogurt with Functional Ingredients from Various Sources That Help Treat Leaky Gut on Intestinal Barrier Dysfunction in Caco-2 Cells.

The impact of yogurts made with starter culture bacteria (L. bulgaricus and S. thermophilus) and supplemented with ingredients (maitake mushrooms, quercetin, L-glutamine, slippery elm bark, licorice root, N-acetyl-D-glucosamine, zinc orotate, and marshmallow root) that can help treat leaky gut were investigated using the Caco-2 cell monolayer as a measure of intestinal barrier dysfunction. Milk from the same source was equally dispersed into nine pails, and the eight ingredients were randomly allocated to the eight pails. The control had no ingredients. The Caco-2 cells were treated with isoflavone genistein (negative control) and growth media (positive control). Inflammation was stimulated using an inflammatory cocktail of cytokines (interferon-γ, tumor necrosis factor-α, and interleukin-1β) and lipopolysaccharide. The yogurt without ingredients (control yogurt) was compared to the yogurt treatments (yogurts with ingredients) that help treat leaky gut. Transepithelial electrical resistance (TEER) and paracellular permeability were measured to evaluate the integrity of the Caco-2 monolayer. Transmission electron microscopy (TEM), immunofluorescence microscopy (IM), and real-time quantitative polymerase chain reaction (RTQPCR) were applied to measure the integrity of tight junction proteins. The yogurts were subjected to gastric and intestinal digestion, and TEER was recorded. Ferrous ion chelating activity, ferric reducing potential, and DPPH radical scavenging were also examined to determine the yogurts' antioxidant capacity. Yogurt with quercetin and marshmallow root improved the antioxidant activity and TEER and had the lowest permeability in fluorescein isothiocyanate (FITC)-dextran and Lucifer yellow flux among the yogurt samples. TEM, IM, and RTQPCR revealed that yogurt enhanced tight junction proteins' localization and gene expression. Intestinal digestion of the yogurt negatively impacted inflammation-induced Caco-2 barrier dysfunction, while yogurt with quercetin, marshmallow root, maitake mushroom, and licorice root had the highest TEER values compared to the control yogurt. Yogurt fortification with quercetin, marshmallow root, maitake mushroom, and licorice root may improve functionality when dealing with intestinal barrier dysfunction.

Protective Mechanisms of Liquid Formulations for Gastro-Oesophageal Reflux Disease in a Human Reconstructed Oesophageal Epithelium Model.

PurposeA novel experimental design based on a human-reconstructed oesophageal epithelium (HO2E) model has been applied to quantitively assess the properties of a set of liquid formulations, Device A (Gerdoff® Protection), Device B (Esoxx® One), and Device C (Marial® gel) developed to form a temporary physical barrier on the oesophageal epithelium and modify epithelial permeability so to protect the oesophageal mucosa from refluxate components.MethodsThe formulations were applied to a prewetted HO2E model for 15 min. Then, a 0.5% caffeine solution was applied, and its penetration kinetics was assessed at 1 h and 2 h in acidic environments (pH= 3.3) to mirror exposure of the oesophageal mucosa to acidic reflux in GORD patients. Caffeine permeated into the basolateral compartment (evaluated by HPLC-UV) and Lucifer yellow (LY) permeability were quantified 15 min after application of the caffeine in acidic environments.ResultsAt the 15 min timepoint, Device A reduced caffeine permeation by 77.2% and LY flux by 30.4% compared to the untreated control and with a faster mode of action than that of the other liquid formulations. Transepithelial caffeine flux was reduced, albeit with different timing and efficiency, by all three compounds up to the end of the 2 hour experiment. At 1 h, Device A reduced the caffeine flux by 79.2%; Device B, by 67.2%; and Device C, by 37%.ConclusionThese results confirm the ability of the medical devices tested to interact with the oesophageal epithelium and create a temporary physical protective film for up to 2 hours after their application. The results underline differences in the mechanism of action of the three medical devices, with Device A performing faster than the other formulations. The overall results support the relevance of the reconstructed mucosal model to investigate oesophageal epithelium–product interactions and precisely differentiate liquid formulation performance.

Reparative Effects of Ethanol-Induced Intestinal Barrier Injury by Flavonoid Luteolin via MAPK/NF-κB/MLCK and Nrf2 Signaling Pathways.

Luteolin, a dietary flavonoid, has gained increasing interest as an intestinal protectant. This study aimed to evaluate the reparative effect of luteolin against ethanol-induced intestinal barrier damage in a Caco-2 cell monolayer model and the potential mechanisms. Luteolin attenuated ethanol-induced intestinal barrier injury, by increasing transepithelial monolayer resistance (TEER, 27.75 ± 14.75% of the ethanol group, p < 0.01), reducing Lucifer yellow flux (13.21 ± 1.23% of ethanol group, p < 0.01), and upregulating the expression of tight junction (TJ) proteins zonulin occludin-1 (ZO-1), occludin, and claudin-1 (37.963 ± 8.62%, 17.69 ± 7.35%, and 29.40 ± 8.08% of the ethanol group, respectively, p < 0.01). Further mechanistic studies showed that luteolin suppressed myosin light chain 2 (MLC) phosphorylation, myosin light chain kinase (MLCK) activation, nuclear factor kappa-B (NF-κB) nuclear translocation, and mitogen-activated-protein-kinase (MAPK) phosphorylation. Moreover, luteolin also acted as antioxidants indirectly by upregulating antioxidant-responsive-element (ARE) and nuclear factor (erythroid-derived 2)-like 2 (Nrf2) nuclear translocation to relieve ethanol-induced oxidative damage and TJ dysfunction. The results of the study indicate that luteolin may play an effective role in relieving intestinal barrier damage, and this effect is at least partially due to its indirect antioxidant capacity.

Evaluation of Human Esophageal Epithelium Permeability in Presence of Different Formulations Containing Hyaluronic Acid and Chondroitin Sulphate.

PurposeNew medical devices that contain hyaluronic acid (HA) and chondroitin sulphate (CS), with or without antacid components, have been developed for the treatment of gastroesophageal reflux disease (GERD) with the aim of improving oesophageal mucosal defences by creating a film on the oesophageal mucosa and acting as a mechanical barrier against the noxious components of refluxate, both acidic and basic.MethodsThe film-forming and protective efficacy of medical device A based on HA and CS plus aluminium hydroxide, device B combining HA and CS with magnesium trisilicate and device C with only the combination of HA and CS was tested on a reconstructed human oesophageal epithelium (HO2E/S/5) as a biological model in 2 different pH environments, neutral and acidic, to mimic realistic conditions. Caffeine penetration kinetics and Lucifer yellow (LY) permeability modifications induced by these products were compared to those induced by a negative control series (saline solution, code NC) and positive control series (white Vaseline, code V) under neutral and acidic pH conditions.ResultsUnder neutral and acidic pH conditions, compared to the negative control, all the products tested reduced (>80% and 85–90%, respectively) the caffeine passage, and no significant difference was observed among the products tested. Under neutral and acidic conditions, the LY permeabilities registered with device A and device C were not different from that registered with the negative control, while an LY flux% increase was calculated after 2 hrs of treatment (21.1%) with device B under acidic conditions.ConclusionThese results confirm the ability of the products tested to interact with the oesophageal epithelium in order to adhere and create a stable protective film for at least 2 hours after their homogeneous distribution on the epithelium surface. Further clinical studies are needed to test these devices in the topical treatment of gastroesophageal reflux symptoms.

Hsa-miR-375 promotes the progression of inflammatory bowel disease by upregulating TLR4.

To elucidate the biological function of hsa-miR-375 in the progression of inflammatory bowel disease (IBD) and the potential mechanism. Intestinal mucosa tissues of 26 IBD patients and 30 healthy volunteers who underwent colonoscopy were harvested for determining hsa-miR-375 level by quantitative Real-time polymerase chain reaction (qRT-PCR). Binding of hsa-miR-375 to toll-like receptor 4 (TLR4) was verified by the dual-luciferase reporter gene assay. Changes in the viability and apoptosis in Caco-2 cells influenced by hsa-miR-375 were examined by cell counting kit-8 (CCK-8) assay and flow cytometry, respectively. The regulatory effect of hsa-miR-375 on the intestinal epithelial barrier was examined by detecting transepithelial electrical resistance (TEER) and lucifer yellow flux. Relative levels of TLR4, nuclear factor-kappa B (NF-κB), zonula occludens-1 (ZO-1), occludin and inflammatory factors in Caco-2 cells were detected by qRT-PCR, Western blot and enzyme-linked immunosorbent assay (ELISA). Hsa-miR-375 was downregulated in intestinal mucosa tissues of patients with Crohn's disease (CD) and ulcerative colitis (UC). Knockdown of hsa-miR-375 decreased viability and TEER, but elevated apoptotic rate and lucifer yellow flux. Overexpression of hsa-miR-375 achieved the opposite trends. TLR4 was the direct downstream of hsa-miR-375, and its level was negatively mediated by hsa-miR-375. In addition, TLR4 level in Caco-2 cells was upregulated after LPS induction, while hsa-miR-375 level was unchangeable. Knockdown of hsa-miR-375 upregulated NF-κB and pro-inflammatory factors TNF-α, IL-1β, IL-6 and IL-8, and downregulated ZO-1, occludin and anti-inflammatory factor IL-10. Hsa-miR-375 is involved in the pathogenesis of IBD by upregulating TLR4 and inducing NF-κB activation.

L-Arginine supplementation prevents intestinal epithelial barrier breakdown under heat stress conditions by promoting nitric oxide synthesis

Heat stress (HS) induced by exposure to high ambient temperatures or prolonged excessive physical activities is known to primarily induce deleterious effects on the intestinal integrity by disrupting junctional complexes. Considering the association of l-arginine (l-Arg) with the improvement of gut function, the hypothesis of this study was to assess whether l-Arg supplementation can prevent the intestinal barrier disruption under HS conditions and to understand whether the l-Arg–induced effects are associated with maintaining nitric oxide (NO) as the major product of l-Arg metabolism. For this study, human colorectal adenocarcinoma (Caco-2) cells grown on Transwell inserts were pretreated with different l-Arg concentrations (0.4, 1, and 4 mmol/L), and after exposure to HS, markers of intestinal barrier integrity, stress-related markers, and NO levels were determined. l-Arg deprivation markedly increased the mRNA expression of heat shock protein 70 and heme-oxygenase-1 under HS conditions. The HS-induced drop in transepithelial electrical resistance values and increase in Lucifer Yellow permeability could be prevented by 4 mmol/L l-Arg supplementation. In turn, l-Arg mitigated the downregulation and delocalization of adherens junction protein E-cadherin in HS-exposed cells. NO and inducible NO synthase levels were significantly decreased in HS-exposed cells, whereas pretreatment with 4 mmol/L l-Arg prevented this decrease. Inhibition of inducible NO synthase by the NO synthase inhibitor l-NG-nitroarginine methyl ester abrogated the effect of l-Arg on preserving intestinal integrity under HS conditions as measured by transepithelial electrical resistance, Lucifer Yellow flux, and E-cadherin expression. In summary, l-Arg supplementation protects the intestinal epithelial integrity, at least partly, by maintaining NO synthesis under HS conditions.

Polyphenol-Rich Propolis Extracts Strengthen Intestinal Barrier Function by Activating AMPK and ERK Signaling.

Propolis has abundant polyphenolic constituents and is used widely as a health/functional food. Here, we investigated the effects of polyphenol-rich propolis extracts (PPE) on intestinal barrier function in human intestinal epithelial Caco-2 cells, as well as in rats. In Caco-2 cells, PPE increased transepithelial electrical resistance and decreased lucifer yellow flux. PPE-treated cells showed increased expression of the tight junction (TJ) loci occludin and zona occludens (ZO)-1. Confocal microscopy showed organized expressions in proteins related to TJ assembly, i.e., occludin and ZO-1, in response to PPE. Furthermore, PPE led to the activation of AMPK, ERK1/2, p38, and Akt. Using selective inhibitors, we found that the positive effects of PPE on barrier function were abolished in cells in which AMPK and ERK1/2 signaling were inhibited. Moreover, rats fed a diet supplemented with PPE (0.3% in the diet) exhibited increased colonic epithelium ZO-1 expression. Overall, these data suggest that PPE strengthens intestinal barrier function by activating AMPK and ERK signaling and provide novel insights into the potential application of propolis for human gut health.

Establishment of Caco-2 cell monolayer model and barrier permeability

Objective To establish and evaluate intestinal epithelial barrier model using Caco-2 cell so as to play a foundation for next study of barrier permeability. Methods Caco-2 cells were cultured in vitro then seeded into Transwell cell culture inserts.The permeability of the intestinal epithelial barrier was detected by transepithelial electrical resistance(TEER)and lucifer yellow flux, and verified by transmission electron microscope.Different concentrations of PAF(0, 50, 100, and 200 nmol/L)were exposed for 24 hours to Caco-2 monolayer when cultured 21 days.The tight junction was observed under transmission electron microscope.Assessment of ZO-1 protein localization and expression were detected by immunofluorescence and Western blot analysis. Results Cultured Caco-2 cell confluencd as monolayer with time passed.From 5th day, TEER increased, then reached 600Ω·cm2 at 15th day and lasted to 21st day, there was little flux of lucifer yellow, transmission electron microscopy also found cells differentiated better, had well-arranged villi and polarity alined as monolayer, forming completed tight junction which was the marker of intestinal epithelial barrier model in vitro.TEER decreased and lucifer yellow flux increased in cells exposed to PAF.The permeability reached the peak when exposed to 100 nmol/L PAF(P< 0.01), tight junction disrupted, ZO-1 protein expression downregulated, abnormal localization and distribution was assessed by immunofluorescence staining. Conclusion Cultured Caco-2 cells for 2-3w can be used to study intestinal epithelial barrier as a model in vitro.PAF increased intestinal epithelial permeability, which would correlate to the decreased protein expression and abnormal distribution of ZO-1. Key words: Human colon carcinoma cell line(Caco-2); Permeability; Transepithelial electrical resistance; Platelet-activating factor(PAF); ZO-1

Y-27632 inhibits ethanol-induced increase in intestinal epithelial barrier permeability

The present study aimed to investigate the effect of Y-27632, an inhibitor of the Rho-associated protein kinase (ROCK), which belongs to a family of downstream effectors of activated RhoA, on the ethanol-induced increase in permeability of the intestinal epithelial barrier (IEB). The in vitro model of IEB was established usiung Caco-2 cells, and the cells were pretreated with Y-27632 prior to treatment with ethanol for 60 min. Transepithelial resistance (TEER) and paracellular marker Lucifer yellow flux measurements were performed to assess the IEB permeability. The localization and expression of tight junction (TJ)-associated proteins were detected by immunofluorescence and western blot analysis, respectively. Y-27632 partially inhibited epithelial leakage and restored normal TEER values in the IEB. Immunofluorescence and western blot analysis results indicated that ethanol induces a shift from the insoluble to the soluble fractions of claudin-1, and that the ethanol-induced decreased expression of the zonula occludens-1 (ZO-1) protein is restored by Y-27632. In conclusion, our results suggest that ROCK may play a key role in the ethanol-induced increase of IEB permeability.

Protective Effect of Intestinal Trefoil Factor on Injury of Intestinal Epithelial Tight Junction Induced by Platelet Activating Factor

Intestinal barrier dysfunction plays an important role in the pathogenesis of inflammatory bowel disease (IBD). To evaluate the effect of intestinal trefoil factor (ITF) on increased intestinal permeability and its association with tight junction proteins, an in vitro intestinal epithelia barrier model was established with Caco-2 cells and treated with platelet-activating factor (PAF). We found that exposing cells to 0.3 M ITF (30 min before or 30 min after PAF treatment) attenuated the PAF-induced changes in transepithelial electrical resistance and Lucifer yellow flux. A quantitative RT-PCR and western blot analysis revealed that ITF suppressed PAF-induced downregulation of tight junction proteins claudin-1 and ZO-1 expression; furthermore, an abnormal localization and distribution of these proteins was inhibited, as assessed by immunofluorescence staining. These results suggest that ITF decreases mucosal permeability and shows potential as a therapy for treating IBD.

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.

The Electrophysiological Properties of the Mucosal Barrier in the Injured Mouse Jejunum in Ussing Chambers

We previously suggested in the in vitro mouse jejunum that when the primary epithelial barrier on the villi was largely disabled, a secondary mucosal barrier substitutes for the primary one. This study showed that the mucosal Ca(2+) removal did not further deteriorate the histology but increased transmural conductance and lucifer yellow flux, and decreased forskolin-induced potential difference. It is suggested that in this injury model the postulated secondary barrier is Ca(2+)-dependent and likely to be present on or immediately beneath the mucosal surface.

In Vitro Diffusion Barriers of the Mouse Jejunum in Ussing Chambers

Epithelial cells and their intercellular junctions play a primary role in the intestinal mucosal diffusion barrier to the invasion of noxious agents. Our previous study has shown that the mouse jejunum, when incubated in Ussing chambers for 4 h, exhibited morphological deterioration of the villi with denudation of the epithelia while it retained cAMP-induced potential difference (PD), suggesting that some subepithelial tissues had taken part in the barrier functions when the primary epithelial barrier on the villous surface was broken. To further characterize the barrier function of the jejunum in Ussing chambers, we measured the unidirectional lucifer yellow flux (J(LY)), which represents the permeation of a medium-sized anion, the transmural electrical conductance (G(t)), which reflects the permeation of electrolytes, mainly NaCl, and forskolin-induced PD. The values of J(LY), G(t) and forskolin-induced PD were not affected by removing the muscularis propria, suggesting that this tissue did not substantially contribute to the barrier. In addition, the values of J(LY), G(t) and forskolin-induced PD were not correlated with the degree of denudation of epithelial cells on the mucosal surface, supporting the notion that the main barrier function is constituted by the subepithelial tissues rather than the epithelium. Loosening the tissue (thereby approximating the free diffusion condition) by removing Ca(2+) from the bathing solution increased both J(LY) and G(t), with the former being larger than the latter. In conclusion, the mucosa propria and/or the submucosa constitute a diffusion barrier that restricted the permeation of lucifer yellow more tightly than NaCl in the injured jejunum incubated in Ussing chambers.

Evidence for restriction of fluid and solute movement across the glomerular capillary wall by the subpodocyte space

The glomerular filtration barrier (GFB) is generally considered to consist of three layers: fenestrated glomerular endothelium, glomerular basement membrane, and filtration slits between adjacent podocyte foot processes. Detailed anatomic examination of the GFB has revealed a novel abluminal structure, the subpodocyte space (SPS), identified as the labyrinthine space between the underside of podocyte cell body/primary processes and the foot processes. The SPS covers 50-65% of the filtration surface of the GFB, indicating that SPS may influence glomerular permeability. We have examined the contribution of the SPS to the permeability characteristics of the GFB using multiphoton microscopy techniques in isolated, perfused glomeruli and in the intact kidney in vivo. SPS were identified using this technique, with comparable dimensions to SPS examined with electron microscopy. The passage of the intermediate-weight molecule rhodamine-conjugated 10-kDa dextran, but not the low-weight molecule lucifer yellow ( approximately 450 Da), accumulated in SPS-covered regions of the GFB, compared with GFB regions not covered by SPS ("naked regions"). Net lucifer yellow flux (taken to indicate fluid flux) through identifiable SPS regions was calculated to be 66-75% of that occurring through naked regions. These observations indicate both ultrafiltration and hydraulic resistance imparted by the SPS, demonstrating the potential physiological contribution of the SPS to glomerular permeability.

Docosahexaenoic acid and eicosapentaenoic acid-enriched phosphatidylcholine liposomes enhance the permeability, transportation and uptake of phospholipids in Caco-2 cells

The influence of docosahexaenoic acid (DHA)- and eicosapentaenoic acid (EPA)-enriched phosphatidylcholine (PC) on the permeability, transport and uptake of phospholipids was evaluated in Caco-2 cells. The cells were grown on permeable polycarbonate transwell filters, thus allowing separate access to the apical and basolateral chambers. The monolayers of the cells were used to measure lucifer yellow permeability and transepithelial electrical resistance (TEER). Transcellular transportation of diphenylhexatriene (DPH) labeled-PC small unilamellar vesicles (SUV) from the apical to basolateral chamber, and uptake of the same SUV was monitored in the cell monolayers. Cell-membrane perturbation was evaluated to measure the release of lactate dehydrogenase and to determine the cell viability with sodium 2-(4-iodophenyl)-3-(4-nitrophenyl) -5-(2, 4-disulfophenyl)-2H-tetrazolium dye reduction assay. The lucifer yellow flux was 1.0 and 1.5 nmol/h/cm(2) with 50 microM PC, and 17.0 and 23.0 nmol/h/cm(2) with 100 microM PC when monolayers of Caco-2 cells were treated with DHA- and EPA-enriched PC, respectively. TEER decreased to 24 and 27% with 50 and 100 microM DHA-enriched PC, and to 25 and 30% with 50 and 100 microM EPA-enriched PC, respectively. Our results show that DHA- and EPA-enriched PC increases tight junction permeability across the Caco-2 cell monolayer whereas soy PC has no effect on tight junction permeability. Transportation and uptake of DHA- and EPA-enriched PC SUV differed significantly (P < 0.01) from those of soy PC SUV at all doses. We found that PC SUV transported across Caco-2 monolayer and was taken up by Caco-2 cells with very slight injury of the cell membrane up to 100 microM PC. Lactate dehydrogenase release and cell viability did not differ significantly between the treatment and control, emphasizing that injury was minimal. Our results suggest that DHA- and EPA-enriched PC enhance the permeability, transport and uptake of PC SUV across monolayers of Caco-2 cells.