Alexandrine During

Carotenoid uptake and secretion by CaCo-2 cells β-carotene isomer selectivity and carotenoid interactions

In presence of oleate and taurocholate, differentiated CaCo-2 cell monolayers on membranes were able to assemble and secrete chylomicrons. Under these conditions, both cellular uptake and secretion into chylomicrons of β-carotene (β-C) were curvilinear, time-dependent (2–16 h), saturable, and concentration-dependent (apparent Km of 7–10 μM) processes. Under linear concentration conditions at 16 h incubation, the extent of absorption of all-trans β-C was 11% (80% in chylomicrons), while those of 9-cis- and 13-cis-β-C were significantly lower (2–3%). The preferential uptake of the all-trans isomer was also shown in hepatic stellate HSC-T6 cells and in a cell-free system from rat liver (microsomes), but not in endothelial EAHY cells or U937 monocyte-macrophages. Moreover, extents of absorption of α-carotene (α-C), lutein (LUT), and lycopene (LYC) in CaCo-2 cells were 10%, 7%, and 2.5%, respectively. Marked carotenoid interactions were observed between LYC/β-C and β-C/α-C. The present results indicate that β-C conformation plays a major role in its intestinal absorption and that cis isomer discrimination is at the levels of cellular uptake and incorporation into chylomicrons. Moreover, the kinetics of cellular uptake and secretion of β-C, the inhibition of the intestinal absorption of one carotenoid by another, and the cellular specificity of isomer discrimination all suggest that carotenoid uptake by intestinal cells is a facilitated process.

Dietary polyphenols can modulate the intestinal inflammatory response

Inflammatory bowel diseases (IBD) arise from multiple causes, including environmental factors, gut microflora, immunity, and genetic predispositions. In the course of IBD, immune homeostasis and intestinal mucosa barrier integrity are impaired. Among natural preventive treatments that have been identified to date, polyphenols appear as promising candidates. They have been shown to protect against several diseases, including cardiovascular diseases and cancers, and they have anti-inflammatory properties in non-intestinal models. This paper will review the literature that has described to date some effects of polyphenols on intestinal inflammation. Studies, conducted using in vivo and in vitro models, provide evidence that pure polyphenolic compounds and natural polyphenolic plant extracts can modulate intestinal inflammation.

Modulation of signalling nuclear factor-kappa B activation pathway by polyphenols in human intestinal Caco-2 cells

Recent studies support beneficial effects of polyphenols in various chronic inflammatory diseases, for example, the inflammatory bowel diseases. Inhibition of NF-kappaB activation by polyphenols could explain part of their anti-inflammatory properties, but few data are available on the intestine. The purpose of the present study was thus to investigate the effects of some polyphenols on NF-kappaB activation using human intestinal Caco-2 cells. Effects of standard polyphenols (50 mumol/l) were studied on different cellular events associated with NF-kappaB activation: (i) NF-kappaB activity using cells transiently transfected with a NF-kappaB-luciferase construct and stimulated by inflammatory agents (IL-1beta, TNF-alpha or lipopolysaccharides (LPS)); (ii) phosphorylation of the inhibitor of kappaB (IkappaB-alpha) analysed by Western blot; (iii) secretion of IL-8 quantified by ELISA assay. Results showed that chrysin and ellagic acid inhibited NF-kappaB activity, whereas genistein and resveratrol increased it. These effects were independent of the nature of the inducer, indicating that polyphenols may modulate NF-kappaB activation by acting on a common event to the cytokine- and LPS-mediated cascades. Chrysin strongly reduced (2.5-fold) IL-1beta-induced IkappaB-alpha phosphorylation, whereas ellagic acid increased it (1.7-fold). Ellagic acid, genistein and epigallocatechin gallate reduced (4- to 8-fold) IL-1beta-induced IL-8 secretion, while resveratrol promoted (1.7-fold) the secretion. Chrysin also diminished IL-8 secretion by 1.6-fold (but P>0.05). The data indicate that polyphenols can modulate the NF-kappaB activation pathway in the intestine. Chrysin could block NF-kappaB activation via the inhibition of IkappaB-alpha phosphorylation. The other molecular targets of the active polyphenols are still to be identified.

Mechanisms of provitamin A (carotenoid) and vitamin A (retinol) transport into and out of intestinal Caco-2 cells

The purpose of this study was to compare the mechanisms of intestinal retinol (ROL) and carotenoid transport. When differentiated Caco-2 cells were incubated with ROL for varying times, cellular ROL plateaued within 2 h, whereas retinyl ester (RE) formation increased continuously. ROL and RE efflux into basolateral medium (BM) increased linearly with time, ROL in the nonlipoprotein fraction and REs in chylomicrons (CMs). In contrast to carotenoids, ROL uptake was proportional to ROL concentration (0.5–110 μM). ROL efflux into BM occurred via two processes: a) a saturable process at low concentrations (<10 μM) and b) a nonsaturable process at higher concentrations. When ROL-loaded cells were maintained on retinoid-free medium, free ROL, but not REs, was secreted into BM. Glyburide significantly reduced ROL efflux but not ROL uptake. Inhibition of ABCA1 protein expression by small interfering RNAs decreased ROL efflux but not carotenoid efflux. Scavenger receptor class B type I (SR-BI) inhibition did not affect ROL transport but decreased carotenoid uptake. The present data suggest that a) ROL enters intestinal cells by diffusion, b) ROL efflux is partly facilitated, probably by the basolateral transporter ABCA1, and c) newly synthesized REs, but not preformed esters, are incorporated into CM and secreted. In contrast to ROL transport, carotenoid uptake is mediated by the apical transporter SR-BI, and carotenoid efflux occurs exclusively via their secretion in CM.

Inhibition of inflammatory mediators by polyphenolic plant extracts in human intestinal Caco-2 cells.

The mitogen-activated protein kinases (MAPK) and nuclear factor kappaB (NF-kappaB) are involved in transduction cascades that play a key role in inflammatory response. We tested the ability of preselected natural polyphenolic extracts (grape seed, cocoa, sugar cane, oak, mangosteen and pomegranate) to modulate intestinal inflammation using human intestinal Caco-2 cells treated for 4h with these extracts and then stimulated by cytokines for 24 or 48h. The effect of polyphenolic extracts, at 50 micromol of gallic acid equivalent/l, was investigated on inflammation-related cellular events: (i) NF-kappaB activity (cells transfected with a NF-kappaB-luciferase construct), (ii) activation of Erk1/2 and JNK (western blotting), (iii) secretion of interleukin 8 (IL-8) (ELISA), (iv) secretion of prostaglandin (PG) E(2) (ELISA), (v) production of NO (Griess method). Results show that: (i) sugar cane, oak and pomegranate extracts inhibited NF-kappaB activity (from 1.6 to 1.9-fold) (P<0.001); (ii) pomegranate slightly inhibited Erk1/2 activation (1.3-fold) (P=0.008); (iii) oak and pomegranate decreased NO synthesis by 1.5-fold (P<0.001) and that of IL-8 by 10.3 and 6.7-fold respectively; (iv) pomegranate and cocoa decreased PGE(2) synthesis by 4.6 (P<0.0001) and 2.2-fold (P=0.001), respectively. We suggest that pomegranate extract could be particularly promising in dietary prevention of intestinal inflammation.

Physio-pathological parameters affect the activation of inflammatory pathways by deoxynivalenol in Caco-2 cells

The intake of deoxynivalenol (DON), a mycotoxin contaminating cereal food items, causes gastro-intestinal illness in human and animal. This study investigated whether intracellular inflammatory cascades (MAPKs and NF-κB), cell maturity (proliferating vs. differentiated), cell state (control vs. inflamed) and exposure duration (chronic vs. acute) affect IL-8 secretion and PGE-2 synthesis in Caco-2 cells exposed to plausible intestinal concentrations (50, 500 and 5000 ng/ml) of DON. IL-8 secretion and PGE-2 synthesizing capacity were dose-dependently upregulated in differentiated Caco-2 cells exposed to DON during 24h, reaching an increase of ∼25 and 1.7-fold respectively, whereas transcript level of IL-8 and COX-2 were increased by ∼40 and 17-fold. Similar results were obtained with proliferating cells. The upregulation decreased upon simultaneous incubation with inhibitors of MAPKs ERK1/2 or p38 or of transcription factor NF-κB. IL-8 secretion and PGE-2 synthesizing capacity increased respectively by ∼15 and 2-fold after chronic 21 day incubation with DON (50 ng/ml). IL-8 production was exacerbated (∼510-fold versus negative control) upon simultaneous exposure to inflammatory stimuli. These results suggest activation of inflammatory pathways in intestinal epithelial cells exposed chronically or acutely to DON. The sensitivity to DON, whereas not affected by cell differentiation, is exacerbated by the presence of additional stimuli.