Fátima Martel

Chemopreventive effect of dietary polyphenols in colorectal cancer cell lines.

Colorectal cancer (CRC) is the second most fatal and the third most diagnosed type of cancer worldwide. Despite having multifactorial causes, most CRC cases are mainly determined by dietary factors. In recent years, a large number of studies have attributed a protective effect to polyphenols and foods containing these compounds (fruits and vegetables) against CRC. Indeed, polyphenols have been reported to interfere with cancer initiation, promotion, and progression, acting as chemopreventive agents. The aim of this review is to summarize the main chemopreventive properties of some polyphenols (quercetin, rutin, myricetin, chrysin, epigallocatechin-3-gallate, epicatechin, catechin, resveratrol, and xanthohumol) against CRC, observed in cell culture models. From the data reviewed in this article, it can be concluded that these compounds inhibit cell growth, by inducing cell cycle arrest and/or apoptosis; inhibit proliferation, angiogenesis, and/or metastasis; and exhibit anti-inflammatory and/or antioxidant effects. In turn, these effects involve multiple molecular and biochemical mechanisms of action, which are still not completely characterized. Thus, caution is mandatory when attempting to extrapolate the observations obtained in CRC cell line studies to humans.

Molecular cloning and characterization of two novel transport proteins from rat kidney

The recent cloning of renal transport systems for organic anions and cations (OAT1, OCT1, and OCT2) opened the possibility to search, via polymerase chain reaction (PCR) homology screening, for novel transport proteins. Two integral membrane proteins, UST1 and UST2, were cloned from rat kidney. RT‐PCR revealed that UST1 is confined to the kidney whereas UST2 mRNA was detected in all tested tissues. Sequence analyses suggest that UST1 and UST2, together with four related transporters, comprise, within the major facilitator superfamily, a so far unrecognized transporter family, termed amphiphilic solute facilitator (ASF) family. Characteristic signatures for the ASF family were identified.

The Extraneuronal Transporter for Monoamine Transmitters Exists in Cells Derived from Human Central Nervous System Glia

From studies on sympathetically innervated peripheral tissues it is well known that both neuronal and nonneuronal transport systems contribute to the inactivation of released monoamine transmitters. The close proximity between synapses and glia cell processes in the CNS leads to the so far unresolved question whether non‐neuronal transporters are involved in the inactivation of centrally released monoamine transmitters such as noradrenaline, dopamine and 5‐hydroxytryptamine. 1‐Methyl‐4‐phenylpyridinium (MPP+) is a prototypical substrate of the extraneuronal monoamine transporter (uptake2). [3H]MPP+ was found to accumulate in various human glioma cell lines. [3H]MPP+transport was characterized in more detail in HTZ146 human glioma cells. The Ki values of various compounds for the inhibition of initial rates of [3H]MPP+ transport into HTZ146 cells were closely correlated with the known Ki values for the inhibition of the extraneuronal monoamine transporter (P < 0.001, r= 0.991, n= 7). The rank order of inhibitory potencies was decynium 22 > corticosterone > cyanine 863 > O‐methylisoprenaline > quinine > clonidine > quinidine. [3H]MPP+ accumulation was investigated not only in various CNS tumour cell lines but also in primary cultures of human astrocytes and rat cerebral cortex slices. In all tested experimental systems, accumulation was sensitive to cyanine‐related inhibitors of the extraneuronal monoamine transporter. These findings suggest that the extraneuronal monoamine transporter exists in glia cells. Furthermore, it was shown that MPP+ is able to make use of the extraneuronal monoamine transporter not only to enter but also to leave glia cells. This finding suggests that the extraneuronal monoamine transporter may play a key role in the mechanism of 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) neurotoxicity.

Impact of Gestational Diabetes Mellitus in the Maternal-to-Fetal Transport of Nutrients

Gestational diabetes mellitus (GDM) is a metabolic disorder prevalent among pregnant women. This disease increases the risk of adverse perinatal outcomes and diseases in the offspring later in life. The human placenta, the main interface between the maternal and fetal blood circulations, is responsible for the maternal-to-fetal transfer of nutrients essential for fetal growth and development. In this context, the aim of this article is to review the latest advances in the placental transport of macro and micronutrients and how they are affected by GDM and its associated conditions, such as elevated levels of glucose, insulin, leptin, inflammation, and oxidative stress. Data analyzed in this article suggest that GDM and its associated conditions, particularly high levels of glucose, leptin, and oxidative stress, disturb placental nutrient transport and, consequently, fetal nutrient supply. As a consequence, this disturbance may contribute to the fetal and postnatal adverse health outcomes associated with GDM.

Absorption of anthocyanins through intestinal epithelial cells – Putative involvement of GLUT2

Anthocyanins bioavailability is a major issue regarding their biological effects and remains unclear due to few data available on this matter. This work aimed to evaluate the absorption of anthocyanins at the intestine using Caco-2 cells. Anthocyanin extract, rich in malvidin-3-glucoside, was obtained from red grape skins and tested on Caco-2 cells. The absorption of anthocyanins, in absence or presence of 1% ethanol, was detected by HPLC/DAD/LC-MS. Our results showed that this transport was significantly increased in the presence of ethanol especially after 60 min of incubation. In addition, cells that were pretreated for 96 h with anthocyanins (200 microg/mL) showed an increase of their own transport (about 50% increase). Expression of glucose transporters sodium-dependent glucose transporter 1, facilitative glucose transporters 5, and facilitative glucose transporters 2 was assessed by RT-PCR. It was found that facilitative glucose transporters 2 expression was increased (60%) in Caco-2 cells pretreated with anthocyanins, by comparison with controls. When the effect of anthocyanin extract on (3)H-2-deoxy-D-glucose uptake was tested, an inhibitory effect was observed (about 60% decrease). However, the malvidin aglycone was tested and had no effect. In conclusion, anthocyanins could be absorbed through Caco-2 cells, and can interfere with their own transport and also with glucose intestinal uptake.

Apical uptake of organic cations by human intestinal Caco-2 cells: putative involvement of ASF transporters

Abstract. The aim of this work was to characterise the intestinal absorption of organic cations, by testing the possibility of involvement of known members of the amphiphilic solute facilitator (ASF) family in this process. For that purpose, the characteristics of the uptake of 1-methyl-4-phenylpyridinium, a model organic cation, at the brush-border membrane of Caco-2 cells were compared with those of the extraneuronal monoamine transporter (EMT)-mediated transport.Uptake of [3H]MPP+ by Caco-2 and 293hEMT cells showed pH-dependence: it was significantly reduced (to 86% and 62% of control, respectively) when the pH of the extracellular medium was decreased to 6.2, and increased (to 116% and 136% of control, respectively) when the extracellular pH was increased to 8.2. Uptake of [3H]MPP+ by Caco-2 cells and 293hEMT cells showed potential-dependence: substitution of KCl for NaCl in the incubation medium resulted in a reduction in the inward transport of [3H]MPP+ (to 70% and 40% of control, respectively). Uptake of [3H]MPP+ by Caco-2 and 293hEMT cells showed only little dependence on Na+: substitution of NaCl of the incubation media with LiCl resulted in a small decrease (of 19% and 14%, respectively) in [3H]MPP+ uptake. However, when NaCl was substituted with choline chloride, a significant reduction in [3H]MPP+ uptake by Caco-2 and 293hEMT cells (of 56% and 68%, respectively) was observed. The effect of various compounds on initial rates of [3H]MPP+ uptake into Caco-2 and 293hEMT cells was tested. All compounds tested interacted with the specific [3H]MPP+ uptake in both cell lines. There was no correlation between the IC50s in relation to inhibition of [3H]MPP+ uptake into Caco-2 cells and into 293hEMT cells. Reverse transcriptase-polymerase chain reaction indicates that mRNA of hEMT and of the human organic cation transporter 1 (hOCT1) are present in Caco-2 cells.In conclusion, our results suggest that uptake of organic cations at the brush-border membrane of Caco-2 cells may occur through two distinct Na+-independent transporters belonging to the ASF family: hEMT and hOCT1.

Folates and aging: Role in mild cognitive impairment, dementia and depression

In almost all tissues, including the brain, folates are required for one-carbon transfer reactions, which are essential for the synthesis of DNA and RNA nucleotides, the metabolism of amino acids and the occurrence of methylation reactions. The aim of this paper is to review the impact of folate status on the risk of development of neuropsychiatric disorders in older individuals. The prevalence of folate deficiency is high among individuals aged ≥ 65 years mainly due to reduced dietary intake and intestinal malabsorption. Population-based studies have demonstrated that a low folate status is associated with mild cognitive impairment, dementia (particularly Alzheimers disease) and depression in healthy and neuropsychiatric diseased older individuals. The proposed mechanisms underlying that association include hyperhomocysteinemia, lower methylation reactions and tetrahydrobiopterin levels, and excessive misincorporation of uracil into DNA. However, currently, there is no consistent evidence demonstrating that folic acid supplementation improves cognitive function or slows cognitive decline in healthy or cognitively impaired older individuals. In conclusion, folate deficiency seems to be an important contributor for the onset and progression of neuropsychiatric diseases in the geriatric population but additional studies are needed in order to increase the knowledge of this promising, but still largely unexplored, area of research.

Transport of small organic cations in the rat liver

The kidneys and the liver are the principal organs for the inactivation of circulating organic cations. Recently, an organic cation transporter (OCT1) has been cloned from rat kidney. In order to answer the question whether OCT1 is involved also in hepatic uptake of organic cations, the pharmacological characteristics of organic cation transport in hepatocytes were compared to the characteristics of transiently expressed OCT1.Primary cultures of rat hepatocytes avidly accumulated the small organic cation 3H-1-methyl-4-phenylpyridinium (3H-MPP+). At equilibrium, the hepatocytes accumulated 3H-MPP+ 56-fold. Initial rates of specific 3H-MPP+ transport in hepatocytes were saturable. The half-saturating concentration was 13 μmol/l. 3H-MPP+ transport was sensitive to quinine (Ki = 0.79 μmol/l) and cyanine863 (Ki = 0.097 µmol/l). Quinine and cyanine863 are known inhibitors of type I hepatic transport of cationic drugs and of renal excretion of organic cations, respectively. To compare the functional characteristics of 3H-MPP+ transport in hepatocytes with those of OCT1, OCT1 has been heterologously expressed and characterized in a mammalian cell line (293 cells). Initial rates of 3H-MPP+ transport were saturable, the Km being 13 μmol/l. The rank order of inhibitory potencies of various inhibitors was almost identical in hepatocytes and 293 cells transiently transfected with OCT1. There was a positive correlation between the Kis for the inhibition of 3H-MPP+ transport in isolated hepatocytes and transfected 293 cells (r = 0.85; P<0.01; n = 8).The results indicate that OCT1 is functionally expressed not only in the kidney but also in hepatocytes where it is responsible for the transport of small organic cations which, in the past, have been classified as type I substrates.

Inward transport of [3H]-1-methyl-4-phenylpyridinium in rat isolated hepatocytes: putative involvement of a P-glycoprotein transporter.

1 The liver has an important role in the detoxification of organic cations from the circulation. [3H]‐1‐methyl‐4‐phenylpyridinium ([3H]‐MPP+), a low molecular weight organic cation, is efficiently taken up and accumulated by rat hepatocytes through mechanisms partially unknown. 2 The aim of the present work was to characterize further the uptake of MPP+ by rat isolated hepatocytes. The putative interactions of a wide range of drugs, including inhibitors/substrates of P‐glycoprotein, were studied. 3 The uptake of MPP+ was investigated in rat freshly isolated hepatocytes (incubated in Krebs‐Henseleit medium with 200 nM [3H]‐MPP+ for 5 min) and in the rat liver in situ (perfused with Krebs‐Henseleit/BSA medium with 200 nM [3H]‐MPP+ for 30 min). [3H]‐MPP+ accumulation in the cells and in tissue was determined by liquid scintillation counting. 4 Verapamil (100 μm), quinidine (100 μm), amiloride (1 mM), (+)‐tubocurarine (100 μm), vecuronium (45 μm), bilirubin (200 μm), progesterone (200 μm), daunomycin (100 μm), vinblastine (100 μm), cyclosporin A (100 μm) and cimetidine (100 μm) had a significant inhibitory effect on the accumulation of [3H]‐MPP+ in isolated hepatocytes. Tetraethylammonium (100 μm) had no effect. 5 In the rat perfused liver, both cyclosporin A (100 μm) and verapamil (100 μm) had much less marked inhibitory effects as compared to their effects on isolated hepatocytes (0% against 35% and 45% against 96% of inhibition, respectively). 6 Inhibition of alkaline phosphatase activity by increasing or decreasing the pH of the incubation medium or by the presence of vanadate (1 mM) or homoarginine (500 μm) led to a significant increase in the accumulation of [3H]‐MPP+ in isolated hepatocytes. 7 It was concluded that, in addition to the type I organic cation hepatic transporter, [3H]‐MPP+ is taken up by rat isolated hepatocytes through P‐glycoprotein, a canalicular transport system that usually excretes endobiotics and xenobiotics. We proposed that the reversal of transport through P‐glycoprotein may be related to the loss of efficiency of alkaline phosphatase in isolated hepatocytes.

Effect of polyphenols on the intestinal and placental transport of some bioactive compounds

Polyphenols are a group of widely distributed phytochemicals present in most foods of vegetable origin. A growing number of biological effects have been attributed to these molecules in the past few years and only recently has their interference with the transport capacity of epithelial barriers received attention. This review will present data obtained concerning the effect of polyphenols upon the transport of some compounds (organic cations, glucose and the vitamins thiamin and folic acid) at the intestinal and placental barriers. Important conclusions can be drawn: (i) different classes of polyphenols affect transport of these bioactive compounds at the intestinal epithelia and the placenta; (ii) different compounds belonging to the same phenolic family often possess opposite effects upon transport of a given molecule; (iii) the acute and chronic/short-term and long-term exposures to polyphenols do not produce parallel results and, therefore, care should be taken when extrapolating results; (iv) the effect of polyphenolics in combination may be very different from the expected ones taking into account the effect of each of these compounds alone, and so care should be taken when speculating on the effect of a drink based on the effect of one component only; (v) care should be taken in drawing conclusions for alcoholic beverages from results obtained with ethanol alone. Although most of the data reviewed in the present paper refer to in vitro experiments with cell-culture systems, these studies raise a concern about possible changes in the bioavailability of substrates upon concomitant ingestion of polyphenols.