Angela Conte

Antiglycative and neuroprotective activity of colon-derived polyphenol catabolites

SCOPE Dietary flavonoids and allied phenolic compounds are thought to be beneficial in the control of diabetes and its complications, because of their ability to inhibit oxidative stress, protein glycation and to act as neuroprotectants. Following ingestion by humans, polyphenolic compounds entering the large intestine undergo extensive metabolism by interaction with colonic microbiota and it is metabolites and catabolites of the parent compounds that enter the circulatory system. The aim of this study was to investigate the inhibitory activity of some colonic microbiota-derived polyphenol catabolites against advanced glycation endproducts formation in vitro and to determine their ability, at physiological concentrations, to counteract mild oxidative stress of cultured human neuron cells. METHODS AND RESULTS This study demonstrated that ellagitannin-derived catabolites (urolithins and pyrogallol) are the most effective antiglycative agents, whereas chlorogenic acid-derived catabolites (dihydrocaffeic acid, dihydroferulic acid and feruloylglycine) were most effective in combination in protecting neuronal cells in a conservative in vitro experimental model. CONCLUSION Some polyphenolic catabolites, generated in vivo in the colon, were able in vitro to counteract two key features of diabetic complications, i.e. protein glycation and neurodegeneration. These observations could lead to a better control of these events, which are usually correlated with hyperglycemia.

Nitric oxide synthase activity in molluscan hemocytes

The hemocytes of the freshwater snail Viviparus ater have nitric oxide synthase (NOS) activity, as demonstrated by [3H]citrulline and nitrite+nitrate formation. The enzyme is NADPH dependent and is competitively inhibited by the mammalian NOS inhibitor (K i = 4.7 μM). The K m for l‐arginine is 2.5 μM. 70% of the total activity is observed at very low free Ca2+ concentration (3 nM). LPS treatment increased total NOS activity 2.4 fold. The activity is partly present in the non‐soluble fraction of hemocytes (24% and 8% in non‐stimulated and LPS‐stimulated snails, respectively). An antiserum to the C‐terminal synthetic pentadecapeptide of the rat cerebellar NOS inhibited the enzyme activity in a concentration‐dependent manner. This is the first biochemical demonstration of the existance of NOS activity in molluscan hemocytes, the cells responsible for defence mechanisms.

Synergistic protection of PC12 cells from β-amyloid toxicity by resveratrol and catechin

beta-Amyloid peptide (beta-AP) elicits a toxic effect on neurons in vitro and in vivo. Many environmental factors including antioxidants, metal ions and proteoglycans modify beta-AP toxicity. We have investigated on PC12 cells, the protective effect from beta-AP (1-41) of two plant polyphenols, resveratrol and catechin. PC12 cells treated with 10(-6)M beta-AP (1-41) for 16h decrease the cell viability at 24% of the control with an IC(50) value of 1.1+/-0.14 x 10(-8)M. Twenty-five micromolar resveratrol and 50 microM catechin protect PC12 cells from beta-AP (1-41) toxicity with the IC(50) value increased at 2.2+/-0.19 x 10(-7)M and at 8.9+/-0.7 x 10(-8)M, respectively. While the protective effect is concentration dependent for catechin, resveratrol shows a concentration-dependent biphasic effect. Up to 50 microM concentration, resveratrol protects PC12 cells from beta-AP (1-41) toxicity. At concentration higher than 50 microM, an inhibitory activity on cell proliferation appears. This antiproliferative effect is shown also in the absence of beta-AP (1-41). Resveratrol and catechin have a synergistic protective action. In the presence of 50 microM catechin and 10 microM resveratrol or 25 microM resveratrol and 10 microM catechin, the toxicity determined by 10(-7)M beta-AP (1-41) is almost completely abolished. Catechin is more effective than resveratrol in protecting PC12 cells from the toxicity of hydrogen peroxide. The protection from Oxygen Reactive Species (ROS) toxicity is concentration dependent for both resveratrol and catechin. In this case the protection is merely additive and the synergistic effect is not observed. These results demonstrate that resveratrol and catechin protect PC12 cells from beta-AP (1-41) toxicity and that their protective effect is synergistic. Such a protective effect probably is not due only to their antioxidant activity. The different chemical and biological activity shown by these compounds on several cell types and the complexity of the beta-AP (1-41) toxicity may explain the synergistic protective effect and suggest that the utilization of different compounds with synergistic activity may protect more effectively from complex mechanisms of toxicity.

Effect of dietary melanoidins on lipid peroxidation during simulated gastric digestion: their possible role in the prevention of oxidative damage.

The ability of high molecular weight melanoidins extracted from coffee, barley coffee, and dark beer to inhibit lipid peroxidation during simulated gastric digestion of turkey meat has been investigated. Results showed that melanoidins decrease the synthesis of lipid hydroperoxides and secondary lipoxidation products. Coffee melanoidins at 3 mg/mL reversed the reaction and broke down hydroperoxides to concentrations lower than the initial value. Barley coffee and dark beer melanoidins were less effective, and even at 12 mg/mL did not reverse the reaction. The proposed mechanism of action involved Fe(2+) chelating capacity, heme-binding ability, and radical-scavenging activity. Melanoidins were characterized for their content in total proteins, carbohydrates, and phenolics, and the relationship between the chemical composition and the antioxidant activity of dietary melanoidins was investigated. Coffee melanoidins, which contain more phenolics and proteins with respect to the other melanoidins, showed greater antioxidant activity with respect to the other melanoidins tested.

Nitric oxide: an ancestral immunocyte effector molecule.

The presence and the role of nitric oxide synthase (NOS) were investigated in the molluscan hemocytes by immunocytochemical, biochemical and functional approaches. Using an anti-NOS polyclonal antibody, immunoreactivity was observed in the hemocytes, and this reactivity increased after stimulation of the animals with Escherichia coli, indicating that this enzyme is inducible. The NOS inducibility was also histochemically demonstrated by detection of NADPH-diaphorase activity. Biochemical studies show that the enzyme is 70% cytoplasmatic and 30% membrane bound and that the inducible form is mainly cytoplasmatic. The nitrite + nitrate and citrulline formation, the inhibition by N omega-nitro-L-arginine, the Km value for arginine, the calcium and co-enzyme dependence show that the molluscan NOS shares the same properties as the NOS isoenzymes so far studied. However, it cannot be identified with any of these enzymes. It appears to be in some way similar to an inducible form of human hepatocyte NOS. Also cytokines are able to induce NOS. In vitro studies have shown that hemocytes produce nitric oxide (NO), a bactericide substance, and that there is a relationship between the NO system and phagocytosis. The presence of NO in the invertebrate hemocyte demonstrates that critical molecules have been conserved over the course of evolution.

From balsamic to healthy: Traditional balsamic vinegar melanoidins inhibit lipid peroxidation during simulated gastric digestion of meat

In this work traditional balsamic vinegar (TBV) melanoidins were characterized for chemical composition and antioxidant activity and their anti-peroxidative effect during an in vitro gastric digestion of turkey meat was studied. The most important constituents of TBV melanoidins were carbohydrates (51% w/w) of which glucose (35% w/w) and fructose (10% w/w) are the main representatives, hydroxymethylfurfural (7.2% w/w), phenolic groups (4.6% w/w) and proteins (1.2% w/w). The antioxidant capacity of melanoidins was studied, measuring lipid hydroperoxides and secondary lipoxidation products formed during in vitro gastric digestion of turkey meat. The most important mechanisms in their antioxidant activity resulted radical scavenging and Fe(2+)-chelating activities. Pepsin inhibiting ability has been excluded. TBV melanoidins were also able to bind heme under gastric conditions potentially preventing its absorption and prooxidant and cytotoxic effects. Our results support the idea that TBV melanoidins may have a role in oxidative damage prevention. Fe(2+)-chelating and heme-binding activities as well as mechanisms of antioxidant activity of TBV melanoidins were also compared with coffee, barley coffee and dark beer melanoidins.

SNPs in Neurotrophin System Genes and Alzheimer's Disease in an Italian Population

Increasing evidence suggests a role for nerve growth factor (NGFB), brain-derived neurotrophic factor (BDNF), and their receptors, nerve growth factor receptor (NGFR), and neurotrophin tyrosine kinase receptors 1 and 2 (NTRK1 and NTRK2), in Alzheimers disease (AD). However, genetic association between the neurotrophin system genes and AD has been poorly investigated. We genotyped 21 single nucleotide polymorphisms (SNPs) within these genes in a population of Italian AD patients and healthy controls. We found an allele-wise association of rs2072446 on NGFR with familial AD (fAD, p = 0.047), and a genotype-wise association of rs2289656 on NTRK2 with sporadic AD (sAD, p = 0.0036). rs6336 on NTRK1 resulted associated to early-onset sAD in both allele-wise (p = 0.028) and genotype-wise (p = 0.014) analysis, while rs1048218 on BDNF showed allele-wise association with late-onset sAD (p = 0.047). A trend to association with sAD and/or fAD was observed for other SNPs. Our results suggest that genetic variants of neurotrophin system genes might confer susceptibility to AD.

Reversing of chlorambucil resistance by ethacrynic acid in a B-CLL patient

Summary We evaluated the reversing activity of ethacrynic acid in a B‐CLL patient resistant to chlorambucil. The glutathione S‐transferase (GST) activity, measured in peripheral blood lymphocytes, resulted extremely elevated. Etha crynic acid, at pharmacological concentrations, partially reversed chlorambucil resistance and this result appeared related to the increased GST levels.

The Type and Concentration of Milk Increase the in Vitro Bioaccessibility of Coffee Chlorogenic Acids

Coffee with different types and concentrations of milk was digested with pepsin (2 h) and pancreatin (2 h) to simulate gastropancreatic digestion. Chlorogenic acids (CGAs) were determined by high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry in ultrafiltrate (cutoff 3 kDa) to evaluate their bioaccessibility. After digestion, bioaccessible CGAs decreased from 80.2 to 53.0 and 69.5 μmol/200 mL in coffee without milk and coffee-whole milk, respectively. When whole, semiskimmed, skimmed, or diluted milk were present, the increase in bioaccessibility was dependent on fat content (r = 0.99, p < 0.001). No relationship was observed between bioaccessibility and proteins, carbohydrates, and calcium content. The addition of milk to coffee caused an immediate decrease in the bioaccessibility due to CGAs binding to proteins. After digestion, 86-94% of bound CGAs remained in the high molecular weight fraction. Casein bound 5-caffeoylquinic acid with high affinity (K(D) of 37.9 ± 2.3 μmol/L; n = 0.88 ± 0.06).

Effects of Different Low-Frequency Electromagnetic Fields on Lymphocyte Activation: At which Cellular Level?

It has been shown that low-frequency electromagnetic fields may inhibit or enhance the lymphocyte response to many mitogens. How this happens is not known, and many reports suggest that alterations of surface receptors may be involved. Results presented here indicate that cellular activation is inhibited by a high-intensity EMF after the early phases of signal transduction, but it may be enhanced by a low-intensity EMF.