Biomedical subjects

M Nardini

Publications and source records attributed to M Nardini.

At least 19 recordsLinked to original sources

PubMed2004

Non-cycloplegic refractive screening can identify infants whose visual outcome at 4 years is improved by spectacle correction.

The Second Cambridge Population Infant Vision Screening Programme using the VPR-1 videorefractor without cycloplegia was undertaken in order to identify those infants with refractive errors who were potentially amblyogenic or strabismogenic. Infants identified at eight months were entered into a control trial of treatment with partial spectacle correction and underwent a long-term follow-up that monitored a wide range of visual, visuoperceptual, visuocognitive, visuomotor, linguistic and social development. In the present paper, the authors report on the outcome measures of visual acuity and strabismus. Poor acuity was defined as a best-corrected acuity of 6/12 or worse on crowded letters or 6/9 or worse on single letters, at age 4 years. Acuity was measured in 79 infants who were significantly hyperopic and/or anisometropic at 11-12 months of age, 23 who showed hyperopia of +3D but less than +3.5D, 196 control subjects, 14 controls with refractive errors, and 126 others who showed an accommodative lag on screening but were not significantly hyperopic on first retinoscopy. There was a poorer acuity outcome in the untreated group of hyperopes compared to controls (p < 0.0001) and to the children who were compliant in spectacle wear (p < 0.001) or who were prescribed spectacles (p < 0.05). Children who were significantly hyperopic at eight months were also more likely to be strabismic by 5.5 years compared to the emmetropic control group (p < 0.001). However, the present study did not find a significant difference in the incidence of strabismus between corrected and uncorrected hyperopic infants. Children who were not refractively corrected for significant hyperopia were four times more likely to have poor acuity at 5.5 years than infants who wore their hyperopic correction, supporting the findings of the First Cambridge Population Infant Vision Screening Programme.

Accommodation, Ocular↗

PubMed2003

Determination of aminoethylcysteine ketimine decarboxylated dimer in human plasma and cultured cells by high-performance liquid chromatography with electrochemical detection.

Aminoethylcysteine ketimine decarboxylated dimer (AECK-DD) is a natural compound with antioxidant properties of a new family of sulfur-containing amino acids. It has been detected in human urine and plasma, in mammalian cerebellum and, more recently, in dietary vegetables. In the present study, a simple, highly sensitive method using a high-performance liquid chromatography system with electrochemical detection (ECD) has been developed. The method showed excellent precision and accuracy. It has been found to be about 100-fold more sensitive than gas chromatographic method and 2000-fold more sensitive in respect to the liquid chromatography method with UV detection. The method showed the required features of specificity and sensitivity to detect aminoethylcysteine ketimine decarboxylated dimer in human plasma and in cultured cells after in vitro supplementation.

Amino Acids, Sulfur↗

PubMed2002

Absorption of phenolic acids in humans after coffee consumption.

Despite extensive literature describing the biological effects of polyphenols, little is known about their absorption from diet, one major unresolved point consisting of the absorption of the bound forms of polyphenols. In this view, in the present work we studied the absorption in humans of phenolic acids from coffee, a common beverage particularly rich in bound phenolic acids, such as caffeic acid, ferulic acid, and p-coumaric acid. Coffee brew was analyzed for free and total (free + bound) phenolic acids. Chlorogenic acid (5'-caffeoylquinic acid), a bound form of caffeic acid, was present in coffee at high levels, while free phenolic acids were undetectable. After alkaline hydrolysis, which released bound phenolic acids, ferulic acid, p-coumaric acid, and high levels of caffeic acid were detected. Plasma samples were collected before and 1 and 2 h after coffee administration and analyzed for free and total phenolic acid content. Two different procedures were applied to release bound phenolic acids in plasma: beta-glucuronidase treatment and alkaline hydrolysis. Coffee administration resulted in increased total plasma caffeic acid concentration, with an absorption peak at 1 h. Caffeic acid was the only phenolic acid found in plasma samples after coffee administration, while chlorogenic acid was undetectable. Most of caffeic acid was present in plasma in bound form, mainly in the glucuronate/sulfate forms. Due to the absence of free caffeic acid in coffee, plasma caffeic acid is likely to be derived from hydrolysis of chlorogenic acid in the gastrointestinal tract.

Absorption↗

PubMed2001

Crystallization and preliminary X-ray analysis of neural haemoglobin from the nemertean worm Cerebratulus lacteus.

The nemertean worm Cerebratulus lacteus neural tissue haemoglobin (109 amino acids, the shortest known haemoglobin) has been overexpressed in Escherichia coli, purified and crystallized. A highly redundant native data set has been collected at the Cu K(alpha) wavelength to 2.05 A resolution. The crystals belong to the orthorhombic P2(1)2(1)2(1) space group, with unit-cell parameters a = 42.5, b = 43.1, c = 60.2 A and one molecule per asymmetric unit. The anomalous difference Patterson map clearly reveals the position of the haem Fe atom, thus paving the way for MAD/SAD structure determination.

Animals↗

PubMed2001

Modulation of ceramide-induced NF-kappaB binding activity and apoptotic response by caffeic acid in U937 cells: comparison with other antioxidants.

Ceramide acts as second messenger in the signal transduction triggered by a variety of stress stimuli and extracellular agents. Stress response through ceramide is involved in the development of many human diseases, such as atherosclerosis, inflammation, neurodegenerative disorders, and acquired immunodeficiency syndrome. Dietary polyphenols have been reported to exert a beneficial effect on the onset and development of most of these human chronic-degenerative pathologies. However, the mechanisms underlying this beneficial effect are mostly not understood at the present. To investigate the ability of polyphenols in modulating fundamental cellular functions, we studied the effect of caffeic acid, a widespread phenolic acid largely present in human diet, in the modulation of ceramide-induced signal transduction pathway leading to apoptosis in U937 cells, in comparison with other established antioxidants of nutritional interest (N-acetylcysteine, d-alpha-tocopherol acetate and ascorbic acid). Our results indicate that caffeic acid efficiently inhibits both ceramide-induced NF-kappaB binding activity and apoptosis at micromolar concentration. Other antioxidants tested are totally ineffective in inhibiting apoptosis, although affecting NF-kappaB activation. Caffeic acid was found to inhibit protein tyrosine kinase activity, suggesting that this mechanism can be on the basis of the inhibition of apoptosis. Our results suggest that dietary caffeic acid might modulate ceramide-induced signal transduction pathway and NF-kappaB activation through either antioxidant and nonantioxidant mechanisms.

Antioxidants↗

PubMed2001

A syndrome characterized by psychiatric disorders, recurrent mucosal infections and natural immunity deficits: clinical approach.

The authors summarize their own previous work on the identification of a subset of patients characterized by psychiatric disorders, recurrency of mucosal infections and impaired natural immunity. The diagnostic approach to these patients based on the close collaboration between infectivologists, immunologists and psychiatrists is described with the aim to find out combined treatments for the amelioration of clinical manifestations.

Humans↗

PubMed2001

Inhibition of neutrophil apoptosis by acrolein: a mechanism of tobacco-related lung disease?

Cigarette smoking is known to contribute to inflammatory diseases of the respiratory tract by promoting recruitment of inflammatory-immune cells such as neutrophils and perhaps by altering neutrophil functional properties. We investigated whether acrolein, a toxic unsaturated aldehyde found in cigarette smoke, could directly affect neutrophil function. Exposure of freshly isolated human neutrophils to acrolein markedly inhibited spontaneous neutrophil apoptosis as indicated by loss of membrane asymmetry and DNA fragmentation and induced increased neutrophil production of the chemokine interleukin-8 (IL-8). Acrolein (1--50 microM) was found to induce marked activation of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinases (MAPKs), and inhibition of p38 MAPK activation by SB-203580 prevented acrolein-induced IL-8 release. However, inhibition of either ERK or p38 MAPK did not affect acrolein-dependent inhibition of apoptosis. Acrolein exposure prevented the activation of caspase-3, a crucial step in the execution of neutrophil apoptosis, presumably by direct inhibition of the enzyme. Our results indicate that acrolein may contribute to smoke-induced inflammatory processes in the lung by increasing neutrophil recruitment and reducing neutrophil clearance by apoptosis.

Acrolein↗

PubMed2001

mRNA expression of serotonin receptors of type 2C and 5A in human resting lymphocytes.

We investigated the presence of mRNA for serotonin receptors of type 2C (5-HT(2C)) in resting lymphocytes by means of RT-PCR and Southern blotting analyses, given their possible role in the pathophysiology of anxiety and eating disorders. At the same time, we explored also the presence of the specific mRNA for 5-HT(5A) receptors, a novel subtype for which still no functional data exist. Healthy subjects and patients with obsessive-compulsive or bipolar disorders were included in the study. The results showed the presence of the specific mRNAs for both 5-HT(2C) and 5-HT(5A) receptors in resting lymphocytes of the three groups of subjects. An additional band was also observed after the amplification of the 5-HT(5A) cDNA in each sample. These findings, while revealing the presence of 5-HT(2C) and 5-HT(5A) receptor mRNAs in an easily available tissue, can be considered preliminary for future quantitative analyses in patients with different psychiatric conditions.

Adult↗

PubMed2000

Crystal structure of pseudomonas aeruginosa lipase in the open conformation. The prototype for family I.1 of bacterial lipases.

The x-ray structure of the lipase from Pseudomonas aeruginosa PAO1 has been determined at 2.54 A resolution. It is the first structure of a member of homology family I.1 of bacterial lipases. The structure shows a variant of the alpha/beta hydrolase fold, with Ser(82), Asp(229), and His(251) as the catalytic triad residues. Compared with the "canonical" alpha/beta hydrolase fold, the first two beta-strands and one alpha-helix (alphaE) are not present. The absence of helix alphaE allows the formation of a stabilizing intramolecular disulfide bridge. The loop containing His(251) is stabilized by an octahedrally coordinated calcium ion. On top of the active site a lid subdomain is in an open conformation, making the catalytic cleft accessible from the solvent region. A triacylglycerol analogue is covalently bound to Ser(82) in the active site, demonstrating the position of the oxyanion hole and of the three pockets that accommodate the sn-1, sn-2, and sn-3 fatty acid chains. The inhibited enzyme can be thought to mimic the structure of the tetrahedral intermediate that occurs during the acylation step of the reaction. Analysis of the binding mode of the inhibitor suggests that the size of the acyl pocket and the size and interactions of the sn-2 binding pocket are the predominant determinants of the regio- and enantio-preference of the enzyme.

Amino Acid Sequence↗

PubMed2000

Functional and crystallographic characterization of Salmonella typhimurium Cu,Zn superoxide dismutase coded by the sodCI virulence gene.

The functional and three-dimensional structural features of Cu,Zn superoxide dismutase coded by the Salmonella typhimurium sodCI gene, have been characterized. Measurements of the catalytic rate indicate that this enzyme is the most efficient superoxide dismutase analyzed so far, a feature that may be related to the exclusive association of the sodCI gene with the most pathogenic Salmonella serotypes. The enzyme active-site copper ion is highly accessible to external probes, as indicated by quenching of the water proton relaxation rate upon addition of iodide. The shape of the electron paramagnetic resonance spectrum is dependent on the frozen or liquid state of the enzyme solution, suggesting relative flexibility of the copper ion environment. The crystal structure (R-factor 22.6%, at 2.3 A resolution) indicates that the dimeric enzyme adopts the quaternary assembly typical of prokaryotic Cu,Zn superoxide dismutases. However, when compared to the structures of the homologous enzymes from Photobacterium leiognathi and Actinobacillus pleuropneumoniae, the subunit interface of Salmonella Cu,Zn superoxide dismutase shows substitution of 11 out of 19 interface residues. As a consequence, the network of structural water molecules that fill the dimer interface cavity is structured differently from the other dimeric bacterial enzymes. The crystallographic and functional characterization of this Salmonella Cu,Zn superoxide dismutase indicates that structural variability and catalytic efficiency are higher in prokaryotic than in the eukaryotic homologous enzymes.

Amino Acid Sequence↗

PubMed2000

In vitro inhibition of the activity of phosphorylase kinase, protein kinase C and protein kinase A by caffeic acid and a procyanidin-rich pine bark (Pinus marittima) extract.

Caffeic acid (CA) is a common constituent of human diet while pine bark extract (PBE) is utilized either as nutritional supplement or as phytochemical remedy for different diseases. CA and PBE, are reported as efficient antioxidants and more recently have been described to modulate cellular response to oxidative challenge and to possess many other biological activities, i.e. anti-inflammatory, antimutagenic, antitumoral effects. In order to investigate in depth the mechanism of action of these polyphenols, the effects of CA and PBE on the activity of some protein kinases involved in the regulation of fundamental cellular processes were studied in vitro: phosphorylase kinase (PhK), protein kinase A (PKA), protein kinase C (PKC). PBE at the concentration of 20 microg/ml (corresponding to 69 microM catechin equivalents) inhibited PKA, PhK and PKC by about 90, 59, 57%, respectively, while 100 microM CA inhibited by 37, 52 and 54%, respectively. Considerable inhibitions have been still observed at even lower concentrations of CA and PBE. For PhK and PKA, the inhibition follows a non-competitive mechanism. CA also inhibits PKC activity in a partially purified cellular extract. The results suggest a possible involvement of CA and PBE in modulation of cellular functions.

Caffeic Acids↗

PubMed2000

Directed evolution of an enantioselective lipase.

BACKGROUND: The biocatalytic production of enantiopure compounds is of steadily increasing importance to the chemical and biotechnological industry. In most cases, however, it is impossible to identify an enzyme that possesses the desired enantioselectivity. Therefore, there is a strong need to create by molecular biological methods novel enzymes which display high enantioselectivity. RESULTS: A bacterial lipase from Pseudomonas aeruginosa (PAL) was evolved to catalyze with high enantioselectivity the hydrolysis of the chiral model substrate 2-methyldecanoic acid p-nitrophenyl ester. Successive rounds of random mutagenesis by ep-PCR and saturation mutagenesis resulted in an increase in enantioselectivity from E=1.1 for the wild-type enzyme to E=25.8 for the best variant which carried five amino acid substitutions. The recently solved three-dimensional structure of PAL allowed us to analyze the structural consequences of these substitutions. CONCLUSIONS: A highly enantioselective lipase was created by increasing the flexibility of distinct loops of the enzyme. Our results demonstrate that enantioselective enzymes can be created by directed evolution, thereby opening up a large area of novel applications in biotechnology.

Amino Acid Sequence↗

PubMed2000

Normal emmetropization in infants with spectacle correction for hyperopia.

PURPOSE: The development of emmetropic refraction is known to be under visual control. Does partial spectacle correction of infants' refractive errors, which has been shown to have beneficial effects in reducing strabismus and amblyopia, impede emmetropization? The purpose of the present study was to perform the first longitudinal controlled trial to investigate this question in human subjects. METHODS: Children identified as having significant hyperopia in a population screening program at age 8 to 9 months were assigned to treated (partial spectacle correction) or untreated groups. A control group of infants with no significant refractive errors at screening was also recruited. Measurements of retinoscopic refraction under cycloplegia were taken at 4- to 6-month intervals up to the age of 36 months, and changes in refraction of 148 subjects were analyzed longitudinally. RESULTS: Refractive error decreased toward low hyperopic values between 9 and 36 months in both hyperopic groups. By 36 months, this reduction of hyperopia showed no overall difference between children who were treated with partial spectacle correction and those who were not. Despite the improvement, both hyperopic groups' mean refractive error at 36 months remained higher than that of the control group. When infants in all three groups were considered together, the rate of reduction of refractive error was, on average, a linear function of the initial level of hyperopia. CONCLUSIONS: The benefits of spectacle correction for infants with hyperopia can be achieved without impairing the normal developmental regulation of refraction.

Accommodation, Ocular↗

PubMed1999

The x-ray structure of epoxide hydrolase from Agrobacterium radiobacter AD1. An enzyme to detoxify harmful epoxides.

Epoxide hydrolases catalyze the cofactor-independent hydrolysis of reactive and toxic epoxides. They play an essential role in the detoxification of various xenobiotics in higher organisms and in the bacterial degradation of several environmental pollutants. The first x-ray structure of one of these, from Agrobacterium radiobacter AD1, has been determined by isomorphous replacement at 2.1-A resolution. The enzyme shows a two-domain structure with the core having the alpha/beta hydrolase-fold topology. The catalytic residues, Asp107 and His275, are located in a predominantly hydrophobic environment between the two domains. A tunnel connects the back of the active-site cavity with the surface of the enzyme and provides access to the active site for the catalytic water molecule, which in the crystal structure, has been found at hydrogen bond distance to His275. Because of a crystallographic contact, the active site has become accessible for the Gln134 side chain, which occupies a position mimicking a bound substrate. The structure suggests Tyr152/Tyr215 as the residues involved in substrate binding, stabilization of the transition state, and possibly protonation of the epoxide oxygen.

Amino Acid Sequence↗

PubMed1999

Alpha/beta hydrolase fold enzymes: the family keeps growing.

The alpha/beta hydrolase fold is a typical example of a tertiary fold adopted by proteins that have no obvious sequence similarity, but nevertheless, in the course of evolution, diverged from a common ancestor. Recently solved structures demonstrate a considerably increased variability in fold architecture and substrate specificity, necessitating the redefinition of the minimal features that distinguish the family.

Aspartic Acid↗

PubMed1999

Benzoic and cinnamic acid derivatives as antioxidants: structure-activity relation.

The antioxidant activity of four derivatives of benzoic acid was systematically compared with the activity of the four homologous derivatives of cinnamic acid. The couples of compounds differed for the kind of aromatic substitution (p-hydroxy, p-hydroxymethoxy, p-hydroxydimethoxy, dihydroxy). The antioxidant activity was measured using (i) a competition kinetic test, to measure the relative capacity to quench peroxyl radical and (ii) the in vitro oxidative modification of human low-density lipoprotein (LDL), initiated by 2,2'-azobis(amidinopropane) dihydrochloride or catalyzed by Cu(II). In both models, cinnamic acids were more efficient than their benzoic counterparts. As for the influence of the aromatic substitution, in the kinetic test the antioxidant activity increased in the sequence p-hydroxy < p-hydroxymethoxy < dihydroxy < p-hydroxydimethoxy. In contrast, in the LDL system, the dihydroxy acids had an antioxidant capacity equal to or higher than that of the p-hydroxydimethoxy acids.

Antioxidants↗

PubMed1998

Antioxidant Activity of Different Phenolic Fractions Separated from an Italian Red Wine.

Using liquid/liquid extraction, three fractions were obtained from an Italian red wine containing single polyphenolic subfractions: (1) phenolic acids and quercetin-3-glucuronide, (2) catechins and quercetin-3-glucoside, and (3) anthocyanins. Beside the scavenging capacity of the different fractions against hydroxyl and peroxyl radicals, the in vitro inhibition of low density lipoprotein oxidation and platelet aggregation (two main events in the pathogenesis of atherosclerosis) were tested. The antioxidant activity of the fractions has been compared with that of the original red wine before and after dealcoholization. The anthocyanin fraction was the most effective both in scavenging reactive oxygen species and in inhibiting lipoprotein oxidation and platelet aggregation. This higher activity can be explained by both its high concentration in red wine and its antioxidant efficiency, which, at least for peroxyl radical scavenging, was three times as high as that of the other two fractions. Our results suggest that anthocyanins could be the key component in red wine in light of the protection against cardiovascular diseases, although this hypothesis needs in vivo evidence.

Journal Article↗

PubMed1998

Effect of caffeic acid on tert-butyl hydroperoxide-induced oxidative stress in U937.

Nonvitamin phenolic compounds are ubiquitous in food plants and therefore potentially present in human plasma in a diet-dependent concentration. The aim of this study was to evaluate the ability of caffeic acid, a phenolic acid with antioxidant activity, to affect cellular response in U937 human monocytic cells to t-butyl hydroperoxide-induced oxidative stress. In our experimental conditions caffeic acid was incorporated into cells without any cytotoxic effect. Caffeic acid-treated cells showed an increased resistance to oxidative challenge, as revealed by an higher percent of survival and the maintenance of an higher proliferative capacity in respect to control cells. This effect seems to be due to the ability of caffeic acid to reduce glutathione depletion and to inhibit lipid peroxidation during tBOOH treatment. It can be concluded that caffeic acid exerts an antioxidant action inside the cell, responsible for the observed modulation of the cellular response to oxidative challenge. Due to its presence in the diet, therefore, caffeic acid may play a role in the modulation of oxidative processes in vivo.

Antioxidants↗