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P Giusti

Publications and source records attributed to P Giusti.

At least 19 recordsLinked to original sources
PubMed2006

Transglutaminase reactivity with gelatine: perspective applications in tissue engineering.

Gelatine was crosslinked by means of an enzymatic treatment using tissue transglutaminase (tTGase) (Sigma) and microbial transglutaminase (mTGase) (Ajinomoto) which catalyses the formation of isopeptide bonds between the gamma-carbonyl group of a glutamine residue and the epsilon-amino group of a lysine residue. The reaction is an interesting alternative to the traditional glutaraldehyde crosslinking, which has several drawbacks (e.g., in medical application) due to the toxicity of the chemical reagent. To further investigate the possibility to utilize the modified protein for tissue engineering application, TGase crosslinked gelatine was incorporated in a gellan matrix, a polysaccharide, to enhance the stability in aqueous media. Films obtained by casting were characterized by thermal analysis, chemical imaging, swelling behaviour and cell adhesion.

Animals↗
PubMed2005

The relevance of the transfer of molecular information between natural and synthetic materials in the realisation of biomedical devices with enhanced properties.

Past and recent attempts to introduce in synthetic polymers molecular information from natural substances through simple blending, template polymerization and molecular imprinting are reviewed. The most promising approaches that can open the way to the realisation of new materials with improved biocompatibility, antibody- or enzyme-like performances are analysed more deeply. The realisation of bioartificial blends from natural and synthetic polymers, molecularly imprinted nanospheres or membranes that can act as recognition element in (bio)sensing devices, as synthetic enzymes or as key constituents of body fluids purification tools is presented in order to make the reader aware of the fascinating possibilities that these techniques make available to the biomedical science and engineering in the close future. The last part of the paper describes recent attempts to insert recognition elements for large molecules as proteins, DNA segments, viruses or whole cells in synthetic polymer systems, in order to develop new systems in the treatments of diseases and for tissue-engineering applications.

Animals↗
PubMed2005

Molecularly imprinted bioartificial membranes for the selective recognition of biological molecules. Part 2: release of components and thermal analysis.

Molecularly imprinted membranes imprinted for a large-molecular-weight protein were realised using a blend of natural and synthetic polymers. Bioartificial membranes of synthetic (poly(ethylene-co-vinyl alcohol)-EVAL, Clarene) and biological (Dextran) polymers, molecularly imprinted with alpha-amylase as the template, were prepared and investigated. Dimethyl sulfoxide (DMSO) solutions of the alpha-amylase template, Clarene and Dextran were mixed under stirring in the desired proportions and dipped in DMSO (solvent)/water (non solvent) mixture, to obtain the phase separation. The release of Clarene, Dextran and alpha-amylase in the inversion baths was quantified by spectrophotometric methods and final composition of membranes was established. To study the interactions between the polymer components and between polymeric materials and the template, differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA) were carried out. Results indicated that stable and continuous bioartificial membranes of Clarene and Dextran can be obtained, whereby calorimetric analysis suggested the presence of high interaction between alpha-amylase and the Clarene component.

Biocompatible Materials↗
PubMed2004

Acrylic polymeric nanospheres for the release and recognition of molecules of clinical interest.

Cross-linked poly(methylmethacrylate-co-methacrylic acid) nanospheres were imprinted with theophylline through template radical polymerisation in diluted acetonitrile solution. This study will focus on the effect of functional monomer nature used (methylmethacrylate and/or methacrylic acid) in the recognition and in the release of template in order to develop a material with combined properties of drug delivery and rebinding for clinical applications. After template extraction the nanospheres showed satisfactory recognition properties (up to 1mg template/g of polymer). Moreover polymers prepared selectively removed theophylline with a theophylline rebinding of 5.1 times higher than that of caffeine, a compound of similar structure. Drug release properties were also satisfactory (up to 95% of loaded theophylline in 7 days).

Acrylic Resins↗
PubMed2004

Molecularly imprinted bioartificial membranes for the selective recognition of biological molecules.

Membranes of a synthetic (poly(ethylene-co-vinyl alcohol), Clarene) and a biological (dextran) polymer, imprinted with alpha-amylase, of different compositions were prepared by the phase-inversion process. Membrane morphologies were shown to be dependent on the synthetic-biological components composition. The removal of the template from the membranes was performed by extraction with water, while an aqueous solution of alpha-amylase was permeated across the membranes under pressure to obtain the rebinding of the template. The selectivity of alpha-amylase-imprinted membranes was investigated by the same uptake experiment using pepsin, albumin and amyloglucosidase, and the rebinding of these proteins was compared with that of the print molecule. Before and after template extraction and after the rebinding experiment, kinetic measurements of the imprinting molecule were conducted to estimate the activity of the enzyme immobilised in the polymer matrix. Results obtained revealed that the immobilised enzyme maintains a good functionality while in the membrane compared to the free enzyme and the imprinted 'bioartificial' dextran and Clarene membranes, obtained by the phase-inversion method, can establish efficient interaction with alpha-amylase as template molecule, as confirmed by the fair selectivity in rebinding tests.

Carbohydrate Sequence↗
PubMed2004

Cerebellar granular cell cultures as an in vitro model for antidepressant drug-induced neurogenesis.

Both preclinical and clinical evidence suggested that antidepressant drugs upregulate hippocampal cell proliferation and neurogenesis. In addition, direct evidence was recently published that hippocampal de novo cell proliferation is necessary for antidepressant action. Within this frame, we used primary cultures of rat cerebellar granule cells (CGC) as an in vitro model of central nervous system (CNS) to investigate whether a neurogenic response could be elicited also in the cerebellum, upon chronic treatment with selective serotonin reuptake inhibitors (SSRIs). Furthermore, we assayed the presence of neural precursor cells in CGC, possibly responsive to proliferation and differentiation stimuli. We found that 1 microM fluoxetine increased cell proliferation, as assayed by [3H]-thymidine incorporation. CGC immunocytochemical analysis with neural cell-specific markers revealed the presence of granule neurons, glial cells, and a cell component that we named "round cells." Because only round cells displayed proliferation ability, as revealed by 5-bromo-2'-deoxyuridine (BrdU) labeling, they were further characterized. For this purpose, round cells were isolated and expanded by culturing in a serum-free medium, containing basic fibroblast growth factor (bFGF), before immunocytochemical analysis. We found that round cells were not immunoreactive for glial, neuronal, and oligodendrocyte markers, whereas they were immunoreactive for several immature neuronal markers. Accordingly, round cells could be induced to differentiate into astrocytes, neurons, and oligodendrocytes, either by withdrawing the mitogen bFGF or by exposing them to fluoxetine. These findings suggest that round cells in CGC possess the features and potentials of neural precursors, able to differentiate in mature neural cells upon a pharmacological simulum.

Animals↗
PubMed2001

Melatonin protects against 6-OHDA-induced neurotoxicity in rats: a role for mitochondrial complex I activity.

Unilateral injection into the right substantia nigra of the catecholaminergic neurotoxin 6-hydroxydopamine (6-OHDA) produces extensive loss of dopaminergic cells ('hemi-parkinsonian rat'). The pineal hormone melatonin, which is a potent antioxidant against different reactive oxygen species and has been reported to be neuroprotective in vivo and in vitro, was evaluated for potential anti-Parkinson effects in this model. Imbalance in dopaminergic innervation between the striata produced by intranigral administration of 6-OHDA results in a postural asymmetry causing rotation away from the nonlesioned side. Melatonin given systemically prevented apomorphine-induced circling behavior in 6-OHDA-lesioned rats. Reduced activity of mitochondrial oxidative phosphorylation enzymes has been suggested in some neurodegenerative diseases; in particular, selective decrease in complex I activity is observed in the substantia nigra of Parkinson's disease patients. Analysis of mitochondrial oxidative phosphorylation enzyme activities in nigral tissue from 6-OHDA-lesioned rats by a novel BN-PAGE histochemical procedure revealed a clear loss of complex I activity, which was protected against in melatonin-treated animals. A good correlation between behavioral parameters and enzymatic (complex I) analysis was observed independent of melatonin administration. A deficit in mitochondrial complex I could conceivably contribute to cell death in parkinsonism via free radical mechanisms, both directly via reactive oxygen species production and by decreased ATP synthesis and energy failure. Melatonin may have potential utility in the treatment of neurodegenerative disorders where oxidative stress is a participant.

Adenosine Triphosphatases↗
PubMed2001

Bioartificial polymeric materials based on polysaccharides.

Bioartificial polymeric materials, based on blends of polysaccharides with synthetic polymers such as poly(vinyl alcohol) (PVA) and poly(acrylic acid) (PAA), were prepared as films or hydrogels. The physico-chemical, mechanical, and biological properties of these materials were investigated by different techniques such as differential scanning calorimetry, dynamic mechanical thermal analysis, scanning electron microscopy, and in vitro release tests, with the aim of evaluating the miscibility of the polymer blends and to establish their potential applications. The results indicate that while dextran is perfectly miscible with PAA, dextran/PVA, chitosan/PVA, starch/PVA, and gellan/PVA blends behave mainly as two-phase systems, although interactions can occur between the components. Cross-linked starch/PVA films could be employed as dialysis membranes: they showed transport properties comparable to, and in some cases better than, those of currently used commercial membranes. Hydrogels based on dextran/PVA and chitosan/PVA blends could find applications as delivery systems. They appeared able to release physiological amounts of human growth hormone, offering the possibility to modulate the release of the drug by varying the content of the biological component.

Biocompatible Materials↗
PubMed2000

Intracellular glutathione levels determine cerebellar granule neuron sensitivity to excitotoxic injury by kainic acid.

Glutathione (GSH) is a key component of the cellular defence cascade against injury caused by reactive oxygen species. Kainic acid (KA) is a potent central nervous system excitotoxin. KA-elicited neuronal death may result from the generation of ROS. The present study was undertaken to characterize the role of GSH in KA-induced neurotoxicity. Cultures of cerebellar granule neurons were prepared from 8-day-old rats, and used at 8, 14 and 20 days in vitro (DIV). Granule neurons displayed a developmental increase in their sensitivity to KA injury, as quantified by an ELISA-based assay with the tetrazolium salt MTT. At DIV 14 and 20, a 30-min challenge with KA (500 microM) reduced cell viability by 45% after 24 h, significantly greater (P<0.01) than the 22% cell loss with DIV 8 cultures. Moreover acute (30 min) KA exposure concentration-dependently reduced intracellular GSH and enhanced reactive oxygen species generation (evaluated by 2', 7'-dichlorofluorescein diacetate). In comparison to control, KA (500 microM) lowered GSH levels in DIV 8 granule neurons by 16% (P=0. 0388), and by 36% (P=0.0001) in both DIV 14 and DIV 20 neurons, after 30 min. Preincubation of granule neurons with the membrane permeant GSH delivery agent, GSH ethyl ester (5 mM), for 30 min significantly increased intracellular GSH content. Importantly, GSH ethyl ester reduced the toxic effects of KA, becoming significant at 1 mM (P=0.007 vs. KA-treated group), and was maximal at >/=2.5 mM (P<0.0001). GSH ethyl ester displayed a similar dose-dependence in its ability to counteract KA-induced depletion of cellular GSH. The data strengthen the notion that cellular GSH levels have a fundamental role in KA-induced neurotoxicity.

Animals↗
PubMed2000

Benzodiazepine receptor affinities, behavioral, and anticonvulsant activity of 2-aryl-2,5-dihydropyridazino[4,3-b]indol- 3(3H)-ones in mice.

The anticonvulsant properties of 1,4-benzodiazepines (BDZs), pyrazoloquinolones (CGS), 2-aryl-2,5-dihydropyridazino[4, 3-b]indol-3(3H)-ones (PIs) 1 1i 1d 1f 1e 1b 1c 1h, and 1a, the latter being inactive against audiogenic seizures. Some PIs 1 and abecarnil showed anticonvulsant properties against seizures induced by PTZ with a potency lower than that observed in audiogenic seizures. The pharmacological actions of 1d, 1f, and 1i were significantly reduced by a treatment with flumazenil (8.24 micromol/kg IP), suggesting a clear involvement of benzodiazepine mechanisms in the anticonvulsant activity of these compounds or their metabolites. The anticonvulsant activity of 1d, 1f, and 1i was also evaluated against seizures induced by two beta-carbolines namely methyl-beta-carboline-3-carboxylate (beta-CCM) and methyl-6, 7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM), in DBA/2 mice: they gave better protection against seizures induced by beta-CCM than the ones by DMCM. The potency of various BDZs and PIs as inhibitors of specific [3H]flumazenil binding to neuronal membranes, was also evaluated. The radioligand binding study, carried out on stable cell lines expressing definite combinations of benzodiazepine receptor subunits, demonstrated that 1b, 1e, 1d, and 1h have preferential interaction with alpha(1), beta(3), gamma(2), receptor subtypes.

Animals↗
PubMed1999

A new place conditioning paradigm to study tolerance to opiates in mice.

Tolerance to the rewarding properties of morphine was investigated in mice using a new conditioned place preference (CPP) procedure. Four pairings of morphine with specific environmental cues induced a significant CPP for the drug-paired cues. Further opiate conditioning trials in the presence of the same environmental cues revealed no change in the drug-induced CPP on repeated test sessions. Subsequent exposure of the same animals to conditioning trials by pairing morphine with a set of novel environmental cues showed that the opiate was still able to produce a CPP in mice treated with a total of 16 morphine injections. The present CPP paradigm may prove useful to investigate tolerance to the rewarding properties of drugs of abuse.

Animals↗
PubMed1999

Further evidences for neuroprotective effects of melatonin.

The physiological roles of the pineal hormone melatonin are still not completely clarified. Recently it has been shown that melatonin is a potent, endogenous scavenger of reactive oxygen species suggesting that it might interfere with neurodegenerative processing involving free-radical formation and excitatory aminoacid release. These neuroprotective effects of melatonin may result, at least in part, from a sparing of glutathione reductase, which is decreased following administration of the neurotoxic agent kainate (KA) in rats. Moreover, KA causes a rapid decrease in glutathione (GSH) content of cultured cerebellar granule neurons but not in astrocytes. These cell types both express functional KA receptors, but only the former is sensitive to reactive oxygen species-dependent KA injury. Melatonin counteracts the changes in GSH, induced by KA, in cultured cerebellar granule neurons.

Animals↗
PubMed1999

Abecarnil enhances recovery from diazepam tolerance.

Treatment with diazepam (25 mg/kg; p.o., twice-daily for 17 days) induced tolerance to the anticonvulsant effect of diazepam against bicuculline-induced convulsions in mice. Cross-tolerance was observed to the anticonvulsant action of clonazepam, imidazenil but not abecarnil. While substitution of clonazepam (12 mg/kg; p.o., twice-daily for 15 days) for diazepam did not affect tolerance to diazepam, substitution of imidazenil (17 mg/kg; p.o., twice-daily for 15 days) for diazepam significantly increased sensitivity to the anticonvulsant effect of diazepam, although tolerance was not abolished. Tolerance to diazepam progressively decreased either after suspension of diazepam administration or replacement treatment with abecarnil (20 mg/kg; p.o., twice-daily). Complete recovery of diazepam efficacy was detected after 8 and 15 days of administration of abecarnil and vehicle, respectively. Binding experiments using [3H]-flumazenil showed that Kd values did not differ among treatment groups. A significant decrease in Bmax (-42%) was observed in the cortex of diazepam-tolerant mice whether or not also treated with imidazenil and clonazepam. Conversely, chronically diazepam-treated mice, that further received abecarnil for either 8 or 15 days or vehicle for 15 days showed Bmax values similar to those of vehicle-treated mice never exposed to diazepam. Results suggest that repeated abecarnil administration to diazepam-tolerant mice can facilitate re-adaptation of receptors to the diazepam-free state. It is proposed that replacement therapy with abecarnil after long-term treatment with conventional benzodiazepines (BDZs) may provide a novel approach for reducing tolerance to their anticonvulsant effects.

Animals↗
PubMed1999

Effect of acute and chronic tramadol on [3H]-norepinephrine-uptake in rat cortical synaptosomes.

1 Tramadol hydrochloride is a centrally acting opioid analgesic whose efficacy and potency is only five to ten times lower than that of morphine. Opioid, as well as non-opioid mechanisms, may participate in the analgesic activity of tramadol. 2 [3H]-NE uptake in isolated rat cortical synaptosomes was studied in the presence of tramadol, desipramine, methadone, and morphine. Desipramine and tramadol inhibited synaptosomal [3H]-NE uptake with apparent Kis of 7.3 +/- 0.66 and 1.4 +/- 0.0045 microM, respectively. Methadone was active at a 10-fold higher concentration (Ki: 87 +/- 5.6 microM). In contrast, morphine essentially failed to inhibit [3H]-5-HT uptake (Ki: 0.75 +/- 0.40 M). 3 Methadone, morphine, and tramadol were active in the hot plate test with ED50s of 6.2, 9.3, and 40 mg kg-1, respectively. 4 [3H]-NE uptake was examined in synaptosomes prepared from rats 30 min after receiving a single dose of morphine, methadone or tramadol. Only tramadol (31 mg kg-1, i.p.) decreased uptake of the transmitter, with an ED50 equal to that in the hot plate test. 5 Animals were chronically treated for 15 days with increasing doses of tramadol (20 to 125 mg kg-1, i.p.). Twenty-four hours after the last drug injection, a challenge dose of tramadol (40 mg kg-1, i.p.) was administered. Chronic tramadol was still able to reduce [3H]-NE uptake by 35%. 6 These results further support the hypothesis that [3H]-NE uptake inhibition may contribute to the antinociceptive effects of tramadol. The lack of tolerance in [3H]-NE uptake, together with the absence of behavioural alteration after chronic tramadol treatment proposes that tramadol holds potential over classical opioids in the treatment of pain disorders.

Analgesics, Opioid↗
PubMed1999

Excitotoxicity, oxidative stress, and the neuroprotective potential of melatonin.

The brain consumes large quantities of oxygen relative to its contribution to total body mass. This, together with its paucity of oxidative defense mechanisms, places this organ at risk for damage mediated by reactive oxygen species. The pineal secretory product melatonin possesses broad-spectrum free radical scavenging and antioxidant activities, and prevents kainic acid-induced neuronal lesions, glutathione depletion, and reactive oxygen species-mediated apoptotic nerve cell death. Melatonin's action is thought to involve electron donation to directly detoxify free radicals such as the highly toxic hydroxyl radical, which is a probable end-product of the reaction between NO. and peroxynitrite. Moreover, melatonin limits NO.-induced lipid peroxidation, inhibits cerebellar NO. synthase, scavenges peroxynitrite, and alters the activities of enzymes that improve the total antioxidative defense capacity of the organism. Melatonin function as a free radical scavenger and antioxidant is likely facilitated by the ease with which it crosses morphophysiological barriers, e.g., the blood-brain barrier, and enters cells and subcellular compartments. Pinealectomy, which eliminates the nighttime rise in circulating and tissue melatonin levels, worsens both reactive oxygen species-mediated tissue damage and brain damage after focal cerebral ischemia and excitotoxic seizures. That melatonin protects against hippocampal neurodegeneration linked to excitatory synaptic transmission is fully consistent with the last study. Conceivably, the decreased melatonin secretion that is documented to accompany the aging process may be exaggerated in populations with dementia.

Animals↗
PubMed1998

Acetylcholinesterase inhibitors: synthesis and structure-activity relationships of omega-[N-methyl-N-(3-alkylcarbamoyloxyphenyl)- methyl]aminoalkoxyheteroaryl derivatives.

Acetylcholinesterase (AChE) inhibitors are one of the most actively investigated classes of compounds in the search for an effective treatment of Alzheimer's disease. This work describes the synthesis, AChE inhibitory activity, and structure-activity relationships of some compounds related to a recently discovered series of AChE inhibitors: the omega-[N-methyl-N-(3-alkylcarbamoyloxyphenyl)methyl]aminoalkoxy xanthen-9-ones. The influence of structural variations on the inhibitory potency was carefully investigated by modifying different parts of the parent molecule, and a theoretical model of the binding of one representative compound to the enzyme was developed. The biological properties of the series were investigated in some detail by considering not only the activity on isolated enzyme but the selectivity with respect to butyrylcholinesterase (BuChE) and the in vitro inhibitory activity on rat cerebral cortex as well. Some of the newly synthesized derivatives, when tested on isolated and/or AChE-enriched rat brain cortex fraction, displayed a selective inhibitory activity and were more active than physostigmine. In particular, compound 13, an azaxanthone derivative, displayed the best rat cortex AChE inhibition (190-fold higher than physostigmine), as well as a high degree of enzyme selectivity (over 60-fold more selective for AChE than for BuChE). When tested in the isolated enzyme, compound 13 was less active, suggesting some differences either in drug availability/biotransformation or in the inhibitor-sensitive residues of the enzyme when biologically positioned in rat brain membranes.

Acetylcholinesterase↗