Biomedical subjects

F Bortolotti

Publications and source records attributed to F Bortolotti.

At least 19 recordsLinked to original sources

PubMed2006

Molecular mechanism involved in the transport of a prodrug dopamine glycosyl conjugate.

We have previously demonstrated that dopamine conjugation to glucose allows it to induce therapeutic effects against Parkinson's disease after intravenous administration. In this paper we demonstrate that, unlike dopamine, the prodrug glu-dopamine is a transportable substrate of glucose transporters. Towards this, the effect of glucose-conjugation on the affinity and uptake of dopamine have been assessed in vitro, using human retinal pigment epithelium (HRPE) cells. Glucose transporter-mediated uptake was measured using [(3)H]3-O-methylglucose ([(3)H]3-O-MG) as the tracer. The uptake was found to be rapid and hyperbolically related to its concentrations (K(t)=7.8+/-1.2mM and V(max)=54+/-2 nmol/min mg protein). Inhibition experiments showed that dopamine was able to interact with glucose carriers only when conjugated to glucose (IC(50)=2.6+/-0.6mM). HPLC analysis of HRPE cell extracts showed that both dopamine and the prodrug permeate the cell, but only the uptake of the prodrug is inhibitable by glucose. This confirms that glucose transporters mediate the transport of the prodrug glu-dopamine, but not of dopamine. HRPE cells is therefore proposed as a promising model for in vitro studies involving the glucose transporter-mediated transport of drugs and their conjugates.

3-O-Methylglucose↗

PubMed2006

A novel multicompartimental system based on aminated poly(vinyl alcohol) microspheres/succinoylated pullulan microspheres for oral delivery of anionic drugs.

Poly(vinyl alcohol) (PVA) microspheres were prepared by dispersion reticulation with glutaraldehyde and further aminated. These microspheres were firstly loaded with diclofenac (DF) and then entrapped in cellulose acetate butyrate (CAB) microcapsules by an o/w solvent evaporation technique for intestinal delivery of drug. The encapsulated PVA microspheres due to their low swelling degree in intestinal fluids, do not have enough force to produce the disruption of CAB shell, therefore different amounts of succinoylated pullulan microspheres (SP-Ms) (exchange capacity up to 5.2 meq/g) were co-encapsulated. The SP-Ms do not swell in acidic pH, but swell up to 20-times in intestinal fluids causing the rupture of CAB shell and facilitating the escape of loaded PVA microspheres.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2006

Cationic liposomes as potential carriers for ocular administration of peptides with anti-herpetic activity.

In the present study the preparation, characterization and activity of cationic liposomes containing the secretory form of herpes simplex virus type 1 (HSV-1) glycoprotein B (gB1s) or two related polylysine rich peptides, namely DTK1 and DTK2, were described. The immunotherapeutic potential of these HSV antigens containing liposomes was examined with a rabbit ocular model of HSV-1 infection. Our study indicates that the liposomes (i) are able to encapsulate quantitatively gB1s and around 30% the DTK peptides, (ii) are characterized by dimensions compatible with ocular applications and (iii) can release the peptide comparably to the free solution. In addition, neutralization studies demonstrated that an anti-DTK specific polyclonal antiserum can inhibit HSV-1 infection, indicating that such peptides could be a good immunogen/antigen in an anti-HSV vaccine formulation. Although the vaccination protocol did not induce protection against the eye disease, a significative protection against a lethal ocular challenge was detectable together with the absence of reactivation episodes from latency on the survived animals. In this respect, the use of cationic liposomes coupled to gB1s and DTK peptides, as a local ocular vaccine, could represent an interesting approach in order to obtain a possible efficacy in protecting animals against a subsequent HSV-1 ocular challenge.

Animals↗

PubMed2006

Evolution of hepatitis C virus quasispecies in children with chronic hepatitis C.

BACKGROUND: Hepatitis C virus (HCV) circulates as a mixture of different but closely related genomes: this quasispecies nature could be essential for virus persistence and could induce resistance to interferon therapy. Since little is known on the behavior of HCV quasispecies in children and adolescents with chronic hepatitis C, we analyzed the virus population in six untreated children during a 5-year follow-up. METHODS: Six children aged 1-8 years, infected early in life with HCV, were included in the study. From each of them, 2 or 3 sequential serum samples obtained over a 5-year follow-up period were examined. The HCV quasispecies heterogeneity and diversity in the E2 hypervariable region-1 (HVR-1) were analyzed among samples by the heteroduplex mobility assay, and the distance between variants was estimated by the heteroduplex mobility ratio (HMR). RESULTS: The HCV population was initially highly homogeneous in all six children. During follow-up, diversification of HVR-1 leading to a more complex viral population occurred in all cases, and was particularly evident in the three older children (HMR: 0.82-0.54). Changes in the HVR-1 sequence occurred without relation to the profile of ALT and HCV-RNA levels. CONCLUSIONS: HCV quasispecies diversification is a common event during chronic hepatitis C in childhood. Host and environmental pressure could be major determinants. The increasing viral heterogeneity could impair the response to antiviral therapy, thus indicating a rationale for early antiviral treatment in children with chronic hepatitis C.

Child↗

PubMed2005

Nanoparticle formulation may affect the stabilization of an antiischemic prodrug.

The prodrug 5'-octanoyl-CPA (Oct-CPA) of the antiischemic N6-cyclopentyladenosine (CPA) has been encapsulated by nanoprecipitation in poly(lactic acid) nanoparticles, which have been recovered by gel-filtration, ultra-centrifugation or dialysis. We have analysed how different surfactants and purification methods can influence the nanoparticle characteristics. The particle sizes have been obtained by scanning electron microscope, whereas a SdFFF system was employed to detect their distributions. The Oct-CPA release from nanoparticles and stabilities in human blood of free and encapsulated prodrug have been analysed by HPLC techniques. The effects of nanoparticles on CPA interaction toward adenosine A1 receptor (its action site) have been analysed using radiolabelled drugs. The smallest nanoparticles and the best degree of homogeneity have been obtained using sodium cholate; the best recovery has been achieved by dialysis, whereas gel-filtration and ultra-centrifugation have induced the greatest removal of surfactants. The release of Oct-CPA was better controlled from the nanoparticles obtained using Pluronic F68 and purified by gel-filtration or ultra-centrifugation. Similarly, these nanoparticles better increased the stability of the prodrug in human blood. In particular, the nanoparticles purified by ultra-centrifugation induced a strong stability to a fraction of the encapsulated Oct-CPA. Any interference by unloaded nanoparticles has been registered for CPA-adenosine A1 receptor interaction.

Adenosine↗

PubMed2005

Spontaneous regression of hepatitis B virus-associated cirrhosis developed in childhood.

We reported two cases of hepatitis B virus infection-related cirrhosis developed during childhood and followed up for more than 20 years. Both the subjects remained untreated, and ultimately regression of cirrhosis was documented by clinical (including ultrasound) and histological examination. Recent studies have already suggested that hepatitis B virus-related cirrhosis may regress after treatment, but this is the first demonstration that hepatitis B virus-associated cirrhosis developed in childhood may be a spontaneously reversible process. Subsidence of virus replication and of necro-inflammatory process and the efficiency of liver regeneration and repair might contribute to this favourable outcome.

Age Factors↗

PubMed2005

HIV protease inhibitors: synthesis and activity of N-aryl-N'-hydroxyalkyl hydrazide pseudopeptides.

We describe the synthesis and activities of a series of pseudopeptides containing an N-aryl-N'-hydroxyalkyl hydrazide core structure to inhibit human immunodeficiency virus protease and viral replication. Of the series, compound Hmb-Leu-N(Bzl)-N(CH2-CH-OH)-rPro-Boc (24) displayed the greatest inhibitory potency (IC50 < 1 microM) and exhibited enzymatic resistance and stability in vitro.

HIV Infections↗

PubMed2005

Caffeine intake induces an alteration in human neutrophil A2A adenosine receptors.

Caffeine is the most widely used drug in the world and acts mainly through antagonism of the effects mediated by the adenosine receptor subtypes A1, A2A, A2B and A3. We determined whether repeated caffeine administration at different doses and for different periods of time (400 or 600 mg/day for 1 week and 400 mg/day for 2 weeks) alters human neutrophil A2A adenosine receptor density and function. Saturation binding assays showed an increase in affinity (K(D)) and density (B(max)) of A2A adenosine receptors after caffeine intake. These changes were accompanied by increases in cAMP accumulation and decreases in superoxide anion production after stimulation of the A2A receptor subtype using the agonist 5'-N-ethylcarboxamidoadenosine (NECA). Binding and functional changes of A2A receptors returned to baseline after 48 h of caffeine withdrawal. The findings are consistent with a potential anti-inflammatory effect of caffeine mediated by neutrophil A2A receptors.

Adenosine A2 Receptor Agonists↗

PubMed2005

Interferon treatment in children with chronic hepatitis C: long-lasting remission in responders, and risk for disease progression in non-responders.

BACKGROUND AND AIM: Large interferon-based therapeutic trials are still lacking in children with hepatitis C and the long-term safety and efficacy of interferon is unknown. This study describes the outcome of hepatitis C in 43 children enrolled in an open-label interferon trial, and were followed up to 66 months after stopping treatment. PATIENTS AND METHODS: All patients received interferon alfa2a (5MU/m(2)) thrice weekly for 6 months; children with genotype 1b received 3MU/m(2) thrice weekly for 6 additional months. RESULTS: Nine children discontinued interferon for adverse events and three were not compliant to treatment. Eight (19%, intention to treat analysis), including 2/20 (10%) with genotype 1b and 6/12 (50%) with genotypes 2 or 3, were sustained responders 12 months after stopping therapy. During further follow-up (mean+/-S.D.: 44.7+/-14.6 months), response was maintained; two non-responders cleared viremia, while a young boy progressed to cirrhosis. CONCLUSIONS: Small sample size and therapy withdrawal are the major limitations in the interpretation of our results. Nevertheless, our data, suggesting that response to interferon in children with hepatitis C is genotype-related and stable, agree with the results of large studies in adults. The outcome in non-responders was variable, including persistence of viremia and mild-moderate cytolysis (most cases), progression to cirrhosis, or eventual sustained viremia clearance.

Adolescent↗

PubMed2005

Hepatitis C virus (HCV) genotypes in 373 Italian children with HCV infection: changing distribution and correlation with clinical features and outcome.

BACKGROUND AND AIMS: Little is known of hepatitis C virus (HCV) genotypes in HCV infected children. This retrospective, multicentre study investigated genotype distribution and correlation with clinical features and outcome in a large series of Italian children. METHODS: Between 1990 and 2002, 373 HCV RNA positive children, consecutively recruited in 15 centres, were assayed for genotypes by a commercial line probe assay. RESULTS: The following genotype distribution pattern was recorded: genotype 1b = 41%; 1a = 20%; 2 = 17%; 3 = 14.5%; 4 = 5%; other = 2.5%. The prevalence of genotypes 1b and 2 decreased significantly (p<0.001) among children born from 1990 onwards compared with older children (46% v 70%) while the rate of genotypes 3 and 4 increased significantly (from 8% to 30%). Children infected with genotype 3 had the highest alanine aminotransferase levels and the highest rate of spontaneous viraemia clearance within the first three years of life (32% v 3% in children with genotype 1; p<0.001). Of 96 children enrolled in interferon trials during the survey, 22% definitely lost HCV RNA, including 57% of those with genotypes 2 and 3. CONCLUSION: HCV genotypes 1 and 2 are still prevalent among infected adolescents and young adults in Italy but rates of infection with genotypes 3 and 4 are rapidly increasing among children. These changes could modify the clinical pattern of hepatitis C in forthcoming years as children infected with genotype 3 have the best chance of spontaneous viraemia clearance early in life, and respond to interferon in a high proportion of cases.

Adolescent↗

PubMed2004

Vitamin C and 6-amino-vitamin C conjugates of diclofenac: synthesis and evaluation.

Diclofenac (Diclo), its ascorbic acid (AA) or 6-amino-AA (AA-NH2) pro-drugs (AA-Diclo or AA-NH-Diclo) were prepared and evaluated on human retinal pigment epithelium (HRPE) cells to investigate their ability to interact with the vitamin C transporter SVCT2 and their cellular uptake. Furthermore, stabilities in physiological fluids of these compounds were investigated. For kinetic experiments, AA-Diclo was incubated in Tris-HCl buffer, human plasma or whole blood. The extracted samples were analysed by HPLC. AA-Diclo was hydrolysed following first order kinetics in buffer, plasma (t1/2 about 10 h) and whole blood (t1/2 about 3.5 h). Transport and inhibition assays were performed by adding [14C]AA and the above-mentioned unlabelled compounds to plated HRPE cells. Intracellular accumulation was measured incubating HRPE cells with increasing concentrations of unlabelled compounds, following by HPLC analysis. Diclo resulted as a non-competitive inhibitor of AA-transport, showing a Na+-dependent and ascorbate-independent uptake. AA-Diclo behaved as a competitive inhibitor, but it was not transported into cells, whereas its analogue AA-NH-Diclo showed a decreased inhibitory activity. Stability studies suggest AA-Diclo as a potential candidate to enhance the Diclo short half life in vivo. The discovery of a Na+-dependent transporter for Diclo on HRPE cells opens new perspectives for targeting diclofenac into the brain.

Ascorbic Acid↗

PubMed2003

Amphiphilic association systems for Amphotericin B delivery.

The present study describes the production and characterization of amphiphilic association systems for Amphotericin B (AMB). In particular, three different classes of microemulsions and different monoglyceride-water systems were produced. Formulations were characterized for macroscopic aspect, pH, rheology, mean size and size distribution, both in the absence and in the presence of AMB. AMB solubility was investigated in the different formulations by HPLC studies. The formulations increased AMB solubility up to 20-fold with respect to the single oil and aqueous phases employed for microemulsion production.AMB diffusion studies from two microemulsions taken as models were performed in a Franz cell system using a nylon membrane. The physical and chemical stability of AMB-containing amphiphilic association systems were investigated for three months after production. For physical stability studies both the macroscopic aspect, droplet mean size and dimensional distribution were analysed. For chemical stability studies, the AMB content of the formulations was quantified by HPLC analysis. Microemulsions and monoglyceride-water systems were free from phase separation for up to three months and in some cases the AMB content was unchanged even after three months.

Administration, Topical↗

PubMed2003

Guidelines for the screening and follow-up of infants born to anti-HCV positive mothers.

Hepatitis C virus infection in infancy largely depends on vertical transmission. The transfer of hepatitis C virus from mother to child is almost invariably restricted to children whose mother is viremic, and the rate of transmission seems to be influenced by maternal virus load, although, in the single patient, the levels of viremia cannot be used as predictors of pediatric infection. In fact, the flow-chart for screening children at risk for vertically transmitted hepatitis C virus infection takes into account maternal viremia. In children born to anti-hepatitis C virus antibody positive, hepatitis C virus-RNA negative mothers, alanine aminotransferase and anti-hepatitis C virus should be investigated at 18-24 months of life. If alanine aminotransferase values are normal and anti-hepatitis C virus is undetectable, follow-up should be interrupted. In children born to hepatitis C virus-RNA positive mothers, alanine aminotransferase and hepatitis C virus RNA should be investigated at 3 months of age: (1) hepatitis C virus-RNA positive children should be considered infected if viremia is confirmed by a second assay performed within the 12th month; (2) hepatitis C virus-RNA negative children with abnormal alanine aminotransferase should be tested again for viremia at 6-12 months, and for anti-hepatitis C virus at 18 months; (3) hepatitis C virus-RNA negative children with normal alanine aminotransferase should be tested for anti-hepatitis C virus and alanine aminotransferase at 18-24 months, and should be considered non-infected if alanine aminotransferase is normal and anti-hepatitis C virus undetectable; (4) anti-hepatitis C virus seropositivity beyond the 18th month in a never-viremic child with normal alanine aminotransferase is likely consistent with past hepatitis C virus infection.

Breast Feeding↗

PubMed2002

Determination of CDT, a marker of chronic alcohol abuse, for driving license issuing: immunoassay versus capillary electrophoresis.

The present work is aimed at a validation of the carbohydrate-deficient transferrin (CDT) determination in real cases by comparison between the a commercial immunometric method and a method based on capillary electrophoresis. Overall, 650 serum samples from subjects applying to re-obtain their driving license, previously withdrawn for "drunk driving", were investigated. A highly significant correlation (P < 0.001) was found between the results from immunoassay and capillary electrophoresis. However, particularly in the samples with CDT values around the cut-off or moderately elevated, a wide dispersion of the correlation data was found. This finding stresses the need to confirm by alternative techniques all the results from CDT immunoassays. For this purpose, capillary electrophoresis, because of its inherent characteristics of high selectivity, easy operation, high productivity and low operative costs looks well-suited for becoming the method of choice.

Adult↗

PubMed2002

Capillary zone electrophoresis and artificial neural networks for estimation of the post-mortem interval (PMI) using electrolytes measurements in human vitreous humour.

Determination of electrolyte concentrations (mainly potassium) in vitreous humour has long been considered an important tool in human death investigations for the estimation of the post-mortem interval (PMI). On the basis of its well known potential in ion analysis, capillary zone electrophoresis (CZE) has recently been applied to achieve a rapid and simultaneous determination of inorganic ions in this extracellular fluid. In the present work, artificial neural networks (ANN) were applied for modelling of the relationship of multicomponent CZE analysis of K+, NH4+, Na+, and Ba2+ ions in vitreous humour with PMI. In a study based on 61 cases with different causes of death and a known PMI ranging from 3 to 144 h, the use of ANNs considering all inorganic ion data from the human vitreous humour, achieved a substantial improvement of post-mortem interval prediction. Good linear correlation was observed (r2 = 0.98) and in comparison to the traditional linear least squares (LLS) method applied only to K+ levels in the vitreous humour, the prediction of PMI with ANN was improved by a factor of 5 from approximately +/- 15 h to less than 3 h.

Autopsy↗

PubMed2001

Potassium concentration differences in the vitreous humour from the two eyes revisited by microanalysis with capillary electrophoresis.

This paper presents a study of the variability of potassium concentrations in the vitreous humour of the two eyes of the same body at identical postmortem interval. The study was carried out by collecting microsample amounts (50 microl) of vitreous humour and by using an original method of capillary electrophoresis with indirect detection. The electrophoretic separations were carried out in a pH 4.5 running buffer composed of 5 mmol/l imidazole, 5 mmol/l 18-crown-6 ether and 6 mmol/l alpha-hydroxybutyric acid. Detection was by indirect UVabsorption at 214 nm. Vitreous humour samples were collected from 57 medico-legal autopsies or external examinations of cases of sudden natural or violent deaths. All samples prior to analysis were diluted 1:20 with a 40 microg/ml aqueous solution of barium, the used internal standard, and finally injected by nitrogen pressure. The mean concentrations of potassium measured in the two eyes of all the cases included in the present study ranged from 4.1 to 23.5 mmol/l with the postmortem interval values varying from 7 to 144 h. A highly significant (P<0.0001) linear correlation was found between these two parameters as described by the equation: y=0.1698x+2.3587, r=0.89. The intra-eye variability of potassium concentrations was low with an average RSD of 3.89% (+/- 1.83 SD) (48 eyes, five samples per eye). No statistically significant difference was found between the potassium concentrations in the two eyes of the same subject in a group of 24 cases, excepting a single case.

Electrophoresis, Capillary↗

PubMed2001

Use of beta-cyclodextrin in the capillary zone electrophoretic separation of the components of clandestine heroin preparations.

The present paper describes the methodological optimization and validation of a capillary zone electrophoresis method for the rapid determination of heroin, secondary products and additives present in clandestine heroin samples, by using 20 mM beta-cyclodextrins in phosphate buffer, pH 3.23. Applied potential was 15 kV and separation temperature was 24 degrees C; detection was by UV absorption at 200 nm wavelength. Heroin samples were first dissolved in CHCl3-MeOH (96:4, v/v) and injected by pressure (0.5 p.s.i., 3 s; 1 p.s.i.=6894.76 Pa) after evaporation of the organic mixture and reconstitution in aqueous buffer. Under the described conditions, phenylethylamine (internal standard), morphine, monoacetylmorphine, heroin, acetylcodeine, papaverine, codeine and narcotine were baseline resolved in less than 10 min. The limit of detection was better than 1 microg/ml for each analyte. The study of the intra-day and day-to-day precision showed, in terms of migration times, RSDs < or = 0.71% and, in terms of peak areas, RSDs < or = 3.2%. Also, the evaluation of linearity and analytical accuracy of the method provided good results for all the analytes investigated, thus allowing its application to real cases of seized controlled drug preparations.

Cyclodextrins↗

PubMed2001

Hepatitis B virus infection in native versus immigrant or adopted children in Italy following the compulsory vaccination.

BACKGROUND: Compulsory vaccination of children against hepatitis B virus (HBV) infection was introduced in Italy in 1991. PATIENTS AND METHODS: To evaluate the current importance of pediatric HBV infection, we studied 359 HBsAg-positive children admitted to 16 centers in Italy from 1991 to 1998. 185 patients were natives of Italy and 174 (39 immigrants and 135 adopted) came from highly endemic countries (eastern Europe: 60.9%, Asia: 16.7%, Africa: 14.9% and Central and South America: 5.7%). RESULTS: Transaminase Levels were moderately altered in both Italian (mean 134 UI/L) and foreign children (mean 168 UI/L). In total, 77% of ItaLian children and 88% of foreign children tested HBeAg positive. High transaminase levels and HBeAg positivity were more frequent in adopted children. Follow-up of 317 patients showed that the incidence of HBeAg/anti-HBe serum conversion was similar in all cohorts, but in adopted children it occurred at an earlier age and was associated with HBsAg clearance in 5%. CONCLUSION: HBV is not frequent in Italian children today, but it is common among children coming from highly endemic areas. The vaccination of nonimmune native populations must be strongly recommended.

Adolescent↗