Biomedical subjects

P Calvo

Publications and source records attributed to P Calvo.

At least 19 recordsLinked to original sources

PubMed2006

Compatibility and stability of the novel anticancer agent ES-285 x HCl formulated with 2-hydroxypropyl-beta-cyclodextrin in infusion devices.

ES-285 x HCl is a novel marine-derived anticancer agent isolated from the clam Spisula polynyma. The compound is pharmaceutically formulated as a lyophilised product containing 25 or 50 mg ES-285 x HCl and 500 or 1000 mg 2-hydroxypropyl-beta-cyclodextrin per dosage unit and requires reconstitution with sterile water for injection before intravenous administration. The aim of this study was to determine the stability and compatibility of ES-285 x HCl in infusion devices. ES-285 x HCl was shown to be stable at concentrations of 10-1400 microg/ml after dilution in 5% dextrose in water and compatible with PE infusion containers and PE and silicone tubing. No sorption on- or into the administration set was observed at concentrations equal to or above 20 microg/ml. In conclusion, ES-285 x HCl infusion solutions can be administered without stability or sorption problems using a PE infusion container and PE or silicone tubing in concentrations equal or above 20 microg/ml in 3-hour or 24-hour infusion administration schedules.

2-Hydroxypropyl-beta-cyclodextrin↗

PubMed2003

Myoglobinuria and COX deficiency in a patient taking cerivastatin and gemfibrozil.

The authors describe a patient who presented with myoglobinuria after starting cerivastatin-gemfibrozil therapy. Muscle histochemistry revealed ragged-red fibers and cytochrome c oxidase negative (COX) fibers, and biochemistry showed a defect of COX activity. Immunoblot analysis showed a 60% reduction of COX I and COX II polypeptides. Cerivastatin myotoxicity might be related to a depletion of essential metabolites needed to anchor COX subunit I to mitochondrial membrane.

Aged↗

PubMed2002

Design of biodegradable particles for protein delivery.

Major research issues in protein delivery include the stabilization of proteins in delivery devices and the design of appropriate protein carriers in order to overcome mucosal barriers. We have attempted to combine both issues through the conception of new biodegradable polymer nanoparticles: (i) poly(ethylene glycol) (PEG)-coated poly(lactic acid) (PLA) nanoparticles, chitosan (CS)-coated poly(lactic acid-glycolic acid (PLGA) nanoparticles and chitosan (CS) nanoparticles. These nanoparticles have been tested for their ability to load proteins, to deliver them in an active form, and to transport them across the nasal and intestinal mucosae. Additionally, the stability of some of these nanoparticles in simulated physiological fluids has been studied. Results showed that the PEG coating improves the stability of PLA nanoparticles in the gastrointestinal fluids and helps the transport of the encapsulated protein, tetanus toxoid, across the intestinal and nasal mucosae. Furthermore, intranasal administration of these nanoparticles provided high and long-lasting immune responses. On the other hand, the coating of PLGA nanoparticles with the mucoadhesive polymer CS improved the stability of the particles in the presence of lysozyme and enhanced the nasal transport of the encapsulated tetanus toxoid. Finally, nanoparticles made solely of CS were also stable upon incubation with lysozyme. Moreover, these particles were very efficient in improving the nasal absorption of insulin as well as the local and systemic immune responses to tetanus toxoid, following intranasal administration. In summary, these results show that a rational modification in the composition and structure of the nanoparticles, using safe materials, increases the prospects of their usefulness for mucosal protein delivery and transport.

Animals↗

PubMed2001

PEGylated polycyanoacrylate nanoparticles as vector for drug delivery in prion diseases.

PEGylated polymeric nanoparticles are hereby presented as a potential efficient drug carrier for the delivery of active therapeutic molecules in prion experimental diseases. Based on their blood long-circulating characteristics, these PEGylated particles made by the amphiphilic copolymer poly [methoxy poly(ethylene glycol) cyanoacrylate-co-hexadecyl cyanoacrylate] (PEG-PHDCA), showed comparatively conventional non-PEGylated nanoparticles, a higher uptake by the spleen and the brain which are both the target tissues of PrPres accumulation in scrapie infected animals.

Animals↗

PubMed2001

Polysaccharide colloidal particles as delivery systems for macromolecules.

Mucosal delivery of complex molecules such as peptides, proteins, oligonucleotides, and plasmids is one of the most intensively studied subjects. The use of colloidal carriers made of hydrophilic polysaccharides, i.e. chitosan, has arisen as a promising alternative for improving the transport of such macromolecules across biological surfaces. This article reviews the approaches which have aimed to associate macromolecules to chitosan in the form of colloidal structures and analyzes the evidence of their efficacy in improving the transport of the associated molecule through mucosae and epithelia. Chitosan has been shown to form colloidal particles and entrap macromolecules through a number of mechanisms, including ionic crosslinking, desolvation, or ionic complexation, though some of these systems have been realized only in conjunction with DNA molecules. An alternative involving the chemical modification of chitosan has also been useful for the association of macromolecules to self-assemblies and vesicles. To date, the in vivo efficacy of these chitosan-based colloidal carriers has been reported for two different applications: while DNA-chitosan hybrid nanospheres were found to be acceptable transfection carriers, ionically crosslinked chitosan nanoparticles appeared to be efficient vehicles for the transport of peptides across the nasal mucosa. The potential applications and future prospects of these new systems for mucosal delivery of macromolecules are highlighted at the end of the chapter.

Administration, Intranasal↗

PubMed2001

Development and characterization of CyA-loaded poly(lactic acid)-poly(ethylene glycol)PEG micro- and nanoparticles. Comparison with conventional PLA particulate carriers.

Cyclosporin A (CyA) loaded poly(lactic acid)-poly(ethylene glycol) (PLA-PEG) micro- and nanoparticles have been developed using an emulsion-solvent evaporation method. Physico-chemical properties, peptide loading content and in vitro release profiles of these novel CyA carriers were compared with those corresponding to conventional PLA micro- and nanoparticles. Results obtained confirm the previously described disposition of PEG chains on the surface of the PLA-PEG formulations. In addition, they revealed the presence of CyA molecules on the surface of both PLA and PLA-PEG systems. Further determination of the surface chemical composition by electron spectroscopy for chemical analysis (ESCA) allowed us to quantify the amount of CyA in the nanospheres' top layers, this amount being higher for nanoparticles than for microparticles, and higher for the PLA systems than for those based on PLA-PEG. In vitro release experiments revealed that PLA-PEG particles provided a more adequate control of CyA release than conventional PLA micro- and nanoparticles. Physico-chemical characterization of the systems during the release studies showed that the developed PLA and PLA-PEG micro- and nanoparticles were not degraded, which suggest a diffusion-mediated release mechanism. Furthermore, we have hypothesized that the hydrophilic outer shell of PEG provides a stationary layer for the diffusion of CyA.

Cyclosporine↗

PubMed2001

Long-circulating PEGylated polycyanoacrylate nanoparticles as new drug carrier for brain delivery.

PURPOSE: The aim of this study was to evaluate the ability of long-circulating PEGylated cyanoacrylate nanoparticles to diffuse into the brain tissue. METHODS: Biodistribution profiles and brain concentrations of [14C]-radiolabeled PEG-PHDCA, polysorbate 80 or poloxamine 908-coated PHDCA nanoparticles, and uncoated PHDCA nanoparticles were determined by radioactivity counting after intravenous administration in mice and rats. In addition, the integrity of the blood-brain barrier (BBB) after nanoparticles administration was evaluated by in vivo quantification of the diffusion of [14C]-sucrose into the brain. The location of fluorescent nanoparticles in the brain was also investigated by epi-fluorescent microscopy. RESULTS: Based on their long-circulating characteristics, PEGylated PHDCA nanoparticles penetrated into the brain to a larger extent than all the other tested formulations. Particles were localized in the ependymal cells of the choroid plexuses, in the epithelial cells of pia mater and ventricles, and to a lower extent in the capillary endothelial cells of BBB. These phenomena occurred without any modification of BBB permeability whereas polysorbate 80-coated nanoparticles owed, in part, their efficacy to BBB permeabilization induced by the surfactant. Poloxamine 908-coated nanoparticles failed to increase brain concentration probably because of their inability to interact with cells. CONCLUSIONS: This study proposes PEGylated poly (cyanoacrylate) nanoparticles as a new brain delivery system and highlights two requirements to design adequate delivery systems for such a purpose: a) long-circulating properties of the carrier, and b) appropriate surface characteristics to allow interactions with BBB endothelial cells.

Acrylic Resins↗

PubMed2000

Analysis of splicing of four mouse JNK/SAPKalpha variants.

The JNK/SAPK (c-Jun NH2-terminal kinase/stress-activated protein kinase) cascade is activated by a variety of stress stimuli and by the inflammatory cytokines interleukin-I (IL-I) and tumor necrosis factor alpha (TNFalpha). Four splice variants of the mouse JNK/SAPKalpha isoform, which differ in a region located in subdomains IX-X of the protein, were previously identified. Analysis of the sequence of the central region of the mouse JNK/SAPKalpha gene indicates that splice variants I and II are generated by a typical alternative splicing mechanism, while splice variants III and IV are generated by a less common mechanism, where alternative 3' splice sites located inside an exon (cryptic sites) are selected. The major splice variants alphaI and all have a wide and similar distribution in hippocampus, cerebral cortex, caudate-putamen, amygdala and the granule cell layer of cerebellum, although their expression is specifically regulated in certain cell types.

Alternative Splicing↗

PubMed1999

Effect of surgical stress on benzodiazepine receptors as a consequence of placebo pellet implantation in rat: an autoradiographic study.

This paper reports modifications in benzodiazepine (BZ) receptors induced by minimally invasive surgery, such as pellet implantation, a widely used surgical process for chronic drug administration. The intrinsic stress induced by this manipulation on BZ receptors was analysed in an autoradiographic saturation study determining the affinity (K(D)) and total number (Bmax) of binding sites of a number of brain areas from the mesencephalon, cerebral cortex, hippocampus, and cerebellum. The radioligand used for the study was [3H]flunitrazepam, which permitted overall characterization of BZ binding sites. Use of the specific BZ1 agonist zolpidem as an inhibitor of this radioligand permitted the direct characterization of subtype 2 (BZ2) and the indirect characterization of subtype 1 receptor (BZ1). Significant increases in Bmax and K(D) values were observed in pellet-implanted animals with respect to those not implanted. The results support the notion of an up-regulation of these receptors, mainly in BZ1 receptors, following sustained desensitization as a result of the surgical stress induced by pellet implantation.

Animals↗

PubMed1999

Enhancement of nasal absorption of insulin using chitosan nanoparticles.

PURPOSE: To investigate the potential of chitosan nanoparticles as a system for improving the systemic absorption of insulin following nasal instillation. METHODS: Insulin-loaded chitosan nanoparticles were prepared by ionotropic gelation of chitosan with tripolyphosphate anions. They were characterized for their size and zeta potential by photon correlation spectroscopy and laser Doppler anemometry, respectively. Insulin loading and release was determined by the microBCA protein assay. The ability of chitosan nanoparticles to enhance the nasal absorption of insulin was investigated in a conscious rabbit model by monitoring the plasma glucose levels. RESULTS: Chitosan nanoparticles had a size in the range of 300-400 nm, a positive surface charge and their insulin loading can be modulated reaching values up to 55% [insulin/nanoparticles (w/w): 55/100]. Insulin association was found to be highly mediated by an ionic interaction mechanism and its release in vitro occurred rapidly in sink conditions. Chitosan nanoparticles enhanced the nasal absorption of insulin to a greater extent than an aqueous solution of chitosan. The amount and molecular weight of chitosan did not have a significant effect on insulin response. CONCLUSIONS: Chitosan nanoparticles are efficient vehicles for the transport of insulin through the nasal mucosa.

Absorption↗

PubMed1998

Cloning of chicken and mouse alpha 1b adrenergic receptor.

A partial cDNA encoding most of the third intracellular loop of the chicken alpha 1b adrenergic receptor subtype, obtained by reverse transcription-polymerase chain reaction (RT-PCR) techniques using degenerate primers derived from mammalian sequences, was used to isolate an alpha 1b adrenergic receptor cDNA from brain. The cDNA encodes a potential protein of 507 amino acids and Northern hybridization of poly(A)+ RNA from chicken brain of different developmental stages detected a single 3.5 kb transcript. Analysis of receptor expression indicated that the alpha 1b adrenergic receptor is widely distributed in chicken tissues, specially kidney and liver. cDNA and genomic clones encoding sequences of the mouse alpha 1b adrenergic receptor were also isolated.

Amino Acid Sequence↗

PubMed1998

Chronic alcoholization in rats by free-choice ingestion of a hydroalcoholic solution.

The present work addresses a schedule designed to render rats alcoholic by offering them free access to a 20% (v/v) ethanol solution over 6 weeks. A solid diet and water (for controls) were fully available ad lib. in the animals' cages. The treatment was observed to achieve some effects suitable for the chronic alcoholization of rats: (1) Ethanol furnished the 36% of the calories of the diet of ethanol-treated rats; (2) The daily ethanol intake achieved was 12.05 +/- 1.18 g ethanol/kg weight; (3) The morning blood ethanol level was 0.45 g/litre; (4) Ethanol-treated rats deprived of ethanol showed several withdrawal symptoms. These results agree with those obtained by standard liquid diets, suggesting that the current protocol can be used to achieve a status of chronic alcoholization in rats.

Alcoholism↗

PubMed1998

Efficacy of topical cyclosporine-loaded nanocapsules on keratoplasty rejection in the rat.

PURPOSE: To evaluate the efficacy of a topical formulation of nanocapsules loaded with 1% cyclosporine A (CsA), which has been previously demonstrated to enhance CsA corneal penetration, compared to 1% CsA in migliol oil on a penetrating keratoplasty rejection model in the rat. METHODS: Lewis rats received orthotopic corneal allografts from inbred Fisher donors. Rats were treated with 10 microliters of the following topical solutions four times daily for 30 days, starting one day before surgery: Group 1 (n = 9), 1% CsA-loaded nanocapsules; group 2 (n = 13), 1% CsA dissolved in migliol oil; group 3 (n = 12), migliol oil; group 4 (n = 13), no treatment. Rejection index, mean survival time and rejection percentage were calculated for each group, and CsA levels in blood were measured. RESULTS: The rejection percentage was 84.6% for group 2, 91.7% for group 3, and 100% for groups 1 and 4, with no significant differences among groups. Mean graft survival time was 7.3 days for group 1, 15.5 days for group 2, 8.36 days for group 3, and 7.69 days for group 4, with significant differences between group 2 and the other groups. Systemic CsA levels were only detectable in group 2. CONCLUSIONS: CsA formulated in migliol oil delayed corneal rejection onset, but blood levels were evident in this group. CsA loaded-nanocapsules showed no effect on rejection and the drug was not detectable in blood. These data, along with the current concepts on corneal graft rejection immunology, suggest that the immunomodulatory effect of topical CsA in the prevention of corneal graft rejection may be systemically-mediated.

Administration, Topical↗

PubMed1997

Effects of chronic ethanol treatment and ethanol withdrawal on [3H]SCH23390 binding to rat striatal membranes.

The effects of chronic ethanol administration and ethanol withdrawal on the kinetic and pharmacological properties of [3H]SCH23390 binding sites and the labelling of central dopamine D-1 receptors were studied in the striatum of the rat. Chronic 40 day ethanol treatment produced a statistically significant decrease (p < 0.05) in maximum binding (Bmax) on striatal dopamine D-1 receptors of the rat, KD remaining unaltered. The withdrawal of ethanol did not affect the kinetic binding parameters. The rank order of potency in displacing the specific [3H]SCH23390 binding of several dopamine antagonists, agonists and serotonin-related drugs was consistent with the pharmacological profile of dopamine D-1 receptors. Chronic ethanol treatment led to a statistically significant increase in receptor affinity (lower Ki than controls) for (+)-butaclamol (p < 0.05). Ethanol withdrawal for 24 hr increased the affinity of [3H]SCH23390-labeled binding sites for norepinephrine. The addition of 0.03-0.68 M ethanol in vitro had no significant effects on [3H]SCH23390 binding in striatal preparations taken from both control and ethanol-treated rats. The results show that rat striatal [3H]SCH23390-labelled binding sites are affected by different conditions of ethanol exposure, possibly suggesting the medication of striatal dopamine pathways in the responses to ethanol intoxication.

Animals↗

PubMed1997

Effect of lysozyme on the stability of polyester nanocapsules and nanoparticles: stabilization approaches.

The efficacy of colloidal particles as drug carriers is closely related to their interaction with proteins and enzymes in different body fluids. In the present work, we analysed the interaction phenomenon between lysozyme (LZM), a positively charged enzyme that is highly concentrated in mucosas, and two different drug carriers: nanocapsules made of an oily core coated by the polymer poly-epsilon-caprolactone (PECL) and nanoparticles made solely of PECL. Results showed that the interaction of LZM with these colloidal drug carriers is highly affected by their surface charge. Nanocapsules, because of their important negative charge (-40 mV), adsorbed a great amount of LZM, which is positively charged. This adsorption process, which was also evidenced by the significant reduction of the nanocapsules' negative surface charge, led to the degradation of the polymer coating and the aggregation of the nanocapsules. In contrast, nanoparticles had a low negative surface charge (-8 mV) and adsorbed only a small amount of LZM, which did not cause the destabilization of the system. Furthermore, the molecular weight of the polymer forming the nanoparticles did not change. Finally, it was observed that the destabilizing effects caused by the adsorption of LZM onto the nanocapsules can be prevented by previous adsorption of the cationic poly(amino acid) poly-L-lysine. Using this approach the adsorption of LZM was hindered and its consequences avoided.

Adsorption↗

PubMed1997

Phosphorylation of the third intracellular loop of the mouse alpha1b-adrenergic receptor by cAMP-dependent protein kinase.

The third intracellular loop of adrenergic receptors has been implicated in their interaction with guanine nucleotide-binding proteins (G proteins). One of the mechanisms involved in the modulation of receptor function is the phosphorylation of specific residues by intracellular kinases. alpha1b-Adrenergic receptor is phosphorylated in vitro by cAMP-dependent protein kinase (PKA), although its physiological effect remains to be determined. We have produced fusion proteins formed by glutathione S-transferase and sequences of the third intracellular loop of mouse alpha1a-, alpha1b-, and alpha1d-adrenergic receptor subtypes, and used them as substrates for PKA. Only the fusion protein containing the alpha1b sequence was phosphorylated in vitro by this kinase. Site-directed mutagenesis of a serine (homologue to serine 278 of the rat sequence, RSS) to an alanine residue precluded phosphorylation by PKA.

Amino Acid Sequence↗

PubMed1997

Chitosan and chitosan/ethylene oxide-propylene oxide block copolymer nanoparticles as novel carriers for proteins and vaccines.

PURPOSE: The aim of this study was to investigate the interaction between the components of novel chitosan (CS) and CS/ethylene oxide-propylene oxide block copolymer (PEO-PPO) nanoparticles and to evaluate their potential for the association and controlled release of proteins and vaccines. METHODS: The presence of PEO-PPO on the surface of the nanoparticles and its interaction with the CS was identified by X-ray photoelectron spectroscopy (XPS). The mechanism of protein association was elucidated using several proteins, bovine serum albumin (BSA), and tetanus and diphtheria toxoids, and varying the formulation conditions (different pH values and concentrations of PEO-PPO), and the stage of protein incorporation into the nanoparticles formation medium. RESULTS: BSA and tetanus and diphtheria toxoids were highly associated with CS nanoparticles partly due to electrostatic interactions between the carboxyl groups of the protein and the amine groups of CS. PEO-PPO also interacted electrostatically with CS, thus competing with the proteins for association with CS nanoparticles. A visible amount of PEO-PPO was projected towards the outer phase of the nanoparticles. Proteins were released from the nanoparticles at an almost constant rate, the intensity of which was closely related to the protein loading. Furthermore, the tetanus vaccine was released in the active form for at least 15 days. CONCLUSIONS: CS and CS/PEO-PPO nanoparticles prepared by a very mild ionic crosslinking technique are novel and suitable systems for the entrapment and controlled release of proteins and vaccines.

Animals↗

PubMed1997

alpha-Tubulin of Histriculus cavicola (Ciliophora; Hypotrichea).

An alpha-tubulin gene fragment amplified by PCR from the hypotrichous ciliate Histriculus cavicola has been sequenced. This fragment, 1,182 bp long, contains an in-frame "stop" codon (UAA), which in other hypotrichous species codes for a glutamine residue. The comparison of the alpha-tubulin genes from several ciliates classes have revealed amino acid positions which could serve to distinguish these taxonomic groups.

Amino Acid Sequence↗