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Biomedical subjects

C Perez

Publications and source records attributed to C Perez.

At least 19 recordsLinked to original sources
PubMed2006

Intron retention as an alternative splice variant of the rat urocortin 1 gene.

Urocortin 1, highly conserved metazoan gene of the corticotropin-releasing hormone family, is a simple gene structured in two exons and the corresponding intron. The urocortin 1 prepropeptide is entirely coded in the second exon. Preliminary non-isotopic in situ hybridization experiments with an oligonucleotide complementary to an intron sequence of the urocortin 1 gene showed a significant cytoplasmic-like staining, suggesting the occurrence of an intron-retained urocortin 1 transcript. This observation prompted us to study whether the urocortin 1 gene presents alternative splicing by intron retention event. Confocal fluorescent in situ hybridization for urocortin 1 RNA and the use of the specific DNA dye TOPRO-3 allowed us to show significant expression of the intron-retained urocortin 1 transcript that did not colocalize with TOPRO-3 staining indicating a cytoplasmic localization for the intron-retained urocortin 1 transcript. The natural occurrence of a polyadenylated intron-retained urocortin 1 RNA was further documented by reverse transcriptase polymerase chain reaction (PCR), primed with oligo(dT), of total RNA extracted from three brain regions, a midbrain region containing the Edinger-Westphal nucleus, cerebellum and prefrontal cortex. In the three brain regions studied, it was possible to amplify both intron-less as well as intron-retained urocortin 1 transcripts. The use of PCR primers that simultaneously amplify both urocortin 1 transcripts allowed us to show that the expression of both urocortin 1 transcripts differs among the brain regions analyzed, suggesting a tissue specific regulation of this alternative splicing. In silico analysis of the five known mammalian urocortin 1 genomic sequences showed high conservation of the urocortin 1 intron sequence. Further studies should investigate the regulation of this intron retention event and its consequence for the functionality of the urocortin 1 gene.

Alternative Splicing↗
PubMed2006

Apolipoprotein D expression in substantia nigra of Parkinson disease.

Apolipoprotein D (apo D), a lipocalin transporter of small hydrophobic molecules could play an important role in several neurodegenerative diseases. However, its role in those diseases remains unclear. Increments of apo D have been reported in relation with injury and degeneration in the nervous system. Recently increases of apo D level have been reported in schizophrenia, a neuropathologic disease where the oxidative stress and lipid abnormalities may be involved. Apo D could act as a sequestering molecule binding excess of arachidonic acid in cells. In order to determine the relationship between apo D expression and other neurodegenerative pathologies related to oxidative damage, we studied the presence of apo D in the substantia nigra of control and Parkinson disease (PD) subjects. We found dopaminergic neurons were not immunoreactive for apo D, control or PD subjects. However, surrounding glial cells showed immunostaining for apo D and signal increases in PD cases. These findings support the role of apolipoprotein D in neuroprotection and the importance of glia in the amount of this protein in the central nervous system.

Aged↗
PubMed2006

Historical and current perspectives of neuroactive compounds derived from Latin America.

Plants and invertebrates in Latin America have contributed to a great extent in the use, discovery and development of novel neuroactive tools. Significantly, these neuroactive drugs have proven to be particularly important for our current understanding of the physiology and pharmacology of the nervous system. In addition, these discoveries have helped to build the modern and successful pharmacological business that we know today. For example, curare helped to introduce the use of muscle relaxing agents into modern surgical techniques. The discovery of cocaine from the leaves of Peruvian coca plants was instrumental in the discovery of local anesthetics. The search and discovery for useful neuroactive compounds derived from Latin America has also been ongoing in other areas and new applications for quinine, capsaicin and epibatidine were recently described. Besides these organic compounds, several peptides produced by spiders and other invertebrates to hunt their prey also induce effects in channels and membrane receptors at very low concentrations, indicating their high potency and selectivity. It is likely that new pharmaceutics will be developed from these molecules. The interest to renew the search for new compounds is timely, since largely unexplored lands, such as the Amazon and Patagonia, hold an important number of plants and animals that contain exciting new active compounds. With the introduction of new techniques to isolate, identify and characterize the molecular targets and actions of chemical entities, together with the need for more potent and selective compounds to treat neurological conditions, it is necessary to broaden the current exploratory effort in order to find more beneficial uses.

Animals↗
PubMed2006

Directly observed treatment for tuberculosis in pharmacies compared with self-administered therapy in Spain.

OBJECTIVES: To compare directly observed treatment (DOT) of tuberculosis through pharmacy offices with self-administered treatment (SAT) in patients at risk for non-adherence. METHODS: Prospective study for DOT (1999-2002) and retrospective study for SAT (1996-1998) in patients at risk for non-adherence (human immunodeficiency virus [HIV] infection, alcoholism, illicit drug use, immigrant or homeless status and/or previous failure to complete). Patients in the DOT programme received medication as out-patients twice a week in pharmacies that supervised adherence and provided socio-sanitary support to patients. RESULTS: There were 101 and 112 patients in the DOT and SAT groups, respectively. Demographic and clinical characteristics were similar in both groups. Differences were observed in risk factors for non-adherence (more immigrants and fewer intravenous drug users in the DOT vs. the SAT groups; P < 0.05). In the DOT group, 76 patients (75.2%) completed treatment and were cured compared to only 30 patients (26.7%) in the SAT group (P < 0.001). Implementation of DOT increased the cost of treatment by 400 Euro per patient compared to SAT. CONCLUSION: In patients at risk for non-adherence, DOT implemented through pharmacy offices was better than SAT; however, completion rates were still low.

Adolescent↗
PubMed2005

The nucleotide receptor P2Y13 is a key regulator of hepatic high-density lipoprotein (HDL) endocytosis.

Cell surface receptors for high-density lipoprotein (HDL) on hepatocytes are major partners in the regulation of cholesterol homeostasis. We recently identified a cell surface ATP synthase as a high-affinity receptor for HDL apolipoprotein A-I (apoA-I) on human hepatocytes. Stimulation of this ectopic ATP synthase by apoA-I triggered a low-affinity-receptor-dependent HDL endocytosis by a mechanism strictly related to the generation of ADP. This suggests that nucleotide G-protein-coupled receptors of the P2Y family are molecular components in this pathway. Only P2Y1 and P2Y13 are present on the membrane of hepatocytes. Using both a pharmacological approach and small interference RNA, we identified P2Y13 as the main partner in hepatic HDL endocytosis, in cultured cells as well as in situ in perfused mouse livers. We also found a new important action of the antithrombotic agent AR-C69931MX as a strong activator of P2Y13-mediated HDL endocytosis.

Adenosine Diphosphate↗
PubMed2004

Current-induced nanomagnet dynamics for magnetic fields perpendicular to the sample plane.

We present electrical measurements of high-frequency magnetic dynamics excited by spin-polarized currents in Co/Cu/Ni(80)Fe20 nanopillar devices, with a magnetic field applied perpendicular to the sample layers. As a function of current and magnetic field, the dynamical phase diagram contains several distinguishable precessional modes and also static magnetic states. Using detailed comparisons with numerical simulations, we provide rigorous tests of the theory of spin-transfer torques.

Journal Article↗
PubMed2004

Well diffusion for antifungal susceptibility testing.

INTRODUCTION: The increasing clinical and microbiologic resistance of Candida spp. isolates to several antifungal agents is becoming a serious problem. It is now reasonable to propose the use of antifungal susceptibility testing in Candida spp. isolates from patients who have failed conventional therapy, before the selection of an empirical therapy. METHODS: One hundred and fifty eight isolates of Candida spp. were evaluated simultaneously by broth microdilution (NCCLS standard) and well diffusion testing (WD), a diffusion method similar to disc diffusion. RESULTS: According to the Wilcoxon Signed Ranks test performed, there was no significant difference (p>0.05) between both methodologies for all antifungal agents tested (fluconazole, itraconazole, posaconazole, caspofungin and amphotericin B, with C. tropicalis, C. krusei, C. dubliniensis, C. guillermondii, C. parapsilosis, C. albicans and C. glabrata). A significant difference was observed when comparing well diffusion with NCCLS for fluconazole WD 80% (p=0.008) in C. glabrata, as well as WD 80% (p=0.002) and WD 50% (p=0.002) in C. albicans. CONCLUSIONS: The well diffusion test is simple, easy to reproduce, inexpensive, easy both to read and interpret, and has a good correlation to the reference NCCLS microdilution test and may represent an alternative method for antifungal drug susceptibility testing of Candida spp., mainly in laboratories with few resources.

Antifungal Agents↗
PubMed2004

Transfer and stable transgene expression of a mammalian artificial chromosome into bone marrow-derived human mesenchymal stem cells.

Mammalian artificial chromosomes (ACEs) transferred to autologous adult stem cells (SCs) provide a novel strategy for the ex vivo gene therapy of a variety of clinical indications. Unlike retroviral vectors, ACEs are stably maintained, autonomous, and nonintegrating. In this report we assessed the delivery efficiency of ACEs and evaluated the subsequent differentiation potential of ACE-transfected bone marrow-derived human mesenchymal stem cells (hMSCs). For this, an ACE carrying multiple copies of the red fluorescent protein (RFP) reporter gene was transferred under optimized conditions into hMSCs using standard cationic transfection reagents. RFP expression was detectable in 11% of the cells 4-5 days post-transfection. The RFP-expressing hMSCs were enriched by high-speed flow cytometry and maintained their potential to differentiate along adipogenic or osteogenic lineages. Fluorescent in situ hybridization and fluorescent microscopy demonstrated that the ACEs were stably maintained as single chromosomes and expressed the RFP transgenes in both differentiated cultures. These findings demonstrate the potential utility of ACEs for human adult SC ex vivo gene therapy.

Adipocytes↗
PubMed2004

Properties of sodium caseinate film-forming dispersions and films.

The flow properties of sodium caseinate (NaCAS) film-forming dispersions were studied as a function of protein and glycerol concentration. Apparent viscosity vs. concentration profiles showed a disruption at about 10% (wt/wt) NaCAS. This was interpreted in terms of protein-water and protein-protein interactions. The presence of glycerol in the NaCAS film-forming suspensions reduced the viscosity of aqueous NaCAS as a consequence of decreasing protein-protein and protein-solvent interactions through the hydrogen bonding formation between the glycerol, the polypeptide chain of NaCAS, and water molecules. For higher concentration of glycerol, viscosity of NaCAS dispersions increased as a result of increasing total solid content. The glass transition temperature for solid films of 10% (wt/wt) NaCAS decreased with the glycerol content. The glycerol-protein interactions reduced the protein-protein interaction and, thus, increased the intermolecular spacing, leading to higher protein chain mobility and water vapor permeability.

Caseins↗
PubMed2002

Induction of micronuclei in V79 cells after combined treatments with heterocyclic aromatic amines.

Heterocyclic aromatic amines (HAs) appear in foods rich in proteins when subjected to different cooking processes. These amines have been demonstrated to be mutagenic in bacteria; in eucaryotic cells, controversial results have been referred. The objective of this study is to evaluate the clastogenic and/or aneugenic capacity of three HAs--2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3-methylimidazo[4,5-f]quinoxaline (IQx), and 2-amino-3-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)--in isolated as well as in combined treatments. The micronucleus test in vitro was used on V79 cells in the presence and absence of metabolic activation. The duration of the treatment was 2 h, and cytochalasin B was added for 21 h to stop cytokinesis; then, micronuclei (MN) were counted in binucleated cells. In the presence of metabolic activation, the three amines showed a significant increase in the number of MN with respect to the negative control. The PhIP amine presented the highest values and it also resulted slightly active in the absence of metabolic activation, although these differences have not been considered to be significant. The combined treatments of these amines have shown that the effects attributed to them when administered together are those that are expected for a possible additive effect; the effect attributed to each HA separately is not potentiated nor inhibited.

Cell Line↗
PubMed2002

Retrofitting of a satellite repeat DNA-based murine artificial chromosome (ACes) to contain loxP recombination sites.

A satellite DNA-based mammalian artificial chromosome (ACes) was generated and subsequently modified by targeting of a loxP-red fluorescent protein (RFP) expression cassette via homologous recombination into a ribosomal DNA (rDNA)-containing locus. Clones containing correctly targeted ACes were identified by PCR from populations of RFP-expressing cells enriched by FACS sorting and were further characterized by fluorescent in situ hybridization. The targeted ACes maintained its ability to be purified to near homogeneity. Studies are currently underway to further characterize the functionality, carrying capacity, stability and transfectability of this modified ACes.

Animals↗
PubMed2002

Modulation of mutagenic activity in meat samples after deep-frying in vegetable oils.

Previous studies have been carried out on the influence of frying fats on the formation of food mutagens, but most of them have been performed on model systems or under cooking conditions that are more frequent in northern countries. The objective of this work was to study the overall mutagenic activity generated in hamburgers and frankfurters deep-fried under cooking conditions that are normal practice in Spain and other Mediterranean countries, in order to determine if there was any modulation of the mutagenic activity with respect to other cooking conditions previously studied. Hamburgers were prepared from beef purchased in a butcher's shop. Frankfurters as well as the oils [olive, marc olive ('orujo'), sunflower and soya bean oil] and butter were purchased in a local supermarket. The samples were fried in a teflon-coated frying pan at 170-180 degrees C for 10, 20 or 30 min. The mutagens were extracted and the mutagenic activity evaluated using the Salmonella mammalian microsome assay with strain TA98. Two independent assays were carried out for each experimental condition. All the hamburgers showed a mutagenic activity that was more than four times higher than that of the controls. Frankfurters showed a lower mutagenic activity than hamburgers (fried under the same conditions) because they have a lower protein content and a higher fat content. Hamburgers fried in olive oil for 10 min showed a significant increase in the number of revertants with respect to the other oils, probably due to the fact that the temperature reached was approximately 10 degrees C higher. Longer frying times significantly increased the number of revertants in samples fried in oils, except in olive oil, probably due to its lower content of polyunsaturated fatty acids.

Animals↗
PubMed2001

Poly(lactic acid)-poly(ethylene glycol) nanoparticles as new carriers for the delivery of plasmid DNA.

The purpose of the present work was to produce and characterize poly(lactic acid)-poly(ethylene glycol) (PLA-PEG) nanoparticles (size lower than 300 nm) containing a high loading of plasmid DNA in a free form or co-encapsulated with either poly(vinyl alcohol) (PVA) or poly(vinylpyrrolidone) (PVP). The plasmid alone or with PVA or PVP was encapsulated by two different techniques: an optimized w/o/w emulsion-solvent evaporation technique as well as by a new w/o emulsion-solvent diffusion technique. Particle size, zeta potential, plasmid DNA loading and in vitro release were determined for the three plasmid-loaded formulations. The influence of the initial plasmid loadings (5, 10, 20 microg plasmid DNA/mg PLA-PEG) on those parameters was also investigated. The plasmid loaded into the nanoparticles and released in vitro was quantified by fluorimetry and the different molecular forms were identified by gel electrophoresis. PLA-PEG nanoparticles containing plasmid DNA in a free form or co-encapsulated with PVA or PVP were obtained in the range size of 150-300 nm and with a negative zeta potential, both parameters being affected by the preparation technique. Encapsulation efficiencies were high irrespective of the presence of PVA or PVP (60-90%) and were slightly affected by the preparation technique and by the initial loading. The final plasmid DNA loading in the nanoparticles was up to 10-12 microg plasmid DNA/mg polymer. Plasmid DNA release kinetics varied depending on the plasmid incorporation technique: nanoparticles prepared by the w/o diffusion technique released their content rapidly whereas those obtained by the w/o/w showed an initial burst followed by a slow release for at least 28 days. No significant influence of the plasmid DNA loading and of the co-encapsulation of PVP or PVA on the in vitro release rate was observed. In all cases the conversion of the supercoiled form to the open circular and linear forms was detected. In conclusion, plasmid DNA can be very efficiently encapsulated, either in a free form or in combination with PVP and PVA, into PLA-PEG nanoparticles. Additionally, depending on the processing conditions, these nanoparticles release plasmid DNA either very rapidly or in a controlled manner.

Chemical Phenomena↗
PubMed2001

Bends in human mitotic metaphase chromosomes revisited: 15q11-13 is the most frequent non-random autosomal bend in blood cultures.

We have investigated the preferential bending of some chromosome sites in blood cultures from normal and chromosomally abnormal subjects. A total of 2,262 centromeric and 2,718 non-centromeric bends were recorded, and 69 non-centromeric sites were found not to bend at random. 15q11-13 bending was found to be the most frequent non-random autosomal bend. Bends on chromosomes may be remnants of a folded chromosome state in the nucleus, and may facilitate the preferential involvement of some chromosomal bands in structural reorganizations such as the isoacentric fragments, or contribute to the high frequency of interstitial deletions and isodicentric inversion duplications involving the 15q11-13 region.

Blood Cells↗
PubMed2001

A role of the C-terminal part of p44 in the promoter escape activity of transcription factor IIH.

The p44 subunit plays a crucial role in the overall activity of the transcription/DNA repair factor TFIIH: on the one hand its N-terminal domain interacts with and regulates the XPD helicase (, ); on the other hand, as shown in the present study, it participates with the promoter escape reaction. Mutagenesis along with recombinant technology using the baculovirus/insect cells expression system allowed us to define the function of the two structural motifs of the C-terminal moiety of p44: mutations within the C4 zinc finger motif (residues 291-308) prevent incorporation of the p62 subunit within the core TFIIH. Double mutations in the RING finger motif (residues 345-385) allow the synthesis of the first phosphodiester bond by RNA polymerase II, but prevent its escape from the promoter. This highlights the role of transcription factor IIH in the various steps of the transcription initiation process.

Animals↗
PubMed2001

Randomized phase III intergroup trial of isotretinoin to prevent second primary tumors in stage I non-small-cell lung cancer.

BACKGROUND: Promising data have suggested that retinoid chemoprevention may help to control second primary tumors (SPTs), recurrence, and mortality of stage I non-small-cell lung cancer (NSCLC) patients. METHODS: We carried out a National Cancer Institute (NCI) Intergroup phase III trial (NCI #I91-0001) with 1166 patients with pathologic stage I NSCLC (6 weeks to 3 years from definitive resection and no prior radiotherapy or chemotherapy). Patients were randomly assigned to receive a placebo or the retinoid isotretinoin (30 mg/day) for 3 years in a double-blind fashion. Patients were stratified at randomization by tumor stage, histology, and smoking status. The primary endpoint (time to SPT) and the secondary endpoints (times to recurrence and death) were analyzed by log-rank test and the Cox proportional hazards model. All statistical tests were two-sided. RESULTS: After a median follow-up of 3.5 years, there were no statistically significant differences between the placebo and isotretinoin arms with respect to the time to SPTs, recurrences, or mortality. The unadjusted hazard ratio (HR) of isotretinoin versus placebo was 1.08 (95% confidence interval [CI] = 0.78 to 1.49) for SPTs, 0.99 (95% CI = 0.76 to 1.29) for recurrence, and 1.07 (95% CI = 0.84 to 1.35) for mortality. Multivariate analyses showed that the rate of SPTs was not affected by any stratification factor. Rate of recurrence was affected by tumor stage (HR for T(2) versus T(1) = 1.77 [95% CI = 1.35 to 2.31]) and a treatment-by-smoking interaction (HR for treatment-by-current-versus-never-smoking status = 3.11 [95% CI = 1.00 to 9.71]). Mortality was affected by tumor stage (HR for T(2) versus T(1) = 1.39 [95% CI = 1.10 to 1.77]), histology (HR for squamous versus nonsquamous = 1.31 [95% CI = 1.03 to 1.68]), and a treatment-by-smoking interaction (HR for treatment-by-current-versus-never-smoking = 4.39 [95% CI = 1.11 to 17.29]). Mucocutaneous toxicity (P<.001) and noncompliance (40% versus 25% at 3 years) were higher in the isotretinoin arm than in the placebo arm. CONCLUSIONS: Isotretinoin treatment did not improve the overall rates of SPTs, recurrences, or mortality in stage I NSCLC. Secondary multivariate and subset analyses suggested that isotretinoin was harmful in current smokers and beneficial in never smokers.

Adult↗