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F Núñez

Publications and source records attributed to F Núñez.

At least 19 recordsLinked to original sources
PubMed2007

Transcriptomal profiling of the cellular transformation induced by Rho subfamily GTPases.

We have used microarray technology to identify the transcriptional targets of Rho subfamily guanosine 5'-triphosphate (GTP)ases in NIH3T3 cells. This analysis indicated that murine fibroblasts transformed by these proteins show similar transcriptomal profiles. Functional annotation of the regulated genes indicate that Rho subfamily GTPases target a wide spectrum of functions, although loci encoding proteins linked to proliferation and DNA synthesis/transcription are upregulated preferentially. Rho proteins promote four main networks of interacting proteins nucleated around E2F, c-Jun, c-Myc and p53. Of those, E2F, c-Jun and c-Myc are essential for the maintenance of cell transformation. Inhibition of Rock, one of the main Rho GTPase targets, leads to small changes in the transcriptome of Rho-transformed cells. Rock inhibition decreases c-myc gene expression without affecting the E2F and c-Jun pathways. Loss-of-function studies demonstrate that c-Myc is important for the blockage of cell-contact inhibition rather than for promoting the proliferation of Rho-transformed cells. However, c-Myc overexpression does not bypass the inhibition of cell transformation induced by Rock blockage, indicating that c-Myc is essential, but not sufficient, for Rock-dependent transformation. These results reveal the complexity of the genetic program orchestrated by the Rho subfamily and pinpoint protein networks that mediate different aspects of the malignant phenotype of Rho-transformed cells.

Amino Acid Substitution↗
PubMed2004

Characterization of molds from dry-cured meat products and their metabolites by micellar electrokinetic capillary electrophoresis and random amplified polymorphic DNA PCR.

Molds are common contaminants of dry-cured meat products in which mycotoxins could be synthesized if stored under favorable conditions. Thus, efficient and accurate characterization of the toxigenic molds from dry-cured meat products is necessary. A micellar electrokinetic capillary chromatography (MECC) method was tested to analyze secondary metabolites produced by 20 mold strains commonly found in dry-cured meat products. In addition, their random amplified polymorphic DNA (RAPD) genotypes were determined by using a PCR method. Although peak profiles of the secondary metabolites differed among mold strains of different species, they were similar in the same species. MECC analysis showed that 10 of the 20 molds tested produced mycotoxins, including patulin, penicillic acid, cyclopiazonic acid, mycophenolic acid, aflatoxin B1, sterigmatocystin, and griseofulvin. The RAPD analysis yielded a different pattern for each of the mold species tested. However, strains of the same species showed similar RAPD profiles. A high correlation between RAPD analysis and MECC was observed, since strains of the same species that showed similar RAPD patterns had similar profiles of secondary metabolites. RAPD patterns with primer GO2 and MECC profiles, either singly or combined, could be of great interest to distinguish toxigenic from nontoxigenic molds in dry-cured meat products.

Chromatography, Micellar Electrokinetic Capillary↗
PubMed2001

Evaluation of microbial proteolysis in meat products by capillary electrophoresis.

AIMS: The available methods for evaluating proteolysis in meat products, particularly the contribution of micro-organisms, are expensive, time-consuming and require an unacceptable sample size. To minimize these problems, two capillary electrophoresis-based methods have been developed. METHODS AND RESULTS: Six Gram-positive, catalase-positive cocci, four moulds and three yeasts, isolated from dry-cured ham, were tested on sterile pork slices. Using the Capillary Gel Electrophoresis (CGE) method, changes in sarcoplasmic and myofibrillar proteins due to endogenous and microbial enzymes were detected. The Capillary Zone Electrophoresis (CZE) analysis allowed evaluation of bulk changes by micro-organisms in soluble nitrogen compounds. CONCLUSION: CGE analysis of myofibrillar proteins and CZE determination of soluble nitrogen compounds have proved to be valuable tools for evaluating proteolytic activity of endogenous and microbial origin. SIGNIFICANCE AND IMPACT OF THE STUDY: The CGE and CZE methods developed can be used for a rapid and sensitive analysis of proteolysis in meat products.

Animals↗
PubMed2000

Nucleotide excision repair gene (ERCC1) deficiency causes G(2) arrest in hepatocytes and a reduction in liver binucleation: the role of p53 and p21.

A wide range of DNA lesions, both UV and chemically induced, are dealt with by the nucleotide excision repair (NER) pathway. Defects in NER result in human syndromes such as xeroderma pigmentosum (XP), where there is a 1000-fold increased incidence of skin cancer. The ERCC1 protein is essential for NER, but ERCC1 knockout mice are not a model for XP. In the absence of exogenous DNA-damaging agents, these mice are runted and die before weaning, with dramatically accelerated liver polyploidy and elevated levels of p53. Here we present a morphological, immunological, and molecular study to understand the mechanism for the unusual liver pathology in ERCC1-deficient mice. We show that the enlarged ERCC1-deficient hepatocytes are arrested in G(2) and that DNA replication and the normal process of binucleation are both reduced. This is associated with a p53-independent increase in expression of the cyclin-dependent kinase inhibitor p21. The most dramatic feature of the ERCC1-deficient liver phenotype, the accelerated polyploidy, is not rescued by p53 deficiency, but we show that p53 is responsible for the reduced DNA replication and binucleation. We consider that the liver phenotype is a response to unrepaired endogenous DNA damage, which may reflect an additional non-NER-related function for the ERCC1 protein.

Animals↗
PubMed2000

Effects of substrate, water activity, and temperature on growth and verrucosidin production by Penicillium polonicum isolated from dry-cured ham.

Penicillium polonicum, a common mold on dry-cured meat products, is able to produce verrucosidin, a potent neurotoxin. The ability of P. polonicum isolated from dry-cured ham to grow and produce verrucosidin from 4 to 40 degrees C at water activities (a(w)) of 0.99, 0.97, and 0.95 on malt extract agar (MEA) and a medium made up with meat extract, peptone, and agar (MPA) was evaluated. Verrucosidin was quantified by high-pressure liquid chromatography and mass spectrometry. P. polonicum was able to grow on MEA and MPA at all the a(w) values tested from 4 to 37 degrees C but not at 40 degrees C. The optimal environmental conditions for growth were 20 degrees C, 0.99 a(w) on MEA and 20 to 25 degrees C, 0.97 a(w) on MPA, but the highest amount of verrucosidin was obtained at 25 degrees C, 0.99 a(w) in both media. No direct correlation between extension of mold growth and verrucosidin production was found. Temperature appears to be the most important factor ruling mycelial growth, whereas verrucosidin accumulation is mostly influenced by a(w). However, analysis of variance of the data showed that there was a complex interaction among all the environmental factors (medium, temperature, and a(w)) that significantly (P < 0.0001) affected growth and verrucosidin production. The reduction of a(w) to intermediates values of 0.95 has a stronger effect on growth on MEA than on MPA. Given that the meat-based medium proved to be an appropriate substrate for the biosynthesis of verrucosidin by P. polonicum, the ability of this mold to produce the toxin on meat products should be established.

Animals↗
PubMed1998

Evaluation of proteolytic activity of micro-organisms isolated from dry cured ham.

In order to determine the possible contribution of micro-organisms to the ripening of meat products, 48 cocci, 18 moulds and 20 yeasts isolated from dry-cured Iberian ham were evaluated for proteolytic activity. Two specific methods were used: the ability to hydrolyse myosin in broth and, for those strains showing high activities, hydrolysis on both myofibrillar and sarcoplasmic proteins on pork slices. Moulds and cocci showed the highest proteolytic activity for myosin in broth. Both myofibrillar and sarcoplasmic proteins were recovered at lower rates from inoculated than from sterile incubated pork. The deepest changes in myofibrillar and sarcoplasmic proteins were originated by one strain each of Penicillium chrysogenum and Staphylococcus xylosus, respectively. Only small changes were observed in the concentrations of free amino acids from inoculated pork slices, except for the samples with P. chrysogenum, where there were increases in all free amino acids. Thus, P. chrysogenum makes a significant contribution to proteolysis during the ripening of dry-cured meat products.

Animals↗
PubMed1998

Cells from ERCC1-deficient mice show increased genome instability and a reduced frequency of S-phase-dependent illegitimate chromosome exchange but a normal frequency of homologous recombination.

The ERCC1 protein is essential for nucleotide excision repair in mammalian cells and is also believed to be involved in mitotic recombination. ERCC1-deficient mice, with their extreme runting and polyploid hepatocyte nuclei, have a phenotype that is more reminiscent of a cell cycle arrest/premature ageing disorder than the classic DNA repair deficiency disease, xeroderma pigmentosum. To understand the role of ERCC1 and the link between ERCC1-deficiency and cell cycle arrest, we have studied primary and immortalised embryonic fibroblast cultures from ERCC1-deficient mice and a Chinese hamster ovary ERCC1 mutant cell line. Mutant cells from both species showed the expected nucleotide excision repair deficiency, but the mouse mutant was only moderately sensitive to mitomycin C, indicating that ERCC1 is not essential for the recombination-mediated repair of interstrand cross links in the mouse. Mutant cells from both species had a high mutation frequency and the level of genomic instability was elevated in ERCC1-deficient mouse cells, both in vivo and in vitro. There was no evidence for an homologous recombination deficit in ERCC1 mutant cells from either species. However, the frequency of S-phase-dependent illegitimate chromatid exchange, induced by ultra violet light, was dramatically reduced in both mutants. In rodent cells the G1 arrest induced by ultra violet light is less extensive than in human cells, with the result that replication proceeds on an incompletely repaired template. Illegitimate recombination, resulting in a high frequency of chromatid exchange, is a response adopted by rodent cells to prevent the accumulation of DNA double strand breaks adjacent to unrepaired lesion sites on replicating DNA and allow replication to proceed. Our results indicate an additional role for ERCC1 in this process and we propose the following model to explain the growth arrest and early senescence seen in ERCC1-deficient mice. In the absence of ERCC1, spontaneously occurring DNA lesions accumulate and the failure of the illegitimate recombination process leads to the accumulation of double strand breaks following replication. This triggers the p53 response and the G2 cell cycle arrest, mediated by increased expression of the cyclin-dependent kinase inhibitor p21(cip1/waf1). The increased levels of unrepaired lesions and double strand breaks lead to an increased mutation frequency and genome instability.

Animals↗
PubMed1998

[Efficacy of two gene species in the serological diagnosis of Borrelia burgdorferi infections].

BACKGROUND: There is phenotypic and genetic variability among the species Borrelia burgdorferi that produces Lyme disease. Three gene species and seven serotypes have been defined. AIM: To study the efficacy of two gene species in the serological diagnosis of Borrelia burgdorferi infections in Granada, Spain. MATERIAL AND METHODS: One thousand sixty nine sera coming from 1,251 subjects without Lyme borreliosis were analyzed. These subjects were studied for health or pregnancy controls, differential diagnosis of viral disease, diagnosis of syphilis, neurological or rheumatic diseases. In all samples, antibodies against Borrelia burgdorferi (B31 and Pko strains) and against Treponema pallidum were investigated. Screening tests (ELISA and hemagglutination) were followed by confirmations tests for positive samples (Western Blot IgG strain B31 and FTA-abs respectively). A clinical and laboratory follow up was done for subjects with positive serological tests. RESULTS: The global rate of positive antibodies against Borrelia burgdorferi B31 was 8.31% and against the strain Pko was 0.64%. Western blot was negative in 36% of subjects with positive ELISA B31. The distribution of antibodies against the strain B31 was acute herpes virus infection in 16%, gestation in 3%, HIV infection in 6.4%, T pallidum infection in 36%, rheumatic diseases in 25%, neurological diseases in 17.5% and health controls in 7.4%. The percentage of positive Western Blot analyzes were 0.8, 2.1 and 0.4% respectively. A reversion of positive ELISA tests was observed in 6 subjects. CONCLUSIONS: The disparity in rates of antibodies against Borrelia burgdorferi in different geographic regions may be due to differences in the serological tests used. The high rate of false positive ELISA tests underscores the need to use other serological tests.

Antibodies, Bacterial↗
PubMed1998

[Laboratory diagnosis of Lyme borreliosis].

Some direct methods that can be used for the diagnosis of Lyme borreliosis are the culture, direct visualization or the detection of microbial DNA using polymerase chain reactions, in body tissues or fluids. Unfortunately, all these methods have a low sensitivity. There is a wide assortment of tests and antigens for indirect diagnosis, and the most recommended are ELISA tests and Western blot. The main inconvenient of these tests are the existence of shared serologic reactions, the variability of immune response and the difficult interpretation of results. Therefore, we propose the following guidelines for the diagnosis of Lyme borreliosis: For sero-epidemiological studies and to diagnose infection, antibodies should be determined in subjects with a compatible clinical picture, using an ELISA test that must be positive in at least two separate samples. All positive ELISA results should be confirmed with Western blot analysis, that must be interpreted using established criteria. Polymerase chain reactions should be used when they are available.

Borrelia burgdorferi Group↗
PubMed1996

Detection of p53 protein in oropharyngeal carcinoma. Prognostic implications.

OBJECTIVE: To demonstrate how the detection of p53 protein in formaldehyde-fixed, paraffin-embedded oropharyngeal carcinoma may be used as a factor in estimating prognosis. SETTING: University medical centers. DESIGN: Validation cohort. Formaldehyde-fixed, paraffin-embedded squamous cell carcinoma of the oropharynx tissues from 106 patients who underwent surgical therapy between 1975 and 1988 were immunostained by using M-7001 antibody (IgG class). RESULTS: Overexpression of p53 was observed in 46 tumors (43.4%). The detection of nuclear p53 was significantly associated with an increased risk of recurrence of oropharyngeal carcinoma (P = .05). Similar results were obtained when the presence or absence of p53 in the nuclei of the tumor cells was studied in relation to overall survival (P < .001). In a multivariate analysis stratified according to grade, pathological stage, and lymph node status, nuclear p53 status was an independent predictor of overall survival (P < .001). CONCLUSIONS: In patients with squamous cell carcinoma of the oropharynx, an accumulation of p53 in the tumor cell nuclei detected by immunohistochemical methods predicts a significantly increased risk of death, independent of tumor grade, stage, and lymph node status. The p53 overexpression appears to be a useful prognostic factor.

Adult↗
PubMed1996

Yeast population during ripening of dry-cured Iberian ham.

The relationship between the superficial yeast population and the ripening conditions of Iberian dry cured hams has been studied for three different locations. Tentative identifications were carried out for 836 isolates. Candida zeylanoides was the dominating yeast in early stages, whereas more than 99% of isolates from the surface of matured hams were identified as Debaryomyces hansenii. A great diversity of strains of C. zeylanoides and D. hansenii was found. The characteristic pattern of isolates from the various locations and the selection of various strains of D. hansenii during ripening make the study of the yeasts useful for estimating the progress of maturation.

Animals↗
PubMed1996

Composition and toxigenic potential of the mould population on dry-cured Iberian ham.

The fungal population on dry-cured Iberian ham can be essential to the development of the product's unique characteristics, but health hazards due to mycotoxins may be significant. We examined the natural fungal population of Iberian hams during ripening at three different locations. Chloroform extracts from 59 selected isolates were tested for toxicity to brine shrimp larvae and VERO cells, for mutagenicity in the Ames test and for antimicrobial activity against Staphylococcus aureus. The diversity of moulds increased during ripening. Penicillium commune, Penicillium chrysogenum, Penicillium aurantiogriseum, Penicillium expansum and Penicillium echinulatum dominated most of the ripening time; however, the Eurotium species, particularly E. herbariorum and E. repens, increased in the final product. Using the above tests, most moulds were toxigenic. The toxigenic potential of the fungal population increased as the processing progressed. To minimize health hazards from uncontrolled fungal populations, we identified non toxigenic strains of Penicillium chrysogenum that could be used as starters in dry-cured hams.

Animals↗
PubMed1996

Gram-positive, catalase-positive cocci from dry cured Iberian ham and their enterotoxigenic potential.

Iberian ham is an uncooked, cured meat product ripened under natural uncontrolled conditions for 18 to 24 months. Gram-positive, catalase-positive cocci are the main microbial population in Iberian ham for most of the ripening time. Since some of these organisms are able to produce enterotoxins, adequate characterization and toxicological study are needed. For this, 1,327 gram-positive, catalase-positive cocci, isolated from Iberian hams at different stages and locations, were characterized by physiological and biochemical tests. Selected isolates were further characterized by guanine-cytosine (G+C) content and restriction enzyme analysis of genes coding for 16S rRNA. The toxigenic potential of these organisms was tested with specific DNA gene probes for staphylococcal enterotoxins A, B, C, and D and confirmed by semiquantitative sandwich enzyme immunoassay. The majority of the isolates were identified as Staphylococcus spp. and Micrococcus spp. Non-identified gram-positive, catalase-positive cocci which were moderately halophilic and showed a 42 to 52% G+C content were detected. A great variety of staphylococcal strains were found within the different species at any sampling time. Two strains of Staphylococcus xylosus, one Staphylococcus cohnii strain, and four of the non-identified organisms with 42 to 52% G+C contents hybridized with some of the DNA probes for C and D staphylococcal enterotoxin genes. S. xylosus hybridizing with C-enterotoxin probe reacted with both C and D enterotoxins in the immunological test. In addition, enterotoxin D was confirmed in the nonidentified strains. Some toxigenic organisms were isolated from the final product, posing a health hazard for the consumer.

Animals↗
PubMed1994

Characterization of Staphylococcus spp. and Micrococcus spp. isolated from Iberian ham throughout the ripening process.

The Iberian dry cured ham is an uncooked meat product highly appreciated because of its characteristic flavour. This product is obtained from highly marbled Iberian pig hindlegs after 18-24 months of maturation under natural environmental conditions. The role of Micrococcaceae in the development of the aroma characteristics of this products remains unclear. Identification of Gram-positive, catalase-positive cocci isolated from Mannitol Salt Agar plates showed that Staphylococcus xylosus followed by Staphylococcus equorum are the predominant organisms, even after 16 months of maturing. A remarkable variety of types of both staphylococci and micrococci are detected at any sampling time. The metabolic activities of these organisms could contribute to the characteristics of the final product.

Animals↗
PubMed1994

[Laparoscopy assisted vaginal hysterectomy at the Dr. Sótero del Río Hospital].

Twenty three cases of vaginal hysterectomy performed between june 1993 and june 1994 with the aid of laparoscopy were analyzed in Dr. Sótero del Río Hospital. Average ages, parity, date of previous surgeries and associated pathology do not differ from the general population of the patients of the hospital. The results obtained in our series show that the procedure presents comparative advantages if compared with the classical abdominal hysterectomy by shortening the intrahospitalary stay, improving the postsurgical pain and the ventilating mechanic and by contributing to a faster return to work. The complication observed are normal for this type of surgery, however, a proper previous training in laparoscopic surgery and vaginal surgery is essential to limit these complications to a minimum.

Adult↗
PubMed1992

Analysis of ras oncogene mutations in human squamous cell carcinoma of the head and neck.

The presence of proto-oncogene mutations at codons 12, 13 and 61 of the Ha-, Ki-, and N-ras in primary head and neck squamous cell carcinoma are analysed in this study. Oncogene ras-specific sequences were amplified by the polymerase chain reaction and probed with mutation specific oligonucleotide probes. Mutations were detected in 8 of 22 samples (36.3%). No mutations were detected on patients' peripheral blood DNA. We found that histologically and clinically, squamous cell carcinomas with or without a ras mutation do not differ significantly from each other.

Carcinoma, Squamous Cell↗