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F Vidal

Publications and source records attributed to F Vidal.

At least 55 records · Page 3Linked to original sources
PubMed2001

Critical-point phase separation in laser ablation of conductors.

Laser ablation due to an ultrashort laser pulse on a massive aluminum target was investigated by means of a one-dimensional fluid code. Clear separation between the ablated matter and the unablated target is seen to occur through spinodal decomposition involving thermodynamic instabilities near the critical point of aluminum. The code also shows that the end of the ablation process is preceded by the ejection of droplets, which form about 15% of the total ejected mass.

Journal Article↗
PubMed2001

Evidence for the involvement of diacylglycerol kinase in the activation of hypoxia-inducible transcription factor 1 by low oxygen tension.

Hypoxia-inducible factor 1 (HIF-1) induces a gene expression program essential for the cellular adaptation to lowered oxygen environments. The intracellular mechanisms by which hypoxia induces HIF-1 remain poorly understood. Here we show that exposure of various cell types to hypoxia raises the intracellular level of phosphatidic acid primarily through the action of diacylglycerol kinase (DGK). Pharmacological inhibition of DGK activity through use of the specific DGK inhibitors and abrogated specifically HIF-1-dependent transcription analyzed with a HIF-1-responsive reporter plasmid. A more detailed analysis revealed that pharmacological inhibition of DGK activity prevented the hypoxia-dependent accumulation of the HIF-1alpha subunit and the subsequent HIF-1-DNA complex formation as well as hypoxia-induced activity of the HIF-1 transactivation domains localized to amino acids 530-582 and 775-826 of the HIF-1alpha subunit. Our results demonstrate for the first time that accumulation of phosphatidic acid through DGK underlines oxygen sensing and provide evidence for the involvement of this lipid kinase in the intracellular signaling that leads to HIF-1 activation.

Cell Nucleus↗
PubMed2001

Assessment of the proportion of transgene-bearing sperm by fluorescence in situ hybridization: a novel approach for the detection of germline mosaicism in transgenic male founders.

Genetic mosaicism is frequent among transgenic animals produced by pronuclear microinjection. A successful method for the screening of founder animals for germline mosaicism prior to mating would greatly reduce the costs associated with the propagation of the transgenic lines, and improve the efficiency of transgenic livestock production. With this aim, we have devised a simple method to detect integrated transgenes in individual spermatozoa using fluorescence in situ hybridization (FISH). The experiments reported here were undertaken to investigate the efficiency of this FISH-based approach to accurately evaluate the proportion of transgene-bearing sperm and to be applied for the detection of potential germline mosaics. Sperm samples from mice homozygous and hemizygous for a beta-lactoglobulin transgene were analyzed in a first set of experiments. A high hybridization efficiency was achieved, and the proportions of transgene-positive sperm cells in both homozygous (94.8-98.2%) and hemizygous (49.8-51.9%) animals were close to the expected frequencies (100 and 50%, respectively). To evaluate the sensitivity of the assay more directly, simulated mosaic samples with 5, 10, 15, 20 and 40% of transgene-bearing spermatozoa were then prepared and analyzed by FISH. Significant differences in the frequency of transgene-positive sperm were observed between all mosaic samples, indicating that even small deviations (5%) from the expected 50% transgene transmission rate in a founder animal could be reliably detected with our assay. Therefore, the method proposed represents a novel approach for the identification of germline mosaic founder males in livestock transgenic projects and a much more economic and faster alternative to breeding.

Animals↗
PubMed2001

Family of SRY/Sox proteins is involved in the regulation of the mouse Msh4 (MutS Homolog 4) gene expression.

The eukaryotic MutHLS-like system plays a crucial role in both mitosis and meiosis. Until now, a number of works have focused on the function of MutS and MutL homologs during mismatch DNA repair. Nevertheless, little is known about the role of these proteins during meiosis. MSH4 is a meiosis specific protein that is necessary for meiotic recombination in Saccharomyces cerevisiae. The human MSH4 protein is only found in testis and ovary. It is involved first in synapsis and second during recombination together with MLH1 (MutL Homolog 1). Here, we report the identification of the mouse Msh4 gene that is located on chromosome 3. We examined the expression of mMsh4 in testes of increasing developmental age and in elutriated germ cells. The pattern of expression during spermatogenesis is consistent with a role for MSH4 both during zygonema and pachynema. We demonstrated a promoter activity of the mMsh4 5'-flanking region by cell transfection experiments with a luciferase reporter gene. We found several SRY/Sox binding sites in this region and co-transfection experiments showed that SRY could down regulate mMsh4 promoter transcriptional activity. We propose that the regulation of mMsh4 expression could be one of the reasons for the persistence of SRY and/or SRY-related proteins in adult testis.

3T3 Cells↗
PubMed2001

Deficit in motor cortical activity for simultaneous bimanual responses.

Reaction time (RT) is known to be longer for simultaneous bimanual responses than for unimanual ones. This phenomenon is called "bilateral deficit". To identify the mechanisms subserving the bilateral deficit, brain electrical activity was examined, with a source derivation method, in 12 right-handed subjects, during the preparation and execution periods of a RT task. The responses were either unilateral or bilateral index finger flexion, performed either in a simple RT condition, with 20% catch trials, or in a choice RT condition. A deficit was observed in RT for the bilateral response for the right-index finger movement. In cerebral electrical activities, no evidence of a correlate of a bilateral deficit was found during the preparatory period. Conversely, during the execution period, an EEG correlate of the bilateral deficit was found. For the right hand, the activation of the sensorimotor area directly involved in the voluntary control was weaker for bilateral than for unilateral contralateral responses. The reasons for such a bilateral command weakness are discussed in the context of our RT task. First, the constraint of synchronisation included in the bilateral response might require an interhemispheric information transmission that resulted in a braking effect. Second, given that an ipsilateral inhibition is present in case of choice between the two hands of one particular unimanual response, and given that this ipsilateral inhibition is also present in case of simple unimanual trials, we hypothesise that a mutual transcallosal inhibitory effect also persists in the bilateral response.

Adult↗
PubMed2001

Numerical chromosome abnormalities in the spermatozoa of the fathers of children with trisomy 21 of paternal origin: generalised tendency to meiotic non-disjunction.

The purpose of this study was the evaluation of aneuploidy frequencies in the spermatozoa of two fathers (DP-4 and DP-5) who had children with Down syndrome (DS) of paternal origin and in whom a previous sperm analysis by fluoresence in situ hybridisation (FISH) had suggested a generalised tendency to meiotic non-disjunction. Sperm samples were simultaneously hybridised with FISH probes for chromosomes 4, 13 and 22. Disomy frequencies for each of the chromosomes and diploidy frequencies were compared with data obtained from nine control donors. Both DS fathers had a statistically significant increase in the frequency of disomy for chromosomes 13 and 22. DP-5 also had an increased frequency of diploid spermatozoa. Our data suggest that the two DS fathers have a generalised susceptibility to meiotic non-disjunction and that acrocentric chromosomes seem to be more sensitive to such disturbance in the meiotic process.

Child↗
PubMed2001

Acrocentric chromosome disomy is increased in spermatozoa from fathers of Turner syndrome patients.

The aim of the present study was to investigate whether there was an increase of aneuploidy in the sperm from fathers of Turner syndrome patients of paternal origin who, in a previous study, showed an elevated incidence of XY meiotic nondisjunction. Sperm disomy frequencies for chromosomes 4, 13, 18, 21 and 22 were assessed by fluorescence in situ hybridisation in four of these individuals. As a group, the Turner syndrome fathers showed a general increase in disomy frequencies for chromosomes 13, 21 and 22, with a statistically significant increase in disomy frequencies for chromosomes 13 and 22 in one of the fathers and for chromosome 21 in two of them. Data from a previous work carried out by us in two fathers of Down syndrome patients of paternal origin also revealed increased sperm disomy frequencies for chromosomes 13, 21 and 22. Pooled as one group, these six fathers of aneuploid offspring of paternal origin had a statistically significant increase in the frequency of nondisjunction for these chromosomes with respect to control individuals. Our findings indicate that there may be an association between fathering aneuploid offspring and increased frequencies of aneuploid spermatozoa. Such increases do not seem to be restricted to the chromosome pair responsible for the aneuploid offspring. Acrocentric chromosomes and other chromosome pairs that usually show only one chiasma during meiosis seem to be more susceptible to malsegregation.

Aneuploidy↗
PubMed2001

Cytogenetic analysis of caprine 2- to 4-cell embryos produced in vitro.

Prepubertal goat in vitro matured/in vitro fertilised oocytes produce only a small percentage of blastocysts. The present study examines the incidence of chromosomal anomalies in 2- to 4-cell embryos in vitro produced (IVP) from prepubertal oocytes fertilised with the semen of two males. Cumulus-oocyte complexes were obtained by slicing ovaries from slaughtered prepubertal goats. Oocytes were matured in TCM199 supplemented with 20% heat inactivated Donor Bovine Serum (DBS), 10 microg/ml FSH + 10 microg/ml LH + 1 microg/ml 17beta-oestradiol for 27 h at 38.5 degrees C in 5% CO2 in air. IVM oocytes were inseminated with the sperm from two males prepared using the swim-up and heparin-capacitation procedures. At 24 h postinsemination (hpi) the oocytes were transferred to 100 microl drops of SOF medium for a further 24 h. At 17 hpi a sample of oocytes was stained with lacmoid to evaluate the nuclear stage after fertilisation. The cleavage rate was determined at 24, 36 and 48 hpi and chromosome slides were prepared according to the gradual-fixation technique and stained with Leishman. A total of 1070 2- to 4-cell embryos from prepubertal goat oocytes were studied, but it was only possible to analyse 241 cytogenetically. Of these, 40% exhibited a normal diploid chromosome complement, 59% were haploid and 1% were triploid. There were significant differences between the two males in sperm oocyte penetration and oocyte cleavage but no differences were found in chromosomal anomalies. In conclusion, the low number of embryos karyotyped and the high number of haploid embryos found in this study suggested a high incidence of abnormal fertilised embryos and deficient cytoplasmic maturation of the oocyte which inhibits sperm head decondensation.

Animals↗
PubMed2001

Incidence of sperm chromosomal abnormalities in a risk population: relationship with sperm quality and ICSI outcome.

BACKGROUND: An increased incidence of chromosome abnormalities has been reported in sperm samples of many infertile men by fluorescence in-situ hybridization (FISH). METHODS: Sperm aneuploidy and diploidy rates for chromosomes 13, 18, 21, X and Y were evaluated in 63 patients with normal karyotypes using dual and triple-colour FISH techniques. Indications for sperm FISH analysis were: recurrent miscarriages of unknown aetiology (RM, n = 40), repeated implantation failure after intracytoplasmic sperm injection (ICSI) (IF, n = 19), previous Down's syndrome pregnancies (n = 3), and meiotic abnormalities (MA, n = 1). Nine healthy normozoospermic donors were also evaluated as a control group. RESULTS: A significant increase in the incidence of sex chromosome disomies was found in the RM, IF and MA groups. Oligoasthenoteratozoospermic patients (n = 21) showed significantly higher rates of diploidy and disomies for sex chromosomes and chromosomes 18 and 21 than normozoospermic patients (n = 14). Thirty-one patients with normal and seven with abnormal FISH results had undergone several ICSI treatments (108 and 23 cycles respectively). Couples with abnormal sperm FISH results showed decreased pregnancy and implantation rates and increased miscarriage rates. CONCLUSIONS: Patients with a clinical background of recurrent miscarriages of unknown aetiology or implantation failure after ICSI are at risk of showing sperm chromosomal abnormalities, the incidence of which is higher in oligoasthenoteratozoospermic patients.

Abortion, Habitual↗
PubMed2001

Meiotic behaviour of the sex chromosomes in three patients with sex chromosome anomalies (47,XXY, mosaic 46,XY/47,XXY and 47,XYY) assessed by fluorescence in-situ hybridization.

Meiotic studies using multicolour fluorescent in-situ hybridization (FISH) and chromosome painting were carried out in three patients with sex chromosome anomalies (47,XXY; 46,XY/47,XXY and 47,XYY). In the two patients with Klinefelter syndrome, although variable percentages of XXY cells (88.5 and 28.3%) could be found in the pre-meiotic stages, none of the abnormal cells entered meiosis, and all pachytenes were XY. However, the abnormal testicular environment of these patients probably resulted in meiotic I non-disjunction, and a certain proportion of post-reductional cells were XY (18.3 and 1.7%). The fact that none of the spermatozoa were XY also suggests the existence of an arrest at the secondary spermatocyte or the spermatid level. In the XYY patient, most (95.9%) premeiotic cells were XYY. The percentage of XYY pachytenes was 57.9%. The sex chromosomes were either in close proximity (XYY) or the X chromosome was separated from the two Ys (X + YY). A high proportion (42.1%) of post-reductional germ cells were XY. However, only 0.11% of spermatozoa were disomic for the sex chromosomes. In this case, the data suggest the existence of an arrest of the abnormal cells at the primary and the secondary spermatocyte or the spermatid level, giving rise to the continuous elimination of abnormal cells in the germ-cell line along spermatogenesis. The fact that the proportion of diploid spermatozoa was only increased in one of the three cases (XXY) is also suggestive of an arrest of the abnormal cell lines in these patients. The two apparently non-mosaic patients were, in fact, germ-cell mosaics. This suggests that the cytogenetic criteria used to define non-mosaic patients may be inadequate; thus, the risk of intracytoplasmic sperm injection in apparently non-mosaics may be lower than expected.

Adult↗
PubMed2001

Gene trap analysis of germ cell signaling to Sertoli cells: NGF-TrkA mediated induction of Fra1 and Fos by post-meiotic germ cells.

Analysis of complex signalisation networks involving distinct cell types is required to understand most developmental processes. Differentiation of male germ cells in adult mammals involves such a cross-talk between Sertoli cells, the somatic component which supports and controls germinal differentiation, and germ cells at their successive maturation stages. We developed a gene trapping strategy to identify genes, which, in Sertoli cells, are either up- or down-regulated by signals emitted by the germinal component. A library of approximately 2,000 clones was constituted from colonies independently selected from the Sertoli line 15P-1 by growth in drug-containing medium after random integration of a promoter-less (beta)geo transgene (neo(r)-lacZ fusion), which will be expressed as a fusion transcript from a 'trapped' cellular promoter, different in each clone. A first screen conducted on 700 events identified six clones in which beta-galactosidase activity was increased and one in which it was repressed upon addition of germ cells. The targeted loci were identified by cloning and sequencing the genomic region 5' of the insert. One of them was identified as the gene encoding Fra1, a component of the AP1 transcription regulatory complex. Accumulation of Fra1 mRNA was induced, both in 15P-1 and in freshly explanted Sertoli cells, by addition of either round spermatids or nerve growth factor (NGF). The effect of NGF was mediated by the TrkA receptor and the ERK1-ERK2 kinase kinase pathway. Fos and Fra1 transcription were induced within the first hour after addition of the neurotrophin, but, unlike what is observed after serum induction in the same cells, a second wave of transcription of Fra1, but not of Fos, started 16 hours later and peaked at higher levels at about 20 hours. These results suggest that AP1 activation may be an important relay in the Sertoli-germ cell cross-talk, and validate the gene trapping approach as a tool for the identification of target genes in cell culture systems.

Animals↗
PubMed2001

An electromyographic investigation of the effect of stimulus-response mapping on choice reaction time.

The activity of the agonist muscles was recorded during the performance of a two-choice visual reaction time (RT) task in which the compatibility of the stimulus-response mapping was manipulated. Correct trials were distinguished according to whether or not the activation of the agonist of the required response was preceded by an activation of the agonist of the nonrequired response. Double activation trials were more numerous for the incompatible than for the compatible mapping. Furthermore, these trials yielded longer RTs than the single muscular activation trials. These results suggest that initial activations of nonrequired responses are more frequently aborted and corrected when the mapping is incompatible than when it is compatible. This finding supports the dimensional overlap model of stimulus-response compatibility (S. Kornblum, T. Hasbroucq, & A. Osman, 1990).

Adolescent↗
PubMed2001

Rapid hemophilia A molecular diagnosis by a simple DNA sequencing procedure: identification of 14 novel mutations.

We here describe a simple, efficient DNA sequencing procedure for hemophilia A molecular diagnosis. In severe patients we first test for the presence of factor VIII gene intron 22 inversion using a recently described single-tube PCR method. In moderate, mild, or inversion-negative severe patients we systematically sequence the promoter, all exons and splice junctions of factor VIII gene. Specially designed primers allow amplification of 23 PCR products under the same salt conditions and thermocycling parameters. The whole sequencing procedure, from blood extraction to mutation identification, can be readily done within 42 h when using regular instruments or in just 14 h when using a high-throughput sequencer. Thus, this is a versatile and cost-effective strategy with little hands-on time requirements. Since its implementation we have identified mutations in 45/46 hemophilia A patients, 14 of which are novel. Once the genetic defect has been identified, accurate genetic counseling is then easily performed.

Amino Acid Substitution↗
PubMed2000

Comparison of immunologic restoration and virologic response in plasma, tonsillar tissue, and cerebrospinal fluid in HIV-1-infected patients treated with double versus triple antiretroviral therapy in very early stages: The Spanish EARTH-2 Study. Early Anti-Retroviral Therapy Study.

The objective of antiretroviral therapy is to obtain an almost complete and durable suppression of viral replication in all compartments to facilitate recovery of the immune system. We assessed the virologic effect in plasma, tonsillar tissue, and cerebrospinal fluid (CSF) in 94 HIV-1-infected patients with CD4 counts >500 x 106 cells per liter and viral load >5000 copies/ml randomly assigned to triple antiretroviral therapy (two nucleoside reverse transcriptase inhibitors (NRTIs) plus one protease inhibitor) versus double therapy (two NRTIs). We also analyzed the immunologic recovery in this cohort of patients. Lymphoid tissue and cerebrospinal fluid viral load, development of genotypic resistance, proliferative responses to HIV-1 specific antigens, and other immunophenotypic markers were analyzed. The proportion of patients who achieved a decrease in HIV RNA levels to <200 copies/ml was significantly greater in the triple therapy group than in the two drug groups (p =.0002 for each pair-wise difference). At week 52, tonsillar tissue HIV RNA from 5 patients treated with triple therapy was lower than the limit of detection, whereas the mean +/- standard error in patients with double therapy (n = 5) was 5.03 +/- 0.34 copies/mg/tissue. In all 10 patients, CSF viral load (VL) was <20 HIV-1 RNA copies/ml at week 52. CSF cell counts and protein levels tended to decrease after 52 weeks of antiretroviral therapy. After 1 year of therapy, 13 of 21 patients (62%) in the double-therapy groups (zidovudine plus lamivudine [n = 9] and stavudine plus lamivudine [n = 12]) had evidence of M184V mutation. None of the 10 samples of patients receiving triple therapy could be amplified because of low HIV RNA levels. The mean increase in CD4 cells at week 52 was greater in the stavudine and lamivudine and indinavir group than in the double-treatment arms (186 versus 67 and 102, respectively; p =.03). In patients treated with triple therapy, the increase in naive T cells (CD4 and CD8) was greater than in patients treated with double therapy. Markers of activation decreased further in patients treated with the regimen that included protease inhibitors. Proliferative responses to HIV-1 p24 antigen were never recovered after double or triple therapy. Our study suggests that even in very early stages of HIV-1 disease only therapy with two NRTIs and one protease inhibitor reduces plasma, lymphoid tissue, and CSF VL to undetectable levels. HIV-1-related immune system abnormalities improved but were still defective after 1 year of antiretroviral therapy.

Adult↗
PubMed2000

The additive factor method in rat information processing.

Sternberg introduced the additive factor method as a tool for discovering and studying in choice reaction time situations in human subjects. Underlying the method is the notion that if information processing consists of successive stages, then different factors affecting different stages must have additive effects on reaction-time measurements. The additive factor method has been extensively used in humans but few attempts have been made to examine whether or not additive reaction time patterns were obtainable in rats. In the present work, the subjects (13 Long-Evans rats) had to press down a lever and to respond to a visual (right or left) stimulus by releasing the lever and introducing the head into a lateral (right or left) operandum. The present findings show an additive pattern of effects of signal luminance and foreperiod duration on the mean reaction time which suggests that we have successfully manipulated two stages of rat information processing - the preprocessing stage and the motor adjustment stage, respectively. Accordingly, the present study sets the basis for the basis for investigating the effects of invasive manipulations (pharmacological and/or lesional) on information processing stages.

Journal Article↗
PubMed2000

Preimplantation genetic diagnosis.

Preimplantation genetic diagnosis (PGD) includes a variety of techniques that have been developed to detect the transmission to the offspring of genetic diseases or of chromosome abnormalities by couples at risk before a pregnancy is established, to avoid these couples the risk of recurrent abortions and/or of repeated terminations of pregnancy. Candidate couples are carriers of gene mutations or of structural chromosome rearrangements, or with recurrent spontaneous abortions of unknown origin. Diagnostic procedures include different modalities of gene amplification using the polymerase chain reaction (PCR) or of fluorescent in situ hybridization (FISH). Embryo biopsies are carried out at the 6-8 cell stage. Healthy embryos are transferred on day 4 or at the blastocyst stage. By now, several hundred healthy children have been born using PGD, and only one diagnostic error has been reported.

Abortion, Habitual↗