Biomedical subjects

J Ortín

Publications and source records attributed to J Ortín.

At least 19 recordsLinked to original sources

PubMed2006

Relevance of dynamic wetting in viscous fingering patterns.

We demonstrate that wetting effects at moving contact lines have a strong impact in viscous fingering patterns. Experiments in a rotating Hele-Shaw (HS) cell, dry or prewetted, show consistent morphological differences. When the wetting fluid invades a dry region, contact angle dynamics yield a kinetic contribution to the interface pressure drop that scales with capillary number as Ca(2/3) but is significantly larger than the Park-Homsy kinetic correction. Numerical results are in very good agreement with experiments and show that standard HS equations work best for prewetted cells.

Journal Article↗

PubMed2006

Fluctuations in Saffman-Taylor fingers with quenched disorder.

We make an experimental characterization of the effect that static disorder has on the shape of a normal Saffman-Taylor finger. We find that static noise induces a small amplitude and long wavelength instability on the sides of the finger. Fluctuations on the finger sides have a dominant wavelength, indicating that the system acts as a selective amplifier of static noise. The dominant wavelength does not seem to be very sensitive to the intensity of static noise present in the system. On the other hand, at a given flow rate, rms fluctuations of the finger width, decrease with decreasing intensity of static noise. This might explain why the sides of the fingers are flat for typical Saffman-Taylor experiments. Comparison with previous numerical studies of the effect that temporal noise has on the Saffman-Taylor finger, leads to conclude that the effect of temporal noise and static noise are similar. The behavior of fluctuations of the finger width found in our experiments, is qualitatively similar to one recently reported, in the sense that, the magnitude of the width fluctuations decays as a power law of the capillary number, at low flow rates, and increases with capillary number for larger flow rates.

Journal Article↗

PubMed2006

[The utility of interictal SPECT in temporal lobe epilepsy].

INTRODUCTION: Interictal brain single photon emission computed tomography (SPECT) is used in the presurgical evaluation of patients with complex partial epilepsy. The aim of the present study was to compare interictal SPECT, MRI and video-electroencephalography (EEG) for seizure focus localization in patients with temporal lobe epilepsy, one year after temporal lobectomy, in order to determine the utility of interictal brain SPECT. PATIENTS AND METHODS: Thirty four consecutive patients with refractory temporal epilepsy were evaluated with video-EEG, MRI and interictal SPECT for seizure focus localization before surgery. Seizure focus was confirmed with the clinical follow-up one year after temporal lobectomy in all patients. MRI and SPECT analysis was performed visually. RESULTS: 31/34 patients were seizure free one year after surgery and the remaining 3 patients remain with seizures occasionally. Video-EEG results coincided with postsurgical seizure focus localization in 31 (91%) patients. MRI localized seizure focus correctly in 30 (88%) patients and was normal in 3 cases. Interictal brain SPECT was normal in 10 patients and showed temporal hypoperfusion consistent with postsurgical seizure focus in 23 (68%) patients. In all patients with abnormalities in the interictal SPECT, seizure focus was identified with video-EEG or MRI. CONCLUSIONS: When MRI and video-EEG localize seizure focus in the same temporal lobe, interictal brain SPECT does not offer any additional information for surgical decision making.

Adolescent↗

PubMed2005

Anomalous roughening of viscous fluid fronts in spontaneous imbibition.

We report experiments on spontaneous imbibition of a viscous fluid by a model porous medium in the absence of gravity. The average position of the interface satisfies Washburn's law. Scaling of the interface fluctuations suggests a dynamic exponent z approximately 3, indicative of global dynamics driven by capillary forces. The complete set of exponents clearly shows that interfaces are not self-affine, exhibiting distinct local and global scaling, both for time (beta = 0.64 +/- 0.02, beta(*) = 0.33 +/- 0.03) and space (alpha = 1.94 +/- 0.20, alpha(loc) = 0.94 +/- 0.10). These values are compatible with an intrinsic anomalous scaling scenario.

Journal Article↗

PubMed2005

Measurements of the bulk and interfacial velocity profiles in oscillating Newtonian and Maxwellian fluids.

We present the dynamic velocity profiles of a Newtonian fluid (glycerol) and a viscoelastic Maxwell fluid (CPyCl-NaSal in water) driven by an oscillating pressure gradient in a vertical cylindrical pipe. The frequency range explored has been chosen to include the first three resonance peaks of the dynamic permeability of the viscoelastic-fluid--pipe system. Three different optical measurement techniques have been employed. Laser Doppler anemometry has been used to measure the magnitude of the velocity at the center of the liquid column. Particle image velocimetry and optical deflectometry are used to determine the velocity profiles at the bulk of the liquid column and at the liquid-air interface respectively. The velocity measurements in the bulk are in good agreement with the theoretical predictions of a linear theory. The results, however, show dramatic differences in the dynamic behavior of Newtonian and viscoelastic fluids, and demonstrate the importance of resonance phenomena in viscoelastic fluid flows, biofluids in particular, in confined geometries.

Journal Article↗

PubMed2004

Nonlinear Saffman-Taylor instability.

We show, both theoretically and experimentally, that the interface between two viscous fluids in a Hele-Shaw cell can be nonlinearly unstable before the Saffman-Taylor linear instability point is reached. We identify the family of exact elastica solutions [Nye et al., Eur. J. Phys. 5, 73 (1984)]] as the unstable branch of the corresponding subcritical bifurcation which ends up at a topological singularity defined by interface pinchoff. We devise an experimental procedure to prepare arbitrary initial conditions in a Hele-Shaw cell. This is used to test the proposed bifurcation scenario and quantitatively asses its practical relevance.

Journal Article↗

PubMed2003

Systematic weakly nonlinear analysis of radial viscous fingering.

We present a weakly nonlinear analysis of the interface dynamics in a radial Hele-Shaw cell driven by both injection and rotation. We extend the systematic expansion introduced in [E. Alvarez-Lacalle et al., Phys. Rev. E 64, 016302 (2001)] to the radial geometry, and compute explicitly the first nonlinear contributions. We also find the necessary and sufficient condition for the uniform convergence of the nonlinear expansion. Within this region of convergence, the analytical predictions at low orders are compared satisfactorily to exact solutions and numerical integration of the problem. This is particularly remarkable in configurations (with no counterpart in the channel geometry) for which the interplay between injection and rotation allows that condition to be satisfied at all times. In the case of the purely centrifugal forcing we demonstrate that nonlinear couplings make the interface more unstable for lower viscosity contrast between the fluids.

Journal Article↗

PubMed2002

Anomalous roughening of Hele-Shaw flows with quenched disorder.

The kinetic roughening of a stable oil-air interface moving in a Hele-Shaw cell that contains a quenched columnar disorder (tracks) has been studied. A capillary effect is responsible for the dynamic evolution of the resulting rough interface, which exhibits anomalous scaling. The three independent exponents needed to characterize the anomalous scaling are determined experimentally. The anomalous scaling is explained in terms of the initial acceleration and subsequent deceleration of the interface tips in the tracks coupled by mass conservation. A phenomenological model that reproduces the measured global and local exponents is introduced.

Journal Article↗

PubMed2001

Systematic weakly nonlinear analysis of interfacial instabilities in Hele-Shaw flows.

We develop a systematic method to derive all orders of mode couplings in a weakly nonlinear approach to the dynamics of the interface between two immiscible viscous fluids in a Hele-Shaw cell. The method is completely general: it applies to arbitrary geometry and driving. Here we apply it to the channel geometry driven by gravity and pressure. The finite radius of convergence of the mode-coupling expansion is found. Calculation up to third-order couplings is done, which is necessary to account for the time-dependent Saffman-Taylor finger solution and the case of zero viscosity contrast. The explicit results provide relevant analytical information about the role that the viscosity contrast and the surface tension play in the dynamics of the system. We finally check the quantitative validity of different orders of approximation and a resummation scheme against a physically relevant, exact time-dependent solution. The agreement between the low-order approximations and the exact solution is excellent within the radius of convergence, and is even reasonably good beyond this radius.

Journal Article↗

PubMed2001

Three-dimensional reconstruction of a recombinant influenza virus ribonucleoprotein particle.

A three-dimensional structural model of an influenza virus ribonucleoprotein particle reconstituted in vivo from recombinant proteins and a model genomic vRNA has been generated by electron microscopy. It shows a circular shape and contains nine nucleoprotein monomers, two of which are connected with the polymerase complex. The nucleoprotein monomers show a curvature that may be responsible for the formation of helical structures in the full-size viral ribonucleoproteins. The monomers show distinct contact boundaries at the two sides of the particle, suggesting that the genomic RNA may be located in association with the nucleoprotein at the base of the ribonucleoprotein complex. Sections of the three-dimensional model show a trilobular morphology in the polymerase complex that is consistent with the presence of its three subunits.

Image Processing, Computer-Assisted↗

PubMed2001

PA subunit from influenza virus polymerase complex interacts with a cellular protein with homology to a family of transcriptional activators.

The PA subunit of the influenza virus polymerase complex is a phosphoprotein that induces proteolytic degradation of coexpressed proteins. Point mutants with reduced proteolysis induction reconstitute viral ribonucleoproteins defective in replication but not in transcriptional activity. To look for cellular factors that could associate with PA protein, we have carried out a yeast two-hybrid screen. Using a human kidney cDNA library, we identified two different interacting clones. One of them was identified as the human homologue of a previously described cDNA clone from Gallus gallus called CLE. The human gene encodes a protein of 36 kDa (hCLE) and is expressed ubiquitously in all human organs tested. The interaction of PA and hCLE was also observed with purified proteins in vitro by using pull-down and pep-spot experiments. Mapping of the interaction showed that hCLE interacts with PA subunit at two regions (positions 493 to 512 and 557 to 574) in the PA protein sequence. Immunofluorescence studies showed that the hCLE protein localizes in both the nucleus and the cytosol, although with a predominantly cytosolic distribution. hCLE was found associated with active, highly purified virus ribonucleoproteins reconstituted in vivo from cloned cDNAs, suggesting that PA-hCLE interaction is functionally relevant. Searches in the databases showed that hCLE has 38% sequence homology to the central region of the yeast factor Cdc68, which modulates transcription by interaction with transactivators. Similar homologies were found with the other members of the Cdc68 homologue family of transcriptional activators, including the human FACT protein.

Amino Acid Sequence↗

PubMed2000

Ultrastructural and functional analyses of recombinant influenza virus ribonucleoproteins suggest dimerization of nucleoprotein during virus amplification.

Influenza virus ribonucleoproteins (RNPs) were reconstituted in vivo from cloned cDNAs expressing the three polymerase subunits, the nucleoprotein (NP), and short template RNAs. The structure of purified RNPs was studied by electron microscopy and image processing. Circular and elliptic structures were obtained in which the NP and the polymerase complex could be defined. Comparison of the structure of RNPs of various lengths indicated that each NP monomer interacts with approximately 24 nucleotides. The analysis of the amplification of RNPs with different lengths showed that those with the highest replication efficiency contained an even number of NP monomers, suggesting that the NP is incorporated as dimers into newly synthesized RNPs.

Animals↗

PubMed2000

Protein synthesis shut-off induced by influenza virus infection is independent of PKR activity.

The role of PKR activity in influenza virus-induced cell shut-off was studied by infection of PKR(+) or PKR(-) cell cultures and metabolic labeling in vivo. No differences in the synthesis of viral proteins or the decay of cellular protein synthesis were observed. To investigate the relevance of the inhibition of cellular pre-mRNA polyadenylation and nucleocytoplasmic transport in virus-induced shut-off, we carried out similar experiments with mutant viruses lacking C-terminal sequences of NS1 protein. No differences in the shut-off induced by mutant versus wild-type viruses were observed, indicating that these nuclear events are not relevant for shut-off. The analysis of cytoplasmic mRNA stability indicated that the accumulation of viral mRNA during the infection correlated with the progressive decay of cellular mRNA, in both the wild type and an NS1 deletion mutant.

3T3 Cells↗

PubMed2000

The replication activity of influenza virus polymerase is linked to the capacity of the PA subunit to induce proteolysis.

The PA subunit of the influenza virus polymerase complex is a phosphorylated protein that induces a proteolytic process that decreases its own accumulation levels and those of coexpressed proteins. The amino-terminal third of the protein is responsible for the induction of proteolysis. We mutated five potential casein kinase II phosphorylation sites located in the amino-terminal third of the protein. Mutations affecting position 157 almost completely abrogated proteolysis induction, whereas a mutation at position 162 produced a moderate decrease and mutations at positions 151, 200, and 224 did not affect proteolysis induction. Reconstitution of the influenza virus polymerase in vivo with viral model RNA containing the chloramphenicol acetyltransferase (CAT) gene indicated that the CAT activity obtained correlated with the capacity of each PA mutant to induce proteolysis. RNA protection assays of the products obtained with viral polymerase, reconstituted in vivo with model RNAs, indicated that mutations at position 157 led to a selective loss of the ability to synthesize cRNA from the viral RNA template but not to transcribe viral RNA, while a mutation affecting position 162 showed an intermediate phenotype. Collectively, these data provide a link between PA-mediated induction of proteolysis and the replication activity of the polymerase.

Amino Acid Sequence↗

PubMed2000

Eukaryotic translation initiation factor 4GI is a cellular target for NS1 protein, a translational activator of influenza virus.

Influenza virus NS1 protein is an RNA-binding protein whose expression alters several posttranscriptional regulatory processes, like polyadenylation, splicing, and nucleocytoplasmic transport of cellular mRNAs. In addition, NS1 protein enhances the translational rate of viral, but not cellular, mRNAs. To characterize this effect, we looked for targets of NS1 influenza virus protein among cellular translation factors. We found that NS1 coimmunoprecipitates with eukaryotic initiation factor 4GI (eIF4GI), the large subunit of the cap-binding complex eIF4F, either in influenza virus-infected cells or in cells transfected with NS1 cDNA. Affinity chromatography studies using a purified His-NS1 protein-containing matrix showed that the fusion protein pulls down endogenous eIF4GI from COS-1 cells and labeled eIF4GI translated in vitro, but not the eIF4E subunit of the eIF4F factor. Similar in vitro binding experiments with eIF4GI deletion mutants indicated that the NS1-binding domain of eIF4GI is located between residues 157 and 550, in a region where no other component of the translational machinery is known to interact. Moreover, using overlay assays and pull-down experiments, we showed that NS1 and eIF4GI proteins interact directly, in an RNA-independent manner. Mapping of the eIF4GI-binding domain in the NS1 protein indicated that the first 113 N-terminal amino acids of the protein, but not the first 81, are sufficient to bind eIF4GI. The first of these mutants has been previously shown to act as a translational enhancer, while the second is defective in this activity. Collectively, these and previously published data suggest a model where NS1 recruits eIF4GI specifically to the 5' untranslated region (5' UTR) of the viral mRNA, allowing for the preferential translation of the influenza virus messengers.

5' Untranslated Regions↗

PubMed1999

Distinct regions of influenza virus PB1 polymerase subunit recognize vRNA and cRNA templates.

The influenza virus RNA polymerase is a heterotrimer comprising the PB1, PB2 and PA subunits. PB1 is the core of the complex and accounts for the polymerase activity. We have studied the interaction of PB1 with model cRNA template by in vitro binding and Northwestern analyses. The binding to model cRNA was specific and showed an apparent Kd of approximately 7x10(-8) M. In contrast to the interaction with vRNA, PB1 was able to bind equally the 5' and 3' arm of the cRNA panhandle. The N-terminal 139 amino acids of PB1 and sequences between positions 267 and 493 proved positive for binding to cRNA, whereas the interaction with vRNA template previously was mapped to the N- and C-terminal regions. Competition experiments using the 5' and 3' arms of either the vRNA or cRNA panhandle indicated that the N-terminal binding site is shared by both templates. The data indicate that the PB1 RNA-binding sites are constituted by: (i) residues located at the N-terminus (probably common for vRNA and cRNA binding) and, either (ii) residues from the central part of PB1 (for cRNA) or (iii) residues from the C-terminal region of PB1 (for vRNA), and suggest that PB1 undergoes a conformational change upon binding to cRNA versus vRNA templates.

Animals↗

PubMed1999

Interaction of influenza virus NS1 protein and the human homologue of Staufen in vivo and in vitro.

A screening for human proteins capable of interacting with influenza virus NS1 has been carried out using the two-hybrid genetic trap in yeast. A cDNA corresponding to the human homologue of Drosophila melanogaster Staufen protein (hStaufen) was isolated that fulfilled all genetic controls of the two-hybrid protocol. Using a hStaufen cDNA isolated from a lambda human library, the interaction of hStaufen and NS1 proteins was characterised in vivo and in vitro. Co-transfection of NS1 cDNA and a partial cDNA of hStaufen led to the relocalisation of recombinant hStaufen protein from its normal accumulation site in the cytoplasm to the nuclear location of NS1 protein. NS1 and hStaufen proteins could be co-immunoprecipitated from extracts of co-transfected cells and from mixtures of extracts containing either protein, as well as from extracts of influenza virus-infected cells. Furthermore, both proteins co-localised in the ribosomal and polysomal fractions of influenza virus-infected cells. The interaction was also detected in pull-down experiments using a resin containing purified hStaufen and NS1 protein translated in vitro. Deletion mapping of the NS1 gene indicated that a mutant protein containing the N-terminal 81 amino acids is unable to interact with hStaufen, in spite of retaining full RNA-binding capacity. These results are discussed in relation to the possible mechanisms of action of hStaufen and its relevance for influenza virus infection.

Animals↗

PubMed1999

The mammalian staufen protein localizes to the somatodendritic domain of cultured hippocampal neurons: implications for its involvement in mRNA transport.

In hippocampal neurons, certain mRNAs have been found in dendrites (), and their localization and translation have been implicated in synaptic plasticity (). One attractive candidate to achieve transport of mRNAs into dendrites is Staufen (Stau), a double-stranded RNA-binding protein, which plays a pivotal role in mRNA transport, localization, and translation in Drosophila (). Using antibodies raised against a peptide located in the RNA-binding domain IIa and a polyclonal antibody raised against a recently cloned human Staufen homolog, we identify a 65 kDa rat homolog in cultured rat hippocampal neurons. In agreement with the exclusive somatodendritic localization of mRNAs in these cells, we find that Staufen is restricted to the same domain. By immunoelectron microscopy, we show enrichment of the mammalian homolog of Stau (mStau) in the vicinity of smooth endoplasmic reticulum and microtubules near synaptic contacts. Finally, the association of the mStau with neuronal mRNAs is suggested by the colocalization with ribonucleoprotein particles specifically in distal dendrites known to contain mRNA, ribosomes, and translation factors (). These results suggest a role for mStau in the polarized transport and localization of mRNAs in mammalian neurons.

Amino Acid Sequence↗