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Results for “Coupled model”

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PubMed1996

The role of altered [Ca2+]i regulation in apoptosis, oncosis, and necrosis.

Understanding the processes and events that occur when a cell undergoes a prelethal injury or that lead the cell to death following a lethal injury has been the aim of our research for a number of years. Throughout this period much has been learned, recently at rapid rates, not only by us but by many other investigators as well. Based on the data gathered, we proposed a working hypothesis over a decade ago and have since continually updated it as new experimentation is performed. Our laboratory has focused particularly on the role of cytoplasmic ionized calcium ([Ca2+]i) and the effects of its deregulation on prelethal events, including oncosis and apoptosis, and lethal events (necrosis) following cell death. [Ca2+]i appears to be a major link and signalling event. Understanding the mechanisms involved by using a variety of in vivo and in vitro models, coupled with state-of-the-art methodologies, should now allow us to prevent cell death by killing cells when necessary through gene therapy and cancer chemotherapy.

Animals↗

PubMed2005

Disruption of the filamentous actin cytoskeleton is necessary for the activation of capacitative calcium entry in naive smooth muscle cells.

It has been proposed that cytoskeleton plays a key positive role in the activation of capacitative calcium entry (CCE), which supported the secretion-like hypothesis for the mechanisms underlying this process. However, its role on CCE in native smooth muscle is unknown. Here we demonstrate that CCE in isolated gallbladder myocytes was enhanced by cytochalasin D or latrunculin A treatments (agents that cause actin disassembly) whereas it was reduced by jasplakinolide treatment (which causes actin polymerization), suggesting that actin cytoskeleton acts as a barrier in CCE. In addition, we show for the first time that depletion of intracellular Ca2+ stores by thapsigargin and cholecystokinin in BAPTA-loaded cells induced a decrease in F-actin content that was consistent with a link between CCE and actin reorganization. In conclusion, these data suggest an active participation of actin reorganization in the implementation of CCE and support a conformational coupling model for this process in naive smooth muscle cells.

Actin Cytoskeleton↗

PubMed2006

Store-mediated calcium entry in pleural mesothelial cells.

Store-mediated Ca2+ entry is thought as the main pathway for Ca2+ influx in non-excitable cells. Although a role for the actin cytoskeleton in store-mediated Ca2+ entry has been proposed in some cell types, the role of actin cytoskeleton in store-mediated Ca2+ entry is still a controversy. To address this question, the effects of cytoskeletal modifiers on store-mediated Ca2+ entry in pleural mesothelial cells were examined. Thapsigargin (1 microM) induced a sufficient signal for the activation of store-mediated Ca2+ entry in pleural mesothelial cells. In the absence of extracellular Ca2+, thapsigargin induced only a transient elevation of [Ca2+]i. Moreover, re-addition of Ca2+ increased the elevation of [Ca2+]i. Passive elevations in [Ca2+]i without thapsigargin, which is induced from Ca2+ containing solution switch to Ca2+ free solution and re-add Ca2+ containing solution, were not observed in pleural mesothelial cells. Thapsigargin-induced Ca2+ entry was still present after nifedipine (1 microM) treatment. However, SKF96365 (1 microM) blocked thapsigargin-induced Ca2+ entry. Mycalolide B (1 microM) completely disrupts actin cytoskeleton in pleural mesothelial cells, but thapsigargin-induced store-mediated Ca2+ entry was preserved. Jasplakinolide (3 microM) prevented thapsigargin-induced store-mediated Ca2+ entry. These results suggest that store-mediated Ca2+ entry in pleural mesothelial cells may be mediated by a recently proposed secretion-like coupling model for store-mediated Ca2+ entry.

Actins↗

PubMed2005

Transfer of beta subunit regulation from high to low voltage-gated Ca2+ channels.

High voltage-activated Ca(2+) channel expression and gating is controlled by their beta subunits. Although the sites of interaction are known at the atomic level, how beta modulates gating remains to be determined. Using a chimeric approach, beta subunit regulation was conferred to a low voltage-activated channel. Regulation was dependent on a rigid linker connecting the alpha(1) interaction domain to IS6. Chimeric channels also revealed a role for IS6 in channel gating. Taken together, these results support a direct coupling model where beta subunits alter movements in IS6 that occur as the channel transits between closed, open, and inactivated states.

Amino Acid Sequence↗

PubMed2006

Leaching behaviour of low level organic pollutants contained in cement-based materials: experimental methodology and modelling approach.

The aim of this paper is the investigation of the leaching behaviour of different porous materials containing organic pollutants (PAH: naphthalene and phenanthrene). The assessment methodology of long term leaching behaviour of inorganic materials was extended to cement solidified organic pollutants. Based on a scenario-approach considering environmental factors, matrix and pollutants specificities, the applied methodology is composed of adapted equilibrium and dynamic leaching tests. The contributions of different physical and chemical mechanisms were identified and the leaching behaviour was modelled. The physical parameters of the analysed reference and polluted materials are similar. A difference in the pore size distribution appears for higher naphthalene content. The solubility of the PAH contained in the material is affected by the ionic strength and by the presence of a co-solvent; the solution pH does not influence PAH solubility. The solubility of the major mineral species is not influenced by the presence of the two PAH nor by the presence of the methanol as co-solvent in the range of the tested material compositions. In the case of the leaching of a monolith material the main transport mechanism is the diffusion in the porous system. For both mineral and organic species we observed at least two dynamic domains. At the beginning of the leaching process the released flux is due to the surface dissolution and to the diffusion of the main quantity dissolved in the initial pore solution. The second period is governed by a stationary regime between dissolution in pore water and diffusion. The model, coupling transport and chemical phenomena in the pore solution, at the monolith surface and in the leachate simulates satisfactory the release for both mineral and organic species.

Adsorption↗

PubMed2004

Spatially dissociated flow-metabolism coupling in brain activation.

The relationships among cerebral blood flow (CBF), oxygen consumption (CMRO(2)) and glucose use (CMR(glc)) constitute the basis of functional brain-imaging. Here we report spatially dissociated changes of CMRO(2) and CBF during motor activity that lead us to propose a revision of conventional CBF-CMRO(2) coupling models. In the left primary and supplementary motor cortices, CBF and CMRO(2) rose significantly during finger-thumb tapping. However, in the right putamen CBF did not rise, despite a significant increase in CMRO(2). We explain these observations by invoking a central command mechanism that regulates CBF in the putamen in anticipation of movement. By this mechanism, CBF rose in the putamen before the measurements of CBF and CMRO(2) while CMRO(2) rose when actual motion commenced.

Adult↗

PubMed2005

Bandwidths studied as a function of temperature in ammonium halides near Tlambda.

This study gives our analysis of the Raman bandwidths using the soft mode-hard mode coupling model applied to ammonium halides. The temperature dependence of our observed bandwidths for the nu5(174 cm-1) Raman mode of NH4Cl and nu5(177 cm-1) Raman mode of NH4Br, is analyzed close to phase transitions. From our analysis, we obtain beta=0.13 as the values of the critical exponent for the order parameter in the first order phase region for the ammonium halides.

Ammonium Chloride↗

PubMed2006

Density functional theory study of vibronic structure of the first absorption Qx band in free-base porphin.

Harmonic vibrational frequencies and vibronic intensities in the first S(0)-->S(1) (pipi( *)) absorption band of free-base porphin (H(2) P) are investigated by hybrid density functional theory (DFT) with the standard B3LYP functional. The S(0)-S(1) transition probability is calculated using time-dependent DFT with account of Franck-Condon (FC) and Herzberg-Teller (HT) contributions to the electric-dipole transition moments including displacements along all 108 vibrational modes. Two weak wide bands observed in the gas phase absorption spectra of the H(2) P molecule at 626 and 576 nm are interpreted as the 0-0 band of the X(1) A(g)-->1B(3u) transition and the 0-1 band with largest contributions from the nu(10)(a(g))=1610 cm(-1) and nu(19)(b(1g))=1600 cm(-1) modes, respectively, in agreement with previous tentative assignments. Both bands are induced by the HT mechanism, while the FC contributions are negligible. A number of fine structure bands, including combination of two vibrational quanta, are obtained and compared with available spectra from supersonic jet and Shpolskij matrices. Both absorption and fluorescence spectra are interpreted on ground of the linear coupling model and a good fulfillment of the mirror-symmetry rule.

Chemical Phenomena↗

PubMed2005

CIF and other mysteries of the store-operated Ca2+-entry pathway.

The molecular mechanism of the store-operated Ca2+-entry (SOCE) pathway remains one of the most intriguing and long lasting mysteries of Ca2+ signaling. The elusive calcium influx factor (CIF) that is produced upon depletion of Ca2+ stores has attracted growing attention, triggered by new discoveries that filled the gap in the chain of reactions leading to activation of store-operated channels and Ca2+ entry. Ca2+-independent phospholipase A2 emerged as a target of CIF, and a major determinant of the SOCE mechanism. Here, we present our viewpoint on CIF and conformational-coupling models of SOCE from a historical perspective, trying to resolve some of the problem areas, and summarizing our present knowledge on how depletion of intracellular Ca2+ stores signals to plasma membrane channels to open and provide Ca2+ influx that is required for many important physiological functions.

Animals↗

PubMed2003

Location of structural transitions in an isotopically labeled lung surfactant SP-B peptide by IRRAS.

Pulmonary surfactant, a lipid/protein complex that lines the air/water interface in the mammalian lung, functions to reduce the work of breathing. Surfactant protein B (SP-B) is a small, hydrophobic protein that is an essential component of this mixture. Structure-function relationships of SP-B are currently under investigation as the protein and its peptide analogs are being incorporated into surfactant replacement therapies. Knowledge of the structure of SP-B and its related peptides in bulk and monolayer phases will facilitate the design of later generation therapeutic agents. Prior infrared reflection-absorption spectroscopic studies reported notable, reversible surface pressure-induced antiparallel beta-sheet formation in a synthetic peptide derived from human SP-B, residues 9-36 (SP-B(9-36)). In the current work, infrared reflection-absorption spectroscopy is applied in conjunction with isotopic labeling to detect the site and pressure dependence of the conformational change. SP-B(9-36), synthesized with (13)C=O-labeled Ala residues in positions 26, 28, 30, and 32, shifted the beta-sheet marker band to approximately 1600 cm(-1) and thus immediately identified this structural element within the labeled region. Surface pressure-induced alterations in the relative intensities of Amide I band constituents are interpreted using a semiempirical transition dipole coupling model. In addition, electron micrographs reveal the formation of tubular myelin structures from in vitro preparations using SP-B(9-36) in place of porcine SP-B indicating that the peptide has the potential to mimic this property of the native protein.

Amino Acid Sequence↗

PubMed2004

Calcium buffering and excitation-contraction coupling in developing avian myocardium.

This report provides a detailed analysis of developmental changes in cytoplasmic free calcium (Ca(2+)) buffering and excitation-contraction coupling in embryonic chick ventricular myocytes. The peak magnitude of field-stimulated Ca(2+) transients declined by 41% between embryonic day (ED) 5 and 15, with most of the decline occurring between ED5 and 11. This was due primarily to a decrease in Ca(2+) currents. Sarcoplasmic reticulum (SR) Ca(2+) content increased 14-fold from ED5 to 15. Ca(2+) transients in voltage-clamped myocytes after blockade of SR function permitted computation of the fast Ca buffer power of the cytosol as expressed as generalized values of B(max) and K(D). B(max) rose with development whereas K(D) did not change significantly. The computed SR Ca(2+) contribution to the Ca(2+) transient and gain factor for Ca(2+)-induced Ca(2+) release increased markedly between ED5 and 11 and slightly thereafter. These results paralleled the maturation of SR and peripheral couplings reported by others and demonstrated a strong relationship between structure and function in development of excitation-contraction coupling. Modeling of buffer power from estimates of the major cytosolic Ca binding moieties yielded a B(max) and K(D) in reasonable agreement with experiment. From ED5 to 15, troponin C was the major Ca(2+) binding moiety, followed by SR and calmodulin.

Aging↗

PubMed1991

Shape transitions and shape stability of giant phospholipid vesicles in pure water induced by area-to-volume changes.

Shape transformations of vesicles of dimyristoylphosphatidylcholine (= DMPC) and palmitoyloleylphosphatidylcholine (= POPC) in ion-free water were induced by changing the area-to-volume ratio via temperature variations. Depending on the pretreatment we find several types of shape changes for DMPC (in pure water) at increasing area-to-volume ratio: (a) budding transitions leading to the formation of a chain of vesicles at further increase of the area-to-volume ratio, (b) discocyte-stomatocyte transitions, (c) reentrant dumbbell-pear-dumbbell transitions, and (d) spontaneous blebbing and/or tether formation of spherical vesicles. Beside these transitions a more exotic dumbbell-discocyte transition (e) was found which proceeded via local instabilities. Pears, discocytes, and stomatocytes are stable with respect to small temperature variations unless the excess area is close to values corresponding to limiting shapes of budded vesicles where temperature variations of less than or equal to 0.1 degree C lead to spontaneous budding to the inside or the outside. For POPC we observed only budding transitions to the inside leading either to chains of vesicles or to distributions of equally sized daughter vesicles protruding to the inside of the vesicle. Preliminary experiments concerning the effect of solutes are also reported. The first three types of shape transitions can be explained in terms of the bilayer coupling model assuming small differences in thermal expansivities of the two monolayers. This does not hold for the observed instabilities close to the limiting shapes.

Dimyristoylphosphatidylcholine↗

PubMed1993

Nucleation and growth of ice crystals inside cultured hepatocytes during freezing in the presence of dimethyl sulfoxide.

A three-part, coupled model of cell dehydration, nucleation, and crystal growth was used to study intracellular ice formation (IIF) in cultured hepatocytes frozen in the presence of dimethyl sulfoxide (DMSO). Heterogeneous nucleation temperatures were predicted as a function of DMSO concentration and were in good agreement with experimental data. Simulated freezing protocols correctly predicted and explained experimentally observed effects of cooling rate, warming rate, and storage temperature on hepatocyte function. For cells cooled to -40 degrees C, no IIF occurred for cooling rates less than 10 degrees C/min. IIF did occur at faster cooling rates, and the predicted volume of intracellular ice increased with increasing cooling rate. Cells cooled at 5 degrees C/min to -80 degrees C were shown to undergo nucleation at -46.8 degrees C, with the consequence that storage temperatures above this value resulted in high viability independent of warming rate, whereas colder storage temperatures resulted in cell injury for slow warming rates. Cell damage correlated positively with predicted intracellular ice volume, and an upper limit for the critical ice content was estimated to be 3.7% of the isotonic water content. The power of the model was limited by difficulties in estimating the cytosol viscosity and membrane permeability as functions of DMSO concentration at low temperatures.

Animals↗

PubMed2000

Activation of G protein by opioid receptors: role of receptor number and G-protein concentration.

The collision-coupling model for receptor-G-protein interaction predicts that the rate of G-protein activation is dependent on receptor density, but not G-protein levels. C6 cells expressing mu- or delta-opioid receptors, or SH-SY5Y cells, were treated with beta-funaltrexamine (mu) or naltrindole-5'-isothiocyanate (delta) to decrease receptor number. The time course of full or partial agonist-stimulated ¿35SGTPgammaS binding did not vary in C6 cell membranes containing <1-25 pmol/mg mu-opioid receptor, or 1. 4-4.3 pmol/mg delta-opioid receptor, or in SHSY5Y cells containing 0. 16-0.39 pmol/mg receptor. The association of ¿35SGTPgammaS binding was faster in membranes from C6mu cells than from C6delta cells. A 10-fold reduction in functional G-protein, following pertussis toxin treatment, lowered the maximal level of ¿35SGTPgammaS binding but not the association rate. These data indicate a compartmentalization of opioid receptors and G protein at the cell membrane.

Enkephalin, Ala(2)-MePhe(4)-Gly(5)-↗

PubMed2003

Intracellular trafficking of CLN3, the protein underlying the childhood neurodegenerative disease, Batten disease.

Juvenile neuronal ceroid lipofuscinoses (Batten disease) is a progressive neurodegenerative disorder resulting from mutations in the CLN3 gene, which encodes a hydrophobic 438 amino acid protein of unknown function. Prior studies have shown that CLN3 is expressed in multiple tissues, with highest levels in brain and testis. Experiments using cells overexpressing CLN3 indicate that CLN3 is a lysosomal resident protein. However, studies to date have not addressed trafficking of endogenous CLN3. As such, the purpose of the present study was two-fold. First, to develop a culture model to allow evaluation of native CLN3 transport. Second, to utilize available epitope-specific antibodies to determine if CLN3 reaches the plasma membrane en route to the lysosome. Our data using a NCCIT (embryonic testicular carcinoma) cell model coupled with surface biotinylation and antibody trapping demonstrated that at least a proportion of CLN3 trafficks to the lysosome via the cell membrane. Moreover, inhibition of the micro3A subunit of the AP-3 adapter protein complex increased levels of CLN3 at the cell surface.

Amino Acid Sequence↗

PubMed1997

Capacitative calcium entry is inhibited in vascular endothelial cells by disruption of cytoskeletal microfilaments.

The role of an intact cytoskeleton for store-operated ('capacitative') Ca2+ influx was investigated in single cultured vascular endothelial cells. Capacitative Ca2+ entry was measured as changes of cytoplasmic Ca2+ concentration ([Ca2+]i) induced by depletion of Ca2+ stores with thapsigargin. In cells pretreated with cytochalasin D, an agent that disrupts the microfilament network of the cytoskeleton, as confirmed with FITC-phalloidin staining, capacitative Ca2+ entry was inhibited. Cytochalasin D did not affect basal [Ca2+]i nor ATP-induced increases of [Ca2+]i, indicating that release of Ca2+ from intracellular stores through the inositol-phosphate pathway was intact. These results suggest that microfilaments are an integral part of the mechanism for capacitative Ca2+ entry. The necessity for an intact cytoskeleton favors a conformational coupling model for store-operated Ca2+ influx.

Adenosine Triphosphate↗

PubMed2003

Experimental comparison of three monoclonal antibodies for the class-selective immunoextraction of triazines. Correlation with molecular modeling and principal component analysis studies.

The specificity of three immunosorbents (ISs) based on different monoclonal anti-triazine antibodies has been characterized by extraction recoveries studies and with step elution experiments. Both indicated that the anti-dichloroatrazine IS is specific of terbutylazine and cyanazine. The anti-atrazine IS is specific of the chlorotriazines, whereas the anti-ametryn IS can trap all the triazines. This confirms the great influence of the hapten design on the specificity of the resulting antibodies, even if the target molecules are small. Moreover, the anti-ametryn IS is suitable for class-selective extraction of triazines contained in complex matrices. An approach designed to learn more about the specificity for a group of structurally related compounds of antibodies produced with a given compound is proposed and evaluated. Molecular modeling followed by principal component analysis has been used to obtain distribution maps with the relative position of each immunoconjugate and all the triazines. In all three cases, conclusions on specificity made with the analysis of the maps fit well with the experimental results. Consequently, molecular modeling coupled with principal component analysis seems to be a unique, inexpensive, and rapid tool to select the appropriate hapten providing highly specific or class-specific antibodies according to the given problem.

Antibodies, Monoclonal↗

PubMed2003

Surface patterning via evaporation of ultrathin films containing nanoparticles.

The dewetting dynamics of ultrathin films containing potentially surface-active nanoparticles is considered in the presence of evaporation. Evolution equations for the film height and particle surface and bulk concentration are derived using a lubrication model coupled by a constitutive relation for the dependence of the viscosity on local particle concentration. A linear stability analysis and numerical simulations are used to determine how particle mass distribution depends on the various physical parameters such as equilibrium film separation distance, initial packing concentration, rate of evaporation, and particle surface activity. Our results show that when starting from an initially uniform distribution the particles become aligned into distinct "bands" in rectilinear geometry, or "rings" in cylindrical geometry. The functional dependence of the pattern spacing on relevant system parameters is studied and detailed herein.

Journal Article↗