Apoptosis and proliferation of dentate gyrus neurons after single and intermittent limbic seizures

Neuronal apoptosis was observed in the rat dentate gyrus in two experimental models of human limbic epilepsy. Five hours after one hippocampal kindling stimulation, a marked increase of in situ terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling (TUNEL) of fragmented DNA was observed in nuclei located within and on the hilar border of the granule cell layer and in the polymorphic region. Forty kindling stimulations with 5-min interval produced higher numbers of labeled nuclei compared with one stimulation. The increase of TUNEL-positive nuclei was prevented by the protein synthesis inhibitor cycloheximide but not affected by the N-methyl-d-aspartate receptor antagonist MK-801. Kainic acid-induced seizures lead to a pattern of labeling in the hippocampal formation identical to that evoked by kindling. A large proportion of cells displaying TUNEL-positive nuclei was double-labeled by the neuron-specific antigen NeuN, demonstrating the neuronal identity of apoptotic cells. Either 1 or 40 kindling stimulations also gave rise to a marked increase of the number of cells double-labeled with the mitotic marker bromodeoxyuridine and NeuN in the subgranular zone and on the hilar border of the dentate granule cell layer. The present data show that single and intermittent, brief seizures induce both apoptotic death and proliferation of dentate gyrus neurons. We hypothesize that these processes, occurring early during epileptogenesis, are primary events in the development of hippocampal pathology in animals and possibly also in patients suffering from temporal lobe epilepsy.

Direct molecular level measurements of the electrostatic properties of a protein surface

In this work, we used direct measurements with the surface force apparatus to determine the pH-dependent electrostatic charge density of a single binding face of streptavidin. Mean field calculations have been used with considerable success to model electrostatic potential fields near protein surfaces, but these models and their inherent assumptions have not been tested directly at the molecular level. Using the force apparatus and immobilized, oriented monolayers of streptavidin, we measured a pI of 5–5.5 for the biotin-binding face of the protein. This differs from the pI of 6.3 for the soluble protein and confirms that we probed the local electrostatic features of the macromolecule. With finite difference solutions of the linearized Poisson–Boltzmann equation, we then calculated the pH-dependent charge densities adjacent to the same face of the protein. These calculated values agreed quantitatively with those obtained by direct force measurements. Although our study focuses on the pH-dependence of surface electrostatics, this direct approach to probing the electrostatic features of proteins is applicable to investigations of any perturbations that alter the charge distribution of the surfaces of immobilized molecules.

Stretching single-domain proteins: Phase diagram and kinetics of force-induced unfolding

Single-molecule force spectroscopy reveals unfolding of domains in titin on stretching. We provide a theoretical framework for these experiments by computing the phase diagrams for force-induced unfolding of single-domain proteins using lattice models. The results show that two-state folders (at zero force) unravel cooperatively, whereas stretching of non-two-state folders occurs through intermediates. The stretching rates of individual molecules show great variations reflecting the heterogeneity of force-induced unfolding pathways. The approach to the stretched state occurs in a stepwise “quantized” manner. Unfolding dynamics and forces required to stretch proteins depend sensitively on topology. The unfolding rates increase exponentially with force f till an optimum value, which is determined by the barrier to unfolding when f = 0. A mapping of these results to proteins shows qualitative agreement with force-induced unfolding of Ig-like domains in titin. We show that single-molecule force spectroscopy can be used to map the folding free energy landscape of proteins in the absence of denaturants.

Escherichia coli RNA polymerase terminates transcription efficiently at rho-independent terminators on single-stranded DNA templates

Several models have been proposed for the mechanism of transcript termination by Escherichia coli RNA polymerase at rho-independent terminators. Yager and von Hippel (Yager, T. D. & von Hippel, P. H. (1991) Biochemistry 30, 1097–118) postulated that the transcription complex is stabilized by enzyme–nucleic acid interactions and the favorable free energy of a 12-bp RNA–DNA hybrid but is destabilized by the free energy required to maintain an extended transcription bubble. Termination, by their model, is viewed simply as displacement of the RNA transcript from the hybrid helix by reformation of the DNA helix. We have proposed an alternative model where the RNA transcript is stably bound to RNA polymerase primarily through interactions with two single-strand specific RNA-binding sites; termination is triggered by formation of an RNA hairpin that reduces binding of the RNA to one RNA-binding site and, ultimately, leads to its ejection from the complex. To distinguish between these models, we have tested whether E. coli RNA polymerase can terminate transcription at rho-independent terminators on single-stranded DNA. RNA polymerase cannot form a transcription bubble on these templates; thus, the Yager–von Hippel model predicts that intrinsic termination will not occur. We find that transcript elongation on single-stranded DNA templates is hindered somewhat by DNA secondary structure. However, E. coli RNA polymerase efficiently terminates and releases transcripts at several rho-independent terminators on such templates at the same positions as termination occurs on duplex DNAs. Therefore, neither the nontranscribed DNA strand nor the transcription bubble is essential for rho-independent termination by E. coli RNA polymerase.

Neutralizing monoclonal antibodies to hepatocyte growth factor/scatter factor (HGF/SF) display antitumor activity in animal models

The hepatocyte growth factor (HGF/SF) receptor, Met, regulates mitogenesis, motility, and morphogenesis in a cell type-dependent fashion. Activation of Met via autocrine, paracrine, or mutational mechanisms can lead to tumorigenesis and metastasis and numerous studies have linked inappropriate expression of this ligand-receptor pair to most types of human solid tumors. To prepare mAbs to human HGF/SF, mice were immunized with native and denatured preparations of the ligand. Recloned mAbs were tested in vitro for blocking activity against scattering and branching morphogenesis. Our results show that no single mAb was capable of neutralizing the in vitro activity of HGF/SF, and that the ligand possesses a minimum of three epitopes that must be blocked to prevent Met tyrosine kinase activation. In vivo, the neutralizing mAb combination inhibited s.c. growth in athymic nu/nu mice of tumors dependent on an autocrine Met-HGF/SF loop. Importantly, growth of human glioblastoma multiforme xenografts expressing Met and HGF/SF were markedly reduced in the presence of HGF/SF-neutralizing mAbs. These results suggest interrupting autocrine and/or paracrine Met-HGF/SF signaling in tumors dependent on this pathway is a possible intervention strategy.

Immunization with DNA vaccines encoding glycoprotein D or glycoprotein B, alone or in combination, induces protective immunity in animal models of herpes simplex virus-2 disease.

DNA vaccines expressing herpes simplex virus type 2 (HSV-2) full-length glycoprotein D (gD), or a truncated form of HSV-2 glycoprotein B (gB) were evaluated for protective efficacy in two experimental models of HSV-2 infection. Intramuscular (i.m.) injection of mice showed that each construction induced neutralizing serum antibodies and protected the mice from lethal HSV-2 infection. Dose-titration studies showed that low doses (< or = 1 microgram) of either DNA construction induced protective immunity, and that a single immunization with the gD construction was effective. The two DNAs were then tested in a low-dosage combination in guinea pigs. Immune sera from DNA-injected animals had antibodies to both gD and gB, and virus neutralizing activity. When challenged by vaginal infection with HSV-2, the DNA-immunized animals were significantly protected from primary genital disease.

Simulação e modelagem computacional de dados de espalhamento à baixos ângulos - enfoque em estruturas de alta simetria

Esta tese apresenta uma abordagem para a criação rápida de modelos em diferentes geometrias (complexas ou de alta simetria) com objetivo de calcular a correspondente intensidade espalhada, podendo esta ser utilizada na descrição de experimentos de es- palhamento à baixos ângulos. A modelagem pode ser realizada com mais de 100 geome- trias catalogadas em um Banco de Dados, além da possibilidade de construir estruturas a partir de posições aleatórias distribuídas na superfície de uma esfera. Em todos os casos os modelos são gerados por meio do método de elementos finitos compondo uma única geometria, ou ainda, compondo diferentes geometrias, combinadas entre si a partir de um número baixo de parâmetros. Para realizar essa tarefa foi desenvolvido um programa em Fortran, chamado de Polygen, que permite modelar geometrias convexas em diferentes formas, como sólidos, cascas, ou ainda com esferas ou estruturas do tipo DNA nas arestas, além de usar esses modelos para simular a curva de intensidade espalhada para sistemas orientados e aleatoriamente orientados. A curva de intensidade de espalhamento é calculada por meio da equação de Debye e os parâmetros que compõe cada um dos modelos, podem ser otimizados pelo ajuste contra dados experimentais, por meio de métodos de minimização baseados em simulated annealing, Levenberg-Marquardt e algorítmicos genéticos. A minimização permite ajustar os parâmetros do modelo (ou composição de modelos) como tamanho, densidade eletrônica, raio das subunidades, entre outros, contribuindo para fornecer uma nova ferramenta para modelagem e análise de dados de espalhamento. Em outra etapa desta tese, é apresentado o design de modelos atomísticos e a sua respectiva simulação por Dinâmica Molecular. A geometria de dois sistemas auto-organizado de DNA na forma de octaedro truncado, um com linkers de 7 Adeninas e outro com linkers de ATATATA, foram escolhidas para realizar a modelagem atomística e a simulação por Dinâmica Molecular. Para este sistema são apresentados os resultados de Root Mean Square Deviations (RMSD), Root Mean Square Fluctuations (RMSF), raio de giro, torção das hélices duplas de DNA além da avaliação das ligações de Hidrogênio, todos obtidos por meio da análise de uma trajetória de 50 ns.

Cosmological perturbations in coherent oscillating scalar field models

The fact that fast oscillating homogeneous scalar fields behave as perfect fluids in average and their intrinsic isotropy have made these models very fruitful in cosmology. In this work we will analyse the perturbations dynamics in these theories assuming general power law potentials V(ϕ) = λ|ϕ|^n /n. At leading order in the wavenumber expansion, a simple expression for the effective sound speed of perturbations is obtained c_eff^ 2  = ω = (n − 2)/(n + 2) with ω the effective equation of state. We also obtain the first order correction in k^ 2/ω_eff^ 2 , when the wavenumber k of the perturbations is much smaller than the background oscillation frequency, ω_eff. For the standard massive case we have also analysed general anharmonic contributions to the effective sound speed. These results are reached through a perturbed version of the generalized virial theorem and also studying the exact system both in the super-Hubble limit, deriving the natural ansatz for δϕ; and for sub-Hubble modes, exploiting Floquet’s theorem.

USLE and WEPP Soil Erosion Models: Their Applicability and Use in Generating Cover Values to Stabilize Soil in the Los Alamos, New Mexico Area

Soil erosion is a naturally occurring process that involves the detachment, transport, and deposition of soil particles. Disturbances such as thinning and wildfire can reduce cover greatly and increase erosion rates. Forest managers may use erosion prediction tools, such as the Universal Soil Loss Equation (USLE) and Water Erosion Prediction Project (WEPP) to estimate erosion rates and develop techniques to manage erosion. However, it is important to understand the differences and the applications of each model. Erosion rates were generated by each model and the model most applicable to the study site, Los Alamos, New Mexico was determined. It was also used to find the amount of cover needed to stabilize soil. The USLE is a simpler model and less complicated than a computer model like WEPP, and thus easier to manipulate to estimate cover values. Predicted cover values were compared to field cover values. Cover is necessary to establish effective erosion control guidelines.

Response of Catch-Per-Unit-Effort to Spatial and Environmental Factors: an Exploratory Data Analysis Using Generalized Additive Models

Understanding spatial distributions and how environmental conditions influence catch-per-unit-effort (CPUE) is important for increased fishing efficiency and sustainable fisheries management. This study investigated the relationship between CPUE, spatial factors, temperature, and depth using generalized additive models. Combinations of factors, and not one single factor, were frequently included in the best model. Parameters which best described CPUE varied by geographic region. The amount of variance, or deviance, explained by the best models ranged from a low of 29% (halibut, Charlotte region) to a high of 94% (sablefish, Charlotte region). Depth, latitude, and longitude influenced most species in several regions. On the broad geographic scale, depth was associated with CPUE for every species, except dogfish. Latitude and longitude influenced most species, except halibut (Areas 4 A/D), sablefish, and cod. Temperature was important for describing distributions of halibut in Alaska, arrowtooth flounder in British Columbia, dogfish, Alaska skate, and Aleutian skate. The species-habitat relationships revealed in this study can be used to create improved fishing and management strategies.

GE models and algorithms for condensed phase equilibrium data regression in ternary systems: limitations and proposals

Phase equilibrium data regression is an unavoidable task necessary to obtain the appropriate values for any model to be used in separation equipment design for chemical process simulation and optimization. The accuracy of this process depends on different factors such as the experimental data quality, the selected model and the calculation algorithm. The present paper summarizes the results and conclusions achieved in our research on the capabilities and limitations of the existing GE models and about strategies that can be included in the correlation algorithms to improve the convergence and avoid inconsistencies. The NRTL model has been selected as a representative local composition model. New capabilities of this model, but also several relevant limitations, have been identified and some examples of the application of a modified NRTL equation have been discussed. Furthermore, a regression algorithm has been developed that allows for the advisable simultaneous regression of all the condensed phase equilibrium regions that are present in ternary systems at constant T and P. It includes specific strategies designed to avoid some of the pitfalls frequently found in commercial regression tools for phase equilibrium calculations. Most of the proposed strategies are based on the geometrical interpretation of the lowest common tangent plane equilibrium criterion, which allows an unambiguous comprehension of the behavior of the mixtures. The paper aims to show all the work as a whole in order to reveal the necessary efforts that must be devoted to overcome the difficulties that still exist in the phase equilibrium data regression problem.

The imprint of the crustal magnetic field on the thermal spectra and pulse profiles of isolated neutron stars

Isolated neutron stars (NSs) show a bewildering variety of astrophysical manifestations, presumably shaped by the magnetic field strength and topology at birth. Here, using state-of-the-art calculations of the coupled magnetic and thermal evolution of NSs, we compute the thermal spectra and pulse profiles expected for a variety of initial magnetic field configurations. In particular, we contrast models with purely poloidal magnetic fields to models dominated by a strong internal toroidal component. We find that, while the former displays double-peaked profiles and very low pulsed fractions, in the latter, the anisotropy in the surface temperature produced by the toroidal field often results in a single pulse profile, with pulsed fractions that can exceed the 50–60 per cent level even for perfectly isotropic local emission. We further use our theoretical results to generate simulated ‘observed’ spectra, and show that blackbody (BB) fits result in inferred radii that can be significantly smaller than the actual NS radius, even as low as ∼1–2 km for old NSs with strong internal toroidal fields and a high absorption column density along their line of sight. We compute the size of the inferred BB radius for a few representative magnetic field configurations, NS ages and magnitudes of the column density. Our theoretical results are of direct relevance to the interpretation of X-ray observations of isolated NSs, as well as to the constraints on the equation of state of dense matter through radius measurements.

Integration of different models in the design of chemical processes: Application to the design of a power plant

With advances in the synthesis and design of chemical processes there is an increasing need for more complex mathematical models with which to screen the alternatives that constitute accurate and reliable process models. Despite the wide availability of sophisticated tools for simulation, optimization and synthesis of chemical processes, the user is frequently interested in using the ‘best available model’. However, in practice, these models are usually little more than a black box with a rigid input–output structure. In this paper we propose to tackle all these models using generalized disjunctive programming to capture the numerical characteristics of each model (in equation form, modular, noisy, etc.) and to deal with each of them according to their individual characteristics. The result is a hybrid modular–equation based approach that allows synthesizing complex processes using different models in a robust and reliable way. The capabilities of the proposed approach are discussed with a case study: the design of a utility system power plant that has been decomposed into its constitutive elements, each treated differently numerically. And finally, numerical results and conclusions are presented.

Generation of representation models for complex systems using Lagrangian functions

In this article, a new methodology is presented to obtain representation models for a priori relation z = u(x1, x2, . . . ,xn) (1), with a known an experimental dataset zi; x1i ; x2i ; x3i ; . . . ; xni i=1;2;...;p· In this methodology, a potential energy is initially defined over each possible model for the relationship (1), what allows the application of the Lagrangian mechanics to the derived system. The solution of the Euler–Lagrange in this system allows obtaining the optimal solution according to the minimal action principle. The defined Lagrangian, corresponds to a continuous medium, where a n-dimensional finite elements model has been applied, so it is possible to get a solution for the problem solving a compatible and determined linear symmetric equation system. The computational implementation of the methodology has resulted in an improvement in the process of get representation models obtained and published previously by the authors.

Spectral features in isolated neutron stars induced by inhomogeneous surface temperatures

The thermal X-ray spectra of several isolated neutron stars display deviations from a pure blackbody. The accurate physical interpretation of these spectral features bears profound implications for our understanding of the atmospheric composition, magnetic field strength and topology, and equation of state of dense matter. With specific details varying from source to source, common explanations for the features have ranged from atomic transitions in the magnetized atmospheres or condensed surface, to cyclotron lines generated in a hot ionized layer near the surface. Here, we quantitatively evaluate the X-ray spectral distortions induced by inhomogeneous temperature distributions of the neutron star surface. To this aim, we explore several surface temperature distributions, we simulate their corresponding general relativistic X-ray spectra (assuming an isotropic, blackbody emission), and fit the latter with a single blackbody model. We find that, in some cases, the presence of a spurious ‘spectral line’ is required at a high significance level in order to obtain statistically acceptable fits, with central energy and equivalent width similar to the values typically observed. We also perform a fit to a specific object, RX J0806.4−4123, finding several surface temperature distributions able to model the observed spectrum. The explored effect is unlikely to work in all sources with detected lines, but in some cases it can indeed be responsible for the appearance of such lines. Our results enforce the idea that surface temperature anisotropy can be an important factor that should be considered and explored also in combination with more sophisticated emission models like atmospheres.