As vias de sinalização celular por meio de moléculas são um dos principais meios de controle funcional de um organismo. O entendimento das funções de moléculas sinalizadoras facilita a compreensão das vias metabólicas de um organismo, assim possibilitando uma melhor compreensão de vários eventos biológicos e também de várias doenças. A sinalização pelo ácido retinóico (AR), e seus derivados, é responsável pelo controle de várias funções, por exemplo: crescimento celular, diferenciação celular, formação da retina, desenvolvimento cardíaco e também relacionado a várias patologias como diabetes, obesidades, cânceres, e doenças cardiovasculares. A ação do ácido retinóico é controlada em dois níveis: no metabolismo de síntese/degradação e na sua utilização na sinalização para a expressão gênica. A maquinaria que controla o metabolismo inclui as enzimas de síntese do AR (aldeído desidrogenase ALDH) e as enzimas de degradação do AR (Cyp26), que controlam a distribuição espaço-temporal do AR durante a embriogênese. As ALDHs são enzimas NAD(P)+ dependentes, que oxidam uma ampla gama de aldeídos para os seus correspondentes ácidos carboxílicos, sendo ALDH1A2 a principal enzima na transformação de retinal em ácido retinóico. A maquinaria da sinalização celular por AR contém os receptores nucleares controlados por AR (RARs) que estão envolvidos com o controle da transcrição gênica. Os mecanismos de controle de expressão mais comuns são os que ocorrem na fase transcricional. Um desses mecanismos envolve proteínas que se ligam às regiões promotoras de transcrição...

A protoporfirina IX (PpIX) ocupa um lugar especial entre a família das porfirinas devido ao seu importante papel na natureza e diversas aplicações, inclusive em terapia fotodinâmica. Sob ação da luz visível, a PpIX transforma-se em fotoprodutos que podem ser citotóxicos ou fotocitotóxicos, e isto pode, de um lado, aumentar a eficiência do tratamento e, por outro lado, apresentar toxicidade no escuro, prejudicando o paciente. Como as características da sua fototransformação dependem do ambiente onde elas se encontram, tornam-se importantes os estudos dos efeitos do ambiente nas características da sua fototransformação. As propriedades espectroscópicas da PpIX sintética e endógena, extraída das glândulas harderianas de ratos da espécie Rattus novergicus albinus, e seus fotoprodutos e a atividade citotóxica foram estudadas no escuro e sob ação da luz visível em uma cultura de células malignas, em comparação com Photogem® e a porfirina TPPS4. A localização das protoporfirinas e seus fotoprodutos dentro da estrutura celular também foram observadas. Foi notado que existe uma diferença entre os espectros de absorção e fluorescência das PpIX endógena e sintética e também de seus fotoprodutos, e do grau de sua fototransformação. Associou-se estes efeitos à diferença do ambiente onde as porfirinas se encontravam...

Myosins are molecular motors associated with the actin cytoskeleton that participate in the mechanisms of cellular motility. During the development of the nervous system, migration of nerve cells to specific sites, extension of growth cones, and axonal transport are dramatic manifestations of cellular motility. We demonstrate, via immunoblots, the expression of myosin Va during early stages of embryonic development in chicks, extending from the blastocyst period to the beginning of the fetal period. The expression of myosin Va in specific regions and cellular structures of the nervous system during these early stages was determined by immunocytochemistry using a polyclonal antibody. Whole mounts of chick embryos at 24-30-h stages showed intense immunoreactivity of the neural tube in formation along its full extent. Cross-sections at these stages of development showed strong labeling in neuroepithelial cells at the basal and apical regions of the neural tube wall. Embryos at more advanced periods of development (48h and 72 h) showed distinctive immunolabeling of neuroepithelial cells, neuroblasts and their cytoplasmic extensions in the mantle layer of the stratified neural tube wall, and neuroblasts and their cytoplasmic extensions in the internal wall of the optic cup...

Myosins are molecular motors associated with the actin cytoskeleton that participate in the mechanisms of cellular motility. During the development of the nervous system, migration of nerve cells to specific sites, extension of growth cones, and axonal transport are dramatic manifestations of cellular motility. We demonstrate, via immunoblots, the expression of myosin Va during early stages of embryonic development in chicks, extending from the blastocyst period to the beginning of the fetal period. The expression of myosin Va in specific regions and cellular structures of the nervous system during these early stages was determined by immunocytochemistry using a polyclonal antibody. Whole mounts of chick embryos at 24-30-h stages showed intense immunoreactivity of the neural tube in formation along its full extent. Cross-sections at these stages of development showed strong labeling in neuroepithelial cells at the basal and apical regions of the neural tube wall. Embryos at more advanced periods of development (48h and 72 h) showed distinctive immunolabeling of neuroepithelial cells, neuroblasts and their cytoplasmic extensions in the mantle layer of the stratified neural tube wall, and neuroblasts and their cytoplasmic extensions in the internal wall of the optic cup...

The molecular mechanism of how insects recognize intruding microorganisms and parasites and distinguish them from own body structures is not well known. We explored evolutionary adaptations in an insect parasitoid host interaction to identify components that interfere with the recognition of foreign objects and cellular encapsulation. Because some parasitoids provide protection for the developing wasp in the absence of an overt suppression of the insect host defense, we analyzed the surface of eggs and symbiotic viruses for protective properties. Here we report on the molecular cloning of a 32-kDa protein (Crp32) that is one of the major protective components. It is produced in the calyx cells of the female wasp ovaries and attached to the surface of the egg and other particles including polydnaviruses. The recombinant protein confers protection to coated objects in a cellular encapsulation assay suggesting that a layer of Crp32 may prevent cellular encapsulation reactions by a local inactivation of the host defense system.

Methylglyoxal is an α-ketoaldehyde and dicarbonyl formed in cells as a side product of normal metabolism. Endogenously produced dicarbonyls, such as methylglyoxal, are involved in numerous pathogenic processes in vivo, including carcinogenesis and advanced glycation end-product formation; advanced glycation end-products are contributors to the pathophysiology of aging and chronic diabetes. Despite recent advances in understanding of the systemic effects of methylglyoxal, the full significance of this compound remains unknown. Herein we provide evidence that the majority of the methylglyoxal present in vivo is bound to biological ligands. The basis for our finding is an experimental approach that provides a measure of the bound methylglyoxal present in living systems, in this instance Chinese hamster ovary cells; with our approach, as much as 310 μM methylglyoxal was detected, 100- to 1,000-fold more than observed previously in biological systems. Several artifacts were considered before concluding that the methylglyoxal was associated with cellular structures, including phosphate elimination from triose phosphates, carbohydrate degradation under the assay conditions, and interference from the derivatizing agent used as part of the assay procedure. A major source of the recovered methylglyoxal is most probably modified cellular proteins. With methylglyoxal at about 300 μM...

Infection of fibroblast cell cultures with human cytomegalovirus (HCMV) leads to the production of significant amounts of defective enveloped particles, termed dense bodies (DB). These noninfectious structures contain major antigenic determinants which are responsible for induction of both the humoral and the cellular immune response against HCMV. We tested the hypothesis that, by virtue of their unique antigenic and structural properties, DB could induce a significant immune response in the absence of infectious virus. Mice were immunized with gradient-purified DB, which were either left untreated or subjected to sequential rounds of sonication and freeze-thawing to prevent cellular entry. Titers of neutralizing antibodies induced by DB were in a range comparable to levels present in convalescent human sera. The virus-neutralizing antibody response was surprisingly durable, with neutralizing antibodies detected 12 months following primary immunization. The HCMV-specific major histocompatibility complex class I-restricted cytolytic T-cell (CTL) response was assayed using mice transgenic for the human HLA-A2 molecule. Immunization with DB led to high levels of HCMV-specific CTL in the absence of de novo viral protein synthesis. Maximal total cytolytic activity in mice immunized with DB was nearly as efficient as the cytolytic activity induced by a standard immunization with murine cytomegalovirus. Furthermore...

Influenza virus transcription occurs in the nuclei of infected cells, where the viral genomic RNAs are complexed with a nucleoprotein (NP) to form ribonucleoprotein (RNP) structures. Prior to assembly into progeny virions, these RNPs exit the nucleus and accumulate in the cytoplasm. The mechanisms responsible for RNP export are only partially understood but have been proposed to involve the viral M1 and NS2 polypeptides. We found that the drug leptomycin B (LMB), which specifically inactivates the cellular CRM1 polypeptide, caused nuclear retention of NP in virus-infected cells, indicating a role for the CRM1 nuclear export pathway in RNP egress. However, no alteration was seen in the cellular distribution of M1 or NS2, even in the case of a mutant virus which synthesizes greatly reduced amounts of NS2. Furthermore, NP was distributed throughout the nuclei of infected cells at early times postinfection but, when retained in the nucleus at late times by LMB treatment, was redistributed to the periphery of the nucleoplasm. No such change was seen in the nuclear distribution of M1 or NS2 after drug treatment. Similar to the behavior of NP, M1 and NS2 in infected cells, LMB treatment of cells expressing each polypeptide in isolation caused nuclear retention of NP but not M1 or NS2. Conversely...

Herpes simplex virus (HSV) ICP0 can effectively activate gene expression from otherwise silent promoters contained on persisting viral genomes. However, the expression of high levels of ICP0, as from ICP4− HSV type 1 (HSV-1) vectors, results in marked toxicity. We have analyzed the results of ICP0 expressed from an E1− E4− adenovirus vector (AdS.11E4ICP0) in which ICP0 expression is controlled from the endogenous adenoviral E4 promoter. In this system, the expression level of ICP0 was reduced more than 1,000-fold relative to the level of expression from HSV-1 vectors. This low level of ICP0 did not affect cellular division or greatly perturb cellular metabolism as assessed by gene expression array analysis comparing the effects of HSV and adenovirus vector strains. However, this amount of ICP0 was sufficient to quantitatively destroy ND10 structures as measured by promyelocytic leukemia immunofluorescence. The levels of adenovirus-expressed ICP0 were sufficient to activate quiescent viral genomes in trans and promote persistent transgene expression in cis. Moreover, infection of complementing cells with AdS.11E4ICP0 promoted viral growth and resulted in a 20-fold increase in the plaquing efficiency of d109, a virus defective for all five immediate-early genes. Thus...

Autonomous parvovirus minute virus of mice (MVM) DNA replication is strictly dependent on cellular factors expressed during the S phase of the cell cycle. Here we report that MVM DNA replication proceeds in specific nuclear structures termed autonomous parvovirus-associated replication bodies, where components of the basic cellular replication machinery accumulate. The presence of DNA polymerases α and δ in these bodies suggests that MVM utilizes partially preformed cellular replication complexes for its replication. The recruitment of cyclin A points to a role for this cell cycle factor in MVM DNA replication beyond its involvement in activating the conversion of virion single-stranded DNA to the duplex replicative form.

Aromatic diamidines represent a class of DNA minor groove-binding ligands that exhibit high levels of antiparasitic activity. Since the chemotherapy for Chagas' disease is still an unsolved problem and previous reports on diamidines and related analogues show that they have high levels of activity against Trypanosoma cruzi infection both in vitro and in vivo, our present aim was to evaluate the cellular effects in vitro of three reversed amidines (DB889, DB702, and DB786) and one diguanidine (DB711) against both amastigotes and bloodstream trypomastigotes of T. cruzi, the etiological agent of Chagas ' disease. Our data show that the reversed amidines have higher levels of activity than the diguanidine, with the order of trypanocidal activities being as follows: DB889 > DB702 > DB786 > DB711. Transmission electron microscopy analysis showed that the reversed amidines induced many alterations in the nuclear morphology, swelling of the endoplasmic reticulum and Golgi structures, and consistent damage in the mitochondria and kinetoplasts of the parasites. Interestingly, in trypomastigotes treated with the reversed amidine DB889, multiple axoneme structures (flagellar microtubules) were noted. Flow cytometry analysis confirmed that the treated parasites presented an important loss of the mitochondrial membrane potential...

We revisit the problem of finding the conditions under which synchronous probabilistic cellular automata indexed by the line $\mathbb{Z}$, or the periodic line $\cyl{n}$, depending on 2 neighbours, admit as invariant distribution the law of a space-indexed Markov chain. Our advances concerns PCA defined on a finite alphabet, where most of existing results concern size 2 alphabet. A part of the paper is also devoted to the comparison of different structures ($\mathbb{Z}$, $\cyl{n}$, and also some structures constituted with two consecutive lines of the space time diagram) with respect to the property to possess a Markovian invariant distribution.

Quantitative analysis of the dynamics of tiny cellular and sub-cellular structures, known as particles, in time-lapse cell microscopy sequences requires the development of a reliable multi-target tracking method capable of tracking numerous similar targets in the presence of high levels of noise, high target density, complex motion patterns and intricate interactions. In this paper, we propose a framework for tracking these structures based on the random finite set Bayesian filtering framework. We focus on challenging biological applications where image characteristics such as noise and background intensity change during the acquisition process. Under these conditions, detection methods usually fail to detect all particles and are often followed by missed detections and many spurious measurements with unknown and time-varying rates. To deal with this, we propose a bootstrap filter composed of an estimator and a tracker. The estimator adaptively estimates the required meta parameters for the tracker such as clutter rate and the detection probability of the targets, while the tracker estimates the state of the targets. Our results show that the proposed approach can outperform state-of-the-art particle trackers on both synthetic and real data in this regime.

Voronoi tessellations of scale-invariant fractal sets are characterized by topological and metrical properties that are significantly different from those of natural cellular structures. As an example we analyze Voronoi diagrams of intermittent particle distributions generated by a directed percolation process in (2+1) dimensions. We observe that the average area of a cell increases much faster with the number of its neighbours than in natural cellular structures where Lewis' law predicts a linear behaviour. We propose and numerically verify a universal scaling law that relates shape and size of the cells in scale-invariant tessellations. A novel exponent, related to the topological properties of the tessellation, is introduced and estimated numerically.; Comment: RevTeX, 5 pages, including 3 encapsulated postscript figures

Independent of the technology, it is generally expected that future nanoscale devices will be built from vast numbers of densely arranged devices that exhibit high failure rates. Other than that, there is little consensus on what type of technology and computing architecture holds most promises to go far beyond today's top-down engineered silicon devices. Cellular automata (CA) have been proposed in the past as a possible class of architectures to the von Neumann computing architecture, which is not generally well suited for future parallel and fine-grained nanoscale electronics. While the top-down engineered semi-conducting technology favors regular and locally interconnected structures, future bottom-up self-assembled devices tend to have irregular structures because of the current lack precise control over these processes. In this paper, we will assess random dynamical networks, namely Random Boolean Networks (RBNs) and Random Threshold Networks (RTNs), as alternative computing architectures and models for future information processing devices. We will illustrate that--from a theoretical perspective--they offer superior properties over classical CA-based architectures, such as inherent robustness as the system scales up, more efficient information processing capabilities...

Several cellular automata (CA) models have been developed to simulate self-organization of multiple levels of structures. However, they do not obey microscopic reversibility and conservation laws. In this paper, we describe the construction of a reversible lattice molecular automata (RLMA) model, which simulates molecular interaction and self-organization of higher-order structures. The model's strict reversibility entails physically relevant conservation laws, and thus opens a way to precise application and validation of the methods from statistical physics in studying the necessary conditions for such multiple levels of self-organization.; Comment: 29 pages, 20 figures

We prove existence results a la Jeff Smith for left-induced model category structures, of which the injective model structure on a diagram category is an important example. We further develop the notions of fibrant generation and Postnikov presentation from Hess, which are dual to a weak form of cofibrant generation and cellular presentation. As examples, for k a field and H a differential graded Hopf algebra over k, we produce a left-induced model structure on augmented H-comodule algebras and show that the category of bounded below chain complexes of finite-dimensional k-vector spaces has a Postnikov presentation. To conclude, we investigate the fibrant generation of (generalized) Reedy categories. In passing, we also consider cofibrant generation, cellular presentation, and the small object argument for Reedy diagrams.; Comment: 33 pages; v2 fixes an error in the construction of the Postnikov presentation in section 3 and contains several minor improvements suggested by the referee. To appear in the Proceedings of the August 2013 "Women in Topology" workshop at BIRS, which will be published by Contemporary Mathematics

RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are.; Abstract Background Biolistic transfection is proving an increasingly popular method of incorporating DNA or RNA into cells that are difficult to transfect using traditional methods. The technique routinely uses 'microparticles', which are ~1 ?m diameter projectiles, fired into tissues using pressurised gas. These microparticles are efficient at delivering DNA into cells, but cannot efficiently transfect small cells and may cause significant tissue damage, thus limiting their potential usefulness. Here we describe the use of 40 nm diameter projectiles - nanoparticles - in biolistic transfections to determine if they are a suitable alternative to microparticles. Results Examination of transfection efficiencies in HEK293 cells, using a range of conditions including different DNA concentrations and different preparation procedures...

R loop hybridizations and electron microscopy have been used to determine cellular RNA concentrations for cloned genes. In plasmid DNA sequence excess, all the complementary RNA is driven into R loop structures that can be assayed by electron microscopy. To determine the concentration of a particular poly(A)+ RNA, plasmid DNA crosslinked once every 2000-5000 base pairs with trioxsalen and UV light is hybridized in DNA sequence excess to various known amounts of total poly(A)+ RNA, and the R loops are stabilized by treatment with glyoxal. If necessary, excess nonhybridized RNA is removed by Sepharose 2B chromatography, which enables the visualization of less abundant transcripts. Reconstruction experiments demonstrated that electron microscopic determination of the fraction of plasmid DNA molecules containing specific RNA loops gives accurate values of specific RNA weight fractions or concentrations in the total poly(A)+ RNA populations. These methods were also used to determine the concentrations of five RNA species complementary to sequences on TRT3, a recombinant DNA plasmid containing yeast histone 2A and 2B genes and three other nonhistone genes. The methods described allow one to visualize the R loop structures for both abundant and nonabundant transcripts and to estimate concentrations of these RNA species simply by determining the fraction of DNA containing R loops.