Transcriptomics and systems biology analysis in identification of specific pathways involved in cacao resistance and susceptibility to witches' broom disease

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Theoretical study on the molecular mechanism for the reaction of VO2+ with C2H4

The complex reaction between VO2+ ((1)A(1)/(3)A) and C2H4 (Ag-1(g)/(3)A(1)) to yield VO+ ((1)Delta/(3)Sigma) and CH3CHO ('A'/(3)A) has been studied by means of B3LYP/6-31G* and B3LYP/6-311G(2d,p) calculations. The structures of all reactants, products, intermediates, and transition structures of this reaction have been optimized and characterized at the fundamental singlet and first excited triplet electronic states. Crossing points are localized, and possible spin inversion processes are discussed by means of the intrinsic reaction coordinate approach. Relevant stationary points along the most favorable reaction pathways have been studied at the CCSD/6-311G(2d,p)//B3LYP/6-311G(2d,p) calculation level. The theoretical results allow the development of thermodynamic and kinetic arguments about the reaction pathways of the title process. In the singlet state, the first step is the barrierless obtention of a reactant complex associated with the formation of a V-C bond, while in the triplet state a three-membered ring addition complex with the V bonded to the two C atoms is obtained. Similar behavior is found in the exit channels: the product complexes can be formed from isolated products without barriers. The reactant and product complexes are the most stable stationary points in the singlet and triplet electronic states. From the singlet state reactant complex, two reaction pathways are posssible to reach the triplet state product complex. (i) A mechanism in which a hydrogen transfer process is the first and rate limiting step and the second step is an oxygen transfer between vanadium and carbon atoms with a concomitant change in the spin state. The crossing point between singlet and triplet spin states is not kinetically relevant because it takes place at a later stage occurring in the exit channel. (ii) A mechanism in which the first stage renders a four-membered ring between vanadyl cation and the ethylene fragment and an oxygencarbon bond is formed; on going from this minimum to the second transition structure, associated with a carbon-vanadium bond breaking process, the crossing point between singlet and triplet spin states is reached. The final step is the hydrogen transfer between both carbon atoms to yield the product complex. In this case the spin change opens a lower barrier pathway. The transition structures with larger values of relative energies for both reactive channels of VO2+ ((1)A(1)) + C2H4 (Ag-1) --> VO+ ((3)Sigma) + CH3CHO ((1)A') present similar energies, and the two reaction pathways can be considered as competitive.

Expression of genes related to apoptosis, cell cycle and signaling pathways are independent of TP53 status in urinary bladder cancer cells

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Periacinar Retraction Clefting in Nonneoplastic and Neoplastic Prostatic Glands: Artifact or Molecular Involvement

A space between neoplastic acini and prostatic stroma is not rare and studies have interpreted this as an artifact, considering the absence of endothelial cells indicating vascular invasion. Thus, the aims of this work were to characterize and correlate the occurrence and extent of retraction clefting with the reactivities of alpha and beta dystroglycan (alpha DG, beta DG), laminin, matrix metalloproteinase 2 (MMP-2), p63, insulin-like growth factor 1(IGF-1), vimentin, and fibroblast growth factor 2 (FGF-2). The study was based on nonneoplastic and neoplastic prostatic tissues obtained from necropsies and retropubic radical prostatectomies. The results showed that periacinar retraction clefting was significantly more frequent in prostatic carcinoma samples than in normal prostatic acini. Most of the neoplastic acini (72.0%) showed retraction clefting of more than 50% of circumference, which were significantly more frequent in Gleason score 7 and 6. Decreased collagen and reticular and elastic fibers were verified in the stroma around neoplastic acini. Weak and discontinuous alpha DG, beta DG, and laminin immunoreactivities and intensified MMP-2, vimentin, IGF-1 and FGF-2 immunoreactivities were verified in the neoplastic acini; p63 immunoreactivity was negative in all carcinomas. Thus, these findings showed that the lack of epithelial basal cells, DGs, and laminin and increased MMP-2, IGF-1, and FGF-7 could be considered important pathways in periacinar retraction occurrence. This study demonstrated the origin of and the biological mechanisms responsible for periacinar retraction clefting in prostatic carcinoma.

Analysis of plant LTR-retrotransposons at the fine-scale family level reveals individual molecular patterns

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Molecular models of protein targets from Mycobacterium tuberculosis

Structural characterization of enzymes that belong to microbial metabolic pathways is very important for structure-based drug design since some of these proteins may be present in the bacterial genome, but absent in humans. Thus, metabolic pathways became potential targets for drug design. The motivation of this work is the fact that Mycobacterium tuberculosis is the cause of the deaths of millions of people in the world, so that the structural characterization of protein targets to propose new drugs has become essential. DBMODELING is a relational database, created to highlight the importance of methods of molecular modeling applied to the Mycobacterium tuberculosis genome with the aim of proposing protein-ligand docking analysis. There are currently more than 300 models for proteins from Mycobacterium tuberculosis genome in the database. The database contains a detailed description of the reaction catalyzed by each enzyme and their atomic coordinates. Information about structures, a tool for animated gif image, a table with a specification of the metabolic pathway, modeled protein, inputs used in modeling, and analysis methods used in this project are available in the database for download. The search tool can be used for reseachers to find specific pathways or enzymes.

The use of biodiversity as source of new chemical entities against defined molecular targets for treatment of malaria, tuberculosis, and T-cell mediated diseases: a review

The modern approach to the development of new chemical entities against complex diseases, especially the neglected endemic diseases such as tuberculosis and malaria, is based on the use of defined molecular targets. Among the advantages, this approach allows (i) the search and identification of lead compounds with defined molecular mechanisms against a defined target (e.g. enzymes from defined pathways), (ii) the analysis of a great number of compounds with a favorable cost/benefit ratio, (iii) the development even in the initial stages of compounds with selective toxicity (the fundamental principle of chemotherapy), (iv) the evaluation of plant extracts as well as of pure substances. The current use of such technology, unfortunately, is concentrated in developed countries, especially in the big pharma. This fact contributes in a significant way to hamper the development of innovative new compounds to treat neglected diseases. The large biodiversity within the territory of Brazil puts the country in a strategic position to develop the rational and sustained exploration of new metabolites of therapeutic value. The extension of the country covers a wide range of climates, soil types, and altitudes, providing a unique set of selective pressures for the adaptation of plant life in these scenarios. Chemical diversity is also driven by these forces, in an attempt to best fit the plant communities to the particular abiotic stresses, fauna, and microbes that co-exist with them. Certain areas of vegetation (Amazonian Forest, Atlantic Forest, Araucaria Forest, Cerrado-Brazilian Savanna, and Caatinga) are rich in species and types of environments to be used to search for natural compounds active against tuberculosis, malaria, and chronic-degenerative diseases. The present review describes some strategies to search for natural compounds, whose choice can be based on ethnobotanical and chemotaxonomical studies, and screen for their ability to bind to immobilized drug targets and to inhibit their activities. Molecular cloning, gene knockout, protein expression and purification, N-terminal sequencing, and mass spectrometry are the methods of choice to provide homogeneous drug targets for immobilization by optimized chemical reactions. Plant extract preparations, fractionation of promising plant extracts, propagation protocols and definition of in planta studies to maximize product yield of plant species producing active compounds have to be performed to provide a continuing supply of bioactive materials. Chemical characterization of natural compounds, determination of mode of action by kinetics and other spectroscopic methods (MS, X-ray, NMR), as well as in vitro and in vivo biological assays, chemical derivatization, and structure-activity relationships have to be carried out to provide a thorough knowledge on which to base the search for natural compounds or their derivatives with biological activity.

TNF-alpha acts in the hypothalamus inhibiting food intake and increasing the respiratory quotient - Effects on leptin and insulin signaling pathways

Acting in the hypothalamus, tumor necrosis factor-alpha (TNF-alpha) produces a potent anorexigenic effect. However, the molecular mechanisms involved in this phenomenon are poorly characterized. In this study, we investigate the capacity of TNF-alpha to activate signal transduction in the hypothalamus through elements of the pathways employed by the anorexigenic hormones insulin and leptin. High dose TNF-a promotes a reduction of 25% in 12 h food intake, which is an inhibitory effect that is marginally inferior to that produced by insulin and leptin. In addition, high dose TNF-a increases body temperature and respiratory quotient, effects not reproduced by insulin or leptin. TNF-alpha, predominantly at the high dose, is also capable of activating canonical pro-inflammatory signal transduction in the hypothalamus, inducing JNK, p38, and NF kappa B, which results in the transcription of early responsive genes and expression of proteins of the SOCS family. Also, TNF-a activates signal transduction through JAK-2 and STAT-3, but does not activate signal transduction. through early and intermediary elements of the insulin/leptin signaling pathways such as IRS-2, Akt, ERK and FOXO1. When co-injected with insulin or leptin, TNF-a, at both high and low doses, partially impairs signal transduction through IRS-2, Akt, ERK and FOXO1 but not through JAK-2 and STAT-3. This effect is accompanied by the partial inhibition of the anorexigenic effects of insulin and leptin, when the low, but not the high dose of TNF-alpha is employed. In conclusion, TNF-alpha, on a dose-dependent way, modulates insulin and leptin signaling and action in the hypothalamus. (c) Published by Elsevier B.V.

Interaction of pathogenic fungi with host cells: Molecular and cellular approaches

This review provides an overview of several molecular and cellular approaches that are likely to supply insights into the host-fungus interaction. Fungi present intra- and/or extracellular host-parasite interfaces, the parasitism phenomenon being dependent on complementary surface molecules. The entry of the pathogen into the host cell is initiated by the fungus adhering to the cell surface, which generates an uptake signal that may induce its cytoplasmatic internalization. Furthermore, microbial pathogens use a variety of their surface molecules to bind to host extracellular matrix (ECM) components to establish an effective infection. on the other hand, integrins mediate the tight adhesion of cells to the ECM at sites referred to as focal adhesions and also play a role in cell signaling. The phosphorylation process is an important mechanism of cell signaling and regulation; it has been implicated recently in defense strategies against a variety of pathogens that alter host-signaling pathways in order to facilitate their invasion and survival within host cells. The study of signal transduction pathways in virulent fungi is especially important in view of their putative role in the regulation of pathogenicity. This review discusses fungal adherence, changes in cytoskeletal organization and signal transduction in relation to host-fungus interaction. (c) 2005 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.

miRNApath: A database of miRNAs, target genes and metabolic pathways

MicroRNAs (miRNAs) are small non-coding RNAs that regulate target gene expression and hence play important roles in metabolic pathways. Recent studies have evidenced the interrelation of miRNAs with cell proliferation, differentiation, development, and diseases. Since they are involved in gene regulation, they are intrinsically related to metabolic pathways. This leads to questions that are particularly interesting for investigating medical and laboratorial applications. We developed an miRNApath online database that uses miRNA target genes to link miRNAs to metabolic pathways. Currently, databases about miRNA target genes (DIANA miRGen), genomic maps (miRNAMap) and sequences (miRBase) do not provide such correlations. Additionally, miRNApath offers five search services and a download area. For each search, there is a specific type of input, which can be a list of target genes, miRNAs, or metabolic pathways, which results in different views, depending upon the input data, concerning relationships between the target genes, miRNAs and metabolic pathways. There are also internal links that lead to a deeper analysis and cross-links to other databases with more detailed information. miRNApath is being continually updated and is available at http://lgmb.fmrp.usp.br/mirnapath. ©FUNPEC-RP.

Cellular and molecular mechanisms of apoptosis in skeletal muscle atrophy

Apoptosis is necessary for maintaining the integrity of proliferative tissues, such as epithelial cells of the gastrointestinal and integumentary systems. The role of apoptosis in post-mitotic tissues, such as skeletal muscle, is less well defined, but several lines of evidence suggest that it occurs in both myofiber and other interstitial muscle cell types. Apoptosis of myonuclei likely contributes to the loss of muscle mass, but the mechanisms underlying this process are largely unknown. Caspase-dependent as well as caspase-independent pathways have been implicated, and the mode by which atrophy is induced likely determines the apoptotic mechanisms that are utilized. It remains to be determined whether a decrease in apoptosis will alleviate atrophy and distinct research strategies may be required to clarify the different causes of skeletal muscle mass loss. In this review, it was also speculated that apoptosis is a normal regulatory process that the myofiber can use to reduce the number of nuclear domains, thus ensuring optimal cell functions according to the mechanical load imposed on the muscle. ©FUNPEC-RP.

Dynamics of bone healing after osteotomy with piezosurgery or conventional drilling - histomorphometrical, immunohistochemical, and molecular analysis

Background: Piezosurgery is an osteotomy system used in medical and dental surgery. Many studies have proven clinical advantages of piezosurgery in terms of quality of cut, maneuverability, ease of use, and safety. However, few investigations have tested its superiority over the traditional osteotomy systems in terms of dynamics of bone healing. Therefore, the aim of this study was to evaluate the dynamics of bone healing after osteotomies with piezosurgery and to compare them with those associated to traditional bone drilling.Methods: One hundred and ten rats were divided into two groups with 55 animals each. The animals were anesthetized and the tibiae were surgically exposed to create defects 2 mm in diameter by using piezosurgery (Piezo group) and conventional drilling (Drill group). Animals were sacrificed at 3, 7, 14, 30 and 60 days post-surgery. Bone samples were collected and processed for histological, histomorphometrical, immunohistochemical, and molecular analysis. The histological analysis was performed at all time points (n = 8) whereas the histomorphometrical analysis was performed at 7, 14, 30 and 60 days post-surgery (n = 8). The immunolabeling was performed to detect Vascular Endothelial Growth Factor (VEGF), Caspase-3 (CAS-3), Osteoprotegerin (OPG), Receptor Activator of Nuclear Factor kappa-B Ligand (RANKL), and Osteocalcin (OC) at 3, 7, and 14 days (n = 3). For the molecular analysis, animals were sacrificed at 3, 7 and 14 days, total RNA was collected, and quantification of the expression of 21 genes related to BMP signaling, Wnt signaling, inflammation, osteogenenic and apoptotic pathways was performed by qRT-PCR (n = 5).Results: Histologically and histomorphometrically, bone healing was similar in both groups with the exception of a slightly higher amount of newly formed bone observed at 30 days after piezosurgery (p < 0.05). Immunohistochemical and qRT-PCR analyses didn't detect significant differences in expression of all the proteins and most of the genes tested.Conclusions: Based on the results of our study we conclude that in a rat tibial bone defect model the bone healing dynamics after piezosurgery are comparable to those observed with conventional drilling. © 2013 Esteves et al.; licensee BioMed Central Ltd.

Caracterização morfológica, bioquímica e molecular do músculo esquelético sóleo de ratos espontaneamente hipertensos com insuficiência cardíaca

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Atividade alelopática de Copaifera langsdorffii DESF: abordagem fitoquímica e molecular

Pós-graduação em Ciências Biológicas (Botânica) - IBB

Cellular and molecular studies of the effects of a selective COX-2 inhibitor celecoxib in the cardiac cell line H9c2 and their correlation with death mechanisms

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)