A VONTADE SOB O IMPACTO DA PÓS-MODERNIDADE (A FÉ INTERPELANDO A FRAGMENTAÇÃO DA VONTADE)

Este trabalho investiga uma das formas pelas quais a condição contemporânea, assim chamada pós-moderna, exerce significativo impacto sobre a vontade humana. Numa perspectiva fenomenológica e existencial, apoiada primordialmente no pensamento de Paul Ricoeur e Paul Tillich, esta abordagem busca identificar a relação entre a condição pós-moderna e a crise contemporânea da vontade que, como hipótese de tese, encontra-se vinculada à forma pela qual é estabelecida a relação entre vontade e desejo. A pós-modernidade é analisada a partir de seu desenvolvimento sócio-cultural desde a década de 1930 e em interlocução com os pensamentos de Jean-François Lyotard e Jean Baudrillard de forma a evidenciar o desenvolvimento desta relação. Algumas perspectivas filosófico-teológicas são exploradas a partir da situação de impacto aqui em questão. São tentativas de contribuir para que a vontade redescubra seu caráter integralizador e possa considerá-lo tão imprescindível quanto seu ímpeto expansionista e dissipador, uma vez que ambos os movimentos são partes integrantes da coragem e do poder-de-ser que a fundamentam.(AU)

Mapping the active site in vasoactive intestinal peptide to a core of four amino acids: Neuroprotective drug design

The understanding of the molecular mechanisms leading to peptide action entails the identification of a core active site. The major 28-aa neuropeptide, vasoactive intestinal peptide (VIP), provides neuroprotection. A lipophilic derivative with a stearyl moiety at the N-terminal and norleucine residue replacing the Met-17 was 100-fold more potent than VIP in promoting neuronal survival, acting at femtomolar–picomolar concentration. To identify the active site in VIP, over 50 related fragments containing an N-terminal stearic acid attachment and an amidated C terminus were designed, synthesized, and tested for neuroprotective properties. Stearyl-Lys-Lys-Tyr-Leu-NH2 (derived from the C terminus of VIP and the related peptide, pituitary adenylate cyclase activating peptide) captured the neurotrophic effects offered by the entire 28-aa parent lipophilic derivative and protected against β-amyloid toxicity in vitro. Furthermore, the 4-aa lipophilic peptide recognized VIP-binding sites and enhanced choline acetyltransferase activity as well as cognitive functions in Alzheimer’s disease-related in vivo models. Biodistribution studies following intranasal administration of radiolabeled peptide demonstrated intact peptide in the brain 30 min after administration. Thus, lipophilic peptide fragments offer bioavailability and stability, providing lead compounds for drug design against neurodegenerative diseases.

Identification of the binding site for Gqα on its effector Bruton’s tyrosine kinase

Heterotrimeric G proteins and tyrosine kinases are two major cellular signal transducers. Although G proteins are known to activate tyrosine kinases, the activation mechanism is not clear. Here, we demonstrate that G protein Gqα binds directly to the nonreceptor Bruton’s tyrosine kinase (Btk) to a region composed of a Tec-homology (TH) domain and a sarcoma virus tyrosine kinase (Src)-homology 3 (SH3) domain both in vitro and in vivo. Only active GTP-bound Gqα, not inactive GDP-bound Gqα, can bind to Btk. Mutations of Btk that disrupt its ability to bind Gqα also eliminate Btk stimulation by Gqα, suggesting that this interaction is important for Btk activation. Remarkably, the structure of this TH (including a proline-rich sequence) -SH3 fragment of the Btk family of tyrosine kinases shows an intramolecular interaction. Furthermore, the crystal structure of the Src family of tyrosine kinases reveals that the intramolecular interaction of SH3 and its ligand is the major determining factor keeping the kinase inactive. Thus, we propose an activation model that entails binding of Gqα to the TH-SH3 region, thereby disrupting the TH-SH3 intramolecular interaction and activating Btk.

Two-dimensional IR correlation spectroscopy: Sequential events in the unfolding process of the λ Cro-V55C repressor protein

A question often posed in protein folding/unfolding studies is whether the process is fully cooperative or whether it contains sequential elements. To address this question, one needs tools capable of resolving different events. It seems that, at least in certain cases, two-dimensional (2D) IR correlation spectroscopy can provide answers to this question. To illustrate this point, we have turned to the Cro-V55C dimer of the λ Cro repressor, a protein known to undergo thermal unfolding in two discrete steps through a stable equilibrium intermediate. The secondary structure of this intermediate is compatible with that of a partially unfolded protein and involves a reorganization of the N terminus, whereas the antiparallel β-ribbon formed by the C-terminal part of each subunit remains largely intact. To establish whether the unfolding process involves sequential events, we have performed a 2D correlation analysis of IR spectra recorded over the temperature range of 20–95°C. The 2D IR correlation analysis indeed provides evidence for a sequential formation of the stable intermediate, which is created in three (closely related) steps. A first step entails the unfolding of the short N-terminal β-strand, followed by the unfolding of the α-helices in a second step, and the third step comprises the reorganization of the remaining β-sheet and of some unordered segments in the protein. The complete unfolding of the stable intermediate at higher temperatures also undergoes sequential events that ultimately end with the breaking of the H bonds between the two β-strands at the dimer interface.

Characterization of Schizosaccharomyces pombe RNA triphosphatase

RNA triphosphatase catalyzes the first step in mRNA cap formation which entails the cleavage of the β–γ phosphoanhydride bond of triphosphate-terminated RNA to yield a diphosphate end that is then capped with GMP by RNA guanylyltransferase. Here we characterize a 303 amino acid RNA triphosphatase (Pct1p) encoded by the fission yeast Schizosaccharomyces pombe. Pct1p hydrolyzes the γ phosphate of triphosphate-terminated poly(A) in the presence of magnesium. Pct1p also hydrolyzes ATP to ADP and Pi in the presence of manganese or cobalt (Km = 19 µM ATP; kcat = 67 s–1). Hydrolysis of 1 mM ATP is inhibited with increasing potency by inorganic phosphate (I0.5 = 1 mM), pyrophosphate (I0.5 = 0.4 mM) and tripolyphosphate (I0.5 = 30 µM). Velocity sedimentation indicates that Pct1p is a homodimer. Pct1p is biochemically and structurally similar to the catalytic domain of Saccharomyces cerevisiae RNA triphosphatase Cet1p. Mechanistic conservation between Pct1p and Cet1p is underscored by a mutational analysis of the putative metal-binding site of Pct1p. Pct1p is functional in vivo in S.cerevisiae in lieu of Cet1p, provided that it is coexpressed with the S.pombe guanylyltransferase. Pct1p and other yeast RNA triphosphatases are completely unrelated, mechanistically and structurally, to the metazoan RNA triphosphatases, suggesting an abrupt evolutionary divergence of the capping apparatus during the transition from fungal to metazoan species.

Neural fate of seen and unseen faces in visuospatial neglect: A combined event-related functional MRI and event-related potential study

To compare neural activity produced by visual events that escape or reach conscious awareness, we used event-related MRI and evoked potentials in a patient who had neglect and extinction after focal right parietal damage, but intact visual fields. This neurological disorder entails a loss of awareness for stimuli in the field contralateral to a brain lesion when stimuli are simultaneously presented on the ipsilateral side, even though early visual areas may be intact, and single contralateral stimuli may still be perceived. Functional MRI and event-related potential study were performed during a task where faces or shapes appeared in the right, left, or both fields. Unilateral stimuli produced normal responses in V1 and extrastriate areas. In bilateral events, left faces that were not perceived still activated right V1 and inferior temporal cortex and evoked nonsignificantly reduced N1 potentials, with preserved face-specific negative potentials at 170 ms. When left faces were perceived, the same stimuli produced greater activity in a distributed network of areas including right V1 and cuneus, bilateral fusiform gyri, and left parietal cortex. Also, effective connectivity between visual, parietal, and frontal areas increased during perception of faces. These results suggest that activity can occur in V1 and ventral temporal cortex without awareness, whereas coupling with dorsal parietal and frontal areas may be critical for such activity to afford conscious perception.

Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands.

Precise mapping of DNA methylation patterns in CpG islands has become essential for understanding diverse biological processes such as the regulation of imprinted genes, X chromosome inactivation, and tumor suppressor gene silencing in human cancer. We describe a new method, MSP (methylation-specific PCR), which can rapidly assess the methylation status of virtually any group of CpG sites within a CpG island, independent of the use of methylation-sensitive restriction enzymes. This assay entails initial modification of DNA by sodium bisulfite, converting all unmethylated, but not methylated, cytosines to uracil, and subsequent amplification with primers specific for methylated versus unmethylated DNA. MSP requires only small quantities of DNA, is sensitive to 0.1% methylated alleles of a given CpG island locus, and can be performed on DNA extracted from paraffin-embedded samples. MSP eliminates the false positive results inherent to previous PCR-based approaches which relied on differential restriction enzyme cleavage to distinguish methylated from unmethylated DNA. In this study, we demonstrate the use of MSP to identify promoter region hypermethylation changes associated with transcriptional inactivation in four important tumor suppressor genes (p16, p15, E-cadherin, and von Hippel-Lindau) in human cancer.

High mobility group I(Y)-like DNA-binding domains on a bacterial transcription factor.

The bacterium Myxococcus xanthus responds to blue light by producing carotenoids. It also responds to starvation conditions by developing fruiting bodies, where the cells differentiate into myxospores. Each response entails the transcriptional activation of a separate set of genes. However, a single gene, carD, is required for the activation of both light- and starvation-inducible genes. Gene carD has now been sequenced. Its predicted amino acid sequence includes four repeats of a DNA-binding domain present in mammalian high mobility group I(Y) proteins and other nuclear proteins from animals and plants. Other peptide stretches on CarD also resemble functional domains typical of eukaryotic transcription factors, including a very acidic region and a leucine zipper. High mobility group yI(Y) proteins are known to bind the minor groove of A+T-rich DNA. CarD binds in vitro an A+T-rich element that is required for the proper operation of a carD-dependent promoter in vivo.

Active barnase variants with completely random hydrophobic cores.

The central structural feature of natural proteins is a tightly packed and highly ordered hydrophobic core. If some measure of exquisite, native-like core packing is necessary for enzymatic function, this would constitute a significant obstacle to the development of novel enzymes, either by design or by natural or experimental evolution. To test the minimum requirements for a core to provide sufficient structural integrity for enzymatic activity, we have produced mutants of the ribonuclease barnase in which 12 of the 13 core residues have together been randomly replaced by hydrophobic alternatives. Using a sensitive biological screen, we find that a strikingly high proportion of these mutants (23%) retain enzymatic activity in vivo. Further substitution at the 13th core position shows that a similar proportion of completely random hydrophobic cores supports enzyme function. Of the active mutants produced, several have no wild-type core residues. These results imply that hydrophobicity is nearly a sufficient criterion for the construction of a functional core and, in conjunction with previous studies, that refinement of a crudely functional core entails more stringent sequence constraints than does the initial attainment of crude core function. Since attainment of crude function is the critical initial step in evolutionary innovation, the relatively scant requirements contributed by the hydrophobic core would greatly reduce the initial hurdle on the evolutionary pathway to novel enzymes. Similarly, experimental development of novel functional proteins might be simplified by limiting core design to mere specification of hydrophobicity and using iterative mutation-selection to optimize core structure.

Resolution of Holliday junctions by eukaryotic DNA topoisomerase I.

The Holliday junction, a key intermediate in both homologous and site-specific recombination, is generated by the reciprocal exchange of single strands between two DNA duplexes. Resolution of the junctions can occur in two directions with respect to flanking markers, either restoring the parental DNA configuration or generating a genetic crossover. Recombination can be regulated, in principle, by factors that influence the directionality of the resolution step. We demonstrate that the vaccinia virus DNA topoisomerase, a eukaryotic type I enzyme, catalyzes resolution of synthetic Holliday junctions in vitro. The mechanism entails concerted transesterifications at two recognition sites, 5'-CCCTT decreases, that are opposed within a partially mobile four-way junction. Cruciforms are resolved unidirectionally and with high efficiency into two linear duplexes. These findings suggest a model whereby type I topoisomerases may either promote or suppress genetic recombination in vivo.

Os efeitos previdenciários do reconhecimento de vínculo empregatício pela justiça do trabalho

A Constituição da República, de 1988, previu em seu artigo 201, que a Previdência Social seria organizada sob a forma de regime geral, de caráter contributivo e de filiação obrigatória. Em regra, o trabalho remunerado enseja a filiação obrigatória e automática do trabalhador, assim como o surgimento de sua obrigação de contribuir para o custeio das prestações previdenciárias. Caso o empregador não registre o empregado e promova o recolhimento das contribuições previdenciárias, o trabalhador poderá ter limitada ou excluída sua proteção previdenciária. Mesmo reconhecido o vínculo de emprego no processo do trabalho, o Instituto Nacional da Seguridade Social (INSS) condiciona o aproveitamento previdenciário desse tempo de trabalho e de contribuição à apresentação de início de prova material. Essa exigência, por vezes, cria situação de contradição: há sentença trabalhista de reconhecimento de vínculo de emprego, com execução e recolhimento de contribuições previdenciárias, mas o INSS não reconhece o tempo de contribuição correspondente e nega ao trabalhador proteção previdenciária. A presente dissertação analisa se o reconhecimento de vínculo empregatício pela Justiça do Trabalho é suficiente para que se reconheça o direito do trabalhador à proteção previdenciária, partindo da premissa que o segurado empregado apenas tem de demonstrar sua filiação, não sendo prejudicado pelo descumprimento de obrigações previdenciárias de seu empregador.

Análise de grandezas cinemáticas e dinâmicas inerentes à hemiparesia através da descoberta de conhecimento em bases de dados

Em virtude de uma elevada expectativa de vida mundial, faz-se crescente a probabilidade de ocorrer acidentes naturais e traumas físicos no cotidiano, o que ocasiona um aumento na demanda por reabilitação. A terapia física, sob o paradigma da reabilitação robótica com serious games, oferece maior motivação e engajamento do paciente ao tratamento, cujo emprego foi recomendado pela American Heart Association (AHA), apontando a mais alta avaliação (Level A) para pacientes internados e ambulatoriais. No entanto, o potencial de análise dos dados coletados pelos dispositivos robóticos envolvidos é pouco explorado, deixando de extrair informações que podem ser de grande valia para os tratamentos. O foco deste trabalho consiste na aplicação de técnicas para descoberta de conhecimento, classificando o desempenho de pacientes diagnosticados com hemiparesia crônica. Os pacientes foram inseridos em um ambiente de reabilitação robótica, fazendo uso do InMotion ARM, um dispositivo robótico para reabilitação de membros superiores e coleta dos dados de desempenho. Foi aplicado sobre os dados um roteiro para descoberta de conhecimento em bases de dados, desempenhando pré-processamento, transformação (extração de características) e então a mineração de dados a partir de algoritmos de aprendizado de máquina. A estratégia do presente trabalho culminou em uma classificação de padrões com a capacidade de distinguir lados hemiparéticos sob uma precisão de 94%, havendo oito atributos alimentando a entrada do mecanismo obtido. Interpretando esta coleção de atributos, foi observado que dados de força são mais significativos, os quais abrangem metade da composição de uma amostra.

Exposição a substâncias organocloradas em São Paulo: níveis séricos em doadores de sangue e fatores associados

A utilização de pesticidas organoclorados é motivo de preocupação das entidades ligadas à área de saúde em todo o mundo. Apesar de as formas de contaminação serem bem conhecidas, não há um controle eficaz na fiscalização do seu uso no Brasil. Sabe-se que altos níveis séricos destes compostos nos organismos de seres humanos e animais acarretam sérios problemas de saúde. Tendo em vista essa realidade, foi realizado, em 2009, o Projeto Piloto do I Inquérito Nacional de Populações Expostas a Substâncias Químicas, cujo subprojeto \"doadores de sangue\" teve como objetivo mensurar as concentrações de substâncias químicas no sangue de 547 residentes da região metropolitana de São Paulo, dentre elas os pesticidas organoclorados. Este trabalho teve como objetivos avaliar as concentrações dos pesticidas hexaclorobenzeno (HCB), alfa-HCH, ?-HCH, beta-HCH, beta-HCH, heptacloro, heptacloro epóxido, dieldrin, mirex, o,p\'-DDT, p,p\'-DDT, o,p\'-DDE, p,p\'-DDE, o,p\'-DDD e p,p\'-DDD nesta população e compará-las com as encontradas em outros países e determinar fatores associados aos níveis mais elevados destas substâncias. O método analítico utilizado foi de cromatografia a gás. Os resultados deste estudo indicam que a população adulta de São Paulo não está exposta a níveis preocupantes de pesticidas organoclorados, pois dentre os compostos analisados, apenas o beta-HCH e o p,p\'-DDE tiveram um número significante de amostras acima do limite de quantificação, 10,7% e 31,2% das amostras respectivamente. Quando utilizada a metade do limite de quantificação para substituir os valores abaixo do limite de quantificação do método, o valor médio encontrado para o beta-HCH foi de 0,028 ug/dL e para o p,p\'-DDE foi de 0,045 ug/dL. Este estudo propôs dois modelos multivariados para explicar os fatores associados aos compostos beta-HCH e p,p\'-DDE no sangue dos doadores. Segundo o modelo de Regressão Logística Ordinal Multivariado, os fatores associados a níveis mais altos de beta-HCH foram ter idade entre 26 e 45 anos e ser do sexo feminino. Para o p,p\'-DDE os fatores associados a níveis mais altos foram ter idade entre 26 e 45 anos, ser do sexo feminino e ter trabalhado com pesticidas, enquanto receber renda mensal de 3 a 5 salários mínimos e consumir derivados de origem animal uma ou mais vezes por semana foram associados a níveis mais baixos de p,p\'-DDE. Segundo o modelo de Regressão Linear Múltipla, os fatores associados a níveis mais altos de beta-HCH foram o sexo feminino, ter contato prévio com pesticidas na região agrícola, ter trabalhado com pesticidas em campanhas de saúde pública, ter trabalhado em empresas de capacitores ou transformadores, ter trabalhado em indústrias de solventes clorados, ter renda mensal de 3 a 5 salários mínimos, consumo de carnes uma ou duas vezes por semana e consumo de frutos do mar uma ou duas vezes por semana, enquanto consumo frequente de cerveja e ter renda mensal de 1 a 3 salários mínimos foram associado a níveis menores de beta-HCH. Já para o p,p\'-DDE, os fatores associados a níveis mais elevados foram ser do sexo feminino, ser não branco, ter trabalhado com pesticidas e consumir água de fontes que não sejam minerais ou de rede, enquanto o consumo frequente de bebidas alcoólicas foi associado a níveis mais baixos de p,p\'-DDE

Del síndrome al sinthome

Currently, the concept of symptom is based on the notion of singularity (from a base perspective, underlined by Freud, regarding the persistence of symptomatic residue). This indicates that the demise of the symptom will never be complete, since the demand drive will always persist and will not cease to search for satisfaction.Let us then, insist on this matter, on the existence of an incurable residue in the symptom (which entails a particular relationship between the subject and its own pleasure), resisting sense and interpretation. The following paper has been elaborated following a diachronic trajectory of psychoanalytic theory, which allows establishing pauses, outlining the most important shifts produced in Freudian and Lacanian elaborations, respectively. Starting from Freud‘s productions, as main fulcrum, the Lacanian approach of the symptom will be introduced to link to the proposal of the sinthome proposed by Lacan. Freud will explain symptoms through the theory of trauma; those will find themselves hinged on mnemic traces, which will make the analysis of the patient‘s produced associations a crucial activity, to comprehend the etiology of the symptoms and the development of the cure. The clinical practice of this period may be summarized as ―the unconscious is susceptible to become conscious‖, aiming to the discovery and/or decoding of the symptoms, as long as they carry meaning. All of this at the same time, will be the base of future elaborations...

Riesgos naturales en la actividad turística

El propósito de la presente aportación se centra en la consideración de los riesgos de origen natural en la actividad turística, desde una doble vertiente: el efecto de los riesgos naturales, sobre todo los de carácter excepcional, en el desarrollo de las prácticas turísticas, y la consideración del papel que corresponde al turismo en la configuración de modelos territoriales sobre territorios en riesgo. Como enmarque general, el tema de estudio es analizado como una de las dimensiones dentro de las interacciones entre medio ambiente y sistemas de ocupación del territorio, bien entendido que el desarrollo del turismo, como manifestación de las intervenciones del grupo humano en un territorio, implica y genera nuevos modelos de organización y gestión del suelo. En definitiva, formas de afectar a la relación del hombre con su entorno.