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1.
Comput Methods Programs Biomed ; 208: 106280, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34333204

RESUMO

BACKGROUND AND OBJECTIVES: while traditional sleep staging is achieved through the visual - expert-based - annotation of a polysomnography, it has the disadvantages of being unpractical and expensive. Alternatives have been developed over the years to relieve sleep staging from its heavy requirements, through the collection of more easily assessable signals and its automation using machine learning. However, these alternatives have their limitations, some due to variabilities among and between subjects, other inherent to their use of sub-discriminative signals. Many new solutions rely on the evaluation of the Autonomic Nervous System (ANS) activation through the assessment of the heart-rate (HR); the latter is modulated by the aforementioned variabilities, which may result in data and concept shifts between what was learned and what we want to classify. Such adversary effects are usually tackled by Transfer Learning, dealing with problems where there are differences between what is known (source) and what we want to classify (target). In this paper, we propose two new kernel-based methods of transfer learning and assess their performances in Rapid-Eye-Movement (REM) sleep stage detection, using solely the heart rate. METHODS: our first contribution is the introduction of Kernel-Cross Alignment (KCA), a measure of similarity between a source and a target, which is a direct extension of Kernel-Target Alignment (KTA). To our knowledge, KCA has currently never been studied in the literature. Our second contribution is two alignment-based methods of transfer learning: Kernel-Target Alignment Transfer Learning (KTATL) and Kernel-Cross Alignment Transfer Learning (KCATL). Both methods differ from KTA, whose traditional use is kernel-tuning: in our methods, the kernel has been fixed beforehand, and our objective is the improvement of the estimation of unknown target labels by taking into account how observations relate to each other, which, as it will be explained, allows to transfer knowledge (transfer learning). RESULTS: we compare performances with transfer learning (KCATL, KTATL) to performances without transfer using a fixed classifier (a Support Vector Classifier - SVC). In most cases, both transfer learning methods result in an improvement of performances (higher detection rates for a fixed false-alarm rate). Our methods do not require iterative computations. CONCLUSION: we observe improved performances using our transfer methods, which are computationally efficient, as they only require the computation of a kernel matrix and are non-iterative. However, some optimisation aspects are still under investigation.


Assuntos
Aprendizado de Máquina , Fases do Sono , Frequência Cardíaca , Humanos , Polissonografia
2.
Plant Methods ; 16(1): 152, 2020 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-33292327

RESUMO

BACKGROUND: Hormones are crucial to plant life and development. Being able to follow the plants hormonal response to various stimuli and throughout developmental processes is an important and increasingly widespread tool. The phytohormone cytokinin (CK) has crucial roles in the regulation of plant growth and development. RESULTS: Here we describe a version of the CK sensor Two Component signaling Sensor (TCS), referred to as TCSv2. TCSv2 has a different arrangement of binding motifs when compared to previous TCS versions, resulting in increased sensitivity in some examined tissues. Here, we examine the CK responsiveness and distribution pattern of TCSv2 in arabidopsis and tomato. CONCLUSIONS: The increased sensitivity and reported expression pattern of TCSv2 make it an ideal TCS version to study CK response in particular hosts, such as tomato, and particular tissues, such as leaves and flowers.

3.
Curr Biol ; 29(24): 4183-4192.e6, 2019 12 16.
Artigo em Inglês | MEDLINE | ID: mdl-31761704

RESUMO

Mechanisms through which the evolution of gene regulation causes morphological diversity are largely unclear. The tremendous shape variation among plant leaves offers attractive opportunities to address this question. In cruciferous plants, the REDUCED COMPLEXITY (RCO) homeodomain protein evolved via gene duplication and acquired a novel expression domain that contributed to leaf shape diversity. However, the molecular pathways through which RCO regulates leaf growth are unknown. A key question is to identify genome-wide transcriptional targets of RCO and the DNA sequences to which RCO binds. We investigate this question using Cardamine hirsuta, which has complex leaves, and its relative Arabidopsis thaliana, which evolved simple leaves through loss of RCO. We demonstrate that RCO directly regulates genes controlling homeostasis of the hormone cytokinin to repress growth at the leaf base. Elevating cytokinin signaling in the RCO expression domain is sufficient to both transform A. thaliana simple leaves into complex ones and partially bypass the requirement for RCO in C. hirsuta complex leaf development. We also identify RCO as its own target gene. RCO directly represses its own transcription via an array of low-affinity binding sites, which evolved after RCO duplicated from its progenitor sequence. This autorepression is required to limit RCO expression. Thus, evolution of low-affinity binding sites created a negative autoregulatory loop that facilitated leaf shape evolution by defining RCO expression and fine-tuning cytokinin activity. In summary, we identify a transcriptional mechanism through which conflicts between novelty and pleiotropy are resolved during evolution and lead to morphological differences between species.


Assuntos
Citocininas/metabolismo , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cardamine/genética , Cardamine/metabolismo , Citocininas/genética , Evolução Molecular , Duplicação Gênica/genética , Regulação da Expressão Gênica de Plantas/genética , Homeostase , Proteínas de Plantas/metabolismo , Fatores de Transcrição/metabolismo
4.
Methods Mol Biol ; 1871: 279-293, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30276746

RESUMO

Staphylococcus aureus is a major human pathogen that can readily acquire antibiotic resistance. For instance, methicillin-resistant S. aureus represents a major cause of hospital- and community-acquired bacterial infections. In this chapter, we first provide a detailed protocol for obtaining unbiased and reproducible S. aureus metabolic profiles. The resulting intracellular metabolome is then analyzed in an untargeted manner by using both hydrophilic interaction liquid chromatography and pentafluorophenyl-propyl columns coupled to high-resolution mass spectrometry. Such analyses are done in conjunction with our in-house spectral database to identify with high confidence as many meaningful S. aureus metabolites as possible. Under these conditions, we can routinely monitor more than 200 annotated S. aureus metabolites. We also indicate how this protocol can be used to investigate the metabolic differences between methicillin-resistant and susceptible strains.


Assuntos
Antibacterianos/farmacologia , Cromatografia Líquida , Espectrometria de Massas , Metaboloma/efeitos dos fármacos , Metabolômica/métodos , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/metabolismo , Cromatografia Líquida/métodos , Análise de Dados , Resistência Microbiana a Medicamentos , Humanos , Espectrometria de Massas/métodos , Resistência a Meticilina , Reprodutibilidade dos Testes , Fluxo de Trabalho
5.
EMBO Rep ; 18(7): 1213-1230, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28600354

RESUMO

Auxin acts synergistically with cytokinin to control the shoot stem-cell niche, while both hormones act antagonistically to maintain the root meristem. In aluminum (Al) stress-induced root growth inhibition, auxin plays an important role. However, the role of cytokinin in this process is not well understood. In this study, we show that cytokinin enhances root growth inhibition under stress by mediating Al-induced auxin signaling. Al stress triggers a local cytokinin response in the root-apex transition zone (TZ) that depends on IPTs, which encode adenosine phosphate isopentenyltransferases and regulate cytokinin biosynthesis. IPTs are up-regulated specifically in the root-apex TZ in response to Al stress and promote local cytokinin biosynthesis and inhibition of root growth. The process of root growth inhibition is also controlled by ethylene signaling which acts upstream of auxin. In summary, different from the situation in the root meristem, auxin acts with cytokinin in a synergistic way to mediate aluminum-induced root growth inhibition in Arabidopsis.


Assuntos
Alumínio/farmacologia , Arabidopsis/efeitos dos fármacos , Citocininas/fisiologia , Ácidos Indolacéticos/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Citocininas/biossíntese , Etilenos/metabolismo , Regulação da Expressão Gênica de Plantas , Meristema/efeitos dos fármacos , Meristema/genética , Meristema/fisiologia , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Raízes de Plantas/fisiologia , Transdução de Sinais , Estresse Fisiológico
6.
Methods Mol Biol ; 1497: 81-90, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-27864760

RESUMO

Cytokinins are classical plant hormones that control numerous developmental processes throughout the plant life cycle. Cytokinin-responsive cells activate transcription via a phospho-relay signaling network. Type-B nuclear RESPONSE REGULATOR (RR) proteins mediate transcriptional activation as the final step in the signaling cascade. They bind to promoters of immediate-early target genes via a conserved Myb-related DNA-binding domain. To monitor transcriptional activation in response to a cytokinin stimulus, we have constructed a synthetic promoter, TCS (two-component signaling sensor) that harbors the concatemerized binding motifs for activated type-B RR in an optimized configuration. Here, we describe our protocols for imaging TCSn::GFP expression in transgenic Arabidopsis plants. The use of the fluorescent reporter GFP allows the visualization of cytokinin-responding cells by fluorescent microscopy without the need for tissue processing steps, or staining reactions. This method is fast and with a low risk of artifacts. However, since cytokinin signaling integrates various environmental information including light, nutrient status, and biotic and abiotic stress, special care needs to be devoted to the control of growth conditions.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Citocininas/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Ligação a DNA/genética , Corantes Fluorescentes/metabolismo , Regulação da Expressão Gênica de Plantas/genética , Microscopia de Fluorescência/métodos , Reguladores de Crescimento de Plantas/genética , Regiões Promotoras Genéticas/genética , Transdução de Sinais/genética , Coloração e Rotulagem/métodos , Ativação Transcricional/genética
7.
Science ; 353(6303): 1027-1030, 2016 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-27701112

RESUMO

Morphogenetic signals control the patterning of multicellular organisms. Cytokinins are mobile signals that are perceived by subsets of plant cells. We found that the responses to cytokinin signaling during Arabidopsis development are constrained by the transporter PURINE PERMEASE 14 (PUP14). In our experiments, the expression of PUP14 was inversely correlated to the cytokinin signaling readout. Loss of PUP14 function allowed ectopic cytokinin signaling accompanied by aberrant morphogenesis in embryos, roots, and the shoot apical meristem. PUP14 protein localized to the plasma membrane and imported bioactive cytokinins, thus depleting apoplastic cytokinin pools and inhibiting perception by plasma membrane-localized cytokinin sensors to create a sink for active ligands. We propose that the spatiotemporal cytokinin sink patterns established by PUP14 determine the cytokinin signaling landscape that shapes the morphogenesis of land plants.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Citocininas/metabolismo , Proteínas de Transporte de Nucleobases/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Membrana Celular/enzimologia , Ligantes , Meristema/crescimento & desenvolvimento , Meristema/metabolismo , Proteínas de Transporte de Nucleobases/genética , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Brotos de Planta/genética , Brotos de Planta/crescimento & desenvolvimento , Brotos de Planta/metabolismo , Transdução de Sinais
8.
Rev Col Bras Cir ; 43(4): 292-4, 2016.
Artigo em Inglês, Português | MEDLINE | ID: mdl-27679951

RESUMO

Treatment of bronchial fistula (BF) after pulmonary lobectomy is a challenge. Often, patients require long hospital stay, have recurrent empyema and pneumonia, are susceptible to sepsis, often need broad-spectrum antibiotics, as well as various surgical approaches. With the advent and growing evidence of the benefits of negative pressure therapy (NPT), its use in some patients with BF has been reported with encouraging results concerning its feasibility and cost-effectiveness. The aim of this study was to demonstrate the application of NPT as a resource for BF treatment and comparatively analyze the overall cost of treatment. RESUMO O tratamento de fístula brônquica (FB) após lobectomia pulmonar é um desafio. Muitas vezes, o paciente demanda longo tempo de internação, apresenta recidivas de empiema e pneumonia, pode evoluir para sepse, frequentemente necessita de antibioticoterapia de amplo espectro, bem como de várias abordagens cirúrgicas. Com o advento e acúmulo de evidências dos benefícios da terapia por pressão negativa (TPN), seu uso em alguns pacientes com FB tem sido relatado com resultados animadores relativos à sua viabilidade e ao seu custo-efetividade. O objetivo deste estudo foi demonstrar a aplicação de TPN como recurso para tratamento da FB e analisar comparativamente o custo global do seu tratamento.


Assuntos
Fístula Brônquica/cirurgia , Análise Custo-Benefício , Tratamento de Ferimentos com Pressão Negativa/economia , Estudos de Viabilidade , Humanos , Masculino , Pessoa de Meia-Idade
9.
J Proteome Res ; 14(11): 4863-75, 2015 Nov 06.
Artigo em Inglês | MEDLINE | ID: mdl-26502275

RESUMO

Staphylococcus aureus can cause a variety of severe disease patterns and can readily acquire antibiotic resistance; however, the mechanisms by which this commensal becomes a pathogen or develops antibiotic resistance are still poorly understood. Here we asked whether metabolomics can be used to distinguish bacterial strains with different antibiotic susceptibilities. Thus, an efficient and robust method was first thoroughly implemented to measure the intracellular metabolites of S. aureus in an unbiased and reproducible manner. We also placed special emphasis on metabolome coverage and annotation and used both hydrophilic interaction liquid chromatography and pentafluorophenyl-propyl columns coupled to high-resolution mass spectrometry in conjunction with our spectral database developed in-house to identify with high confidence as many meaningful S. aureus metabolites as possible. Overall, we were able to characterize up to 210 metabolites in S. aureus, which represents a substantial ∼50% improvement over previously published data. We then preliminarily compared the metabolic profiles of 10 clinically relevant methicillin-resistant and susceptible strains harvested at different time points during the exponential growth phase (without any antibiotic exposure). Interestingly, the resulting data revealed a distinct behavior of "slow-growing" resistant strains, which show modified levels of several precursors of peptidoglycan and capsular polysaccharide biosynthesis.


Assuntos
Metaboloma , Resistência a Meticilina/fisiologia , Staphylococcus aureus Resistente à Meticilina/metabolismo , Anotação de Sequência Molecular , Peptidoglicano/isolamento & purificação , Polissacarídeos Bacterianos/isolamento & purificação , Cromatografia Líquida/métodos , Bases de Dados Factuais , Farmacorresistência Bacteriana Múltipla/fisiologia , Interações Hidrofóbicas e Hidrofílicas , Espectrometria de Massas/métodos , Staphylococcus aureus Resistente à Meticilina/química , Peptidoglicano/biossíntese , Polissacarídeos Bacterianos/biossíntese
10.
Anal Chem ; 87(11): 5553-60, 2015 Jun 02.
Artigo em Inglês | MEDLINE | ID: mdl-25932746

RESUMO

We have developed and describe here for the first time a highly sensitive method for the fast and unambiguous detection of viable Escherichia coli in food matrices. The new approach is based on using label-free phages (T4), obligate parasites of bacteria, which are attractive for pathogen detection because of their inherent natural specificity and ease of use. A specific immunomagnetic separation was used to capture the progeny phages produced. Subsequently, T4 phage markers were detected by liquid chromatography coupled to targeted mass spectrometry. Combining the specificity of these three methodologies is of great interest in developing an alternative to conventional time-consuming culture-based technologies for the detection of viable bacteria for industrial applications. First, optimization experiments with phage T4 spiked in complex matrices (without a phage amplification event) were performed and demonstrated specific, sensitive, and reproducible phage capture and detection in complex matrices including Luria-Bertani broth, orange juice, and skimmed milk. The method developed was then applied to the detection of E. coli spiked in foodstuffs (with a phage amplification event). After having evaluated the impact of infection duration on assay sensitivity, we showed that our assay specifically detects viable E. coli in milk at an initial count of ≥1 colony-forming unit (cfu)/mL after an 8-h infection. This excellent detection limit makes our new approach an alternative to PCR-based assays for rapid bacterial detection.


Assuntos
Escherichia coli/fisiologia , Microbiologia de Alimentos/métodos , Separação Imunomagnética , Espectrometria de Massas por Ionização por Electrospray/métodos , Bacteriófago T4/química , Bacteriófago T4/isolamento & purificação , Escherichia coli/isolamento & purificação , Escherichia coli/virologia , Limite de Detecção , Fatores de Tempo
11.
Poult Sci ; 94(4): 574-8, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25701206

RESUMO

Gait problems constitute an important and chronic welfare restriction for broiler chickens. The objective of the present study was to determine if adrenal gland morphology indicates chronic welfare restrictions in broiler chickens, using gait problems as the stressor. Sixty-six birds raised on a commercial unit were selected at 40 d of age and separated into groups according to gait score. One group was apparently healthy birds (AH) with gait scores of 0 to 2, and the other group had birds with gait problems (GP) that showed gait scores of 4 to 5. Birds were slaughtered and weighed, and their adrenal glands were measured and weighed; proportions of medullary and adrenocortical tissues, and lymphatic tissue and blood vessels were studied. GP birds had lower BW when compared to AH birds, and when adrenal gland weight values were adjusted to BW, a greater relative adrenal weight was observed for the GP group. Adrenals from GP birds also presented a higher proportion of blood vessels when compared to AH birds. These results might indicate increased adrenal activity and evidence of the inflammatory process as a consequence of chronic stress. Results showed that gait problems caused significant adrenal gland changes, suggesting a possible role for the study of adrenal gland morphology as an indicator of chronic welfare problems in broiler chickens.


Assuntos
Córtex Suprarrenal/anatomia & histologia , Medula Suprarrenal/anatomia & histologia , Bem-Estar do Animal , Galinhas/anatomia & histologia , Galinhas/fisiologia , Animais , Peso Corporal , Marcha , Masculino , Estresse Fisiológico
12.
Burns ; 41(2): e15-8, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25440855

RESUMO

Decompensation of epilepsy in burned patients may be caused by several factors. Burn is a classic etiology of systemic inflammatory response syndrome, and evolves into two physiological phases. The first 48h after injury corresponds to the first phase involving severe hypovolemic shock. The second phase corresponds to the hypermetabolic response to burns. Altered pharmacokinetics of anticonvulsant drugs is observed. Albumin and other plasma proteins are reduced, leading to increased free fraction of phenytoin, resulting in greater clearance and a lower total drug concentration. Associated with metabolic changes of burned patient, this fact predisposes to seizures in epileptic burned patients. The authors present the case of an epileptic 36-year-old-woman who developed recurrent seizures after a thermal injury, despite using the same medications and doses of anticonvulsant drugs of last 12 years, with controlled epilepsy.


Assuntos
Queimaduras/complicações , Epilepsia/etiologia , Adulto , Anticonvulsivantes/farmacocinética , Queimaduras/metabolismo , Epilepsia/tratamento farmacológico , Feminino , Humanos , Fenitoína/farmacocinética
13.
PLoS One ; 9(10): e109875, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25299686

RESUMO

With the growing availability of genomic sequence information, there is an increasing need for gene function analysis. Antibody-mediated "silencing" represents an intriguing alternative for the precise inhibition of a particular function of biomolecules. Here, we describe a method for selecting recombinant antibodies with a specific purpose in mind, which is to inhibit intrinsic protein-protein interactions in the cytosol of plant cells. Experimental procedures were designed for conveniently evaluating desired properties of recombinant antibodies in consecutive steps. Our selection method was successfully used to develop a recombinant antibody inhibiting the interaction of ARABIDOPSIS HISTIDINE PHOSPHOTRANSFER PROTEIN 3 with such of its upstream interaction partners as the receiver domain of CYTOKININ INDEPENDENT HISTIDINE KINASE 1. The specific down-regulation of the cytokinin signaling pathway in vivo demonstrates the validity of our approach. This selection method can serve as a prototype for developing unique recombinant antibodies able to interfere with virtually any biomolecule in the living cell.


Assuntos
Proteínas de Arabidopsis/genética , Citosol/metabolismo , Fosfotransferases/genética , Mapas de Interação de Proteínas/genética , Proteínas Quinases/genética , Anticorpos/administração & dosagem , Anticorpos/imunologia , Arabidopsis/genética , Proteínas de Arabidopsis/biossíntese , Proteínas de Arabidopsis/imunologia , Citosol/imunologia , Regulação da Expressão Gênica de Plantas , Inativação Gênica/imunologia , Fosfotransferases/biossíntese , Fosfotransferases/imunologia , Mapas de Interação de Proteínas/imunologia , Proteínas Quinases/biossíntese , Proteínas Quinases/imunologia , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/imunologia , Transdução de Sinais
14.
Rev. bras. cir. plást ; 29(1): 114-119, jan.-mar. 2014.
Artigo em Inglês, Português | LILACS | ID: biblio-99

RESUMO

Introdução: Apesar dos grandes avanços em seu tratamento, infecção de pele com queimadura continua a ser um grande desafio. O objetivo deste estudo é avaliar os aspectos microbiológicos do primeiro ano de funcionamento de uma unidade de queimadura em um Hospital Universitário. Métodos: Estudo retrospectivo. Dados microbiológicos foram coletados e analisados a partir de pacientes internados na Unidade de Queimadura (UTQ) do Hospital São Paulo, Hospital Universitário da Escola Paulista de Medicina (EPM) da Universidade Federal de São Paulo (UNIFESP), entre junho de 2009 e julho de 2010. Resultados: O tempo médio de permanência hospitalar foi de 13,8 dias, com uma taxa de mortalidade de 5,9%. A média da superfície corpórea queimada foi de 10,3%. Avaliou-se 159 culturas de 101 pacientes. Culturas de sangue foram as mais solicitadas (41%). Também foram acessadas 245 culturas de vigilância, coletadas de 75 pacientes. A análise microbiológica revelou um índice de positividade total de 34,5%. Os agentes mais prevalentes foram Staphylococcus coagulase-negativo - CoNS - (33%), Pseudomonas aeruginosa (24%), Acinetobacter spp. (22%) e Klebsiella pneumoniae (5%). Conclusão: A avaliação microbiológica do primeiro ano de funcionamento da UTQ da EPM/ UNIFESP revelou que, embora o agente mais prevalente tenha sido a CoNS, bacilos Gram negativos ainda são muito prevalentes, como a Pseudomonas aeruginosa e a Acinetobacter baumannii. Apesar de pouco tempo de operação, observou-se um grande número de microrganismos multirresistentes, que pode ser explicado por longa exposição a agentes antimicrobianos e alta taxa de transferência de outros hospitais.


Introduction: Despite great advances in treatment, burned skin infection remains a major challenge. The aim of this study is to evaluate the microbiological aspects of the first year's operation of a Burn Unit in a University Hospital. Methods: Retrospective study. Microbiological data were collected and analyzed from patients admitted to the Burn Unit of São Paulo Hospital, a University Hospital of the Paulista Medical School (EPM) of the Federal University of São Paulo (UNIFESP) from June 2009 to July 2010. Results: The average length of stay was 13.8 days with a mortality rate of 5.9%, and median of TBSA was 10.3%. Evaluated 159 cultures from 101 patients. Blood cultures were the most requested (41%). It was also accessed 245 surveillance cultures collected from 75 patients. The microbiological analysis revealed a total positivity rate of 34,5%. The most prevalent agents were Coagulase-negative Staphylococcus - CoNS - (33%), Pseudomonas aeruginosa (24%), Acinetobacter spp. (22%) and Klebsiella pneumoniae (5%). Conclusion: The microbiological evaluation of the first year's activity of EPM/UNIFESP Burn Care Unit revealed that, although the most prevalent agent was CoNS, Gram negative bacilli are still very prevalent, such as Pseudomonas aeruginosa and Acinetobacter baumannii. Despite the short time of operation, was observed large number of multiresistant microorganisms which can be explained by long exposure to antimicrobials and high transfer rate from other hospitals.


Assuntos
Humanos , Masculino , Feminino , História do Século XXI , Unidades de Queimados , Queimaduras , Epidemiologia , Bancos de Espécimes Biológicos , Anti-Infecciosos , Infecções Bacterianas , Infecções Bacterianas/microbiologia , Infecções Bacterianas/patologia , Unidades de Queimados/normas , Unidades de Queimados/estatística & dados numéricos , Queimaduras/cirurgia , Queimaduras/complicações , Queimaduras/microbiologia , Queimaduras/epidemiologia , Estudos Epidemiológicos , Epidemiologia/normas , Epidemiologia/estatística & dados numéricos , Estudos Retrospectivos , Bancos de Espécimes Biológicos/normas , Estudo de Avaliação , Pacientes Internados , Pacientes Internados/estatística & dados numéricos , Anti-Infecciosos/isolamento & purificação , Anti-Infecciosos/análise , Anti-Infecciosos/uso terapêutico
15.
J Proteome Res ; 13(3): 1450-65, 2014 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-24517284

RESUMO

According to the World Health Organization, food safety is an essential public health priority. In this context, we report a relevant proof of feasibility for the indirect specific detection of bacteria in food samples using unlabeled phage amplification coupled to ESI mass spectrometry analysis and illustrated with the model phage systems T4 and SPP1. High-resolving power mass spectrometry analysis (including bottom-up and top-down protein analysis) was used for the discovery of specific markers of phage infection. Structural components of the viral particle and nonstructural proteins encoded by the phage genome were identified. Then, targeted detection of these markers was performed on a triple quadrupole mass spectrometer operating in the selected reaction monitoring mode. E. coli at 1 × 10(5), 5 × 10(5), and 1 × 10(6) CFU/mL concentrations was successfully detected after only a 2 h infection time by monitoring phage T4 structural markers in Luria-Bertani broth, orange juice, and French bean stew ("cassoulet") matrices. Reproducible detection of nonstructural markers was also demonstrated, particularly when a high titer of input phages was required to achieve successful amplification. This strategy provides a highly time-effective and sensitive assay for bacterial detection.


Assuntos
Bacillus subtilis/isolamento & purificação , Bebidas/análise , Citrus sinensis , Colífagos/genética , Escherichia coli/isolamento & purificação , Produtos da Carne/análise , Sequência de Aminoácidos , Animais , Bacillus subtilis/virologia , Bebidas/microbiologia , Escherichia coli/virologia , Análise de Alimentos , Humanos , Lisogenia , Produtos da Carne/microbiologia , Dados de Sequência Molecular , Biblioteca de Peptídeos , Espectrometria de Massas por Ionização por Electrospray , Suínos , Proteínas Virais/genética
16.
Development ; 140(22): 4544-53, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24194471

RESUMO

The plant life cycle alternates between a diploid sporophytic and a haploid gametophytic generation. The female gametophyte (FG) of flowering plants is typically formed through three syncytial mitoses, followed by cellularisation that forms seven cells belonging to four cell types. The specification of cell fates in the FG has been suggested to depend on positional information provided by an intrinsic auxin concentration gradient. The goal of this study was to develop mathematical models that explain the formation of this gradient in a syncytium. Two factors were proposed to contribute to the maintenance of the auxin gradient in Arabidopsis FGs: polar influx at early stages and localised auxin synthesis at later stages. However, no gradient could be generated using classical, one-dimensional theoretical models under these assumptions. Thus, we tested other hypotheses, including spatial confinement by the large central vacuole, background efflux and localised degradation, and investigated the robustness of cell specification under different parameters and assumptions. None of the models led to the generation of an auxin gradient that was steep enough to allow sufficiently robust patterning. This led us to re-examine the response to an auxin gradient in developing FGs using various auxin reporters, including a novel degron-based reporter system. In agreement with the predictions of our models, auxin responses were not detectable within the FG of Arabidopsis or maize, suggesting that the effects of manipulating auxin production and response on cell fate determination might be indirect.


Assuntos
Ácidos Indolacéticos/metabolismo , Magnoliopsida/metabolismo , Modelos Biológicos , Óvulo Vegetal/metabolismo , Transportadores de Cassetes de Ligação de ATP/metabolismo , Arabidopsis/citologia , Arabidopsis/efeitos dos fármacos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Padronização Corporal/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Polaridade Celular/efeitos dos fármacos , Simulação por Computador , Difusão , Ácidos Indolacéticos/farmacologia , Magnoliopsida/citologia , Magnoliopsida/efeitos dos fármacos , Óvulo Vegetal/citologia , Óvulo Vegetal/efeitos dos fármacos , Vacúolos/efeitos dos fármacos , Vacúolos/metabolismo , Zea mays/citologia , Zea mays/efeitos dos fármacos , Zea mays/metabolismo
17.
Protein Eng Des Sel ; 26(11): 725-34, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24006372

RESUMO

A single strategy to select RNA polymerase from bacteriophage T7 (T7 RNAP) mutants in Escherichia coli with enhanced thermostability or enzymatic activity is described. T7 RNAP has the ability to specifically transcribe genes under control of T7 phage promoter. By using random mutagenesis of the T7 RNAP gene in combination with an appropriate screening at 25 and 42°C, we have generated and selected E.coli clones with temperature-sensitive phenotype in the presence of chloramphenicol. The resistance to chloramphenicol used to select these clones results from expression control of the chloramphenicol acetyl transferase gene by the T7 promoter. In a second phase, and using the thermosensitive T7 RNAP variants as template, a new round of random mutagenesis was performed. Combined to an appropriate screening strategy, 11 mutations (second-site T7 RNAP revertants) that restore the initial resistance to chloramphenicol at 42°C were identified. Nine of these mutations increase the thermal resistance of the wild-type T7 RNA. They include the five mutations previously described using different approaches and four novel mutations. One improves T7 RNA catalytic activity and one has no positive effect on the natural enzyme but increases the activity of some combined mutants. Additive effects of mutations amount to an increase of as much as 10°C in T1/2 compared with the wild-type enzyme and up to a 2-fold activity enhancement.


Assuntos
RNA Polimerases Dirigidas por DNA , Proteínas Virais , RNA Polimerases Dirigidas por DNA/química , RNA Polimerases Dirigidas por DNA/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Estabilidade Enzimática/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Temperatura Alta , Modelos Moleculares , Mutação/genética , Fenótipo , Plasmídeos/genética , Proteínas Virais/química , Proteínas Virais/genética , Proteínas Virais/metabolismo
18.
Acta cir. bras ; 28(8): 551-558, Aug. 2013. graf, tab
Artigo em Inglês | LILACS | ID: lil-680608

RESUMO

PURPOSE: To evaluate the gene expression of KGF, TNF-alpha and IL-1 beta in skin fibroblasts and keratinocytes cultured from burned patients. METHODS: Three patients with large burns and three patients with small burns, as well as two controls, were included. The cell culture was initiated by the enzymatic method. After extraction and purification of mRNA, qPCR was used to assess the gene expression of KGF, TNF-alpha and IL-1 beta. RESULTS: The expression of KGF was increased on average 220-fold in large burns and 33.33-fold in small burns in fibroblasts, and 11.2-fold in large burns and 3.45-fold in small burns in keratinocytes compared to healthy patients (p<0.05). Expression of TNF-alpha was not observed. IL-1 beta is down-regulated in fibroblasts of burned patients, and much more repressed in small burns (687-fold, p<0.05). In keratinocytes, the repression of IL-1 beta expression occurs in patients with small burns (28-fold), while patients with large burns express this gene intensively (15-fold). CONCLUSIONS: The study showed a quantitative pattern in the expression of KGF gene, which is more expressed according to the size of the burn. TNF-alpha was not expressed. A qualitative pattern in the expression of IL-1 beta gene was demonstrated.


Assuntos
Adulto , Feminino , Humanos , Masculino , Queimaduras/genética , /genética , Fibroblastos/metabolismo , Interleucina-1beta/genética , Fator de Necrose Tumoral alfa/genética , Estudos de Casos e Controles , Células Cultivadas , /análise , Expressão Gênica , Interleucina-1beta/análise , Reação em Cadeia da Polimerase , Pele/citologia , Pele/lesões , Fatores de Tempo , Fator de Necrose Tumoral alfa/análise
19.
Acta Cir Bras ; 28(8): 551-8, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23896833

RESUMO

PURPOSE: To evaluate the gene expression of KGF, TNF-alpha and IL-1 beta in skin fibroblasts and keratinocytes cultured from burned patients. METHODS: Three patients with large burns and three patients with small burns, as well as two controls, were included. The cell culture was initiated by the enzymatic method. After extraction and purification of mRNA, qPCR was used to assess the gene expression of KGF, TNF-alpha and IL-1 beta. RESULTS: The expression of KGF was increased on average 220-fold in large burns and 33.33-fold in small burns in fibroblasts, and 11.2-fold in large burns and 3.45-fold in small burns in keratinocytes compared to healthy patients (p<0.05). Expression of TNF-alpha was not observed. IL-1 beta is down-regulated in fibroblasts of burned patients, and much more repressed in small burns (687-fold, p<0.05). In keratinocytes, the repression of IL-1 beta expression occurs in patients with small burns (28-fold), while patients with large burns express this gene intensively (15-fold). CONCLUSIONS: The study showed a quantitative pattern in the expression of KGF gene, which is more expressed according to the size of the burn. TNF-alpha was not expressed. A qualitative pattern in the expression of IL-1 beta gene was demonstrated.


Assuntos
Queimaduras/genética , Fator 7 de Crescimento de Fibroblastos/genética , Fibroblastos/metabolismo , Interleucina-1beta/genética , Fator de Necrose Tumoral alfa/genética , Adulto , Estudos de Casos e Controles , Células Cultivadas , Feminino , Fator 7 de Crescimento de Fibroblastos/análise , Expressão Gênica , Humanos , Interleucina-1beta/análise , Masculino , Reação em Cadeia da Polimerase , Pele/citologia , Pele/lesões , Fatores de Tempo , Fator de Necrose Tumoral alfa/análise
20.
Motriz rev. educ. fís. (Impr.) ; 19(2): 450-459, abr.-jun. 2013. ilus, tab
Artigo em Português | LILACS | ID: lil-678319

RESUMO

O objetivo deste estudo foi comparar o efeito agudo dos exercícios de alongamento estático e do alongamento dinâmico na impulsão vertical e amplitude do movimento de 22 jogadores de futebol profissional do sexo masculino. Observou-se que após a intervenção do alongamento dinâmico, o grupo experimental (n=13) apresentou um aumento significativo na impulsão vertical (p=0,002), enquanto que após o alongamento estático, não houve alteração significativa (p=0,343), assim como na condição sem alongamento (p>0,05). Com relação aos níveis de amplitude de movimento (n=13), não houve diferenças significativas no sentar e alcançar (p=0,263) nas três condições analisadas: alongamento estático, dinâmico e controle. Os resultados sugerem que o alongamento dinâmico é o mais indicado para ser realizado antes de atividades que exijam potência muscular, entretanto as intervenções propostas neste estudo não evidenciaram aumento na amplitude de movimento.


This study aimed to compare the acute effects of static and dynamic stretching exercises on the vertical jump performance and range of motion of 22 male professional soccer players. After the dynamic stretching intervention, the experimental group (n=13) showed a significant increase in vertical jump performance (p=0,002), but no significant changes were observed after static stretching (p=0,343) as well as after tests without stretching intervention (p>0,05). Regarding range of motion (n=13), there were no significant differences in the sit and reach (p=0,263) under the three conditions examined: static stretching, dynamic stretching, and control (no stretching). These results suggest that dynamic stretching is the most suitable type of stretching to perform before warm-up routines that require muscle power. However, none of the interventions proposed in this study resulted in an increase in range of motion.


Assuntos
Humanos , Masculino , Adulto , Futebol/fisiologia , Amplitude de Movimento Articular
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