RESUMO
The fungal infection causing white-nose disease in hibernating bats in North America has resulted in dramatic population declines of affected species, since the introduction of the causative agent Pseudogymnoascus destructans. The fungus is native to the Palearctic, where it also infects several bat species, yet rarely causes severe pathology or the death of the host. Pseudogymnoascus destructans infects bats during hibernation by invading and digesting the skin tissue, resulting in the disruption of torpor patterns and consequent emaciation. Relations among pathogen, host, and environment are complex, and individuals, populations, and species respond to the fungal pathogen in different ways. For example, the Nearctic Myotis lucifugus responds to infection by mounting a robust immune response, leading to immunopathology often contributing to mortality. In contrast, the Palearctic M. myotis shows no significant immunological response to infection. This lack of a strong response, resulting from the long coevolution between the hosts and the pathogen in the pathogen's native range, likely contributes to survival in tolerant species. After more than 15 years since the initial introduction of the fungus to North America, some of the affected populations are showing signs of recovery, suggesting that the fungus, hosts, or both are undergoing processes that may eventually lead to coexistence. The suggested or implemented management methods of the disease in North America have encompassed, for example, the use of probiotics and fungicides, vaccinations, and modifying the environmental conditions of the hibernation sites to limit the growth of the pathogen, intensity of infection, or the hosts' responses to it. Based on current knowledge from Eurasia, policy makers and conservation managers should refrain from disrupting the ongoing evolutionary processes and adopt a holistic approach to managing the epizootic.
Vista paleártica de una enfermedad fúngica de murciélagos Resumen La enfermedad fúngica que produce el síndrome de nariz blanca en murciélagos en hibernación en Norte América ha resultado en declinaciones poblacionales dramáticas en las especies afectadas desde la introducción del agente causante, Pseudogymnoascus destructans. El hongo es nativo del Paleártico, donde también infecta a varias especies de murciélagos; sin embargo, raramente causa patología severa o la muerte del hospedero. Pseudogymnoascus destructans infecta a los murciélagos durante la hibernación invadiendo y digiriendo el tejido de la piel, lo que resulta en la disrupción de los patrones de torpor y la consecuente emaciación. Las relaciones entre el patógeno, el huésped y el ambiente son complejas, y los individuos, las especies y poblaciones responden al patógeno fúngico de distintas maneras. Por ejemplo, Myotis lucifugus, especie del Neártico, responde a la infección montando una respuesta inmune robusta, produciendo una inmunopatología que a menudo contribuye a la mortalidad. En contraste, M. myotis del Paleártico no presenta respuesta inmunológica significativa a la infección. La falta de una fuerte respuesta, resultado de la larga coevolución entre hospederos y el patógeno en el rango nativo de distribución del patógeno, probablemente contribuye a la supervivencia en especies tolerantes. Después de más de 15 años desde la introducción del hongo en Norte América, algunas de las poblaciones afectadas están mostrando señales recuperación, lo que sugiere que el hongo, hospederos, o ambos, están pasando por procesos que eventualmente pueden conducir a la coexistencia. Los métodos de manejo de la enfermedad sugeridos o implementados en Norte América han abarcado, por ejemplo, el uso de probióticos y fungicidas, vacunaciones y modificación de las condiciones ambientales de los sitios de hibernación para limitar el crecimiento del patógeno, la intensidad de la infección o las respuestas de los hospederos. Con base en conocimiento actual de Eurasia, los formuladores de políticas y los manejadores de la conservación deberían abstenerse de alterar los procesos evolutivos en curso y adoptar un enfoque holístico para gestionar la epizootia.
RESUMO
BACKGROUND: Diesel exhaust (DE) induces neutrophilia and lymphocytosis in experimentally exposed humans. These responses occur in parallel to nuclear migration of NF-κB and c-Jun, activation of mitogen activated protein kinases and increased production of inflammatory mediators. There remains uncertainty regarding the impact of DE on endogenous antioxidant and xenobiotic defences, mediated by nuclear factor erythroid 2-related factor 2 (Nrf2) and the aryl hydrocarbon receptor (AhR) respectively, and the extent to which cellular antioxidant adaptations protect against the adverse effects of DE. METHODS: Using immunohistochemistry we investigated the nuclear localization of Nrf2 and AhR in the epithelium of endobronchial mucosal biopsies from healthy subjects six-hours post exposure to DE (PM10, 300 µg/m3) versus post-filtered air in a randomized double blind study, as a marker of activation. Cytoplasmic expression of cytochrome P450s, family 1, subfamily A, polypeptide 1 (CYP1A1) and subfamily B, Polypeptide 1 (CYP1B1) were examined to confirm AhR activation; with the expression of aldo-keto reductases (AKR1A1, AKR1C1 and AKR1C3), epoxide hydrolase and NAD(P)H dehydrogenase quinone 1 (NQO1) also quantified. Inflammatory and oxidative stress markers were examined to contextualize the responses observed. RESULTS: DE exposure caused an influx of neutrophils to the bronchial airway surface (p = 0.013), as well as increased bronchial submucosal neutrophil (p < 0.001), lymphocyte (p = 0.007) and mast cell (p = 0.002) numbers. In addition, DE exposure enhanced the nuclear translocation of the AhR and increased the CYP1A1 expression in the bronchial epithelium (p = 0.001 and p = 0.028, respectively). Nuclear translocation of AhR was also increased in the submucosal leukocytes (p < 0.001). Epithelial nuclear AhR expression was negatively associated with bronchial submucosal CD3 numbers post DE (r = -0.706, p = 0.002). In contrast, DE did not increase nuclear translocation of Nrf2 and was associated with decreased NQO1 in bronchial epithelial cells (p = 0.02), without affecting CYP1B1, aldo-keto reductases, or epoxide hydrolase protein expression. CONCLUSION: These in vivo human data confirm earlier cell and animal-based observations of the induction of the AhR and CYP1A1 by diesel exhaust. The induction of phase I xenobiotic response occurred in the absence of the induction of antioxidant or phase II xenobiotic defences at the investigated time point 6 h post-exposures. This suggests DE-associated compounds, such as polycyclic aromatic hydrocarbons (PAHs), may induce acute inflammation and alter detoxification enzymes without concomitant protective cellular adaptations in human airways.
Assuntos
Antioxidantes , Receptores de Hidrocarboneto Arílico , Animais , Humanos , Receptores de Hidrocarboneto Arílico/genética , Receptores de Hidrocarboneto Arílico/metabolismo , Emissões de Veículos/toxicidade , Citocromo P-450 CYP1A1 , Fator 2 Relacionado a NF-E2/metabolismo , Epóxido Hidrolases , Xenobióticos , PeptídeosRESUMO
Virtual Compton scattering on the proton has been investigated at three yet unexplored values of the four-momentum transfer Q^{2}: 0.10, 0.20, and 0.45 GeV^{2}, at the Mainz Microtron. Fits performed using either the low-energy theorem or dispersion relations allowed the extraction of the structure functions P_{LL}-P_{TT}/ε and P_{LT}, as well as the electric and magnetic generalized polarizabilities α_{E1}(Q^{2}) and ß_{M1}(Q^{2}). These new results show a smooth and rapid falloff of α_{E1}(Q^{2}), in contrast to previous measurements at Q^{2}=0.33 GeV^{2}, and provide for the first time a precise mapping of ß_{M1}(Q^{2}) in the low-Q^{2} region.
RESUMO
Hantavirus infections may cause severe and sometime life-threatening lung failure. The pathogenesis is not fully known and there is an urgent need for effective treatment. We aimed to investigate the association between pulmonary viral load and immune responses, and their relation to disease severity. Bronchoscopy with sampling of bronchoalveolar lavage (BAL) fluid was performed in 17 patients with acute Puumala hantavirus infection and 16 healthy volunteers acting as controls. Lymphocyte subsets, granzyme concentrations, and viral load were determined by flow cytometry, enzyme-linked immunosorbent assay (ELISA), and quantitative reverse transcription polymerase chain reaction (RT-PCR), respectively. Analyses of BAL fluid revealed significantly higher numbers of activated CD8(+) T cells and natural killer (NK) cells, as well as higher concentrations of the cytotoxins granzymes A and B in hantavirus-infected patients, compared to controls. In patients, Puumala hantavirus RNA was detected in 88 % of BAL cell samples and correlated inversely to the T cell response. The magnitude of the pulmonary cytotoxic lymphocyte response correlated to the severity of disease and systemic organ dysfunction, in terms of need for supplemental oxygen treatment, hypotension, and laboratory data indicating renal failure, cardiac dysfunction, vascular leakage, and cell damage. Regulatory T cell numbers were significantly lower in patients compared to controls, and may reflect inadequate immune regulation during hantavirus infection. Hantavirus infection elicits a pronounced cytotoxic lymphocyte response in the lungs. The magnitude of the immune response was associated with disease severity. These results give insights into the pathogenesis and possibilities for new treatments.
Assuntos
Citotoxicidade Imunológica , Síndrome Pulmonar por Hantavirus/patologia , Pulmão/patologia , Virus Puumala/isolamento & purificação , Adulto , Idoso , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/citologia , Broncoscopia , Granzimas/análise , Síndrome Pulmonar por Hantavirus/imunologia , Humanos , Pulmão/virologia , Subpopulações de Linfócitos/citologia , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de Doença , Carga ViralRESUMO
Cardiopulmonary diseases are major causes of death worldwide, but currently recommended strategies for diagnosis and prevention may be outdated because of recent changes in risk factor patterns. The Swedish CArdioPulmonarybioImage Study (SCAPIS) combines the use of new imaging technologies, advances in large-scale 'omics' and epidemiological analyses to extensively characterize a Swedish cohort of 30 000 men and women aged between 50 and 64 years. The information obtained will be used to improve risk prediction of cardiopulmonary diseases and optimize the ability to study disease mechanisms. A comprehensive pilot study in 1111 individuals, which was completed in 2012, demonstrated the feasibility and financial and ethical consequences of SCAPIS. Recruitment to the national, multicentre study has recently started.
Assuntos
Doenças Cardiovasculares , Doença Pulmonar Obstrutiva Crônica , Doenças Cardiovasculares/diagnóstico , Doenças Cardiovasculares/epidemiologia , Doenças Cardiovasculares/etiologia , Doenças Cardiovasculares/metabolismo , Doenças Cardiovasculares/terapia , Feminino , Técnicas Genéticas , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Proteômica/métodos , Saúde Pública/métodos , Saúde Pública/estatística & dados numéricos , Doença Pulmonar Obstrutiva Crônica/diagnóstico , Doença Pulmonar Obstrutiva Crônica/epidemiologia , Doença Pulmonar Obstrutiva Crônica/etiologia , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/terapia , Fatores de Risco , Fatores Socioeconômicos , Suécia/epidemiologiaRESUMO
Air pollution is now recognized as an important independent risk factor for cardiovascular morbidity and mortality and may be responsible for up to 3 million premature deaths each year worldwide. The mechanisms underlying the observed effects are poorly understood but are likely to be multifactorial. Here, we review the acute and chronic effects of air pollution exposure on the cardiovascular system and discuss how these effects may explain the observed increases in cardiovascular morbidity and mortality.
Assuntos
Poluição do Ar/efeitos adversos , Doenças Cardiovasculares/etiologia , Exposição Ambiental/efeitos adversos , Material Particulado , Velocidade do Fluxo Sanguíneo/fisiologia , Pressão Sanguínea/fisiologia , Endotélio Vascular/fisiopatologia , Fibrinólise/fisiologia , Frequência Cardíaca/fisiologia , Humanos , Inflamação/etiologia , Estresse Oxidativo/fisiologia , Rigidez Vascular/fisiologia , Vasoconstrição/fisiologiaRESUMO
Asthmatics are recognised to be more susceptible than healthy individuals to adverse health effects caused by exposure to the common air pollutant ozone. Ozone has been reported to induce airway neutrophilia in mild asthmatics, but little is known about how it affects the airways of asthmatic subjects on inhaled corticosteroids. We hypothesised that ozone exposure would exacerbate the pre-existent asthmatic airway inflammation despite regular inhaled corticosteroid treatment. Therefore, we exposed subjects with persistent asthma on inhaled corticosteroid therapy to 0.2 ppm ozone or filtered air for 2 h, on 2 separate occasions. Lung function was evaluated before and immediately after exposure, while bronchoscopy was performed 18 h post exposure. Compared to filtered air, ozone exposure increased airway resistance. Ozone significantly enhanced neutrophil numbers and myeloperoxidase levels in airway lavages, and induced a fourfold increase in bronchial mucosal mast cell numbers. The present findings indicate that ozone worsened asthmatic airway inflammation and offer a possible biological explanation for the epidemiological findings of increased need for rescue medication and hospitalisation in asthmatic people following exposure to ambient ozone.
Assuntos
Corticosteroides/uso terapêutico , Asma/induzido quimicamente , Asma/patologia , Inflamação/patologia , Mastócitos/patologia , Oxidantes Fotoquímicos/toxicidade , Ozônio/toxicidade , Administração por Inalação , Corticosteroides/administração & dosagem , Adulto , Asma/tratamento farmacológico , Líquido da Lavagem Broncoalveolar/citologia , Contagem de Células , Feminino , Fluxo Expiratório Forçado , Volume Expiratório Forçado , Humanos , Masculino , Mastócitos/efeitos dos fármacos , Pessoa de Meia-Idade , Peroxidase/metabolismo , Testes de Função Respiratória , Capacidade Vital/efeitos dos fármacos , Adulto JovemRESUMO
BACKGROUND: Endothelial progenitor cells (EPCs) are circulating mononuclear cells that participate in angiogenesis. The aim of this study was to determine the influence of the menstrual cycle on the number and function of EPCs, and to investigate their relationship with circulating concentrations of sex steroids and inflammatory mediators. METHODS: Ten healthy nulliparous, premenopausal, non-smoking women with regular menses were studied over a single menstrual cycle. Venepuncture was performed in the menstrual, follicular, peri-ovulatory and luteal phases. EPCs were quantified by flow cytometry (CD133(+)CD34(+)KDR(+) phenotype) and the colony-forming unit (CFU-EPC) functional assay. Circulating concentrations of estradiol, progesterone and inflammatory mediators (TNF-alpha, IL-6, sICAM-1 and VEGF) were measured by immunoassays. RESULTS: The numbers of CD133(+)CD34(+)KDR(+) cells were higher in the follicular phase (0.99 +/- 0.3 x 10(6) cells/l) compared with the peri-ovulatory phase (0.29 +/- 0.1 x 10(6) cells/l; P < 0.05). In contrast, the numbers of CFU-EPCs did not vary over the menstrual cycle. There were no correlations between EPCs and concentrations of either circulating sex steroids or inflammatory mediators. CONCLUSIONS: CD133(+)CD34(+)KDR(+) cells but not CFU-EPCs vary during the menstrual cycle. Our findings suggest a potential role for circulating EPCs in the normal cycle of physiological angiogenesis and repair of the uterine endometrium that is independent of circulating sex steroids or inflammatory mediators.
Assuntos
Células Endoteliais/patologia , Endotélio Vascular/patologia , Ciclo Menstrual , Células-Tronco/citologia , Antígeno AC133 , Adulto , Antígenos CD/biossíntese , Antígenos CD34/biossíntese , Células Endoteliais/citologia , Endotélio Vascular/citologia , Feminino , Citometria de Fluxo/métodos , Glicoproteínas/biossíntese , Humanos , Imunofenotipagem , Neovascularização Patológica , Peptídeos , Esteroides/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/biossínteseRESUMO
Fourier transform infrared (FT-IR) spectroscopy is a powerful tool for characterizing biological tissues and organisms, but it is plagued by replicate variation of various sources. Here, a method for estimating and correcting unwanted replicate variation in multivariate measurement signals, based on extended multiplicative signal correction (EMSC), is presented. Systematic patterns of unwanted methodological variations are estimated from replicate spectra, modeled by a linear subspace model, and implemented into EMSC. The method is applied to FT-IR spectra of two different sets of microorganisms (different double gene knockout strains of Saccharomyces cerevisiae and different species of Listeria) and compared to other preprocessing methods used in FT-IR absorption spectroscopy of microorganisms. The EMSC replicate correction turns out to perform best among the compared methods.
Assuntos
Algoritmos , Artefatos , Listeria/química , Listeria/isolamento & purificação , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/isolamento & purificação , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Interpretação Estatística de Dados , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
Exposure to particulate matter and ozone cause adverse airway reactions. Individual pollutant effects are often addressed separately, despite coexisting in ambient air. The present investigation was performed to study the effects of sequential exposures to diesel exhaust (DE) and ozone on airway inflammation in human subjects. Healthy subjects underwent bronchoscopy with bronchoalveolar lavage (BAL) and bronchial wash (BW) sampling on two occasions. Once following a DE exposure (with 300 mug.m(-3) particles with a 50% cut-off aerodynamic diameter of 10 mum) with subsequent exposure to O(3) (0.2 ppm) 5 h later. The other bronchoscopy was performed after a filtered air exposure followed by an ozone exposure, using an identical protocol. Bronchoscopy was performed 24 h after the start of the initial exposure. Significant increases in neutrophil and macrophage numbers were found in BW after DE followed by ozone exposure versus air followed by ozone exposure. DE pre-exposure also raised eosinophil protein X levels in BAL compared with air. The present study indicates additive effects of diesel exhaust on the ozone-induced airway inflammation. Together with similar results from a recent study with sequential diesel exhaust and ozone exposures, the present data stress a need to consider the interaction and cumulative effects of different air pollutants.
Assuntos
Poluentes Atmosféricos/efeitos adversos , Exposição por Inalação/efeitos adversos , Ozônio/efeitos adversos , Emissões de Veículos/toxicidade , Adulto , Ciclismo , Testes de Provocação Brônquica , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Neurotoxina Derivada de Eosinófilo/metabolismo , Feminino , Humanos , Inflamação/etiologia , Macrófagos Alveolares , Masculino , NeutrófilosRESUMO
BACKGROUND: Data are lacking from general population studies on how to define changes in lung function after bronchodilation. This study aimed to analyze different measures of bronchodilator response of forced expiratory volume in 1 second (FEV1), forced vital capacity (FVC) and slow vital capacity (SVC). MATERIALS AND METHODS: Data were derived from the Swedish Cardiopulmonary Bioimage Study (SCAPIS) Pilot study. This analysis comprised 1,050 participants aged 50-64 years from the general population. Participants were investigated using a questionnaire, and FEV1, FVC and SVC were recorded before and 15 minutes after inhalation of 400 µg of salbutamol. A bronchodilator response was defined as the relative change from baseline value expressed as the difference in units of percent predicted normal. Predictors of bronchodilator responses were assessed using multiple linear regression models. Airway obstruction was defined as FEV1/FVC ratio below lower limit of normal (LLN) before bronchodilation, and COPD was defined as an FEV1/FVC ratio below LLN after bronchodilation. Physician-diagnosed asthma was defined as an affirmative answer to "Have you ever had asthma diagnosed by a physician?". Asymptomatic never-smokers were defined as those not reporting physician-diagnosed asthma, physician-diagnosed COPD or emphysema, current wheeze or chronic bronchitis and being a lifelong never-smoker. RESULTS: Among all subjects, the greatest bronchodilator responses (FEV1, FVC and SVC) were found in subjects with asthma or COPD. The upper 95th percentile of bronchodilator responses in asymptomatic never-smokers was 8.7% for FEV1, 4.2% for FVC and 5.0% for SVC. The bronchodilator responses were similar between men and women. In a multiple linear regression model comprising all asymptomatic never-smokers, the bronchodilator response of FEV1 was significantly associated with airway obstruction and height. CONCLUSION: When the bronchodilator response in asymptomatic never-smokers is reported as the difference in units of predicted normal, significant reversibility of FEV1, FVC and SVC to bronchodilators is ~9%, 4% and 5%, respectively.
Assuntos
Agonistas de Receptores Adrenérgicos beta 2/administração & dosagem , Albuterol/administração & dosagem , Asma/fisiopatologia , Broncodilatadores/administração & dosagem , Volume Expiratório Forçado/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Doença Pulmonar Obstrutiva Crônica/fisiopatologia , Capacidade Vital/efeitos dos fármacos , Administração por Inalação , Asma/diagnóstico , Asma/epidemiologia , Doenças Assintomáticas , Feminino , Humanos , Modelos Lineares , Pulmão/fisiopatologia , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Doença Pulmonar Obstrutiva Crônica/diagnóstico , Doença Pulmonar Obstrutiva Crônica/epidemiologia , Fumar/efeitos adversos , Inquéritos e Questionários , Suécia/epidemiologia , Fatores de TempoRESUMO
We deleted the PBS2 gene encoding the MAP kinase activator of the osmosignaling HOG pathway in Saccharomyces cerevisiae and examined the effects on the kinetics of the osmoregulatory glycerol response and protein induction during adaptation to 0.7 M NaCl. Changes in protein expression as analyzed by two-dimensional polyacrylamide gel electrophoresis (2D PAGE) demonstrated that for the 29 proteins showing a 6-fold induction in wild-type cells during adaptation to NaCl stress, all displayed a decreased and delayed response in pbs2delta cells. Of the seven proteins that were identified, two were previously not known to be under HOG pathway control: Ald6p, an isoform of aldehyde dehydrogenase and Dak1p, a putative dihydroxyacetone kinase. The presence of a remaining significant induction in pbs2delta cells for about half of the examined proteins indicates existence of alternative osmosignaling pathway(s). Northern analysis of the salt induced transcription of GPD1 and GPP2, encoding the cytosolic glycerol-3-phosphate dehydrogenase and glycerol-3-phosphatase involved in the osmostress induced glycerol production, demonstrated an about 20-fold PBS2-dependent transient activation, in agreement with the previously reported transient nature of the signal transduced by the HOG pathway.
Assuntos
Adaptação Fisiológica/genética , Quinases de Proteína Quinase Ativadas por Mitógeno , Proteínas Quinases/genética , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/metabolismo , Clonagem Molecular , Teste de Complementação Genética , Osmose , Fenótipo , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Saccharomyces cerevisiae/genética , Deleção de Sequência , Cloreto de SódioRESUMO
The Saccharomyces cerevisiae gene PDC5 encodes the minor isoform of pyruvate decarboxylase (Pdc). In this work we show that expression of PDC5 but not that of PDC1, which encodes the major isoform, is repressed by thiamine. Hence, under thiamine limitation both PDC1 and PDC5 are expressed. PDC5 also becomes strongly expressed in a pdc1delta mutant. Two-dimensional gel electrophoresis of whole protein extracts shows that thiamine limitation stimulates the production of THI gene products and of Pdc5p. Deletion of PDC1 only stimulates production of Pdc5p. We conclude that the stimulation of PDC5 expression in a pdc1delta mutant is not due to a response to thiamine limitation.
Assuntos
Regulação Enzimológica da Expressão Gênica , Regulação Fúngica da Expressão Gênica , Piruvato Descarboxilase/metabolismo , Saccharomyces cerevisiae/enzimologia , Tiamina/metabolismo , Eletroforese em Gel Bidimensional , Regiões Promotoras Genéticas , Piruvato Descarboxilase/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Saccharomyces cerevisiae/genéticaRESUMO
The air pollutant ozone induces both airway inflammation and restrictions in lung function. These responses have been proposed to arise as a consequence of the oxidizing nature of ozone, depleting endogenous antioxidant defenses with ensuing tissue injury. In this study we examined the impact of an environmentally relevant ozone challenge on the antioxidant defenses present at the surface of the lung in two groups known to have profound differences in their antioxidant defense network: healthy control (HC) and mild asthmatic (MA) subjects. We hypothesized that baseline differences in antioxidant concentrations within the respiratory tract lining fluid (RTLF), as well as induced responses, would predict the magnitude of individual responsiveness. We observed a significant loss of ascorbate (ASC) from proximal (-45.1%, p <.01) and distal RTLFs (-11.7%, p <.05) in healthy subjects 6 h after the end of the ozone challenge. This was associated (Rs, -0.71, p <.01) with increased glutathione disulphide (GSSG) in these compartments (p =.01 and p <.05). Corresponding responses were not seen in asthmatics, where basal ASC concentrations were significantly lower (p <.01) and associated with elevated concentrations of GSSG (p <.05). In neither group was any evidence of lipid oxidation seen following ozone. Despite differences in antioxidant levels and response, the magnitude of ozone-induced neutrophilia (+20.6%, p <.01 [HC] vs. +15.2%, p =.01 [MA]) and decrements in FEV(1) (-8.0%, p <.01 [HC] vs. -3.2%, p <.05 [MA]) did not differ between the two groups. These data demonstrate significant differences between the interaction of ozone with RTLF antioxidants in MA and HC subjects. These responses and variations in basal antioxidant defense were not, however, useful predictive markers of group or individual responsiveness to ozone.
Assuntos
Antioxidantes/metabolismo , Asma/metabolismo , Dissulfeto de Glutationa/agonistas , Pulmão/metabolismo , Neutrófilos/metabolismo , Ozônio , Adulto , Ácido Ascórbico/antagonistas & inibidores , Ácido Ascórbico/metabolismo , Asma/induzido quimicamente , Asma/diagnóstico , Testes de Provocação Brônquica , Broncoscopia , Método Duplo-Cego , Feminino , Dissulfeto de Glutationa/metabolismo , Humanos , Pulmão/citologia , Pulmão/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Neutrófilos/efeitos dos fármacos , Ozônio/efeitos adversos , Valor Preditivo dos Testes , Testes de Função Respiratória , Sistema Respiratório/citologia , Sistema Respiratório/efeitos dos fármacos , Sistema Respiratório/metabolismoRESUMO
This review describes the metabolic alterations and adaptations of yeast cells in response to osmotic stress. The basic theme of the cellular response is known to be exclusion of the extracellular stress agent salt and intracellular accumulation of the compatible solute glycerol. Molecular details of these basic processes are currently rather well known. However, analysis of expression changes during adaptation to salt has revealed a number of metabolic surprises. These include the induced expression of genes involved in glycerol dissimilation as well as trehalose turnover. The physiological rationale for these responses to osmotic stress is discussed. A model is presented in which it is hypothesised that the two pathways function as glycolytic safety valves during adaptation to stress.
Assuntos
Adaptação Fisiológica , Saccharomyces cerevisiae/metabolismo , Cloreto de Sódio/farmacologia , Concentração Osmolar , Saccharomyces cerevisiae/fisiologia , Equilíbrio HidroeletrolíticoRESUMO
Labelling of Saccharomyces cerevisiae grown in 0.7 M NaCl with 35S-methionine revealed a 5-6 fold lowering of the methionine incorporation into protein, which could not be attributed solely to the approximately 50% longer generation time of cells grown in 0.7 M NaCl. Subsequent studies of the high affinity methionine uptake system showed a strongly reduced uptake of methionine during growth in 0.7 M NaCl medium. This reduced uptake was shown to be strain-independent and caused mainly by an approximately 20-fold lowered maximum velocity (Vmax) of the transport system, while the substrate affinity (Km) displayed only a minor change. A salt-instigated reduction of uptake was furthermore demonstrated for the leucine and histidine high affinity uptake systems and also for a mixture of 15 different amino acids. We therefore suggest that the reduced amino acid uptake is a general phenomenon observed in salt-grown cells.
Assuntos
Metionina/farmacocinética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/metabolismo , Cloreto de Sódio/farmacologia , Aminoácidos/metabolismo , Transporte Biológico/fisiologia , Proteínas de Transporte/metabolismo , Meios de Cultura/farmacologia , Cinética , Pressão Osmótica , Saccharomyces cerevisiae/efeitos dos fármacosRESUMO
Saccharomyces cerevisiae exponentially growing in basic or 0.7 M NaCl medium were isotopically labelled with 35S-methionine, followed by protein separation and quantification by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) combined with computerised image analysis. The electrophoretic separation resolved about 650 proteins of which 13 displayed significant and at least 2-fold changes in rate of synthesis during saline growth. By sequencing of 2D-PAGE resolved proteins, one of the 8 induced spot, p42.9/5.5, was shown to correspond to the full length (containing the N-terminal extension) product of the GPD1 gene encoding the cytoplasmic glycerol 3-phosphate dehydrogenase. The expression of the TDH3 gene, glyceraldehyde 3-phosphate dehydrogenase, and the ENO2 gene, enolase, decreased during growth in NaCl medium, declines hypothesised to have an impact on the flux to glycerol.
Assuntos
Proteínas Fúngicas/biossíntese , Saccharomyces cerevisiae/metabolismo , Sequência de Aminoácidos , Meios de Cultura , Eletroforese em Gel Bidimensional , Proteínas Fúngicas/genética , Proteínas Fúngicas/isolamento & purificação , Genes Fúngicos , Gliceraldeído-3-Fosfato Desidrogenases/biossíntese , Gliceraldeído-3-Fosfato Desidrogenases/genética , Gliceraldeído-3-Fosfato Desidrogenases/isolamento & purificação , Glicerol/metabolismo , Glicerolfosfato Desidrogenase/biossíntese , Glicerolfosfato Desidrogenase/genética , Glicerolfosfato Desidrogenase/isolamento & purificação , Dados de Sequência Molecular , Fragmentos de Peptídeos/biossíntese , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/isolamento & purificação , Fosfopiruvato Hidratase/biossíntese , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/isolamento & purificação , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Cloreto de Sódio , Equilíbrio HidroeletrolíticoRESUMO
In clinical neuroscience as well as in many other clinical disciplines, the completion of the human genome project offers a new possibility to identify and localize the products of the genes, the proteins. Nuclear proteins are synthesized in the cytoplasm and imported into the nucleus by multiple pathways. The mechanisms by which nuclear accumulation of different molecular species occur are unclear but it is apparent that changes in the cellular and molecular events associated with the accumulation of nuclear proteins sometimes precedes transformation of cells into diseased states. The significance of the accumulation and the operation of nuclear proteins remain to be elucidated in detail. Such knowledge will play a key role in the understanding of the regulation of transcription and its disturbances in several of our most devastating diseases. In this paper we present a strategy to identify nuclear associated proteins in small samples by using two-dimensional electrophoresis and mass spectrometry. We have used human blood lymphocytes as a model, but the method should be rather general for any kind of tissue. Twenty two proteins were randomly chosen, and of these 18 proteins were identified by database searching of mass spectrometric data, obtained from in-gel tryptic digests of the spots. Thirteen proteins recently described with nuclear localization and function were identified, and five proteins; calgranulin B, glyceraldehyde-3-phosphate dehydrogenase (G3P2), a TATA-binding protein (ATBP), tubulin beta chain and moesin were also identified as being nuclear associated. The presented data clearly shows of the great role of two-dimensional gel electrophoresis and modern mass spectrometry in the excavation of the protein patterns on the subcellular level, and the ability to use small samples well suited for clinical screening.
Assuntos
Doenças do Sistema Nervoso Central/sangue , Eletroforese em Gel Bidimensional , Linfócitos/metabolismo , Espectrometria de Massas , Proteínas Nucleares/análise , Proteínas Nucleares/sangue , HumanosRESUMO
To determine if diesel exhaust (DE) exposure modifies the antioxidant defense network within the respiratory tract lining fluids, a randomized, single blinded, crossover control study using nasal lavage and flexible video bronchoscopy with bronchial and bronchoalveolar lavage was performed. Fifteen healthy, non-smoking, asymptomatic subjects were exposed to filtered air or diluted diesel exhaust (300mg m(-3) particulates, 1.6ppm nitrogen dioxide) for one hour on 2 separate occasions, at least three weeks apart. To examine the kinetics of any DE-induced antioxidant reactions, nasal lavage fluid and blood samples were collected prior to, immediately after, and 5 1/2 hours post exposure. Bronchoscopy was performed 6 hours after the end of DE exposure. Ascorbic acid, uric acid and reduced glutathione (GSH) concentrations were determined in nasal, bronchial, bronchoalveolar lavage and plasma samples. Malondialdehyde (MDA) and protein carbonyl concentrations were determined in plasma and bronchoalveolar lavage samples. Nasal lavage ascorbic acid concentration increased 10-fold during DE exposure [1.02 (0.26-2.09) Vs 7.13 (4.66-10.79) micromol/L(-1)], but returned to basal levels 5.5 hours post-exposure [0.75 (0.26-1.51) micromol/L(-1)]. There was no significant effect of DE exposure on nasal lavage uric acid or GSH concentration. DE exposure did not influence plasma, bronchial wash, or bronchoalveolar lavage antioxidant concentrations and no change in MDA or protein carbonyl concentrations were found. The physiological response to acute DE exposure is an increase in the level of ascorbic acid in the nasal cavity. This response appears to be sufficient to prevent further oxidant stress in the respiratory tract of normal individuals.
Assuntos
Ácido Ascórbico/metabolismo , Líquidos Corporais/metabolismo , Cavidade Nasal/metabolismo , Emissões de Veículos , Adulto , Antioxidantes/metabolismo , Ácido Ascórbico/sangue , Brônquios/metabolismo , Líquido da Lavagem Broncoalveolar/química , Estudos Cross-Over , Feminino , Glutationa/sangue , Glutationa/metabolismo , Humanos , Cinética , Masculino , Malondialdeído/metabolismo , Irrigação Terapêutica , Ácido Úrico/sangue , Ácido Úrico/metabolismoRESUMO
A cultivation set-up for multiple cultures has been designed that can be used for anaerobic screening for quantitative changes in growth rate or other analyses, e.g. protein composition of different strains. The developed gas distribution system provides a reproducible level of anaerobicity in 30 cultivation flasks and resembles the open system of a high-performance bioreactor in that it ensures cultivation at atmospheric pressure and avoids supersaturation of carbon dioxide. The system is cheap and user-friendly and allows rapid screenings of many strains simultaneously.