Expression patterns of peroxiredoxin genes in bronchial epithelial cells exposed to diesel exhaust particles.

Several mechanisms have been suggested to explain the adverse effects of air pollutants on airway cells. One such explanation is the presence of high concentrations of oxidants and pro-oxidants in environmental pollutants. All animal and plant cells have developed several mechanisms to prevent damage by oxidative molecules. Among these, the peroxiredoxins (PRDXs) are of interest due to a high reactivity with reactive oxygen species (ROS) through the functioning of the thioredoxin/thioredoxin reductase system. This study aimed to verify the gene expression patterns of the PRDX family in bronchial epithelial airway cells (BEAS-2B) cells exposed to diesel exhaust particles (DEPs) at a concentration of 15 µg/mL for 1 or 2 h because this it is a major component of particulate matter in the atmosphere. There was a significant decrease in mRNA fold changes of PRDX2 (0.43 ± 0.34; *p = 0.0220), PRDX5 (0.43 ± 0.34; *p = 0.0220), and PRDX6 (0.33 ± 0.25; *p = 0.0069) after 1 h of exposure to DEPs. The reduction in mRNA levels may consequently lead to a decrease in the levels of PRDX proteins, increasing oxidative stress in bronchial epithelial cells BEAS-2B and thus, negatively affecting cellular functions.

Therapeutic l-asparaginase: upstream, downstream and beyond.

l-asparaginase (l-asparagine amino hydrolase, E.C.3.5.1.1) is an enzyme clinically accepted as an antitumor agent to treat acute lymphoblastic leukemia and lymphosarcoma. It catalyzes l-asparagine (Asn) hydrolysis to l-aspartate and ammonia, and Asn effective depletion results in cytotoxicity to leukemic cells. Microbial l-asparaginase (ASNase) production has attracted considerable attention owing to its cost effectiveness and eco-friendliness. The focus of this review is to provide a thorough review on microbial ASNase production, with special emphasis to microbial producers, conditions of enzyme production, protein engineering, downstream processes, biochemical characteristics, enzyme stability, bioavailability, toxicity and allergy potential. Some issues are also highlighted that will have to be addressed to achieve better therapeutic results and less side effects of ASNase use in cancer treatment: (a) search for new sources of this enzyme to increase its availability as a drug; (b) production of new ASNases with improved pharmacodynamics, pharmacokinetics and toxicological profiles, and (c) improvement of ASNase production by recombinant microorganisms. In this regard, rational protein engineering, directed mutagenesis, metabolic flux analysis and optimization of purification protocols are expected to play a paramount role in the near future.

Can environmental pollution by metals change genetic diversity? Ucides cordatus (Linnaeus, 1763) as a study case in Southeastern Brazilian mangroves.

Industrial areas on estuarine systems are commonly affected by heavy metals, affecting all local biota. Random Amplified Polymorphic DNA (RAPD) was used to evaluate genetic diversity of Ucides cordatus at mangroves in southeastern Brazil (Juréia, J; São Vicente, SV; and Cubatão, C), with distinct pollution levels by metals. The genetic diversity of this species was compared with concentrations of metals (Cd, Pb, Cu, Cr and Hg) in the environment. A pollution gradient was confirmed (SV>C>J), with low levels detected in water, except for mercury in SV. All metals in the sediment samples were below Threshold Effect Level (TEL), without an apparent biological risk to the biota. Genetic distance was very similar between J and C, with SV occurring as an out-group. RAPD was a powerful tool to investigate the effect of metal pollution on genetic diversity of this mangrove crab, and to evaluate the conservation status of the mangrove ecosystem.

Biopharmaceuticals from microorganisms: from production to purification.

The use of biopharmaceuticals dates from the 19th century and within 5-10 years, up to 50% of all drugs in development will be biopharmaceuticals. In the 1980s, the biopharmaceutical industry experienced a significant growth in the production and approval of recombinant proteins such as interferons (IFN α, ß, and γ) and growth hormones. The production of biopharmaceuticals, known as bioprocess, involves a wide range of techniques. In this review, we discuss the technology involved in the bioprocess and describe the available strategies and main advances in microbial fermentation and purification process to obtain biopharmaceuticals.

Crystallization and preliminary X-ray diffraction analysis of NADPH-dependent thioredoxin reductase I from Saccharomyces cerevisiae.

Thioredoxin reductase 1 (Trr1) from Saccharomyces cerevisiae is a member of the family of pyridine nucleotide-disulfide oxidoreductases capable of reducing the redox-active disulfide bond of the cytosolic thioredoxin 1 (Trx1) and thioredoxin 2 (Trx2). NADPH, Trr1 and Trx1 (or Trx2) comprise the thioredoxin system, which is involved in several biological processes, including the reduction of disulfide bonds and response to oxidative stress. Recombinant Trr1 was expressed in Escherichia coli as a His6-tagged fusion protein and purified by nickel-affinity chromatography. The protein was crystallized using the hanging-drop vapour-diffusion method in the presence of PEG 3000 as precipitant after treatment with hydrogen peroxide. X-ray diffraction data were collected to a maximum resolution of 2.4 A using a synchrotron-radiation source. The crystal belongs to the centred monoclinic space group C2, with unit-cell parameters a = 127.97, b = 135.41, c = 75.81 A, beta = 89.95 degrees. The crystal structure was solved by molecular-replacement methods and structure refinement is in progress.

Crystallization and preliminary X-ray crystallographic studies of glutaredoxin 2 from Saccharomyces cerevisiae in different oxidation states.

Glutaredoxins are small (9-12 kDa) heat-stable proteins that are highly conserved throughout evolution; the glutaredoxin active site (Cys-Pro-Tyr-Cys) is conserved in most species. Five glutaredoxin genes have been identified in Saccharomyces cerevisiae; however, Grx2 is responsible for the majority of oxidoreductase activity in the cell, suggesting that its primary function may be the detoxification of mixed disulfides generated by reactive oxygen species (ROS). Recombinant Grx2 was expressed in Escherichia coli as a 6xHis-tagged fusion protein and purified by nickel-affinity chromatography. Prior to crystallization trials, the enzyme was submitted to various treatments with reducing agents and peroxides. Crystals suitable for X-ray diffraction experiments were obtained from untreated protein and protein oxidized with t-butyl hydroperoxide (10 mM). Complete data sets were collected to resolutions 2.15 and 2.05 A for untreated and oxidized Grx2, respectively, using a synchrotron-radiation source. The crystals belong to space group P4(1)2(1)2, with similar unit-cell parameters.

Biopharmaceuticals from microorganisms: from production to purification

ABSTRACT The use of biopharmaceuticals dates from the 19th century and within 5-10 years, up to 50% of all drugs in development will be biopharmaceuticals. In the 1980s, the biopharmaceutical industry experienced a significant growth in the production and approval of recombinant proteins such as interferons (IFN α, β, and γ) and growth hormones. The production of biopharmaceuticals, known as bioprocess, involves a wide range of techniques. In this review, we discuss the technology involved in the bioprocess and describe the available strategies and main advances in microbial fermentation and purification process to obtain biopharmaceuticals.

Biopharmaceuticals from microorganisms: from production to purification

ABSTRACT The use of biopharmaceuticals dates from the 19th century and within 5-10 years, up to 50% of all drugs in development will be biopharmaceuticals. In the 1980s, the biopharmaceutical industry experienced a significant growth in the production and approval of recombinant proteins such as interferons (IFN , , and ) and growth hormones. The production of biopharmaceuticals, known as bioprocess, involves a wide range of techniques. In this review, we discuss the technology involved in the bioprocess and describe the available strategies and main advances in microbial fermentation and purification process to obtain biopharmaceuticals.

Análise contínua de medidas de cateter de artéria pulmonar volumétrico, ecotransesofágico, variações da pressão arterial sistêmica e marcadores de hipoperfusão tissular no choque hemorrágico em suínos / Continuous analyses of pulmonary, volumetric artery catheter parameters, transesophageal echocardiography, pressure pulse variation, and biomarkers of tissue hypoperfusion during hemorrhagic shock in swine: experimental study in swines

INTRODUÇÃO: Diferentes parâmetros hemodinâmicos, incluindo os indicadores estáticos de pré-carga cardíaca como o índice de volume diastólico final ventrículo direito (IVDFVD) e parâmetros dinâmicos como a variação de pressão de pulso (VPP) têm sido usados na tomada de decisão para considerar o processo da expansão volêmica em pacientes em estado grave. O objetivo deste estudo foi comparar a reanimação por fluidos guiados tanto por VPP ou IVDFVD após choque hemorrágico induzido experimentalmente. MÉTODO: vinte e seis suínos anestesiados e ventilados mecanicamente foram alocados em três grupos: controle (Grupo I), VPP (Grupo II) e IVDFVD (Grupo III). Foi induzido choque hemorrágico por retirada de sangue até atingir a pressão arterial média de 40mmhg, que foi mantida por 60 minutos. Parâmetros foram medidos no tempo basal (B), no tempo do choque (Choque 0), sessenta minutos depois do choque (Choque 60), imediatamente depois da ressuscitação com hidroxietilamido 6% (130/0. 4) (R0), uma hora (R60) e duas horas (R120) depois ressuscitação. Os pontos de avaliação da reanimação por fluidos foram determinados pelo retorno aos valores basais iniciais de VPP e IVDFVD. A análise estatística dos dados foi baseada em ANOVA para medidas repetidas seguidos pelo teste de Bonferroni (P<0.05%). RESULTADOS: O volume e tempo para ressuscitação foram maiores no grupo III do que no grupo II (Grupo III = 1305±331ml e Grupo II = 965±245ml; p<0.05 e Grupo III = 24.8± 4.7min e Grupo II = 8.8 ± 1.3 min, p<0.01, respectivamente). Todos os parâmetros estáticos e dinâmicos, bem como os biomarcadores de oxigenação tecidual foram afetados pelo choque hemorrágico e quase todos os parâmetros foram totalmente restaurados após a reanimação em ambos os grupos. CONCLUSÃO: Neste estudo em modelo de choque hemorrágico, a reanimação guiada pelo VPP utilizou menor quantidade de fluido e menor quantidade de tempo do que quando guiado por IVDFVD derivado de cateter de artéria pulmonar.

INTRODUCTION: Different hemodynamic parameters, including static indicators of cardiac preload as right ventricular end-diastolic volume index (RVEDVI) and dynamic parameters as pulse pressure variation (PPV) have been used in the decision-making process regarding volume expansion in critically ill patients. The objective of this study was to compare fluid resuscitation guided by either PPV or RVEDVI after experimentally-induced hemorrhagic shock. METHODS: 26 anesthetized and mechanically ventilated pigs were allocated into control (Group-I), PPV (Group-II) and RVEDVI (Group- III). Hemorrhagic shock was induced by blood withdrawal to target mean arterial pressure of 40mmHg, maintained for 60 minutes. Parameters were measured at baseline, time of shock, sixty minutes after shock, immediately after resuscitation with hydroxyethyl starch 6% (130/0.4), one hour and two hours thereafter. The endpoint of fluid resuscitation was determined as the baseline values of PPV and RVEDVI. Statistical analysis of data was based on ANOVA for repeated measures followed by the Bonferroni test (P<0.05). RESULTS: Volume and time to resuscitation were higher in Group-III than in Group-II (Group-III = 1305±331ml and Group-II = 965±245ml; p<0.05 and Group-IIII = 24.8±4.7min and Group-II = 8.8±1.3 min, p<0.05, respectively). All static and dynamic parameters and biomarkers of tissue oxygenation were affected by hemorrhagic shock and nearly all parameters were restored after resuscitation in both groups. CONCLUSION: In the proposed model of hemorrhagic shock, resuscitation to the established endpoints was achieved within a smaller amount of time and with less volume when guided by PPV than when guided by pulmonary artery catheter-derived RVEDVI.