Mitochondrial bioenergetics and redox state are unaltered in Trypanosoma cruzi isolates with compromised mitochondrial complex I subunit genes

In trypanosomatids the involvement of mitochondrial complex I in NADH oxidation has long been debated. Here, we took advantage of natural Trypanosoma cruzi mutants which present conspicuous deletions in ND4, ND5 and ND7 genes coding for complex I subunits to further investigate its functionality. Mitochondrial bioenergetics of wild type and complex I mutants showed no significant differences in oxygen consumption or respiratory control ratios in the presence of NADH-linked substrates or FADH(2)-generating succinate. No correlation could be established between mitochondrial membrane potentials and ND deletions. Since release of reactive oxygen species occurs at complex I, we measured mitochondrial H(2)O(2) formation induced by different substrates. Significant differences not associated to ND deletions were observed among the parasite isolates, demonstrating that these mutations are not important for the control of oxidant production. Our data support the notion that complex I has a limited function in T. cruzi.

trans, trans-2,4-decadienal induces mitochondrial dysfunction and oxidative stress

Lipid peroxidation produces a large number of reactive aldehydes as secondary products. We have previously shown that the reaction of cytochrome c with trans,trans-2, 4-decadienal (DDE), an aldehyde generated as a product of lipid peroxidation in cell membranes, results in the formation of adducts. Mass spectrometry analysis indicated that His-33, Lys-39, Lys-72 and Lys-100 in cytochrome c were modified by DDE. In the present work, we investigated the effect of DDE on isolated rat liver mitochondria. DDE (162 mu M) treatment increases the rate of mitochondrial oxygen consumption. Extensive mitochondrial swelling upon treatment with DDE (900 nM-162 mu M) was observed by light scattering and transmission electron microscopy experiments. DDE-induced loss of inner mitochondrial membrane potentials, monitored by safranin O fluorescence, was also observed. Furthermore, DDE-treated mitochondria showed an increase in lipid peroxidation, as monitored by MDA formation. These results suggest that reactive aldehydes promote mitochondrial dysfunction.

cis-4-decenoic acid provokes mitochondrial bioenergetic dysfunction in rat brain

Aims: In the present work we investigated the in vitro effect of cis-4-decenoic acid, the pathognomonic metabolite of medium-chain acyl-CoA dehydrogenase deficiency, on various parameters of bioenergetic homeostasis in rat brain mitochondria. Main methods: Respiratory parameters determined by oxygen consumption were evaluated, as well as membrane potential, NAD(P)H content, swelling and cytochrome c release in mitochondrial preparations from rat brain, using glutamate plus malate or succinate as substrates. The activities of citric acid cycle enzymes were also assessed. Key findings: cis-4-decenoic acid markedly increased state 4 respiration, whereas state 3 respiration and the respiratory control ratio were decreased. The ADP/O ratio, the mitochondrial membrane potential, the matrix NAD(P)H levels and aconitase activity were also diminished by cis-4-decenoic acid. These data indicate that this fatty acid acts as an uncoupler of oxidative phosphorylation and as a metabolic inhibitor. cis-4-decenoic acid also provoked a marked mitochondrial swelling when either KCl or sucrose was used in the incubation medium and also induced cytochrome c release from mitochondria, suggesting a non-selective permeabilization of the inner mitochondria! membrane. Significance: It is therefore presumed that impairment of mitochondrial homeostasis provoked by cis-4-decenoic acid may be involved in the brain dysfunction observed in medium-chain acyl-CoA dehydrogenase deficient patients. (C) 2010 Elsevier Inc. All rights reserved.

Myoglobin-H(2)O(2) catalyzes the oxidation of beta-ketoacids to alpha-dicarbonyls: Mechanism and implications in ketosis

Acetoacetate (AA) and 2-methylacetoacetate (MAA) are accumulated in metabolic disorders such as diabetes and isoleucinemia. Here we examine the mechanism of AA and MAA aerobic oxidation initiated by myoglobin (Mb)/H(2)O(2). We propose a chemiluminescent route involving a dioxetanone intermediate whose thermolysis yields triplet alpha-dicarbonyl species (methylglyoxal and diacetyl). The observed ultraweak chemiluminescence increased linearly on raising the concentration of either Mb (10-500 mu M) or AA (10-100 mM). Oxygen uptake studies revealed that MAA is almost a 100-fold more reactive than AA. EPR spin-trapping studies with MNP/MAA revealed the intermediacy of an alpha-carbon-centered radical and acetyl radical. The latter radical, probably derived from triplet diacetyl, is totally suppressed by sorbate, a well-known quencher of triplet carbonyls. Furthermore, an EPR signal assignable to MNP-AA(center dot) adduct was observed and confirmed by isotope effects. Oxygen consumption and a-dicarbonyl yield were shown to be dependent on AA or MAA concentrations (1-50 mM) and on H(2)O(2) or tert-butOOH added to the Mb-containing reaction mixtures. That ferrylMb is involved in a peroxidase cycle acting on the substrates is suggested by the reaction pH profiles and immunospin-trapping experiments. The generation of radicals and triplet dicarbonyl products by Mb/H(2)O(2)/beta-ketoacids may contribute to the adverse health effects of ketogenic unbalance. (C) 2011 Elsevier Inc. All rights reserved.

The isatin-Schiff base copper(II) complex Cu(isaepy)(2) acts as delocalized lipophilic cation, yields widespread mitochondrial oxidative damage and induces AMP-activated protein kinase-dependent apoptosis

We previously demonstrated that Bis[(2-oxindol-3-ylimino)-2-(2-aminoethyl) pyridine-N, N`] copper(II) [Cu(isaepy)(2)] was an efficient inducer of the apoptotic mitochondrial pathway. Here, we deeply dissect the mechanisms underlying the ability of Cu(isaepy)(2) to cause mitochondriotoxicity. In particular, we demonstrate that Cu(isaepy)(2) increases NADH-dependent oxygen consumption of isolated mitochondria and that this phenomenon is associated with oxy-radical production and insensitive to adenosine diphosphate. These data indicate that Cu(isaepy)(2) behaves as an uncoupler and this property is also confirmed in cell systems. Particularly, SH-SY5Y cells show: (i) an early loss of mitochondrial transmembrane potential; (ii) a decrease in the expression levels of respiratory complex components and (iii) a significant adenosine triphosphate (ATP) decrement. The causative energetic impairment mediated by Cu(isaepy)(2) in apoptosis is confirmed by experiments carried out with rho(0) cells, or by glucose supplementation, where cell death is significantly inhibited. Moreover, gastric and cervix carcinoma AGS and HeLa cells, which rely most of their ATP production on oxidative phosphorylation, show a marked sensitivity toward Cu(isaepy)(2). Adenosine monophosphate-activated protein kinase (AMPK), which is activated by events increasing the adenosine monophosphate: ATP ratio, is deeply involved in the apoptotic process because the overexpression of its dominant/negative form completely abolishes cell death. Upon glucose supplementation, AMPK is not activated, confirming its role as fuel-sensing enzyme that positively responds to Cu(isaepy)(2)-mediated energetic impairment by committing cells to apoptosis. Overall, data obtained indicate that Cu(isaepy)(2) behaves as delocalized lipophilic cation and induces mitochondrial-sited reactive oxygen species production. This event results in mitochondrial dysfunction and ATP decrease, which in turn triggers AMPK-dependent apoptosis.

Expression Profile of Rat Hippocampal Neurons Treated with the Neuroprotective Compound 2,4-Dinitrophenol: Up-Regulation of cAMP Signaling Genes

2,4-Dinitrophenol (DNP) is classically known as a mitochondrial uncoupler and, at high concentrations, is toxic to a variety of cells. However, it has recently been shown that, at subtoxic concentrations, DNP protects neurons against a variety of insults and promotes neuronal differentiation and neuritogenesis. The molecular and cellular mechanisms underlying the beneficial neuroactive properties of DNP are still largely unknown. We have now used DNA microarray analysis to investigate changes in gene expression in rat hippocampal neurons in culture treated with low micromolar concentrations of DNP. Under conditions that did not affect neuronal viability, high-energy phosphate levels or mitochondrial oxygen consumption, DNP induced up-regulation of 275 genes and down-regulation of 231 genes. Significantly, several up-regulated genes were linked to intracellular cAMP signaling, known to be involved in neurite outgrowth, synaptic plasticity, and neuronal survival. Differential expression of specific genes was validated by quantitative RT-PCR using independent samples. Results shed light on molecular mechanisms underlying neuroprotection by DNP and point to possible targets for development of novel therapeutics for neurodegenerative disorders.

Sugarcane spirit extracts of oak and Brazilian woods: antioxidant capacity and activity

Sugarcane spirit extracts of six different Brazilian woods for potential use in manufacturing aging casks were compared with similar extracts of five oak samples from different geographic origin and heat treatment regarding: (1) content of phenolics and copper; (2) radical reducing capacity and reactivity toward 2,2-diphenyl-1-picrylhydrazyl radical (DPPH center dot); and (3) effect on the rate of oxygen depletion rate in a peroxidating lipid model system. Total phenolic contents of the Brazilian wood extracts ranged from 0.65 (canela-sassafras) to 6.4 (jatoba) mmol(GAE) L(-1) and from 1.39 to 2.87 mmol(GAE) L(-1) for oak extracts. Flavonoids ranged from 1.54 x 10(-4) (ipe) to 6.5 x 10(-2) (oak) mmol(rutin) L(-1), and tannins from below the detection limit to 0.22 (jatoba) mmol(tannic acid) L(-1). Correlation was observed for the antioxidant capacity versus phenolics/flavonoids/tannins content, where oak extracts exhibit the highest radical scavenging capacity compared to Brazilian woods. Rate constant for radical scavenging by the extracts ranged from 4.9 x 10(3) M(-1) s(-1)(canela-sassafras) to 9.7 x 10(4) M(-1) s(-1) (oak). The oxygen consumption index showed the Brazilian woods amendoim and jatoba to be more efficient inhibitors than the oak extracts for lipid autoxidation initiated by metmyoglobin, despite that the oak extracts seem to be more efficient to scavenge DPPH center dot. No simple correlation with phenolics or copper content could be established, and a prooxidative tendency was observed for the extracts of canela-sassafras, castanheira, and louro-canela.

Efeito do exercício aeróbio moderado em gestantes com sobrepeso avaliado através de teste de exercício submáximo : um ensaio clínico randomizado

Objetivo: Avaliar os efeitos de um programa de exercício aeróbio sobre o condicionamento cardiorrespiratório em gestantes hígidas, de baixo risco, com sobrepeso. Métodos: 92 mulheres gestantes com sobrepeso (índice de massa corporal 26-31kg/m2), idade ≥ 20 anos, idade gestacional ≤ 20 semanas, com ausência de diabetes e hipertensão, foram alocadas aleatoriamente para realizar exercício aeróbio três vezes por semana com uma hora de duração ou para realizar sessões de relaxamento no grupo controle. Foram realizados dois testes de exercício submáximo em esteira, utilizando protocolo de rampa na entrada do estudo e outro teste após 12 semanas. Resultados: Em teste de exercício submáximo 12 semanas após randomização, o consumo de oxigênio (VO2) no limiar anaeróbio aumentou 17% (± 3) no grupo intervenção enquanto reduziu 16% (± 3) no grupo controle, de modo que após 12 semanas de exercício ajustado através da análise de covariância pelo o VO2 no limiar na linha de base, idade gestacional e idade materna foi de 2,68ml/kg/min (IC 95% 1,32-4,03) maior, P = 0,002. Conclusão: Exercício aeróbio realizado em gestantes com sobrepeso produz um aumento no limiar anaeróbio, sobrepondo os efeitos negativos da gestação sobre o condicionamento cardiorrespiratório em mulheres com estilo de vida sedentário.

Comportamento do reator seqüencial em batelada (RSB) sob estado estacionário dinâmico utilizando idade do lodo como parâmetro de controle operacional

O reator seqüencial em batelada (RSB) é uma variante de lodos ativados capaz de promover a remoção da matéria orgânica, a remoção dos nutrientes e a separação da fase sólida da líquida em uma unidade. A valorização das áreas urbanas, a carência de tratamento terciário e a crescente necessidade de redução nas dimensões de estações de tratamento de esgoto devem impulsionar o desenvolvimento de pesquisas sobre RSB em curto espaço de tempo. A partir deste cenário, o presente trabalho teve como objetivo modelar o comportamento do reator seqüencial em batelada a partir da teoria desenvolvida por Marais e colaboradores. Dentro deste contexto, a cinética de oxidação dos compostos orgânicos e do nitrogênio na forma amoniacal foi descrita e modelada. O trabalho experimental foi realizado em duas escalas: bancada e piloto. O experimento em escala de bancada foi dividido em duas fases. Foram utilizados dois RSBs e um sistema de fluxo contínuo. Um reator seqüencial em batelada (RSB1) foi operado com idade de lodo. O outro reator em batelada (RSB2) foi operado em função da relação F/M e o sistema de fluxo contínuo (FC1) por idade de lodo. Estes reatores foram utilizados como controle no monitoramento do RSB1 Na primeira fase, os três sistemas removeram apenas matéria orgânica. Na fase seguinte, removeram matéria orgânica e nitrogênio. A partir dos resultados obtidos em escala de bancada, foi possível concluir que o modelo desenvolvido pode ser aplicado ao reator seqüencial em batelada operando com idade de lodo, permitindo determinar a qualidade do efluente, a produção de lodo e o consumo de oxidante. Além disso, foi possível descrever o comportamento da taxa de consumo de oxigênio em função da oxidação da matéria orgânica biodegradável e da oxidação do nitrogênio na forma amoniacal. O reator seqüencial em batelada operado com idade de lodo (RSB1) alcançou remoção média de matéria orgânica de 90 % nas idades de lodo de 30, 20, 10 e 5 dias. A remoção média de nitrogênio mais elevada foi obtida na idade de lodo de 20 dias e atingiu 87 %. Nas demais idades de lodo a remoção média de nitrogênio variou entre 79 e 42 %. A modelagem do comportamento do reator seqüencial em batelada resultou numa proposta de metodologia para o dimensionamento que tem como finalidade abolir critérios obsoletos e inadequados para o dimensionamento de lodos ativados em batelada No experimento em escala piloto, foram utilizados um reator seqüencial em batelada, denominado RSB, e um sistema de fluxo contínuo com a configuração Bardenpho, denominado FC. Os sistemas de lodos ativados sob investigação foram monitorados em duas idades de lodo: 30 e 10 dias. Os dados do experimento em escala piloto mostraram que os processos físico-químicos e biológicos envolvidos na remoção de matéria orgânica e nitrogênio no RSB foram mais eficientes do que no Bardenpho quando trataram o mesmo esgoto doméstico e foram submetidos às mesmas condições operacionais. No RSB, obteve-se 88 e 89 % de remoção de matéria orgânica nas idades de lodo de 10 e 30 dias, respectivamente. Nesta seqüência das idades de lodo, a eficiência do Bardenpho caiu de 87 para 76 %. O sistema de fluxo contínuo removeu 66 e 52 % do nitrogênio total afluente nas idades de lodo de 10 e 30 dias, respectivamente. A eficiência do RSB na remoção de nitrogênio foi determinada apenas na idade de lodo de 10 dias e alcançou 69 %. A partir dos resultados obtidos em escala de bancada e piloto, constata-se que o reator seqüencial em batelada operando com idade de lodo pode ser utilizado no tratamento de esgoto doméstico e obter eficiência na remoção de matéria orgânica e nitrogênio igual ou superior ao sistema de fluxo contínuo.

Avaliação da biodegradabilidade de petróleo utilizando microrganismos isolados do estuário do Rio Potengi (RN)

The oil activity in the Rio Grande do Norte State (RN) is a permanent threat to coastal ecosystems, particularly mangroves, with the possibility of oil spills. In this context, the objective of this study was to evaluate the potential resistance of the mangrove environment of a possible spill. Were selected and isolated microorganisms degrading oil by the technique of enrichment cultures and formation of a bacterial consortium. The kinetic study of the consortium was held in rotary incubator shaken at 150 rpm and 30° C. Samples were taken at intervals of 4 hours for analysis of cell concentration and surface tension. The biodegradation was monitored using two methods of respirometry: manometric (OxiTop-C ®) and conductivimetry, where the biodegradation of oil was estimated indirectly by oxygen consumption and CO2 production, respectively. Furthermore, it was used a full 2² factorial design with triplicate at central point to the runs that used the conductivimetric methodology.. The technique of enrichment cultures allowed to obtain thirteen bacterial strains. Kinetic study of the consortium, we can showed the absence of the lag phase, reaching a maximum cell concentration of 2.55 g / L at 16 h of cultivation and a reduction on surface tension. When we adopted the methodology of OxiTop-C was detected a band indicating biodegradability (1% oil v/v), however when we used the conductivimetry methodology did not observe any band that would indicate effective biodegradation. By monitoring a process of biodegradation is necessary to observe the methodology will be adopted to evaluate the biodegradation process, since for the same conditions adopted different methodologies can produce different results. The oil-degrading isolates from soils of the mangrove estuary Potengi / RN are largely to be used in bioremediation strategies of these places, in the case of a possible oil spill, or it can be used in the treatment of waste oil generated in saline environments, since they are optimized the conditions of the tests so that the efficiency of biodegradation reach the minimum level suggested by the standarts

Modelos matemáticos para estimativa do consumo máximo de oxigênio pela ventilometria de esforço em indivíduos saudáveis

The relation between metabolic demand and maximal oxygen consumption during exercise have been investigated in different areas of knowledge. In the health field, the determination of maximal oxygen consumption (VO2max) is considered a method to classify the level of physical fitness or the risk of cardiocirculatory diseases. The accuracy to obtain data provides a better evaluation of functional responses and allows a reduction in the error margin at the moment of risk classification, as well as, at the moment of determination of aerobic exercise work load. In Brasil, the use of respirometry associated to ergometric test became an opition in the cardiorespiratory evaluation. This equipment allows predictions concerning the oxyredutase process, making it possible to identify physiological responses to physical effort as the respiratory threshold. This thesis focused in the development of mathematical models developed by multiple regression validated by the stepwise method, aiming to predict the VO2max based on respiratory responses to physical effort. The sample was composed of a ramdom sample of 181 healthy individuals, men and women, that were randomized to two groups: regression group and cross validation group (GV). The voluntiars were submitted to a incremental treadmill test; objetiving to determinate of the second respiratory threshold (LVII) and the Peak VO2max. Using the método forward addition method 11 models of VO2max prediction in trendmill were developded. No significative differences were found between the VO2max meansured and the predicted by models when they were compared using ANOVA One-Way and the Post Hoc test of Turkey. We concluded that the developed mathematical models allow a prediction of the VO2max of healthy young individuals based on the LVII

Physiological responses of the brown mussel Perna perna (Mollusca, Bivalvia) exposed to the anionic surfactant linear alkylbenzene sulphonate (LAS).

The effects of the Linear Alkylbenzene Sulphonate (LAS) were evaluated on the mussel Perna perna, using physiological and genotoxic biomarkers. The Micronuclei (MN) assay was used to estimate effects at nuclear level, whereas the physiological effects were evaluated by measuring the oxygen consumption and ammonia excretion rates. Significant effects were observed for the MN assay and the ammonia excretion rate, even in low concentrations. The oxygen consumption was not affected in the tested concentrations. For MN and ammonia excretion, the animals exposed to intermediate concentrations were not affected, but responded to the higher concentrations, indicating the existence of compensatory mechanisms at physiological level. However, parallel to this study other authors indicate the presence of progressive effects at the cellular level, suggesting that the organisms are not capable to recover of such increasing effects. Additionally, the results show that the levels of LAS observed for Brazilian coastal waters may chronically affect the biota.

Physiological and morphological responses to feeding in broad-nosed caiman (Caiman latirostris)

Broad nosed caiman are ectotherm sauropsids that naturally experience long fasting intervals. We have studied the postprandial responses by measuring oxygen consumption using respirometry, the size changes of the duodenum, the distal small intestine, and the liver, using repeated non-invasive ultrasonography, and by investigating structural changes on the level of tissues and cells by using light- and electron microscopy. The caimans showed the same rapid and reversible changes of organ size and identical histological features, down to the ultrastructure level, as previously described for other ectothermic sauropsids. We found a configuration change of the mucosa epithelium from pseudostratified during fasting to single layered during digestion, in association with hypertrophy of enterocytes by loading them with lipid droplets. Similar patterns were also found for the hepatocytes of the liver. By placing the results of our study in comparative relationship and by utilizing the phylogenetic bracket of crocodiles, birds and squamates, we suggest that the observed features are plesiomorphic characters of sauropsids. By extending the comparison to anurans, we suggest that morphological and physiological adjustments to feeding and fasting described here may have been a character of early tetrapods. In conclusion, we suggest that the ability to tolerate long fasting intervals and then swallow a single large meal as described for many sit-an-wait foraging sauropsids is a functional feature that was already present in ancestral tetrapods.

Effects of leaves of Ipomoea batatas (Convolvulaceae) on nest development and on respiratory metabolism of leaf-cutting ants Atta sexdens L. (Hym., Formicidae)

Laboratory colonies of the leaf-cutting ants Atta sexdens feed daily with leaves of Ipomoea batatas showed ant mortality and a significant decrease in the size of the fungal garden after the second week, with complete depletion of nests after 5 weeks of treatment. The mean oxygen consumption rate of these ants was higher than the control (ants collected from nests feed with leaves of Eucalyptus alba), suggesting a physiological action of the leaves of I. batatas on the ants in addition to the effect of inhibiting the growth of the fungal garden.

Effect of nitrite on larval development of giant river prawn Macrobrachium rosenbergii

The effects of ambient nitrite concentrations on larval development of giant river prawn Macrobrachium rosenbergii were evaluated. The trials were conducted in two phases: phase 1, larvae from stages I through VIII and phase 2, larvae from stage VIII until post-larvae. In both phases larvae were kept in water with nitrite (NO2-N) concentrations of 0, 2, 4, 8 and 16 mg/L. Oxygen consumption was analyzed for larvae in stage II at nitrite concentrations of 0, 4, and 8 mg/L. Survival, weight gain, larval stage index and metamorphosis rate decreased linearly with increasing ambient nitrite concentration. However, there was no significant difference between larvae subjected to 0 and 2 mg/L NO2-N. In phase 1, there was total mortality at 16 mg/L NO2-N, while in phase 2 larval development stopped at stage X in this treatment. The oxygen consumption in stage II increased significantly at NO2-N concentration from 0 to 4 mg/L, but there was no difference between 4 and 8 mg/L NO2-N. In conclusion, increasing ambient nitrite up to 16 mg/L NO2-N delays larval development, reduces larval growth rate and causes mortality, whereas no significant effect occurs for levels below 2 mg/L NO2-N. However, the establishment of a general safe level of nitrite to M rosenbergii hatchery may be difficult due to the great variability in larvae individual sensitivity. (c) 2006 Elsevier B.V. All rights reserved.