Animal and Veterinary Science Center

We investigated the effect of sub-chronic soybean oil (SO) treatment on the insulin secretion and fatty acid composition of islets of Langerhans obtained from Goto-Kakizaki (GK), a model of type 2 diabetes, and normal Wistar rats. We observed that soybean-treated Wistar rats present insulin resistance and defective islet insulin secretion when compared with untreated Wistar rats. The decrease in insulin secretion occurred at all concentrations of glucose and arginine tested. Furthermore we observed that soybean-treated normal islets present a significant decrease in two saturated fatty acids, myristic and heneicosanoic acids, and one monounsaturated eicosenoic acid, and the appearance of the monounsaturated erucic acid. Concerning diabetic animals, we observed that soybean-treated diabetic rats, when compared with untreated GK rats, present an increase in plasma non-fasting free fatty acids, an exacerbation of islet insulin secretion impairment in all conditions tested and a significant decrease in the monounsaturated palmitoleic acid. Altogether our results show that SO treatment results in a decrease of insulin secretion and alterations on fatty acid composition in normal and diabetic islets. Furthermore, the impairment of insulin secretion, islet erucic acid and fasting plasma insulin levels are similar in treated normal and untreated diabetic rats, suggesting that SO could have a deleterious effect on β-cell function and insulin sensitivity.

Growth of potato tuber calli cells (non-green) is inhibited by 4-chloro-2-methylphenoxyacetic acid (MCPA) as a consequence of perturbation of membrane integrity. MCPA also depresses ATP content with simultaneous increase of ADP and AMP, i.e., the energy charge is severely compromised. Cell redox state is also affected by MCPA, as a function of concentration. Up to 60 lM, MCPA stimulates glutathione reductase and glutathione transferase, whereas superoxide dismutase and catalase activities are not affected. However, 120 lM MCPA inhibits all these activities. Cell death challenged by MCPA is putatively related to disturbance of membrane integrity responsible for mitochondrial uncoupling with decrease of the energy charge and subsequent loss of ions and metabolites. Keywords Herbicide Á MCPA Á Potato tuber calli Á Redox state Á Energy charge Á Antioxidant enzymes Abbreviations GST Glutathione S-transferase CAT Catalase GR Glutathione reductase SOD Superoxide dismutase ROS Reactive oxygen species EDTA Ethylenediamine-tetraacetic acid MCPA 4-Chloro-2-methylphenoxyacetic acid Communicated by S. Lewak.

The aim of this study was to analyze the influence of aerobic fitness (AF) on age-related lymphocyte DNA damage in humans, giving special attention to the role of the mitochondrial respiratory chain and hydrogen peroxide production. Considering age and AF (as assessed by VO 2 max), 66 males (19-59 years old) were classified as high fitness (HF) or low fitness (LF) and distributed into one of the following groups: young adults (19-29 years old), adults (30-39 years old), and middle-aged adults (over 40 years old). Peripheral lymphocytes obtained at rest were used to assess DNA damage (strand breaks and formamidopyrimidine DNA glycosylase (FPG) sites through the comet assay), activity of mitochondrial complexes I and II (polarographically measured), and the hydrogen peroxide production rate (assayed by fluorescence). Results revealed a significant interaction between age groups and AF for DNA strand breaks (F=8.415, p=.000), FPG sites (F=11.766, p=.000), mitochondrial complex I activity (F=7.555, p=.000), and H 2 O 2 production (F=7.500, p=.000). Except for mitochondrial complex II activity, the age variation of the remaining parameters was significantly attenuated by HF. Considering each AF level, an increase in DNA strand breaks and FPG sites with age (r=0.655, p=0.000, and r=0.738, p=0.000, respectively) was only observed in LF. Moreover, decreased mitochondrial complex I activity with age (r=−.470, p=.009) was reported in LF. These results allow the conclusion that high AF seems to play a key role in attenuating the biological aging process.

Despite medicinal plants increasing use in diabetes mellitus therapy, recurrently, however, beneficial effects are mainly empirical. In this study, the effects of decoctions of Vaccinium myrtillus L. (bilberry) leaves were investigated on GK rats, a type 2 diabetes mellitus animal model. The possible toxic effects of V. myrtillus over mitochondrial respiratory activity indexes were evaluated. Our results show that V. myrtillus leaf decoctions presented significant benefits on glycaemic control. Moreover, GK rats treated during four weeks with V. myrtillus decoction presented an improvement of mitochondrial respiratory parameters evaluated (RCR and FCCP stimulated respiration) which could be explained due to mitochondrial biogenesis improvement by quercetins present in V. myrtillus leaves.

Despite an increasing use of medicinal plants in diabetes mellitus therapy, their use is usually based on empirical knowledge. This work aims to evaluate the effects of decoctions prepared from Arctium minus Bernh. on glycaemic control of goto-kakizaki (GK) rats, a type 2 diabetes mellitus animal model, and access the potential toxicity of A. minus decoctions, using liver mitochondrial preparations. GK rats were drinking ad libitum A. minus root decoction for an interval of 4 weeks. During this time, food and beverage ingested and weight increase were controlled. Occasional glycaemias were also evaluated twice a week. Intraperitoneal glucose tolerance was determined at the beginning and at the end of the experiment. Four weeks later, GK rats liver mitochondria were isolated and respiratory parameters evaluated. A. minus root decoctions did not significantly affect the glycaemic control and long-lasting treatments induced noxious effects, considering that the liver mitochondria isolated from A. minus treated GK rats presented a significant decrease in mitochondrial respiratory parameters (RCR and FCCP stimulated respiration). This study showed that long lasting treatments with A. minus tea induced deleterious effects on cellular metabolism and, thus, should be avoided in type 2 diabetes long-term therapy.

CESAM Centre for Environmental and Marine Studies www.cesam.ua.pt ... It is know that cyanobacterial taxonomic groups are characterized by particular lipid patterns that can be used as their biological markers. The present study examined the fatty acid composition of nostocacean ...

Herbicide contamination of aquatic ecosystems is a serious global environmental concern. Several herbicides enhance the intracellular formation of reactive oxygen species, and can lead to the damage of macromolecules and to a decrease of oxidant defenses in a wide range of non-target microorganisms including cyanobacteria. The effects of molinate (a thiocarbamate herbicide used for controlling grassy weeds in rice fields) on the activities of antioxidant enzymes such as superoxide dismutase, catalase, ascorbate peroxidase, glutathione reductase, and glutathione S-transferase were evaluated in Nostoc muscorum, a freshwater cyanobacterium with a significant spread in Portuguese rice fields. These were determined in N. muscorum cultures acutely (72 h) exposed to concentrations ranging from 0.75 to 2 mM of molinate. This study also analyzed the effects of molinate on: (1) the nonenzymatic antioxidant contents (reduced and oxidized glutathione, carotenoids, and proline), (2) the oxidative cell damage measured in terms of lipid peroxidation (MDA level) and electrolyte leakage (intactness of plasma membrane), and (3) the total fatty acid profile. The results showed that the activities of all antioxidant enzymes decreased dramatically with the rising concentration of molinate after 72 h. Time-dependent and concentration-dependent increase in MDA and enhanced cell membrane leakage were indicative of lipid peroxidation, formation of free radicals and oxidative damage. Compared to control, 72-h herbicide exposure increased lipid peroxidation by 5.4%, 19% and 28% with 0.75, 1.5 and 2 mM of molinate, respectively. Similarly, herbicide stress induced an increase in electrolyte leakage (5.8%, 29.5% and 30.2% above control, with 0.75, 1.5 and 2 mM of molinate, respectively). The increased production of proline at higher molinate concentrations (the values rose above control by 45%, 95% and 156% with 0.75, 1.5 and 2 mM, respectively) indicated the involvement of this osmoprotectant in a free radical scavenging mechanism. Moreover, a radical decline in both glutathione pool, carotenoids and saturated fatty acids were also observed. The results of the present study lead us to conclude that: (1) both enzymatic and nonenzymatic antioxidative defense system of N. muscorum are dramatically affected by molinate, (2) the herbicide induces peroxidation, (3) it contributes to an increase of the unsaturation level of cell membrane fatty acids. These evidences should be taken in account when using N. muscorum as an environmental indicator species in studies of herbicide biotransformation and biomarker response as well as in environmental monitoring programmes.

The increasing use of C 60 nanoparticles and the diversity of their applications in industry and medicine has led to their production in a large scale. C 60 release into wastewaters and the possible accumulation in the environment has raised concerns about their ecotoxicological impact. In the present study, an aqueous suspension of C 60 nanoparticles was prepared and its potential toxicity studied in laboratory, using a bacterium (Bacillus stearothermophilus) and an aquatic plant (Lemna gibba) as model systems. C 60 nanoparticles inhibited the growth of L. gibba, in contrast to that of the bacterium. Consistently, the ultrastructure and respiratory activity of bacterial cells were not affected by C 60 , but the contents of chlorophylls a and b and chloroplast oxygen production decreased considerably in L. gibba. Altogether, our results suggest that C 60 aqueous dispersions must be viewed as an environmental pollutant, potentially endangering the equilibrium of aquatic ecosystems.

The effects of dicamba, a widely used broadleaf herbicide, on rat liver mitochondrial bioenergetic activities were examined. The results obtained for state 4 respiration indicate not only an uncoupling effect, the result of an increase on the permeability of inner mitochondria membrane to protons, but also a strong inhibitory effect on the redox complexes. State 3 and respiration uncoupled by FCCP (carbonylcyanide p-trifluoromethoxyphenylhydrazone) were inhibited to approximately the same extent, i.e. by about 70%. Depression of respiratory activity is essentially mediated through partial inhibition of mitochondrial complexes II and III. ATPase activity was much less depressed by dicamba than ATP synthase activity. Therefore, a considerable part of the inhibition observed on ATP synthase is the result of an inhibition on the redox complexes. The loss of phosphorylation capacity, induced by dicamba, was in the last analysis not only the result of a direct effect of dicamba on the enzymatic complex (F 0-F 1 ATPase) but also the result of a deleterious effect on the integrity of the mitochondrial membrane, which can promote an inhibition of the respiratory complexes and an increase of the proton permeability of the inner membrane.

Mitochondria generated nitric oxide (NO) regulates several cell functions including energy metabolism, cell cycling, and cell death. Here we report that the NO synthase inhibitors (l-NAME, l-NNA and l-NMMA) administered either in vitro or in vivo induce Ca 2+dependent mitochondrial permeability transition (MPT) in rat liver mitochondria via a mechanism independent on changes in the energy state of the organelle. MPT was determined by the occurrence of cyclosporin A sensitive mitochondrial membrane potential disruption followed by mitochondrial swelling and Ca 2+ release. In in vitro experiments, the effect of NOS inhibitors was dose dependent (1 to 50 μM). In addition to cyclosporin A, l-NAME induced MPT was sensitive to Mg 2+ plus ATP, EGTA, and to a lower degree, to catalase and dithiothreitol. In contrast to l-NAME, its isomer d-NAME did not induce MPT. l-NAME induced MPT was associated with a significant decrease in both the rate of NO generation and the content of membrane protein S-nitrosothiol. Acute and chronic in vivo treatments with l-NAME also promoted MPT and decreased the content of mitochondrial protein Snitrosothiol. SNAP (a NO donor) prevented l-NAME mediated MPT and reversed the decrease in the rate of NO generation and in the content of membrane protein S-nitrosothiol. We propose that S-nitrosylation of critical membrane protein thiols by NO protects against MPT.

The aim of this work was to perform the histological characterization of the C. sativa leaves of three Portuguese cultivars to establish comparison among the leaves of the different quadrants in accord and with the cardinal points of the tree top and among different cultivars of this species, using light microscopy (OM) and scanning electron microscopy (SEM). Measurements were also carried out for the leaf tissue thickness, stomatal density, leaf area in the four tree top quadrants. The leaves turned to the North had lesser thickness of mesophyll mainly due to lower amount of palisade parenchyma. The stomatal density was significantly lower in these leaves, unlike the leaf area that has the highest expression.

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In a previous work the ability of Candida oleophila to use phenolic compounds as sole carbon and energy source at high concentrations without an additional carbon source was reported. C. oleophila grown in bioreactor batch cultures in a diluted and sterilized olive oil mill wastewater (OMW) caused a significant decrease in the total tannins content but no significant alteration was observed in phenolic acid and fatty acid content. Both treated and untreated OMWs were tested to evaluate the capacity in interfering with mitochondrial bioenergetics. Mitochondrial respiration was not affected by treated OMW on the range of used concentrations, contrary to the untreated OMW. Furthermore, mitochondrial membrane potential and respiratory complexes were always significantly less affected by treated OMW in comparison with untreated OMW. However, supplementary treatment should be applied before OMW could be considered non-toxic.

An easy preparation of 4-aminopyrrole-3-carbonitrile derivatives and their transformation into new substituted pyrrolo[3,2-b]pyridines is described. The one-step transformation was carried out, via Friedländer reaction under microwave irradiation and classical heating methods. The use of microwave irradiation led to high conversion and shorter reaction times.

A simple and efficient method has been developed for the synthesis of various pyrazolo[3,4-d]pyrimidines using microwave irradiation under solvent-free conditions. The advantages of applying microwave irradiation compared with the classical method were demonstrated. The structures of all the compounds were confirmed by the usual techniques and in two cases by X-ray analysis. The compounds did not display appreciable ability to inhibit xanthine oxidase activity. Screening for antifungal activity showed that some derivatives were active against four fungi, with more significant results for Botrytis.