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Comparação do tratamento crônico com inibidor da síntese de óxido nítrico ou fluoxetina sobre as alterações cardiovasculares e do tipo depressivas induzidas pelo estresse crônico variável em ratos
(Universidade Federal de São Carlos, 2014-09-05) Almeida, Jeferson de; Crestani, Carlos Cesar; https://lattes.cnpq.br/1117432571971568; https://lattes.cnpq.br/8141647386370567
Here we compared effects of chronic treatment with the preferential nNOS inhibitor 7- nitroindazole (7-NI) with those evoked by the conventional antidepressant fluoxetine on alterations taken as markers of depression (immobility in the forced swimming test(FST), decreased body weight gain, and increased plasma corticosterone) and cardiovascular changes evoked by the chronic variable stress (CVS) in rats. For this, male rats were exposed to a 14- days CVS protocol while being concurrently treated with either 7-NI or fluoxetine. Fluoxetine and 7-NI prevented increased immobility in the FST induced by CVS and reduced plasma corticosterone in stressed rats. Both treatments also prevented CVS-evoked reduction on depressor response to vasodilator agents and baroreflex changes. Nevertheless, none of pharmacological treatments affected body weight reduction induced by CVS. Furthermore, fluoxetine reduced body weight gain in unstressed animals. Fluoxetine and nNOS blockade also induced cardiovascular changes independent of stress exposure, including increase on cardiac parasympathetic tone, intrinsic heart rate, and vascular sympathetic modulation; reduction in sympathetic activity to the heart and the pressor response to vasoconstrictor agents, and impairment of baroreflex activity. These findings provide evidence that both fluoxetine and 7-NI prevent depression-like behavioural changes and cardiovascular dysfunctions associated with CVS. Furthermore, both pharmacological treatments caused cardiovascular changes that were independent of stress exposure, some being beneficial and other potentially increasing vulnerability to development of cardiovascular diseases. Nevertheless, the body weight reduction induced by fluoxetine treatment is especially important considering the weight loss related to depression.
Estudos estruturais e ensaios biológicos de proteínas codificadas por genes de micro-exon (MEG) de Schistosoma mansoni
(Universidade Federal de São Carlos, 2014-09-05) Alves, Natália Oliveira; Marco, Ricardo de; https://lattes.cnpq.br/0229339143607870; https://lattes.cnpq.br/5397307262243397
Schistosoma mansoni is the causative agent of Schistosomiasis, reaching 249 million people in 78 countries. It is believed that this infection is successful due to modulation of the host immune system through proteins secreted by the parasite. Recently a class of proteins derived from micro-exon genes (MEGs) was observed in proteomic analyses of schistosomula and egg secretions of S. mansoni, and also associated with glands of several stages of the parasite life. Furthermore, they found that none of these proteins showed similarity to proteins of organisms from other genres. In this context, aim of this work was the structural study, the search for protein partners and biological assays seeking possible roles for the proteins encoded by genes of micro-exons of S. mansoni: MEG-5, MEG-8.2 e MEG-12. For structural characterization studies we used MEG-5 protein expressed by heterologous system (the expression of MEG-8.2 protein was not obtained) and initially performed tests Circular Dichroism (CD). This essay revealed the predominance of one disordered structure due to the presence of a negative peak in the ranges of 180-200mm and the presence of lower intensity peaks in the range 220- 240mm, indicating that part of the protein may present structure. Such structure was observed in the secondary structure predictions performed in SOPMA and Jpred3 programs, wherein the presence of a predicted alpha-helix in the C-terminal region. To confirm this fact, we performed an analysis of the maximum emission of tryptophan fluorescence by the technique of intrinsic fluorescence, because the protein has a single tryptophan present in the probable alpha-helical region. We note that the maximum fluorescence emission is at 330mm, indicating that it is in more hydrophobic environment. After the structural characterization of MEG-5 protein we seek partners through the assay two-hybrid system performing a scan on leukocyte cDNA libraries, yielding two possible partners. The CHMP1B protein was the one that drew the most attention in for being part of the formation of vesicular body and be related to the processes of antigen presentation. The synthetic peptide MEG-12 has a peculiar characteristic of an amphipathic helix predicted by Jpred3 program, and hemolytic character. This action was best observed by scanning electron microscopy (SEM), to be verifies in a clear disruption of the membrane to the breakup. The peptide is secreted into the esophagus may region associated with the digestion of erythrocytes made by the parasite. Therefore, these two proteins are shown to be important in the host-parasite relationship and possible targets for drugs or creation of more efficient vaccines.
Produção recombinante e caracterização de duas cistatinas de cana-de-açúcar
(Universidade Federal de São Carlos, 2014-09-05) Miguel, Mariana Cardoso; Silva, Flávio Henrique da; https://lattes.cnpq.br/1757309852446263; https://lattes.cnpq.br/7103812372342719
Cystatins are reversible inhibitors of cysteine peptidases. The cystatins found in plants are called phytocystatins, and represent an independent subfamily of the cystatins superfamily. Some studies have reported significant pleiotropic effects for recombinant cystatins expressed in transgenic plants, notably including tolerance phenotypes against attack of herbivorous arthropods and pathogens, and against abiotic and biotic stresses. Besides, the recombinant sugarcane cystatin, CaneCPI-4, showed potential to inhibit development of melanoma cells. Thus, the study and knowledge about phytocystatins, become interesting from agricultural and medicinal point of view. The sugarcane genome project (SUCEST) allowed the identification of about 25 putative cystatins in this plant, which were gathered in 4 groups, by phylogenetic analysis. In this study, we propose a new classification for the putative cystatins found in the SUCEST database. Furthermore, we describe the heterologous expression, purification and characterization of two novel sugarcane cystatins, CaneCPI-5 and CaneCPI-6, which showed different inhibitory activities against human cathepsin B. While protein CaneCPI-6 was not able to inhibit this enzyme efficiently (Ki = 1,83 μM), the protein CaneCPI-5 showed a good inhibitory capacity against the same enzyme (Ki = 6,87 nM). The CaneCPI-5 cystatin was also analyzed against recombinant cathepsin L from the beetle Sphenophorus levis (rSl-CathL), and showed a good inhibitory capacity against this enzyme (Ki = 0,059 nM). Finally, both of proteins, CaneCPI-5 and CaneCPI-6, proved to be thermostable when kept at 100°C for 30 minutes.
Spatial and temporal influences of road duplication on wildlife road kill using habitat suitability models
(Universidade Federal de São Carlos, 2014-09-05) Ciocheti, Giordano; Ribeiro, Milton Cezar; https://lattes.cnpq.br/4158685235743119; https://lattes.cnpq.br/6803662982558040
Urban growth and population growth led to the construction of a gigantic road network around the world. This network is responsible for several impacts on fauna, flora and the environment, such as road kill, isolation of populations, facilitating the establishment of invasive species, river siltation, among others. However, although road ecology has advanced recently, there are still many gaps on how roads affect fauna, as little is known about how effects of changing the structure of highways can modify animal-vehicle collisions. This study aims to evaluate some of the impacts of roads on species of medium and large mammals in fragmented and naturally heterogeneous landscapes. Using a functional group approach based on animal sensitivity to disturbance and displacement capacity, I set out to answer three questions, one in each chapter: 1) the contribution of various landscape indices to predict wildlife road kill; 2) highway duplication and the implementation of wildlife crossing structures alter animal road kill; 3) duplication of roads change the way fauna road kill is correlated with the landscape metrics. To answer the first and third questions, we have developed innovative methods combining road kill data with a multi-scale approach with landscape metrics involving quantity and distance of various landscape elements, such as natural vegetation, cerrado, water, forestry and sugar cane. This method proposed was derived from habitat suitability model, and proved very promising for estimating the probability of animal road kill. Each functional group of species responded differently to landscape elements. Distance and amount of vegetation has been more important to estimate road kill probability of more sensitive mammals, but the amount of sugar cane also contributed to these results. The proposed method is highly replicable and can be easily applied in other regions with other taxa. The second question was addressed in an analytical way, with a conventional hypothesis testing approach. We found that, in general, there was no significant difference between road kill before and after road duplication. However, when considering the functional groups, and even species, some changes were significant for both increasing and reducing road kill. We also found that the proximity of wildlife crossing structures to road kill records did not reduce the frequency of animal-vehicle collision, indicating that such mitigation measures may not have been appropriate to reduce animal road mortality. Finally, in the third chapter we have proposed a new approach to estimate the changes in animal road kill probability before and after duplication of highways. In this chapter we recorded an increase in the probability of road kill after duplication for generalist species with high mobility. The methods proposed here are easy to implement in several actions related to roads, both for seeking their structural improvement and for making them more sustainable for biodiversity.