Camila Pereira Braga

Influence of treatment with quercetin on lipid parameters and oxidative stress of pregnant diabetic rats

Among the numerous coadjuvant therapies that could influence the incidence and progression of diabetic complications, antioxidants and flavonoids are currently being tested in clinical trials. We investigated the effect of quercetin on biochemical parameters in streptozotocin-induced (60 mg/kg body mass, by intraperitoneal injection) diabetic rats. A total of 32 female Wistar rats were distributed among 4 groups as follows: control (G1); control treated with quercetin (G2); diabetic (G3); and diabetic treated with quercetin (G4). Quercetin administered to pregnant diabetic rats controlled dyslipidemia and improved lipid profiles in diabetes mellitus, regulated oxidative stress by reducing the generation of lipid hydroperoxides, and increased the activity of the antioxidant enzyme glutathione peroxidase.

Identification of protein biomarkers of mercury toxicity in fish

Bioaccumulative metals such as mercury are found in increasing amounts in fish and their consumers. In the region of the Madeira River, in the Brazilian Amazon, mercury (Hg) is a predominant contaminant in the aquatic ecosystem. There is therefore a need to find specific biomarkers of mercury toxicity in fish to monitor contaminations. Here, mercury-bound proteins were identified in the liver tissues of fishes Mylossoma duriventre and Brachyplatystoma rousseauxii. Mercury was quantified in liver tissue, pellets and protein spots by graphite furnace atomic absorption. Proteins were fractionated by two-dimensional polyacrylamide gel electrophoresis and identified by mass spectrometry with electrospray ionization. We identified nine proteins linked to mercury and that presented biomarker characteristics of mercury. Among the proteins identified, isoforms of parvalbumin, ubiquitin-40S ribosomal protein S27a, brain-specific angiogenesis inhibitor 1-associated protein 2-like protein 2 and betaine–homocysteine S-methyltransferase 1 are notable for having the molecular function of binding to metallic ions.

Revealing oxidative damage to enzymes of carbohydrate metabolism in yeast: An integration of 2D DIGE, quantitative proteomics and bioinformatics

Clinical usage of lidocaine, a pro‐oxidant has been linked with severe, mostly neurological complications. The mechanism(s) causing these complications is independent of the blockade of voltage‐gated sodium channels. The budding yeast Saccharomyces cerevisiae lacks voltage‐gated sodium channels, thus provides an ideal system to investigate lidocaine‐induced protein and pathway alterations. Whole‐proteome alterations leading to these complications have not been identified. To address this, S. cerevisiae was grown to stationary phase and exposed to an LC50 dose of lidocaine. The differential proteomes of lidocaine treatment and control were resolved 6 h post exposure using 2D DIGE. Amine reactive dyes and carbonyl reactive dyes were used to assess protein abundance and protein oxidation, respectively. Quantitative analysis of these dyes (⩾ 1.5‐fold alteration, p ⩽ 0.05) revealed a total of 33 proteoforms identified by MS differing in abundance and/or oxidation upon lidocaine exposure. Network analysis showed enrichment of apoptotic proteins and cell wall maintenance proteins, while the abundance of proteins central to carbohydrate metabolism, such as triosephosphate isomerase and glyceraldehyde‐3‐phosphate dehydrogenase, and redox proteins superoxide dismutase and peroxiredoxin were significantly decreased. Enzymes of carbohydrate metabolism, such as phosphoglycerate kinase and enolase, the TCA cycle enzyme aconitase, and multiple ATP synthase subunits were found to be oxidatively modified. Also, the activity of aconitase was found to be decreased. Overall, these data suggest that toxic doses of lidocaine induce significant disruption of glycolytic pathways, energy production, and redox balance, potentially leading to cell malfunction and death.

Total Mercury Determination in Muscle and Liver Tissue Samples from Brazilian Amazon Fish Using Slurry Sampling

This paper presents a slurry sampling method for total mercury determination by graphite furnace atomic absorption spectrometry (GFAAS) in tissue of fish from the Amazon. The tissue samples were lyophilized and macerated, and then the slurry samples were prepared by putting 20xa0mg of tissue, added to a solution containing Triton X-100, Suprapur HNO3, and zirconium nitrate directly in sampling vials of a spectrometer. Mercury standard solutions were prepared under the same conditions as the slurry samples. The slurry samples and the mercury standard solutions were sonicated for 20xa0s. Twenty microliters of slurry samples were injected into the graphite tube, which contained an internal wall lined with tungsten carbide. Under these conditions, it was possible to thermally stabilize the mercury up to an atomization temperature of 1700xa0°C. The method was validated by mercury determination in reference materials DORM-4 and DOLT-4. The LOD and LOQ were 0.014 and 0.045xa0mgxa0kg−1, respectively, and recovery percentages in relation to the concentration values were certified in the order of 98%.

Influence of oxidative damage to proteins on meat tenderness using a proteomics approach

The objective of this study was to evaluate the association between oxidative damage to proteins (represented by protein carbonylation) and beef tenderness. Three experimental groups were selected by shear force (SF): tender (38.2u202f±u202f2.9u202fN), intermediate (51.9u202f±u202f6.8u202fN), and tough meat (74.5u202f±u202f7.8u202fN). Two-dimensional electrophoresis with hydrazide fluorophore derivatization was used. The structural proteins actin (ACTA1), myosin (MYL1 and MYL3), desmin (DES) and troponin T (TNNT1 and TNNT3), antioxidant proteins (PRDX1, PRDX2 and PARK7) and heat shock proteins (HSPB1, CRYAB and HSPB6) showed an increase in the oxidative damage in tender meat when compared to the intermediate and tough meat (Pu202f<u202f.05). Decrease in oxidative damage of the metabolic enzymes (TPI1, GAPDH and ENO3) were observed in tender meat group (Pu202f<u202f.05). The present results suggest that oxidation act on the proteins of different metabolic pathways and consequently affect meat tenderness in Angus crossbred cattle.

Characterization of molecular biomarkers of mercury exposure to muscle tissue of Plagioscion squamosissimus and Colossoma macropomum from the Amazon region

Mercury has the ability to bind to a variety of biomolecules, which can compromise its structure and functionality and thus promote its toxic effects. The aim of this study is to identify possible mercury biomarkers in muscle samples of Plagioscion squamosissimus (carnivorous fish) and Colossoma macropomum (omnivorous fish), from the Amazon region. The muscle proteome of fish species was separated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), and the total mercury concentrations in protein spots were determined by graphite furnace atomic absorption spectrometry (GFAAS). The protein spots containing mercury were characterized by electrospray ionization tandem mass spectrometry (ESI-MS/MS). The mercury concentrations in the protein spots were in the range of 1.10u202f±u202f0.02-23.90u202f±u202f0.33u202fμgu202fg-1. The proteins phosphoglycerate mutase 2 (P. squamosissimus), hemoglobin β and cytochrome P450scc (C. macropomum), identified by ESI-MS/MS and showing the highest values of mercury concentration, may be considered possible mercury biomarkers.

Metallomics in Fish

Metallomics allows the integration of traditionally analytical studies with inorganic and biochemical studies. The study of metallomics in living organisms allows us to obtain information about how the metal ion is distributed and coordinated with proteins, the essentiality and/or toxicity, and the individual concentrations of metal species, thus contributing to elucidation of the physiological and functional aspects of these biomolecules. In this context, several lines of research have appeared in the literature with different terms and approaches. For example, metallomic, which deals with the characterization of the total metal/metalloid species present in an organism; metalloprotein, which deals with the characterization of the total elements present in a specific site of an organism (cellular behavior, protein, metalloprotein); and metallomic, which deals with a more in-depth study of metallome. In this area, information is sought on the interactions and functional connections of metal/metalloid species with genes, proteins, metabolites and other biomolecules of the organism and, therefore, the elucidation of the biological role exerted by the metal ions bound to the biomolecules. In this chapter, we will describe techniques used in animal studies.

Identification of Biomarkers of Mercury Contamination in Brachyplatystoma filamentosum of the Madeira River, Brazil, Using Metalloproteomic Strategies

Predator fish can accumulate high levels of mercury, which qualifies them as potential indicators of this toxic metal. The predatory species Brachyplatystoma filamentosum, popularly known as filhote, is among the most consumed species in the Brazilian Amazon. Continuing the metalloproteomic studies of mercury in Amazonian fishes that have been developed in the last 5xa0years, the present paper provides the data of protein characterization associated with mercury in muscle and liver samples of filhote (Brachyplatystoma filamentosum) collected in the Madeira River, Brazilian Amazon. The mercury concentration in the muscle and liver samples was determined by graphite furnace atomic absorption spectrometry (GFAAS). The protein fraction was extracted in an aqueous medium, and later, a fractional precipitation procedure was performed to obtain the protein pellets. Then, the proteome of the tissue samples of this fish species was separated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), and a mercury mapping of the protein spots was carried out by GFAAS after acid digestion. Protein spots that had mercury were characterized by mass spectrometry with electrospray ionization in sequence (ESI-MS/MS) after tryptic digestion. It was possible to characterize 11 mercury-associated protein spots that presented biomarker characteristics and could be used to monitor mercury in fish species of the Amazon region. Thus, the metalloproteomic strategies used in the present study allowed us to characterize 11 mercury-associated protein spots. It should be noted that the protein spots identified as GFRP, TMEM186, TMEM57B, and BHMT, which have coordination sites for elements with characteristics of soft acids, such as mercury, can be used as biomarkers of mercury contamination in monitoring studies of this toxic metal in fish species from the Amazon region.