Browsing by Author "Cordeiro, O."
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- Changes in liver proteome expression of Senegalese sole (Solea senegalensis) in response to repeated handling stressPublication . Cordeiro, O.; Silva, Tomé S.; Alves, Ricardo N.; Costas, B.; Wulff, Tune; Richard, Nadège; Vareilles, Mahaut; Conceição, L. E. C.; Rodrigues, PedroThe Senegalese sole, a high-value flatfish, is a good candidate for aquaculture production. Nevertheless, there are still issues regarding this species’ sensitivity to stress in captivity. We aimed to characterize the hepatic proteome expression for this species in response to repeated handling and identify potential molecular markers that indicate a physiological response to chronic stress. Two groups of fish were reared in duplicate for 28 days, one of them weekly exposed to handling stress (including hypoxia) for 3 min, and the other left undisturbed. Two-dimensional electrophoresis enabled the detection of 287 spots significantly affected by repeated handling stress (Wilcoxon–Mann–Whitney U test, p<0.05), 33 of which could be reliably identified by peptide mass spectrometry. Chronic exposure to stress seems to have affected protein synthesis, folding and turnover (40S ribosomal protein S12, cathepsin B, disulfide-isomerase A3 precursor, cell-division cycle 48, and five distinct heat shock proteins), amino acid metabolism, urea cycle and methylation/folate pathways (methionine adenosyltransferase I α, phenylalanine hydroxylase, mitochondrial agmatinase, serine hydroxymethyltransferase, 3-hydroxyanthranilate 3,4-dioxygenase, and betaine homocysteine methyltransferase), cytoskeletal (40S ribosomal protein SA, α-actin, β-actin, α-tubulin, and cytokeratin K18), aldehyde detoxification (aldehyde dehydrogenase 4A1 family and aldehyde dehydrogenase 7A1 family), carbohydrate metabolism and energy homeostasis (fatty acid-binding protein, enolase 3, enolase 1, phosphoglycerate kinase, glyceraldehyde-3-phosphate dehydrogenase, aconitase 1, mitochondrial ATP synthase α-subunit, and electron-transfer flavoprotein α polypeptide), iron and selenium homeostasis (transferrin and selenium binding protein 1), steroid hormone metabolism (3-oxo-5-β-steroid 4-dehydrogenase), and purine salvage (hypoxanthine phosphoribosyltransferase). Further characterization is required to fully assess the potential of these markers for the monitoring of fish stress response to chronic stressors of aquaculture environment.
- Changes in the soluble bone proteome of reared white seabream (Diplodus sargus) with skeletal deformitiesPublication . Silva, Tomé S.; Cordeiro, O.; Richard, Nadège; Conceição, L. E. C.; Rodrigues, PedroOne of the main constrains for commercial aquaculture production of white seabream (Diplodus sargus) is the high incidence of skeletal malformations in reared fish. The purpose of this study was to obtain a better understanding of the mechanisms involved in the development of these types of skeletal malformations by comparative proteomic analysis of the vertebral column of normal and deformed fish using 2DE for protein separation and MS for protein identification. We observed a 3.2 and 3.4-fold increase in the expression of two tropomyosin isoforms, one of which (tropomyosin-4) is essential for the motility and polarization cycles of osteoclasts. Furthermore, a 1.6, 1.7 and 1.8-fold increase in three parvalbumin spots was detected, suggesting a cellular response to increased intracellular Ca2+ levels. These results can be interpreted as signs of increased cellular activity in the bone of white seabream with skeletal deformities coupled to a higher degree of calcium mobilization, which elicits further studies into the use of these proteins as indicators of skeletal metabolic state.
- Dietary tools to modulate glycogen storage in gilthead seabream muscle: glycerol supplementationPublication . Silva, Tomé S.; Matos, Elisabete; Cordeiro, O.; Colen, Rita; Wulff, Tune; Sampaio, Eduardo; Sousa, Vera; Valente, L. M. P.; Gonçalves, Amparo; Silva, Joana M. G.; Bandarra, N.; Nunes, Maria Leonor; Dinis, Maria Teresa; Dias, J.; Jessen, Flemming; Rodrigues, PedroThe quality and shelf life of fish meat products depend on the skeletal muscle’s energetic state at slaughter, as meat decomposition processes can be exacerbated by energy depletion. In this study, we tested dietary glycerol as a way of replenishing muscle glycogen reserves of farmed gilthead seabream. Two diets were tested in duplicate (n = 42/tank). Results show 5% inclusion of crude glycerol in gilthead seabream diets induces increased muscle glycogen, ATP levels and firmness, with no deleterious effects in terms of growth, proximate composition, fatty acid profile, oxidative state, and organoleptic properties (aroma and color). Proteomic analysis showed a low impact of glycerol-supplementation on muscle metabolism, with most changes probably reflecting increased stress coping capacity in glycerol-fed fish. This suggests inclusion of crude glycerol in gilthead seabream diets (particularly in the finishing phase) seems like a viable strategy to increase glycogen deposition in muscle without negatively impacting fish welfare and quality.
- Dietary vitamin K supplementation and expression of proteins involved in skeleton development of senegalese sole (Solea senegalensis)Publication . Richard, Nadège; Gavaia, Paulo J.; Cordeiro, O.; Silva, Tomé S.; Rodrigues, C.; Rodrigues, Pedro; Conceição, L. E. C.In aquaculture, skeleton deformities are a serious problem as they affect growth, morphology, survival of fish and also increase productions costs and decrease market value of the final products. Since vitamin K is known for playing and important role in bone metabolism, the aim of this study was to investigate the effects of dietary vitamin K1 supplementation on skeleton deformities and on the expression of proteins involved involved in skeleton development of Senegalese sole.
- Effects of genotype and dietary fish oil replacement with vegetable oil on the intestinal transcriptome and proteome of Atlantic salmon (Salmo salar)Publication . Silva, Tomé; Cordeiro, O.; Rodrigues, Pedro; Guy, Derrick R.; Bron, James E.; Taggart, John B.; Bell, J. Gordon; Tocher, Douglas R.Expansion of aquaculture requires alternative feeds and breeding strategies to reduce dependency on fish oil (FO) and better utilization of dietary vegetable oil (VO). Despite the central role of intestine in maintaining body homeostasis and health, its molecular response to replacement of dietary FO by VO has been little investigated. This study employed transcriptomic and proteomic analyses to study effects of dietary VO in two family groups of Atlantic salmon selected for flesh lipid content, 'Lean' or 'Fat'.ResultsMetabolism, particularly of lipid and energy, was the functional category most affected by diet. Important effects were also measured in ribosomal proteins and signalling. The long-chain polyunsaturated fatty acid (LC-PUFA) biosynthesis pathway, assessed by fatty acid composition and gene expression, was influenced by genotype. Intestinal tissue contents of docosahexaenoic acid were equivalent in Lean salmon fed either a FO or VO diet and expression of LC-PUFA biosynthesis genes was up-regulated in VO-fed fish in Fat salmon. Dietary VO increased lipogenesis in Lean fish, assessed by expression of FAS, while no effect was observed on β-oxidation although transcripts of the mitochondrial respiratory chain were down-regulated, suggesting less active energetic metabolism in fish fed VO. In contrast, dietary VO up-regulated genes and proteins involved in detoxification, antioxidant defence and apoptosis, which could be associated with higher levels of polycyclic aromatic hydrocarbons in this diet. Regarding genotype, the following pathways were identified as being differentially affected: proteasomal proteolysis, response to oxidative and cellular stress (xenobiotic and oxidant metabolism and heat shock proteins), apoptosis and structural proteins particularly associated with tissue contractile properties. Genotype effects were accentuated by dietary VO.ConclusionsIntestinal metabolism was affected by diet and genotype. Lean fish may have higher responsiveness to low dietary n-3 LC-PUFA, up-regulating the biosynthetic pathway when fed dietary VO. As global aquaculture searches for alternative oils for feeds, this study alerts to the potential of VO introducing contaminants and demonstrates the detoxifying role of intestine. Finally, data indicate genotype-specific responses in the intestinal transcriptome and proteome to dietary VO, including possibly structural properties of the intestinal layer and defence against cellular stress, with Lean fish being more susceptible to diet-induced oxidative stress.
- Effects of preslaughter stress levels on the post-mortem sarcoplasmic proteomic profile of gilthead seabream musclePublication . Silva, Tomé S.; Cordeiro, O.; Matos, Elisabete; Wulff, Tune; Dias, J.; Jessen, Flemming; Rodrigues, PedroFish welfare is an important concern in aquaculture, not only due to the ethical implications but also for productivity and quality-related reasons. The purpose of this study was to track soluble proteome expression in post-mortem gilthead seabream muscle and to obsere how preslaughter stress affects these post-mortem processes. For the experiment, two groups of gilthead seabream (n = 5) were subjected to distinct levels of preslaughter stress, with three muscle samples being taken from each fish. Proteins were extracted from the muscle samples, fractionated, and separated by 2DE. Protein identification was performed by MALDI-TOF-TOF MS. Analysis of the results indicates changes on several cellular pathways, with some of these changes being attributable to oxidative and proteolytic activity on sarcoplasmic proteins, together with leaking of myofibrillar proteins. These processes appear to have been hastened by preslaughter stress, confirming that it induces clear postmortem changes in the muscle proteome of gilthead seabream.
- Impact of dietary protein hydrolysates on skeleton quality and proteome in Diplodus sargus larvaePublication . Vareilles, Mahaut; Richard, Nadège; Gavaia, Paulo J.; Silva, Tomé S.; Cordeiro, O.; Guerreiro, Inês; Yúfera, M.; Batista, I.; Pires, C.; Pousão-Ferreira, P.; Rodrigues, Pedro; Rønnestad, I.; Fladmark, K. E.; Conceição, L. E. C.In order to investigate the effects of dietary protein hydrolysates (PH) on larval growth performance, skeleton quality and proteome expression, triplicate groups of white seabream (Diplodus sargus) larvae were co-fed from first-feeding with live feed and three microencapsulated diets differing in the molecular weight of their PH fraction (Control – inclusion of CPSP-90; H – inclusion of a high amount in 0.5–30 kDa hydrolysates; L – inclusion of a high amount in <0.5 kDa hydrolysates). At 15 days after hatching (DAH), proteome expression changes were assessed in entire larvae by twodimensional gel electrophoresis and the quality of larval skeleton was analysed at 28 DAH through double staining of cartilage and bone. Dietary PH fractions tested affected growth, the larvae fed diet L being significantly larger than those fed diet H, but it did not affect the incidence of deformed larvae, nor the number of deformities per fish. Two-dimensional analysis of larvae proteome allowed the detection and the comparative quantification of a total of 709 protein spots having a pI between 4 and 7, around half of which had an expression significantly affected by dietary treatment, the main difference being between proteome of Control larvae with those of both groups L and H. From these spots, 52 proteins involved in diverse processes such as cytoskeletal dynamics, energetic, lipoprotein, amino acid (AA), and nucleotide metabolisms, protein chaperoning and degradation, and signal transduction, were identified. This study revealed that the molecular weight of the dietary protein hydrolysate fraction had a minor impact on skeletal deformities in white seabream larvae, but affected growth performance and had a strong impact on larvae whole body proteome.
- Inclusion of protein hydrolysates in the diet of white seabream (Diplodus sargus) larvae: impacts on skeleton quality and larvae proteome expressionPublication . Richard, Nadège; De Vareilles, M.; Gavaia, Paulo J.; Silva, Tomé S.; Cordeiro, O.; Yúfera, M.; Rodrigues, Pedro; Conceição, L. E. C.In order to investigate the effects of dietary protein hydrolysate on larval skeleton quality and larvae proteome expression, triplicate groups of white seabream (Diplodus sargus) larvae were co-fed from first-feeding onwards with live feed and three microencapsulated diets ir differing only on the molecular weight of their protein hydrolysate fraction.
- Metabolic molecular indicators of chronic stress in gilthead seabream (Sparus aurata) using comparative proteomicsPublication . Alves, Ricardo N.; Cordeiro, O.; Silva, Tomé S.; Richard, Nadège; De Vareilles, M.; Marino, G.; Di Marco, Patrizia; Rodrigues, Pedro; Conceição, L. E. C.The aim of this study was to identify possible metabolic molecular indicators of chronic stress in gilthead sea bream Sparus aurata. Two potential stressful conditions were tested: repeated handling and crowding at high stocking density. Gilthead seabream kept under optimized rearing conditions were used as control fish. Cortisol was measured as primary stress indicator and the liver proteome of stressed fish was compared to that of control fish using comparative proteomics. Plasma cortisol levels in sea bream repeatedly handled and crowded at high stocking density were significantly higher than in undisturbed control fish. A total of 560 spots were detected and the statistical analysis revealed a differential expression in about 50% of all detected proteins. Spots with greater than 2-fold or lower than 0.5-fold changes were identified by liquid chromatography–tandem mass spectrometry (LC–MS/MS). Proteins like fatty acid binding protein (lipid transport and antioxidant role), heat shock cognate protein (chaperoning), calmodulin (Ca2+ signaling), mitochondrial porine — voltage-dependent anion channel (lipid oxidation), glutamine synthetase (ammonia metabolism), cofilin and beta-tubulin (cytoskeleton), hemoglobin and several other proteins involved in carbohydrate metabolism (triose-phosphate isomerase, pyruvate dehydrogenase, glyceraldehyde 3-phosphate dehydrogenase, alfa-enolase) were differentially expressed in fish under chronic stress. Some of these proteins may be used in the future as chronic stress and/or part of a panel of welfare biomarkers, after validation studies using RT-PCR and ELISA assays.
- Teleost fish osteocalcin 1 and 2 share the ability to bind the calcium mineral phasePublication . Cavaco, S.; Williamson, M. K.; Rosa, Joana; Roberto, Vania Palma; Cordeiro, O.; Price, P. A.; Cancela, Leonor; Laizé, Vincent; Simes, DThe occurrence of a second osteocalcin (OC2) has been reported in teleost fish, where it coexists with OC1 in some species. While it has been proposed that OC2 gene originated from OC1 through the fish whole-genome duplication event, little information is available on its molecular function and physiological role. The present study brings biological data supporting the presence of OC2 in the mineral phase of teleost fish bone and its association with the mineral phase together with OC1. The occurrence of OC2 forms with different levels of phosphorylation or c-carboxylation, and with amino acid substitutions was observed. Comparative analysis of mature peptide sequences revealed the high conservation existing between OC1 and OC2, in particular within the core c-carboxyglutamic acid domain, and suggests that both protein forms may have the same function, i.e., binding of calcium ions or hydroxyapatite crystals.