Nathalie Marty-Gasset

Proteomic profile evolution during steatosis development in ducks

We investigated a protein profile evolution during steatosis in ducks using 2-dimensional electrophoresis gels to better understand the mechanisms underlying liver steatosis at the level of hepatic proteins in waterfowl. Two-dimensional electrophoresis gels were performed in the liver at different stages of steatosis in the duck. Mule ducks were slaughtered after 0, 14, or 23 meals of overfeeding, according to commercial conditions. Thirty-one proteic spots were differentially expressed between 3 or 2 durations of overfeeding: 3 spots were differentially expressed between the 3 times and 28 spots were differentially expressed between 2 times. The identified proteins (14) could be regrouped into 5 categories: enzymes, translation factors, proteins involved in cell structure, proteins with antioxidant properties, and proteins that can link calcium. This study opens new research areas in the understanding of steatosis in waterfowl, such as cell structure and oxidative stress.

Current advances in proteomic analysis of (fatty) liver.

In this review, an overview on proteomic studies conducted in livers of farm animals is conducted with a special focus on liver steatosis in waterfowl. Several studies had interest in understanding liver metabolism in dairy cows under various conditions (e.g. fasting) or the evolution of liver proteome during embryonic phases or growing periods in chicken. Those studies provide interesting results leading to a better understanding of the liver metabolism. Liver steatosis development in waterfowl represents a special case and a focus on proteomic studies conducted in these birds will be done. Indeed, recent studies aimed at resolving protein evolution during overfeeding in duck. Proteomic analysis combining two complementary approaches (2-dimensional electrophoresis gels and shot gun strategy) in order to better understand the mechanisms underlying the variability of cooking yield of fatty liver will be presented.

Proteome response of fish under multiple stress exposure: Effects of pesticide mixtures and temperature increase.

Aquatic systems can be subjected to multiple stressors, including pollutant cocktails and elevated temperature. Evaluating the combined effects of these stressors on organisms is a great challenge in environmental sciences. To the best of our knowledge, this is the first study to assess the molecular stress response of an aquatic fish species subjected to individual and combined pesticide mixtures and increased temperatures. For that, goldfish (Carassius auratus) were acclimated to two different temperatures (22 and 32°C) for 15 days. They were then exposed for 96h to a cocktail of herbicides and fungicides (S-metolachlor, isoproturon, linuron, atrazine-desethyl, aclonifen, pendimethalin and tebuconazole) at two environmentally relevant concentrations (total concentrations of 8.4μgL-1 and 42μgL-1) at these two temperatures (22 and 32°C). The molecular response in liver was assessed by 2D-proteomics. Identified proteins were integrated using pathway enrichment analysis software to determine the biological functions involved in the individual or combined stress responses and to predict the potential deleterious outcomes. The pesticide mixtures elicited pathways involved in cellular stress response, carbohydrate, protein and lipid metabolisms, methionine cycle, cellular functions, cell structure and death control, with concentration- and temperature-dependent profiles of response. We found that combined temperature increase and pesticide exposure affected the cellular stress response: the effects of oxidative stress were more marked and there was a deregulation of the cell cycle via apoptosis inhibition. Moreover a decrease in the formation of glucose by liver and in ketogenic activity was observed in this multi-stress condition. The decrease in both pathways could reflect a shift from a metabolic compensation strategy to a conservation state. Taken together, our results showed (1) that environmental cocktails of herbicides and fungicides induced important changes in pathways involved in metabolism, cell structure and cell cycle, with possible deleterious outcomes at higher biological scales and (2) that increasing temperature could affect the response of fish to pesticide exposure.

Proteolytic activity alterations resulting from force-feeding in Muscovy and Pekin ducks

We investigated liver protease activity in force-fed and non-force-fed ducks using zymography gels to better understand mechanisms underlying liver steatosis in palmipeds. Male Muscovy and Pekin ducks were slaughtered before and after a short period (13 d) while they were conventionally fed or force fed. The force-fed regimen contained a high level of carbohydrates and was delivered in large doses. Main hepatic proteases (matrix metalloprotease-2, calpains, and cathepsins) were extracted from raw liver and specifically activated within electrophoretic gels. Both force-fed Muscovy and Pekin ducks presented higher liver weights and BW associated with lower matrix metalloprotease-2 and m-calpain hepatic activities. On the other hand, hepatic cathepsin activity was not affected by force feeding. It was concluded that Muscovy and Pekin duck hepatic proteases are affected similarly by the force feeding. Thus, this cannot explain differences observed between Muscovy and Pekin ducks regarding their ability to develop hepatic steatosis generally reported in literature.

Apoptosis during the development of the hepatic steatosis in force-fed ducks and cooking yield implications

ABSTRACT Mule ducks were force‐fed for 12 d to determine whether or not signs of apoptosis could occur during the development of the hepatic steatosis induced by the huge quantities of corn ingested twice daily by the birds. Presence of apoptosis in hepatocytes was assessed through the measurements of increased activities of capsase‐3 +‐7, ‐8, and ‐9. From d 0 of the force‐feeding period until d 8, activities of the different caspases remained at a low level. On the contrary, at d 10 and d 12, activities of all measured caspases dramatically increased, indicating that apoptosis occurred at this stage, which corresponds to the time of accumulation of large quantities of lipids in the hepatic cells. The melting level of the liver issued from force‐feeding (“foie gras”) during cooking is a point of interest for processors because it could degrade the quality of this delicate dish. In this study, we used the levels of caspases activities to improve the predictability of foie gras cooking, in addition to other parameters usually used, such as its weight or lipid content. From this improvement, we suggest that part of the variability of melting during cooking of fatty livers could reside in more or less intense activity of hepatic proteases.

Deciphering mechanisms underlying the genetic variation of general production and liver quality traits in the overfed mule duck by pQTL analyses

BackgroundThe aim of this study was to analyse the mechanisms that underlie phenotypic quantitative trait loci (QTL) in overfed mule ducks by identifying co-localized proteomic QTL (pQTL). The QTL design consisted of three families of common ducks that were progeny-tested by using 294 male mule ducks. This population of common ducks was genotyped using a genetic map that included 334 genetic markers located across 28 APL chromosomes (APL for Anas platyrhynchos). Mule ducks were phenotyped for 49 traits related to growth, metabolism, overfeeding ability and meat and fatty liver quality, and 326 soluble fatty liver proteins were quantified.ResultsOne hundred and seventy-six pQTL and 80 phenotypic QTL were detected at the 5% chromosome-wide significance threshold. The great majority of the identified pQTL were trans-acting and localized on a chromosome other than that carrying the coding gene. The most significant pQTL (1% genome-wide significance) were found for alpha-enolase on APL18 and fatty acid synthase on APL24. Some proteins were associated with numerous pQTL (for example, 17 and 14 pQTL were detected for alpha-enolase and apolipoprotein A1, respectively) and pQTL hotspots were observed on some chromosomes (APL18, 24, 25 and 29). We detected 66 co-localized phenotypic QTL and pQTL for which the significance of the two-trait QTL (2t-QTL) analysis was higher than that of the strongest QTL using a single-trait approach. Among these, 16 2t-QTL were pleiotropic. For example, on APL15, melting rate and abundance of two alpha-enolase spots appeared to be impacted by a single locus that is involved in the glycolytic process. On APLZ, we identified a pleiotropic QTL that modified both the blood level of glucose at the beginning of the force-feeding period and the concentration of glutamate dehydrogenase, which, in humans, is involved in increased glucose absorption by the liver when the glutamate dehydrogenase 1 gene is mutated.ConclusionsWe identified pleiotropic loci that affect metabolic pathways linked to glycolysis or lipogenesis, and in the end to fatty liver quality. Further investigation, via transcriptomics and metabolomics approaches, is required to confirm the biomarkers that were found to impact the genetic variability of these phenotypic traits.