Ontogeny of hepatic metabolism in mule ducks highlights different gene expression profiles between carbohydrate and lipid metabolic pathways.

BACKGROUND: The production of foie gras involves different metabolic pathways in the liver of overfed ducks such as lipid synthesis and carbohydrates catabolism, but the establishment of these pathways has not yet been described with precision during embryogenesis. The early environment can have short- and long-term impacts on the physiology of many animal species and can be used to influence physiological responses that is called programming. This study proposes to describe the basal hepatic metabolism at the level of mRNA in mule duck embryos in order to reveal potential interesting programming windows in the context of foie gras production. To this end, a kinetic study was designed to determine the level of expression of selected genes involved in steatosis-related liver functions throughout embryogenesis. The livers of 20 mule duck embryos were collected every 4 days from the 12th day of embryogenesis (E12) until 4 days after hatching (D4), and gene expression analysis was performed. The expression levels of 50 mRNAs were quantified for these 7 sampling points and classified into 4 major cellular pathways. RESULTS: Interestingly, most mRNAs involved in lipid metabolism are overexpressed after hatching (FASN, SCD1, ACOX1), whereas genes implicated in carbohydrate metabolism (HK1, GAPDH, GLUT1) and development (HGF, IGF, FGFR2) are predominantly overexpressed from E12 to E20. Finally, regarding cellular stress, gene expression appears quite stable throughout development, contrasting with strong expression after hatching (CYP2E1, HSBP1, HSP90AA1). CONCLUSION: For the first time we described the kinetics of hepatic ontogenesis at mRNA level in mule ducks and highlighted different expression patterns depending on the cellular pathway. These results could be particularly useful in the design of embryonic programming for the production of foie gras.

Kinetic study of the expression of genes related to hepatic steatosis, glucose and lipid metabolism, and cellular stress during overfeeding in mule ducks.

Induced by overfeeding, hepatic steatosis is a process exploited for the "foie gras" production in mule ducks. To better understand the mechanisms underlying its development, the physiological responses of mule ducks overfed with corn for a duration of 11 days were analyzed. A kinetic analysis of glucose and lipid metabolism and cell protection mechanisms was performed on 96 male mule ducks during overfeeding with three sampling times (after the 4th, the 12th, and the 22nd meal). Gene expression and protein analysis realized on the liver, muscle, and abdominal fat showed an activation of a cholesterol biosynthetic pathway during the complete overfeeding period mainly in livers with significant correlations between its weight and its cholesterolemia (r = 0.88; P < 0.0001) and between the liver weight and the hmgcr and soat1 expression (r = 0.4, P < 0.0001 and r = 0.67; P < 0.0001, respectively). Results also revealed an activation of insulin and amino acid cells signaling a pathway suggesting that ducks boost insulin sensitivity to raise glucose uptake and use via glycolysis and lipogenesis. Cellular stress analysis revealed an upregulation of key autophagy-related gene expression atg8 and sqstm1(P < 0.0001) during the complete overfeeding period, mainly in the liver, in contrast to an induction of cyp2e1(P < 0.0001), suggesting that autophagy could be suppressed during steatosis development. This study has highlighted different mechanisms enabling mule ducks to efficiently handle the starch overload by keeping its liver in a nonpathological state. Moreover, it has revealed potential biomarker candidates of hepatic steatosis as plasma cholesterol for the liver weight.

Impacts of Embryonic Thermal Programming on the Expression of Genes Involved in Foie gras Production in Mule Ducks.

Embryonic thermal programming has been shown to improve foie gras production in overfed mule ducks. However, the mechanisms at the origin of this programming have not yet been characterized. In this study, we investigated the effect of embryonic thermal manipulation (+1°C, 16 h/24 h from embryonic (E) day 13 to E27) on the hepatic expression of genes involved in lipid and carbohydrate metabolisms, stress, cell proliferation and thyroid hormone pathways at the end of thermal manipulation and before and after overfeeding (OF) in mule ducks. Gene expression analyses were performed by classic or high throughput real-time qPCR. First, we confirmed well-known results with strong impact of OF on the expression of genes involved in lipid and carbohydrates metabolisms. Then we observed an impact of OF on the hepatic expression of genes involved in the thyroid pathway, stress and cell proliferation. Only a small number of genes showed modulation of expression related to thermal programming at the time of OF, and only one was also impacted at the end of the thermal manipulation. For the first time, we explored the molecular mechanisms of embryonic thermal programming from the end of heat treatment to the programmed adult phenotype with optimized liver metabolism.

The Autophagic Flux Inhibitor Bafilomycine A1 Affects the Expression of Intermediary Metabolism-Related Genes in Trout Hepatocytes.

Autophagy is an evolutionarily conserved process of cellular self-eating which emerged these last years as a major adaptive metabolic response to various stresses such as fasting, hypoxia, or environmental pollutants. However, surprisingly very few data is currently available on its role in fish species which are directly exposed to frequent environmental perturbations. Here, we report that the treatment of fasted trout hepatocytes with the autophagy inhibitor Bafilomycine A1 lowered the mRNA levels of many of the gluconeogenesis-related genes and increased those of genes involved in intracellular lipid stores. Concurrently, intracellular free amino acid levels dropped and the expression of the main genes involved in the endoplasmic reticulum (ER) stress exhibited a sharp increase in autophagy inhibited cells. Together these results highlight the strong complexity of the crosstalk between ER, autophagy and metabolism and support the importance of considering this function in future studies on metabolic adaptation of fish to environmental stresses.

Positive Impact of Thermal Manipulation During Embryogenesis on Foie Gras Production in Mule Ducks.

Animal studies have shown that very early life events may have programing effects on adult metabolism and health. In this study, we aim, for the first, time to elucidate the effects of embryonic thermal manipulation (TM) on the performance of overfed mule ducks, in particular for the production of foie gras (fatty liver). We designed three embryonic TMs with different protocols for increasing the incubation temperature during the second part of embryogenesis, to determine whether hepatic metabolism could be "programed" to improve its fattening response to overfeeding at the age of three months. Initial results confirm that an increase in the incubation temperature leads to faster development (observed for all treated groups compared to the control group), and a decrease in the body surface temperature at birth. Thereafter, in a very innovative way, we showed that the three TM conditions specifically increased liver weights, as well as liver lipid content after overfeeding compared to the non-TM control group. These results demonstrate that embryonic TM effectively "programs" the metabolic response to the challenge of force-feeding, resulting in increased hepatic steatosis. Finally, our goal of improving foie gras production has been achieved with three different embryonic thermal stimuli, demonstrating the high reproducibility of the method. However, this repeatability was also perceptible in the adverse effects observed on two groups treated with exactly the same cumulative temperature rise leading to a reduction in hatchability (75 and 76% vs. 82% in control), in addition to an increase in the melting rate after cooking. These results suggest that embryonic thermal programing could be an innovative and inexpensive technique for improving foie gras production, although the specific protocol (duration, level or period of temperature increase), remains to be elucidated in order to avoid adverse effects.