RNA form baker's yeast cultured with and without lipopolysaccharide (LPS) modulates gene transcription in an intestinal epithelial cell model, RTgutGC from rainbow trout (Oncorhynchus mykiss)

The objective of this study was to evaluate if the intestinal RTgutGC cell line could be suitable for research on dietary ingredients and their function as modulators of inflammation during lipopolysaccharide (LPS) induced stress.publishedVersio

Development of a fatty liver model using oleic acid in primary liver cells isolated from Atlantic salmon and the prevention of lipid accumulation using metformin

The following study aimed to develop a fatty liver model in primary hepatocytes isolated from Atlantic salmon. In order to induce the fatty liver, oleic acid (OA) at 0.2 or 0.4 mM was used. Metformin, known to prevent and cure fatty liver in mammalian cells, was used at 1 or 10 mM for 24 hr before addition of OA to test possible prevention effect of metformin on the OA‐induced fatty liver phenotype. Cells grown in 0.2 mM OA did not increase the mean number of lipid droplets, while cells grown in 0.4 mM OA increased the number of lipid droplets within the liver cells (p < 0.0001). Metformin pretreatment prior to OA supplementation reduced the mean number of lipid droplets. Gene expression of ApoB100, CD36 and PPARa increased in cells treated with metformin and most so at 10 mM. On the other hand, gene expression of LXR, SREBP2 and CPT‐1 decreased at both concentrations of metformin, while OA treatment did not affect these genes. Gene expression of IL‐8 increased by 0.4 mM OA (p = 0.002). Metformin reduced the gene expression of IL‐8. Thus, metformin efficiently enhanced the expression of genes related to transport and oxidation of lipids in hepatic cells of salmon, but required higher concentrations of OA and metformin than those required in rodent models to increase and prevent lipid accumulation, respectively.publishedVersio

Differential production of prostaglandins and prostacyclins by liver and head kidney cells from Atlantic salmon challenged with arachidonic and eicosapentaenoic acids

Polyunsaturated fatty acids such as arachidonic and eicosapentaenoic acids are the precursors of eicosanoid metabolites (e.g prostaglandins and prostacyclins) which regulate inflammatory and immune response processes in fish organs. The present research studies the differential production of PGI2, PGI3, PGE2 and PGE3 by primary liver and head kidney cells isolated from salmon and challenged with single or combined ARA and/or EPA. There was a significant increase in the production of PGE2 and PGI3 in both types of cells after exposure to single and combined fatty acids. Increased production of PGE3 was only detected in liver cells after exposure to ARA+EPA. The levels of PGI2 in liver cells were significantly increased after exposure to all the tested fatty acid systems, while the production levels in head kidney cells were only significant after exposure to ARA or ARA+EPA, but not to EPA, where the production was non-significantly decreased compared to the control cells. In general, liver cells synthetized higher prostaglandin levels than prostacyclins, and the opposite was observed in head kidney cells, which synthetized highly remarkable amounts of prostacyclin compared to liver cells. The overall production for both types of cells and various fatty acid systems were characterized by a high proportion of the omega-3 fatty acid metabolites (PGE3+PGI3) compared to the omega-6 counterpart (PGE2+PGI2). Some potential production mechanisms are proposed and comprehensively discussed. The results of the present research are the first to deliver the differential production of prostacyclins and prostaglandins by liver and head kidney cells from salmon, thereby paving the way for understanding the significance of these prostanoids in fish physiology and disease.publishedVersio

Earlier or delayed seasonal broodstock spawning changes nutritional status and metabolic programming of growth for next-generation Atlantic salmon

Atlantic salmon (Salmo salar) breeding companies depend on changing light, temperature and feeding regimes to achieve new generations outside the natural spawning season. However, there have been few conducted trials reported that have studied whether this shift affects important traits. We test whether an induced shift of two months earlier or two months later than normal spawning season affects the nutritional status (folate, methionine, vitamin B12, vitamin B6, free amino acids, N-metabolites and lipids) in broodstock liver and muscle and whether this affects the levels of the same nutrients in the offspring. The results showed significant seasonal differences in the Cahill cycle (glucose-alanine cycle), 1C metabolism and for free amino acids catabolized in the citric acid cycle all which are important for embryonic growth The broodstock nutritional status was reflected in the eggs. Nutritional status of broodstock liver and muscle and newly fertilized eggs showed two general scenarios: Advanced spawning period did not obtain optimal deposition of nutrients in the eggs. Delayed spawning broodstock displayed a metabolic profile which indicated that it had enhanced catabolization of muscle protein which led to accumulation of aminogroups from muscle breakdown to such a degree that these amino groups were increased in the eggs. The total body weight at start-feeding stage revealed best growth for both the normal and late spawning compared to early spawning. We show here that environmental alterations in broodstock husbandry influence the nutrient status of the next generation via nutritional and metabolic programming. This is an important concept which needs more careful awareness as the metabolism compensate and regulate the energy between catabolism and anabolism through the early stages of cell divisions which give rise to changes in permanent traits for the next generation.publishedVersio

Out-of-season spawning affects the nutritional status and gene expression in both Atlantic salmon female broodstock and their offspring

The Atlantic salmon aquaculture industry relies on adjustments of female broodstock spawning season to meet the demand for delivery of embryos outside the natural spawning season. Earlier results from zebrafish have shown that parental micronutrient status program offspring metabolism. Therefore, the main hypothesis of this study was to investigate if out-of-season (off-season) broodstock (spawning in June, in land-based recirculation systems) and their offspring deviate in micronutrient status when compared to broodstock and offspring from normal spawning season. Both seasons of female Atlantic salmon broodstock were fed the same diet and starved for approximately the same time interval prior to spawning. We compared nutrients related to the 1C metabolism (vitamin B12, folate, vitamin B6, methionine), free amino acids (FAAs) and lipid classes in broodstock muscle and liver tissues, and during offspring ontogeny. In general, the off-season broodstock showed higher levels of folate, vitamin B6 and selected FAAs in muscle tissue, and higher levels of folate and lipids (cholesterol and sphingomyelin) in liver tissue compared to normal-season. Furthermore, embryos from off-season had reduced amounts of all the measured lipid classes, like cholesterol and sphingomyelin, and lower levels of one type of folate and changes in FAAs and N-metabolites. We discovered significant differences between the seasons in mRNA levels of genes controlling fatty acid synthesis and 1C metabolism in both broodstock liver and offspring. Moreover, for genes controlling the methylation of DNA; both maintenance and de novo DNA methyltransferases (DNMTs) were expressed at higher levels in off-season compared to normal-season offspring. Our results show, in general that normal spawning season broodstock allocated more nutrients to eggs than off-season. Our results indicate a potential for improved maturation for off-season group to obtain a higher offspring growth potential, and this argues for a reassessment of the nutritional influence from broodstock to offspring and the consequences through nutritional programming.publishedVersio

Methionine deficiency does not increase polyamine turnover through depletion of hepatic S-adenosylmethionine in juvenile Atlantic salmon

During the last few decades, plant protein ingredients such as soya proteins have replaced fishmeal in the diets of aquacultured species. This may affect the requirement and metabolism of methionine as soya contains less methionine compared with fishmeal. To assess whether methionine limitation affects decarboxylated S-adenosylmethionine availability and polyamine status, in the present study, juvenile Atlantic salmon were fed a methionine-deficient plant protein-based diet or the same diet supplemented with dl-methionine for 8 weeks. The test diets were compared with a fishmeal-based control diet to assess their effects on the growth performance of fish. Methionine limitation reduced growth and protein accretion, but when fish were fed the dl-methionine-supplemented diet their growth and protein accretion equalled those of fish fed the fishmeal-based control diet. Methionine limitation reduced free methionine concentrations in the plasma and muscle, while those in the liver were not affected. S-adenosylmethionine (SAM) concentrations were higher in the liver of fish fed the methionine-deficient diet, while S-adenosylhomocysteine concentrations were not affected. Putrescine concentrations were higher and spermine concentrations were lower in the liver of fish fed the methionine-deficient diet, while the gene expression of SAM decarboxylase (SAMdc) and the rate-limiting enzyme of polyamine synthesis ornithine decarboxylase (ODC) was not affected. Polyamine turnover, as assessed by spermine/spermidine acetyltransferase (SSAT) abundance, activity and gene expression, was not affected by treatment. However, the gene expression of the cytokine TNF-α increased in fish fed the methionine-deficient diet, indicative of stressful conditions in the liver. Even though taurine concentrations in the liver were not affected by treatment, methionine and taurine concentrations in muscle decreased due to methionine deficiency. Concomitantly, liver phospholipid and cholesterol concentrations were reduced, while NEFA concentrations were elevated. In conclusion, methionine deficiency did not increase polyamine turnover through depletion of hepatic SAM, as assessed by SSAT activity and abundance.publishedVersio

Effect of dietary replacement of fish meal with insect meal on in vitro bacterial and viral induced gene response in Atlantic salmon (Salmo salar) head kidney leukocytes

Abstract With the fast growth of today's aquaculture industry, the demand for aquafeeds is expanding dramatically. Insects, which are part of the natural diet of salmonids, could represent a sustainable ingredient for aquaculture feed. The aim of the current study was to test how a partial or total replacement of dietary fishmeal with insect meal affect gene responses involved in inflammation, the eicosanoid pathway and stress response in Atlantic salmon (Salmo salar L.) in isolated head kidney leukocytes after exposure to bacterial or viral mimic. Insect meal (IM) was produced from black soldier fly (BSF, Hermetia illucens) larvae. Seawater Atlantic salmon were fed three different diets for 8 weeks; a control diet (IM0, protein from fishmeal and plant based ingredients (25:75) and lipid from fish oil and vegetable oil (33:66); and two insect-meal containing diets, IM66 and IM100, where 66 and 100% of the fishmeal protein was replaced with IM, respectively. Leukocytes were isolated from the head kidney of fish (n = 6) from each of the three dietary groups. Isolated leukocytes were seeded into culture wells and added either a bacterial mimic (lipopolysaccharide, LPS) or a viral mimic (polyinosinic acid: polycytidylic acid, poly I: C) to induce an inflammatory response. Controls (Ctl) without LPS and poly I: C were included. The transcription of interleukins IL-1β, IL-8, IL-10 and TNF-α were elevated in LPS treated leukocytes isolated from salmon fed the three dietary groups (IM0, IM66 and IM100). The inflammatory-related gene expression in head kidney cells were, however, not affected by the pre-fed substitution of fish meal with IM in the diet of salmon. Gene transcriptions of PTGDS and PTGES were neither affected by LPS, poly I: C or the experimental diets fed prior to cell isolation, while salmon fed with IM showed a lower expression of LOX5. The gene expression of TLR22 and C/EBP-β were down-regulated by the LPS treatment in the cells isolated from salmon fed insect-based diets (IM66 and IM100) compared to fish fed the IM0. Similarly, the leukocytes challenged with LPS and isolated from fish fed with IM66 and IM100 down-regulated the expression of Mn-SOD, GPx1, HSP27 and HSP70 compared to salmon fed IM0. In general, these results suggested that replacement of fishmeal with IM in the diets of Atlantic salmon had no effect on the transcription of pro-inflammatory genes in the head kidney cells. There was, however, an effect of dietary IM on the transcription of antioxidant and stress related genes in the leukocytes

Amino Acid Carriers of the Solute Carrier Families 7 (SLC7) and 38 (SLC38) Are Involved in Leucine Sensing in the Brain of Atlantic Salmon (Salmo salar)

Sensing of amino acids in fish brain, especially branched-chain amino acids (BCAA) like leucine, is involved in regulation of feed intake through different mechanisms. However, there is limited information regarding the possible involvement of mechanisms dependent on amino acid carriers of the solute carrier families (SLC) known to be key regulators of intracellular leucine concentration, namely L-type amino acid transporter 1 (LAT1), and sodium-dependent neutral amino acid transporter 2 (SNAT2) and 9,(SNAT9), for which evidence of their participation is available in mammals. Comparative analysis amongst sequences revealed a complex pattern of paralogues in Atlantic salmon, for LAT1 (slc7a5aa, slc7a5ab, slc7a5ba, slc7a5bb, slc7a5ca, and slc7a5cb), SNAT2 (slc38a2a and slc38a2b) and SNAT9 (slc38a9). After establishing phylogenetic relationships of the different paralogues evaluated, samples of the selected brain areas were taken from Atlantic salmon to assess tissue distribution of transcripts. In an additional experiment, fish were fed two diets with different levels of leucine (high leucine: 35 g/kg vs. control leucine: 27.3 g/kg). The high leucine diet resulted in lower feed intake and increased mRNA abundance of specific paralogues of LAT1 (slc7a5aa, slc7a5ab, and slc7a5bb) and SNAT2 (slc38a2a and slc38a2b) though apparently not for SNAT9 in brain areas like hypothalamus and telencephalon involved in food intake regulation. The results obtained suggest a role for members of the SLC family in the anorectic effect of leucine and thus their involvement as additional amino acid sensing mechanism not characterised so far in fish regulation of feed intake.publishedVersio