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

Metabolic and molecular signatures of improved growth in Atlantic salmon (Salmo salar) fed surplus levels of methionine, folic acid, vitamin B6 and B12 throughout smoltification.

A moderate surplus of the 1C nutrients methionine, folic acid, vitamin B6 and B12 above dietary recommendations for Atlantic salmon has shown to improve growth and reduce hepatosomatic index in the on-growing salt water period when fed throughout smoltification. Metabolic properties and molecular mechanisms determining the improved growth are unexplored. Here, we investigate metabolic and transcriptional signatures in skeletal muscle taken before and after smoltification to acquire deeper insight into pathways and possible nutrient-gene-interactions. A control feed (Ctrl) or 1C nutrient surplus feed (1C+) were fed to Atlantic salmon six weeks prior to smoltification until three months after salt water transfer. Both metabolic and gene expression signatures revealed significant 1C nutrient-dependent changes already at pre-smolt, but differences intensified when analysing post-smolt muscle. Transcriptional differences revealed a lower expression of genes related to translation, growth, and amino acid metabolization in post-smolt muscle when fed additional 1C nutrients. The 1C+ group showed less free amino acid and putrescine levels, and higher methionine and glutathione (GSH) amounts in muscle. For Ctrl muscle, the overall metabolic profile suggests a lower amino acid utilization for protein synthesis, and increased methionine metabolization in polyamine and redox homeostasis, whereas transcription changes are indicative of compensatory growth regulation at local tissue level. These findings point to fine-tuned nutrient-gene-interactions fundamental for improved growth capacity through better amino acid utilization for protein accretion when salmon was fed additional 1C nutrients throughout smoltification. It also highlights potential nutritional programming strategies on improved post-smolt growth through 1C+ supplementation before and throughout smoltification.acceptedVersio

Parental micronutrient deficiency distorts liver DNA methylation and expression of lipid genes associated with a fatty-liver-like phenotype in offspring

Micronutrient status of parents can affect long term health of their progeny. Around 2 billion humans are affected by chronic micronutrient deficiency. In this study we use zebrafish as a model system to examine morphological, molecular and epigenetic changes in mature offspring of parents that experienced a one-carbon (1-C) micronutrient deficiency. Zebrafish were fed a diet sufficient, or marginally deficient in 1-C nutrients (folate, vitamin B12, vitamin B6, methionine, choline), and then mated. Offspring livers underwent histological examination, RNA sequencing and genome-wide DNA methylation analysis. Parental 1-C micronutrient deficiency resulted in increased lipid inclusion and we identified 686 differentially expressed genes in offspring liver, the majority of which were downregulated. Downregulated genes were enriched for functional categories related to sterol, steroid and lipid biosynthesis, as well as mitochondrial protein synthesis. Differential DNA methylation was found at 2869 CpG sites, enriched in promoter regions and permutation analyses confirmed the association with parental feed. Our data indicate that parental 1-C nutrient status can persist as locus specific DNA methylation marks in descendants and suggest an effect on lipid utilization and mitochondrial protein translation in F1 livers. This points toward parental micronutrients status as an important factor for offspring health and welfare.publishedVersio

Spawning manipulation, broodfish diet feeding and egg production in farmed Atlantic salmon

Atlantic salmon aquaculture relies on continuous supply of high quality eggs. Broodfish nutrition and manipulation of ovulation time (photoperiod and temperature) are key factors. The optimum feeding period with broodfish diet has not been investigated before. The present study examined how feeding period with broodfish diet (9 vs. 17 months) interacted with manipulation of ovulation time (early (Nov), normal (Dec), late (Feb)) on broodstock egg production capacity and egg quality in two-sea-winter female Atlantic salmon (∼12 kg). All groups were fed until June 2021 when they were transferred to tanks and starved until ovulation.publishedVersio

Micronutrient supplementation affects DNA methylation in male gonads with potential intergenerational epigenetic inheritance involving the embryonic development through glutamate receptor-associated genes

Background DNA methylation has an important role in intergenerational inheritance. An increasing number of studies have reported evidence of germline inheritance of DNA methylation induced by nutritional signals in mammals. Vitamins and minerals as micronutrients contribute to growth performance in vertebrates, including Atlantic salmon (Salmo salar), and also have a role in epigenetics as environmental factors that alter DNA methylation status. It is important to understand whether micronutrients in the paternal diet can influence the offspring through alterations of DNA methylation signatures in male germ cells. Results Here, we show the effect of micronutrient supplementation on DNA methylation profiles in the male gonad through a whole life cycle feeding trial of Atlantic salmon fed three graded levels of micronutrient components. Our results strongly indicate that micronutrient supplementation affects the DNA methylation status of genes associated with cell signalling, synaptic signalling, and embryonic development. In particular, it substantially affects DNA methylation status in the promoter region of a glutamate receptor gene, glutamate receptor ionotropic, NMDA 3A-like (grin3a-like), when the fish are fed both medium and high doses of micronutrients. Furthermore, two transcription factors, histone deacetylase 2 (hdac2) and a zinc finger protein, bind to the hyper-methylated site in the grin3a-like promoter. An estimated function of hdac2 together with a zinc finger indicates that grin3a-like has a potential role in intergenerational epigenetic inheritance and the regulation of embryonic development affected by paternal diet. Conclusions The present study demonstrates alterations of gene expression patterns and DNA methylation signatures in the male gonad when Atlantic salmon are fed different levels of micronutrients. Alterations of gene expression patterns are of great interest because the gonads are supposed to have limited metabolic activities compared to other organs, whereas alterations of DNA methylation signatures are of great importance in the field of nutritional epigenetics because the signatures affected by nutrition could be transferred to the next generation. We provide extensive data resources for future work in the context of potential intergenerational inheritance through the male germline

Micronutrient supplementation affects transcriptional and epigenetic regulation of lipid metabolism in a dose-dependent manner

Micronutrients (vitamins and minerals) have been less well studied compared to macronutrients (fats, proteins, and carbohydrates) although they play important roles in growth, metabolism, and maintenance of tissues. Hence, there is growing interest to understand the influence of micronutrients across various aspects in nutritional research. In the last two decades, aquaculture feeds have been shifted to containing more plant-based materials to meet the increasing demand and maintain the sustainability in the industry. In cultured fish, changing raw materials in feeds alters the requirement levels of micronutrients. A recent whole life cycle feeding trial of Atlantic salmon (Salmo salar) with graded levels of micronutrient packages has concluded that the levels of several B-vitamins and microminerals need to be increased from the current recommendation levels for optimal growth and fish welfare when plant-based diets are used. Here, we show the effect of micronutrient supplementation on transcriptional and epigenetic regulation by using liver samples from the same feeding trial. Specifically, our aim is to reveal the mechanisms of altered cell metabolism, which results in improved growth performance by micronutrient surpluses, at gene expression and DNA methylation levels. Our results strongly indicate that micronutrient supplementation suppresses gene expression in lipid metabolism in a dose-dependent manner and broadly affects DNA methylation in cell adhesion and cell signalling. In particular, it increases DNA methylation levels on the acetyl-CoA carboxylase alpha promoter in a concentration-dependent manner, which further suggests that acetyl-CoA carboxylase alpha is an upstream epigenetic regulator controlling its downstream lipid biosynthesis activities. This study demonstrates a comprehensive analysis to reveal an important role of micronutrients in lipid metabolism through epigenetic control of gene expression

Effects of antiplatelet therapy on stroke risk by brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases: subgroup analyses of the RESTART randomised, open-label trial

Background Findings from the RESTART trial suggest that starting antiplatelet therapy might reduce the risk of recurrent symptomatic intracerebral haemorrhage compared with avoiding antiplatelet therapy. Brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases (such as cerebral microbleeds) are associated with greater risks of recurrent intracerebral haemorrhage. We did subgroup analyses of the RESTART trial to explore whether these brain imaging features modify the effects of antiplatelet therapy

Effects of antiplatelet therapy after stroke due to intracerebral haemorrhage (RESTART): a randomised, open-label trial

Background: Antiplatelet therapy reduces the risk of major vascular events for people with occlusive vascular disease, although it might increase the risk of intracranial haemorrhage. Patients surviving the commonest subtype of intracranial haemorrhage, intracerebral haemorrhage, are at risk of both haemorrhagic and occlusive vascular events, but whether antiplatelet therapy can be used safely is unclear. We aimed to estimate the relative and absolute effects of antiplatelet therapy on recurrent intracerebral haemorrhage and whether this risk might exceed any reduction of occlusive vascular events. Methods: The REstart or STop Antithrombotics Randomised Trial (RESTART) was a prospective, randomised, open-label, blinded endpoint, parallel-group trial at 122 hospitals in the UK. We recruited adults (≥18 years) who were taking antithrombotic (antiplatelet or anticoagulant) therapy for the prevention of occlusive vascular disease when they developed intracerebral haemorrhage, discontinued antithrombotic therapy, and survived for 24 h. Computerised randomisation incorporating minimisation allocated participants (1:1) to start or avoid antiplatelet therapy. We followed participants for the primary outcome (recurrent symptomatic intracerebral haemorrhage) for up to 5 years. We analysed data from all randomised participants using Cox proportional hazards regression, adjusted for minimisation covariates. This trial is registered with ISRCTN (number ISRCTN71907627). Findings: Between May 22, 2013, and May 31, 2018, 537 participants were recruited a median of 76 days (IQR 29–146) after intracerebral haemorrhage onset: 268 were assigned to start and 269 (one withdrew) to avoid antiplatelet therapy. Participants were followed for a median of 2·0 years (IQR [1·0– 3·0]; completeness 99·3%). 12 (4%) of 268 participants allocated to antiplatelet therapy had recurrence of intracerebral haemorrhage compared with 23 (9%) of 268 participants allocated to avoid antiplatelet therapy (adjusted hazard ratio 0·51 [95% CI 0·25–1·03]; p=0·060). 18 (7%) participants allocated to antiplatelet therapy experienced major haemorrhagic events compared with 25 (9%) participants allocated to avoid antiplatelet therapy (0·71 [0·39–1·30]; p=0·27), and 39 [15%] participants allocated to antiplatelet therapy had major occlusive vascular events compared with 38 [14%] allocated to avoid antiplatelet therapy (1·02 [0·65–1·60]; p=0·92). Interpretation: These results exclude all but a very modest increase in the risk of recurrent intracerebral haemorrhage with antiplatelet therapy for patients on antithrombotic therapy for the prevention of occlusive vascular disease when they developed intracerebral haemorrhage. The risk of recurrent intracerebral haemorrhage is probably too small to exceed the established benefits of antiplatelet therapy for secondary prevention

Effects of antiplatelet therapy after stroke due to intracerebral haemorrhage (RESTART): a randomised, open-label trial

Background: Antiplatelet therapy reduces the risk of major vascular events for people with occlusive vascular disease, although it might increase the risk of intracranial haemorrhage. Patients surviving the commonest subtype of intracranial haemorrhage, intracerebral haemorrhage, are at risk of both haemorrhagic and occlusive vascular events, but whether antiplatelet therapy can be used safely is unclear. We aimed to estimate the relative and absolute effects of antiplatelet therapy on recurrent intracerebral haemorrhage and whether this risk might exceed any reduction of occlusive vascular events. Methods: The REstart or STop Antithrombotics Randomised Trial (RESTART) was a prospective, randomised, open-label, blinded endpoint, parallel-group trial at 122 hospitals in the UK. We recruited adults (≥18 years) who were taking antithrombotic (antiplatelet or anticoagulant) therapy for the prevention of occlusive vascular disease when they developed intracerebral haemorrhage, discontinued antithrombotic therapy, and survived for 24 h. Computerised randomisation incorporating minimisation allocated participants (1:1) to start or avoid antiplatelet therapy. We followed participants for the primary outcome (recurrent symptomatic intracerebral haemorrhage) for up to 5 years. We analysed data from all randomised participants using Cox proportional hazards regression, adjusted for minimisation covariates. This trial is registered with ISRCTN (number ISRCTN71907627). Findings: Between May 22, 2013, and May 31, 2018, 537 participants were recruited a median of 76 days (IQR 29–146) after intracerebral haemorrhage onset: 268 were assigned to start and 269 (one withdrew) to avoid antiplatelet therapy. Participants were followed for a median of 2·0 years (IQR [1·0– 3·0]; completeness 99·3%). 12 (4%) of 268 participants allocated to antiplatelet therapy had recurrence of intracerebral haemorrhage compared with 23 (9%) of 268 participants allocated to avoid antiplatelet therapy (adjusted hazard ratio 0·51 [95% CI 0·25–1·03]; p=0·060). 18 (7%) participants allocated to antiplatelet therapy experienced major haemorrhagic events compared with 25 (9%) participants allocated to avoid antiplatelet therapy (0·71 [0·39–1·30]; p=0·27), and 39 [15%] participants allocated to antiplatelet therapy had major occlusive vascular events compared with 38 [14%] allocated to avoid antiplatelet therapy (1·02 [0·65–1·60]; p=0·92). Interpretation: These results exclude all but a very modest increase in the risk of recurrent intracerebral haemorrhage with antiplatelet therapy for patients on antithrombotic therapy for the prevention of occlusive vascular disease when they developed intracerebral haemorrhage. The risk of recurrent intracerebral haemorrhage is probably too small to exceed the established benefits of antiplatelet therapy for secondary prevention