The tolerance of fish to fasting offers a model to study the regulatory mechanisms and changes produced when feeding is restored. Gilthead sea bream juveniles were exposed to a 21-days fasting period ...followed by 2 h to 7-days refeeding. Fasting provoked a decrease in body weight, somatic indexes, and muscle gene expression of members of the Gh/Igf system, signaling molecules (akt, tor and downstream effectors), proliferation marker pcna, myogenic regulatory factors, myostatin, and proteolytic molecules such as cathepsins or calpains, while most ubiquitin-proteasome system members increased or remained stable. In bone, downregulated expression of Gh/Igf members and osteogenic factors was observed, whereas expression of the osteoclastic marker ctsk was increased. Refeeding recovered the expression of Gh/Igf system, myogenic and osteogenic factors in a sequence similar to that of development. Akt and Tor phosphorylation raised at 2 and 5 h post-refeeding, much faster than its gene expression increased, which occurred at day 7. The expression in bone and muscle of the inhibitor myostatin (mstn2) showed an inverse profile suggesting an inter-organ coordination that needs to be further explored in fish. Overall, this study provides new information on the molecules involved in the musculoskeletal system remodeling during the early stages of refeeding in fish.
Gut microbiota plays a fundamental role in maintaining host’s health by controlling a wide range of physiological processes. Administration of probiotics and manipulation of photoperiod have been ...suggested as modulators of microbial composition and are currently undergoing an extensive research in aquaculture as a way to improve health and quality of harvested fish. However, our understanding regarding their effects on physiological processes is still limited. In the present study we investigated whether manipulation of photoperiod and/or probiotic administration was able to alter microbial composition in zebrafish larvae at hatching stage. Our findings show that probiotic does not elicit effects while photoperiod manipulation has a significant impact on microbiota composition. Moreover, we successfully predicted lipid biosynthesis and apoptosis to be modulated by microbial communities undergoing continuous darkness. Interestingly, expression levels of caspase 3 gene (casp3) and lipid-related genes (hnf4a, npc1l1, pparγ, srebf1, agpat4 and fitm2) were found to be significantly overexpressed in dark-exposed larvae, suggesting an increase in the occurrence of apoptotic processes and a lipid metabolism impairment, respectively (p < 0.05). Our results provide the evidence that microbial communities in zebrafish at early life stages are not modulated by a short administration of probiotics and highlight the significant effect that dark photoperiod elicits on zebrafish microbiota and potentially on health.
Control of tissue metabolism and growth involves interactions between organs, tissues, and cell types, mediated by cytokines or direct communication through cellular exchanges. Indeed, over the past ...decades, many peptides produced by adipose tissue, skeletal muscle and bone named adipokines, myokines and osteokines respectively, have been identified in mammals playing key roles in organ/tissue development and function. Some of them are released into the circulation acting as classical hormones, but they can also act locally showing autocrine/paracrine effects. In recent years, some of these cytokines have been identified in fish models of biomedical or agronomic interest. In this review, we will present their state of the art focusing on local actions and inter-tissue effects. Adipokines reported in fish adipocytes include adiponectin and leptin among others. We will focus on their structure characteristics, gene expression, receptors, and effects, in the adipose tissue itself, mainly regulating cell differentiation and metabolism, but in muscle and bone as target tissues too. Moreover, lipid metabolites, named lipokines, can also act as signaling molecules regulating metabolic homeostasis. Regarding myokines, the best documented in fish are myostatin and the insulin-like growth factors. This review summarizes their characteristics at a molecular level, and describes both, autocrine effects and interactions with adipose tissue and bone. Nonetheless, our understanding of the functions and mechanisms of action of many of these cytokines is still largely incomplete in fish, especially concerning osteokines (i.e., osteocalcin), whose potential cross talking roles remain to be elucidated. Furthermore, by using selective breeding or genetic tools, the formation of a specific tissue can be altered, highlighting the consequences on other tissues, and allowing the identification of communication signals. The specific effects of identified cytokines validated through
models or
trials will be described. Moreover, future scientific fronts (i.e., exosomes) and tools (i.e., co-cultures, organoids) for a better understanding of inter-organ crosstalk in fish will also be presented. As a final consideration, further identification of molecules involved in inter-tissue communication will open new avenues of knowledge in the control of fish homeostasis, as well as possible strategies to be applied in aquaculture or biomedicine.
Soybeans are one of the most used alternative dietary ingredients in aquafeeds. However, they contain phytoestrogens like genistein (GE), which can have an impact on fish metabolism and health. This ...study aimed to investigate the in vitro and in vivo effects of GE on lipid metabolism, apoptosis, and autophagy in rainbow trout (
). Primary cultured preadipocytes were incubated with GE at different concentrations, 10 or 100 μM, and 1 μM 17β-estradiol (E2). Furthermore, juveniles received an intraperitoneal injection of GE at 5 or 50 µg/g body weight, or E2 at 5 µg/g. In vitro, GE 100 μM increased lipid accumulation and reduced cell viability, apparently involving an autophagic process, indicated by the higher LC3-II protein levels, and higher
and
transcript levels achieved after GE 10 μM. In vivo, GE 50 µg/g upregulated the gene expression of fatty acid synthase (
) and glyceraldehyde-3-phosphate dehydrogenase in adipose tissue, suggesting enhanced lipogenesis, whereas it increased hormone-sensitive lipase in liver, indicating a lipolytic response. Besides, autophagy-related genes increased in the tissues analyzed mainly after GE 50 µg/g treatment. Overall, these findings suggest that an elevated GE administration could lead to impaired adipocyte viability and lipid metabolism dysregulation in rainbow trout.
The dietary inclusion of plant-based products in fish feeds formulation is required for the sustainable development of aquaculture. Moreover, considering functional diets, hydroxytyrosol, one of the ...major phenolic compounds found in olives (
Olea europaea
), has been identified as a potential candidate to be used in the aquafeeds industry due to its health promoting abilities. The aim of this study was to evaluate the effects of the inclusion of an olive juice extract rich in hydroxytyrosol as an additive (0.52 g HT/kg feed) in a high-fat (24% lipids) diet in gilthead sea bream (
Sparus aurata
) juveniles. Moreover, the experimental diets, with or without the extract, were administered daily at a standard (3% of total biomass in the tank) or restricted ration (40% reduction) for 8–9 weeks. Growth and biometric parameters, insulin-like growth factor 1 (IGF-1) plasma levels and growth hormone/IGF axis-, myogenic- and osteogenic-related genes expression in liver, white muscle and/or bone were analyzed. Moreover,
in vitro
cultures of vertebra bone-derived cells from fish fed the diets at a standard ration were performed at weeks 3 and 9 to explore the effects of hydroxytyrosol on osteoblasts development. Although neither body weight or any other biometric parameter were affected by diet composition after 4 or 8 weeks, the addition of the hydroxytyrosol-rich extract to the diet increased IGF-1 plasma levels, regardless of the ration regime, suggesting an anabolic condition. In muscle, the higher mRNA levels of the binding protein
igfbp-5b
and the myoblast fusion marker
dock5
in fish fed with the hydroxytyrosol-rich diet suggested that this compound may have a role in muscle, inducing development and a better muscular condition. Furthermore in bone, increased osteogenic potential while delayed matrix mineralization after addition to the diet of the olive juice extract was supported by the upregulated expression of
igf-1
and
bmp4
and reduced transcript levels of
osteopontin
. Overall, this study provides new insights into the beneficial use of hydroxytyrosol as a dietary additive in gilthead sea bream functional diets to improve muscle-skeletal condition and, the aquaculture industry.
This study aimed to assess the effects of dietary increasing concentrations of Aloe vera (AV) powder of 0.5%, 2.5% and 5% on the growth performance, hepatic oxidative status, histology, and lipid ...metabolism and cellular signaling pathways-related genes' expression in gilthead sea bream (Sparus aurata). The preliminary phytochemical analysis revealed the richness of the dried AV extract on total phenol content, total flavonoid content, and condensed tannins when compared to the lyophilized sample. The dried extract showed a good DPPH-radical scavenging activity and its profiling by HPLC-DAD-ESI-MS revealed the presence of anthraquinones namely aloin A, aloin B and their hydroxyl (7-hydroxyaloin A and 7-hydroxyaloin B) and methyl-hydroxy (8-O-methyl-7-hydroxyaloin A and 8-O-methyl-7-hydroxyaloin B) derivatives as well as aloeresin A and B. The AV supplementation in fish diet did not affect growth performance (WG, WGR, and SGR) and feed utilization (FI, FCR, FER), and HSI indexes. However, the hepatic insulin-like growth factors (IGF-I and II) levels were significantly enhanced. Genes' expression levels of enzymes or transcription factors involved in lipolysis (lpl, hsl, and atgl), beta-oxidation (pparα, hadh), fatty acid transporters (cd36, fabp11) and lxrα were significantly down-regulated by the two high concentrations of AV powder. In contrast, fatty acid synthase (fas), a key gene of lipogenesis was significantly up regulated by dietary AV 5% powder supplementation. The induction of fas together with the down-regulation of peroxisome proliferator-activated receptor (pparα) and hydroxyacyl-coenzyme A dehydrogenase (hadh) could explain the lipid accumulation resulting in hepatic steatosis, which was confirmed by histological analysis, since the diets at the two higher concentrations (AV 2.5% and AV 5%) induced a significant increase in the number and diameter of hepatic lipid vacuoles in a dose dependent manner. Moreover, the mRNA levels of protein kinase B named (akt), mammalian target of rapamycin (mtor) and extracellular regulated kinase (erk1/2) involved in cell survival and proliferation were decreased by all AV powder supplemented diets. AV 5% increased catalase and glutathione S transferase activities suggesting a cellular strategy to fight against reactive oxygen species (ROS) accumulation. In conclusion, dietary supplementation with AV 0.5% is recommended for gilthead sea bream feed formulation, as it stimulates the igf-i expression. However, higher levels of AV should be avoided as they might cause lipid metabolism disruption, oxidative stress and liver steatosis.
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•The effects of dietary Aloe vera (AV) powder were studied in Gilthead sea bream.•HPLC-DAD-ESI-MS analysis revealed the presence of anthraquinones in (AV) powder•Dietary AV powder reduced genes' expression involved in lipolysis and beta-oxidation•Fatty acid synthase was significantly up regulated by dietary AV powder.•Dietary AV induced oxidative stress and reduced cell survival pathways-related genes expression
This study aimed to evaluate the effect of dietary protein source (fishmeal, FM; or plant-feedstuffs, PF) and dietary protein/carbohydrate (P/CH) ratio on gilthead seabream appetite regulation and ...intermediary metabolism. Additionally, the effect of sampling 5 h after feeding (AF) compared to 24 h AF was also evaluated. Four isolipidic diets were formulated having as major protein sources FM or PF (20% FM and 80% PF), and P/CH ratios of 50/10 or 40/20, being the pregelatinized maize starch the main carbohydrate source (diets FM-P50/CH10; FM-P40/CH20; PF-P50/CH10; PF-P40/CH20). Diets were fed until satiation to 140 g gilthead seabream for 41 days. The expression of appetite regulation genes was assessed at 5 and 24 h AF, while other evaluated parameters were assessed only at 5 h AF. Liver leptin expression was higher at 5 h AF, and brain leptin receptor (lepr) expression was higher at 24 h AF. Brain expression of cocaine- and amphetamine-regulated transcript (cart), leptin and ghrelin receptor (ghrr)-a and liver ghrr-b were also affected by sampling time, but the effects were dependent of the diet provided. FM-based diets promoted the expression of brain cart and leptin (at 24 h AF), and liver growth hormone receptor (ghr)-ii, and increased plasma cholesterol and total lipids levels. Fish fed the PF-based diets had higher liver glycogen content, number and size of adipocytes, and expression of hepatic leptin (at 24 h AF), fatty acid synthase, glucokinase, and target of rapamycin. Regarding dietary P/CH ratio, fish fed the P50/CH10 diets presented higher feed efficiency, plasma triglycerides, and expression of intestine cholecystokinin (at 5 h AF), liver ghrr-b (at 24 h AF), glutamate dehydrogenase and ghr-ii. The protein efficiency ratio, hepatosomatic and visceral indices, plasmatic glucose level, and brain lepr expression (at 5 h AF) were higher in fish fed the P40/CH20 diets. The majority of appetite regulation related-genes were not affected by the use of PF-based diets, while the higher dietary CH seemed to lead to a shorter satiety sensation. PF-based diets promoted liver lipid deposition, hypocholesterolemia, and the activation of glycogenesis pathway, while higher CH content induced an increase in plasma glucose that appeared to be stored as lipids. In conclusion, PF-based diets with up to 20% of CH can be used in gilthead seabream without compromising growth performance and FI, and only slightly modifying appetite and metabolic parameters.
•ghrr-a, leptin, and lepr expression were affected by sampling time.•Response to dietary protein sources took more time to be induced.•High CH-diets promoted a shorter satiety sensation.•PF diets with 20% CH did not affect FI but slightly affected appetite genes expression.
Chaperone-mediated autophagy (CMA) is a major pathway of lysosomal proteolysis critical for cellular homeostasis and metabolism, and whose defects have been associated with several human pathologies. ...While CMA has been well described in mammals, functional evidence has only recently been documented in fish, opening up new perspectives to tackle this function under a novel angle. Now we propose to explore CMA functions in the rainbow trout (RT, Oncorhynchus mykiss), a fish species recognized as a model organism of glucose intolerance and characterized by the presence of two paralogs of the CMA-limiting factor Lamp2A (lysosomal associated membrane protein 2A). To this end, we validated a fluorescent reporter (KFERQ-PA-mCherry1) previously used to track functional CMA in mammalian cells, in an RT hepatoma-derived cell line (RTH-149). We found that incubation of cells with high-glucose levels (HG, 25 mM) induced translocation of the CMA reporter to lysosomes and/or late endosomes in a KFERQ- and Lamp2A-dependent manner, as well as reduced its half-life compared to the control (5 mM), thus demonstrating increased CMA flux. Furthermore, we observed that activation of CMA upon HG exposure was mediated by generation of mitochondrial reactive oxygen species, and involving the antioxidant transcription factor Nfe2l2/Nrf2 (nfe2 like bZIP transcription factor 2). Finally, we demonstrated that CMA plays an important protective role against HG-induced stress, primarily mediated by one of the two RT Lamp2As. Together, our results provide unequivocal evidence for CMA activity existence in RT and highlight both the role and regulation of CMA during glucose-related metabolic disorders.
Abbreviations: AREs: antioxidant response elements; CHC: α-cyano -4-hydroxycinnamic acid; Chr: chromosome; CMA: chaperone-mediated autophagy; CT: control; DMF: dimethyl fumarate; Emi: endosomal microautophagy; HG: high-glucose; HMOX1: heme oxygenase 1; H
2
O
2
: hydrogen peroxide; KFERQ: lysine-phenylalanine-glutamate-arginine-glutamine; LAMP1: lysosomal associated membrane protein 1; LAMP2A: lysosomal associated membrane protein 2A; MCC: Manders' correlation coefficient; Manders' correlation coefficient Mo: morpholino oligonucleotide; NAC: N-acetyl cysteine; NFE2L2/NRF2: NFE2 like bZIP transcription factor 2; PA-mCherry: photoactivable mCherry; PCC: Pearson's correlation coefficient; ROS: reactive oxygen species; RT: rainbow trout; siRNAs: small interfering RNAs; SOD: superoxide dismutase; Tsg101: tumor susceptibility 101; TTFA: 2-thenoyltrifluoroacetone; WGD: whole-genome duplication.
World population is expected to increase to approximately 9 thousand million people by 2050 with a consequent food security decline. Besides, climate change is a major challenge that humanity is ...facing, with a predicted rise in mean sea surface temperature of more than 2°C during this century. This study aims to determine whether a rearing temperature of 19, 24, or 28°C may influence musculoskeletal development and muscle lipid metabolism in gilthead sea bream juveniles. The expression of growth hormone (GH)/insulin-like growth factors (IGFs) system-, osteogenic-, myogenic-, and lipid metabolism-related genes in bone and/or white muscle of treated fish, and the
viability, mineralization, and osteogenic genes expression in primary cultured cells derived from bone of the same fish were analyzed. The highest temperature significantly down-regulated
, the receptor
, and the binding proteins
and
in bone, and in muscle,
and
, suggesting impaired musculoskeletal development. Concerning myogenic factors expression, contrary responses were observed, since the increase to 24°C significantly down-regulated
and
, while at 28°C
and
were significantly up-regulated. Moreover, in the muscle tissue, the expression of the fatty acid transporters
and
, and the lipases
and
resulted significantly increased at elevated temperatures, whereas β-oxidation markers
and
were significantly reduced. Regarding the primary cultured bone-derived cells, a significant up-regulation of the extracellular matrix proteins
, and
expression was found with increased temperatures, together with a gradual decrease in mineralization along with fish rearing temperature. Overall, these results suggest that increasing water temperature in this species appears to induce unfavorable growth and development of bone and muscle, through modulating the expression of different members of the GH/IGFs axis, myogenic and osteogenic genes, while accelerating the utilization of lipids as an energy source, although less efficiently than at optimal temperatures.
High-fat diets (HFDs) enhance fish growth by optimizing nutrient utilization (i.e., protein-sparing effect); however, their potential negative effects have also encouraged the search for feed ...additives. This work has investigated the effects of an extract rich in a polyphenolic antioxidant, hydroxytyrosol (HT), supplemented (0.52 g HT/kg feed) in a HFD (24% lipid) in gilthead sea bream (
). Fish received the diet at two ration levels, standard (3% of total fish weight) or restricted (40% reduction) for 8 weeks. Animals fed the supplemented diet at a standard ration had the lowest levels of plasma free fatty acids (4.28 ± 0.23 mg/dL versus 6.42 ± 0.47 in the non-supplemented group) and downregulated hepatic mRNA levels of lipid metabolism markers (
,
,
,
,
,
,
and
), supporting potential fat-lowering properties of this compound in the liver. Moreover, the same animals showed increased muscle lipid content and peroxidation (1.58- and 1.22-fold, respectively, compared to the fish without HT), suggesting the modulation of body adiposity distribution and an enhanced lipid oxidation rate in that tissue. Our findings emphasize the importance of considering this phytocompound as an optimal additive in HFDs for gilthead sea bream to improve overall fish health and condition.
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