Publications by authors named "Encarnación Capilla"

53 Publications

The probiotic Lactobacillus rhamnosus mimics the dark-driven regulation of appetite markers and melatonin receptors' expression in zebrafish (Danio rerio) larvae: Understanding the role of the gut microbiome.

Comp Biochem Physiol B Biochem Mol Biol 2021 Oct-Dec;256:110634. Epub 2021 Jun 11.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Barcelona 08028, Spain. Electronic address:

The use of probiotics has been recently considered a novel therapeutic strategy to prevent pathologies such as obesity; however, the specific mechanisms of action by which probiotics exert their beneficial effects on metabolic health remain unclear. The aim of the present study was to investigate the short-term effects of a probiotic Lactobacillus rhamnosus supplementation (PROB) on appetite regulation, growth-related markers, and microbiota diversity in zebrafish (Danio rerio) larvae, compared to a group subjected to a constant darkness photoperiod (DARK), as well as to evaluate the effects of both treatments on melatonin receptors' expression. After a 24 h treatment, both PROB and DARK conditions caused a significant increase in leptin a expression. Moreover, mRNA abundances of leptin b and proopiomelanocortin a were elevated in the PROB group, and DARK showed a similar tendency, supporting a negative regulation of appetite markers by the treatments. Moreover, both PROB and DARK also enhanced the abundances of melatonin receptors transcript (melatonin receptor 1 ba and bb) and protein (melatonin receptor 1) suggesting a potential involvement of melatonin in mediating these effects. Nevertheless, treatments did not exhibit a significant effect on the expression of most of the growth hormone/insulin-like growth factor axis genes evaluated. Finally, only the DARK condition significantly modulated gut microbiota diversity at such short time, altogether highlighting the rapid effects of this probiotic on modulating appetite regulatory and melatonin receptors' expression, without a concomitant variation of gut microbiota.
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http://dx.doi.org/10.1016/j.cbpb.2021.110634DOI Listing
September 2021

Musculoskeletal Growth Modulation in Gilthead Sea Bream Juveniles Reared at High Water Temperature and Fed with Palm and Rapeseed Oils-Based Diets.

Animals (Basel) 2021 Jan 21;11(2). Epub 2021 Jan 21.

Departament de Biologia Cellular, Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, 08028 Barcelona, Spain.

The upward trend of seawater temperature has encouraged improving the knowledge of its consequences on fish, considering also the development of diets including vegetable ingredients as an approach to achieve a more sustainable aquaculture. This study aims to determine the effects on musculoskeletal growth of: (1) a high-water temperature of 28 °C (versus 21 °C) in gilthead sea bream juveniles () fed with a diet rich in palm oil and, (2) feeding the fish reared at 28 °C with two other diets containing rapeseed oil or an equilibrated combination of both vegetable oils. Somatic parameters and mRNA levels of growth hormone-insulin-like growth factors (GH-IGFs) axis-, osteogenic-, myogenic-, lipid metabolism- and oxidative stress-related genes in vertebra bone and/or white muscle are analyzed. Overall, the data indicate that high-water rearing temperature in this species leads to different adjustments through modulating the gene expression of members of the GH-IGFs axis (down-regulating , its receptors, and binding proteins) and also, to bone turnover (reducing the resorption-activity genes cathepsin K () and matrix metalloproteinase-9 ()) to achieve harmonic musculoskeletal growth. Moreover, the combination of palm and rapeseed oils seems to be the most beneficial at high-water rearing temperature for both balanced somatic growth and muscular fatty acid uptake and oxidation.
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http://dx.doi.org/10.3390/ani11020260DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7909841PMC
January 2021

Genistein Induces Adipogenic and Autophagic Effects in Rainbow Trout () Adipose Tissue: In Vitro and In Vivo Models.

Int J Mol Sci 2020 Aug 16;21(16). Epub 2020 Aug 16.

Departament de Biologia Cellular, Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, 08028 Barcelona, Spain.

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.
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http://dx.doi.org/10.3390/ijms21165884DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7461592PMC
August 2020

Gilthead seabream (Sparus aurata) in vitro adipogenesis and its endocrine regulation by leptin, ghrelin, and insulin.

Comp Biochem Physiol A Mol Integr Physiol 2020 11 22;249:110772. Epub 2020 Jul 22.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Av. Diagonal 643, 08028 Barcelona, Spain.

Leptin, ghrelin, and insulin influence lipid metabolism and thus can directly affect adipose tissue characteristics, modulating the organoleptic quality of aquaculture fish. The present study explored gilthead seabream (Sparus aurata) cultured preadipocytes development, and the regulation of adipogenesis by those three hormones. Preadipocytes presented a fibroblast-like phenotype during the proliferation phase that changed to round-shaped with an enlarged cytoplasm filled with lipid droplets after complete differentiation, confirming the characteristics of mature adipocytes. peroxisome proliferator-activated receptor-γ (pparγ) expression was higher at the beginning of the culture, while fatty acid synthase and 3-hydroxyacyl-CoA dehydrogenase gradually increased with cell maturation. The expression of lipoprotein lipase-like, lysosomal acid lipase (lipa), fatty acid translocase/cluster of differentiation-36 (cd36), and leptin receptor (lepr) were not affected during cell culture development; and undetectable expression levels were observed for leptin. Concerning regulation, leptin inhibited lipid accumulation significantly reducing pparγ and cd36 gene expression, both in early differentiating and mature adipocytes, while ghrelin decreased the expression of pparγ in the early differentiating phase but did not reduce intracellular lipid content significantly. Additional insulin past the onset of adipogenesis did not affect lipid accumulation either. In conclusion, at present culture conditions leptin has an anti-adipogenic function in differentiating preadipocytes of gilthead seabream and continues exerting this role in mature adipocytes, while ghrelin and insulin do not seem to influence adipogenesis progression. A better understanding of leptin, ghrelin, and insulin impact on the adipogenic process could help in the prevention of fat accumulation, improving aquaculture fish production and quality.
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http://dx.doi.org/10.1016/j.cbpa.2020.110772DOI Listing
November 2020

Short-Term Responses to Fatty Acids on Lipid Metabolism and Adipogenesis in Rainbow Trout ().

Int J Mol Sci 2020 Feb 27;21(5). Epub 2020 Feb 27.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, 08028 Barcelona, Spain.

Fish are rich in n-3 long-chain polyunsaturated fatty acids (LC-PUFA) such as eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids. Due to the increasing use of vegetable oils (VO), their proportion in diets has lowered, affecting lipid metabolism and fillet composition. Rainbow trout cultured preadipocytes were treated with representative FA found in fish oils (EPA and DHA) or VO (linoleic, LA and alpha-linolenic, ALA acids), while EPA and LA were also orally administered, to evaluate their effects on adipogenesis and lipid metabolism. In vitro, all FA increased lipid internalization, with ALA producing the highest effect, together with upregulating the FA transporter . In vivo, EPA or LA increased peroxisome proliferator-activated receptors and transcripts abundance in adipose tissue, suggesting elevated β-oxidation, contrary to the results obtained in liver. Furthermore, the increased expression of FA synthase () and the FA translocase/cluster of differentiation ( in adipose tissue indicated an enhanced uptake of lipids and lipogenesis de novo, whereas stable or low hepatic expression of genes involved in lipid transport and turnover was found. Thus, fish showed a similar tissue metabolic response to the short-term availability of EPA or LA in vivo, while in vitro VO-derived FA demonstrated greater potential inducing fat accumulation.
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http://dx.doi.org/10.3390/ijms21051623DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7084833PMC
February 2020

Photoperiod Manipulation Affects Transcriptional Profile of Genes Related to Lipid Metabolism and Apoptosis in Zebrafish (Danio rerio) Larvae: Potential Roles of Gut Microbiota.

Microb Ecol 2020 May 9;79(4):933-946. Epub 2019 Dec 9.

Dipartimento di Scienze della Vita e dell'Ambiente, Università Politecnica delle Marche, Ancona, Italy.

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.
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http://dx.doi.org/10.1007/s00248-019-01468-7DOI Listing
May 2020

Editorial: Nutritional and Environmental Modulation of the Endocrine System: Effects on Metabolism and Growth.

Front Endocrinol (Lausanne) 2019 4;10:354. Epub 2019 Jun 4.

Department of Cell Biology, Physiology and Immunology, University of Barcelona, Barcelona, Spain.

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http://dx.doi.org/10.3389/fendo.2019.00354DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6558001PMC
June 2019

Gene expression analyses in malformed skeletal structures of gilthead sea bream (Sparus aurata).

J Fish Dis 2019 Aug 10;42(8):1169-1180. Epub 2019 Jun 10.

Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, Barcelona, Spain.

The incidence of skeletal anomalies in reared fish has been translated for years in important economic losses for the aquaculture industry. In the present study, we have analysed the gene expression of extracellular matrix components and transcription factors involved in bone development in gilthead sea bream presenting different skeletal anomalies: lordosis (LD), lordosis-scoliosis-kyphosis (LSK) or opercular, dental or jaw malformations in comparison with control (CT) specimens. Results showed a possible link between the presence of LD and LSK and the significant downregulation of genes involved in osteoblasts' maturation and matrix mineralization (collagen type 1-alpha, osteopontin, osteocalcin, matrix Gla protein and tissue non-specific alkaline phosphatase), as well as in bone resorption (cathepsin K and matrix metalloproteinase 9) compared to CT animals. Contrarily, the key osteogenic transcription factor runx2 was upregulated in the malformed vertebra suggesting impaired determination of mesenchymal stem cells towards the osteoblastic lineage. Despite the gene expression patterns of the other malformed structures were not affected in comparison with CT fish, the results of the present study may contribute in the long term to identify potential candidate gene profiles associated with column deformities that may help reducing the incidence of appearance of skeletal anomalies in this important aquaculture species.
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http://dx.doi.org/10.1111/jfd.13019DOI Listing
August 2019

Fatty acids from fish or vegetable oils promote the adipogenic fate of mesenchymal stem cells derived from gilthead sea bream bone potentially through different pathways.

PLoS One 2019 24;14(4):e0215926. Epub 2019 Apr 24.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Barcelona, Spain.

Fish are rich in n-3 long-chain polyunsaturated fatty acids (LC-PUFA), such as eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids, thus they have a great nutritional value for human health. In this study, the adipogenic potential of fatty acids commonly found in fish oil (EPA and DHA) and vegetable oils (linoleic (LA) and alpha-linolenic (ALA) acids), was evaluated in bone-derived mesenchymal stem cells (MSCs) from gilthead sea bream. At a morphological level, cells adopted a round shape upon all treatments, losing their fibroblastic form and increasing lipid accumulation, especially in the presence of the n-6 PUFA, LA. The mRNA levels of the key transcription factor of osteogenesis, runx2 significantly diminished and those of relevant osteogenic genes remained stable after incubation with all fatty acids, suggesting that the osteogenic process might be compromised. On the other hand, transcript levels of the main adipogenesis-inducer factor, pparg increased in response to EPA. Nevertheless, the specific PPARγ antagonist T0070907 appeared to suppress the effects being caused by EPA over adipogenesis. Moreover, LA, ALA and their combinations, significantly up-regulated the fatty acid transporter and binding protein, fatp1 and fabp11, supporting the elevated lipid content found in the cells treated with those fatty acids. Overall, this study has demonstrated that fatty acids favor lipid storage in gilthead sea bream bone-derived MSCs inducing their fate into the adipogenic versus the osteogenic lineage. This process seems to be promoted via different pathways depending on the fatty acid source, being vegetable oils-derived fatty acids more prone to induce unhealthier metabolic phenotypes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0215926PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6481918PMC
January 2020

Temperature Affects Musculoskeletal Development and Muscle Lipid Metabolism of Gilthead Sea Bream ().

Front Endocrinol (Lausanne) 2019 22;10:173. Epub 2019 Mar 22.

Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, Barcelona, Spain.

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.
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http://dx.doi.org/10.3389/fendo.2019.00173DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6439310PMC
March 2019

A long-term growth hormone treatment stimulates growth and lipolysis in gilthead sea bream juveniles.

Comp Biochem Physiol A Mol Integr Physiol 2019 06 15;232:67-78. Epub 2019 Mar 15.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, 08028 Barcelona, Spain. Electronic address:

The enhancement of the endocrine growth hormone (GH)/insulin-like growth factor I (IGF-I) system by the treatment with a sustained release formulation of a recombinant bovine GH (rBGH), is a good strategy to investigate growth optimization in aquaculture fish species. To further deepen into the knowledge of rBGH effects in fish and to estimate the growth potential of juveniles of gilthead sea bream, the present work evaluated rBGH injection on growth, GH/IGF-I axis and lipid metabolism modulation, and explored the conservation of GH effects provoked by the in vivo treatment using in vitro models of different tissues. The rBGH treatment increased body weight and specific growth rate (SGR) in juveniles and potentiated hyperplastic muscle growth while reducing circulating triglyceride levels. Moreover, the results demonstrated that the in vivo treatment enhanced also lipolysis in both isolated hepatocytes and adipocytes, as well as in day 4 cultured myocytes. Furthermore, these cultured myocytes extracted from rBGH-injected fish presented higher gene expression of GH/IGF-I axis-related molecules and myogenic regulatory factors, as well as stimulated myogenesis (i.e. increased protein expression of a proliferation and a differentiation marker) compared to Control fish-derived cells. These data, suggested that cells in vitro can retain some of the pathways activated by in vivo treatments in fish, what can be considered an interesting line of applied research. Overall, the results showed that rBGH stimulates somatic growth, including specifically muscle hyperplasia, as well as lipolytic activity in gilthead sea bream juveniles.
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http://dx.doi.org/10.1016/j.cbpa.2019.03.012DOI Listing
June 2019

Effects of β-adrenoceptor agonists on gilthead sea bream (Sparus aurata) cultured muscle cells.

Comp Biochem Physiol A Mol Integr Physiol 2019 01 19;227:179-193. Epub 2018 Oct 19.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Barcelona 08028, Spain. Electronic address:

β-adrenoceptors are a subtype of G-protein coupled receptors whose activation leads to increased protein synthesis and decreased degradation in mammalian skeletal muscle, causing hypertrophy. In this study, we compared the effects of the classical β-agonist noradrenaline (NA) with two representatives of a new generation of agonists (formoterol, FOR and salmeterol, SALM) on growth and metabolism of primary cultured muscle cells of gilthead sea bream. Activation of signaling pathways, cell development and expression of relevant genes were analyzed in day 4 myocytes. The three agonists increased either cAMP levels or PKA phosphorylation, plus TOR phosphorylation, and the proportion of proliferating cell nuclear antigen (PCNA)-positive cells, in parallel with pcna mRNA levels. Thus, demonstrating that these cells are β-agonists-responsive, and supporting enhanced cell proliferation. The expression of the myogenic factor myf5 was significantly down-regulated, suggesting that the cells were already destined to the muscular linage; while insulin-like growth factors (igf-1 and igf-2) transcript levels were up-regulated, proposing an additional anabolic effect through their local production. Furthermore, SALM treatment up-regulated expression of the lipases (hsl and lipa) and the β-oxidation marker cpt1a, and all three agonists increased mitochondrial dehydrogenase hadh mRNA levels. These data correspond with a situation of enhanced lipolytic and β-oxidation capacity, a fact supported by the higher glycerol released into the media induced by the agonists. Overall, these results suggest a hyperplastic growth condition and a favorable protein/fat ratio profile upon these treatments; consequently, β-agonists (especially SALM) may be considered good candidates to optimize the growth in this aquaculture species.
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http://dx.doi.org/10.1016/j.cbpa.2018.10.015DOI Listing
January 2019

Ghrelin and Its Receptors in Gilthead Sea Bream: Nutritional Regulation.

Front Endocrinol (Lausanne) 2018 30;9:399. Epub 2018 Jul 30.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Barcelona, Spain.

Ghrelin is involved in the regulation of growth in vertebrates through controlling different functions, such as feed intake, metabolism, intestinal activity or growth hormone (Gh) secretion. The aim of this work was to identify the sequences of and Ghrelin receptors (), and to study their responses to different nutritional conditions in gilthead sea bream () juveniles. The structure and phylogeny of was analyzed, and a tissue screening was performed. The effects of 21 days of fasting and 2, 5, 24 h, and 7 days of refeeding on plasma levels of Ghrelin, Gh and Igf-1, and the gene expression of and members of the Gh/Igf-1 system were determined in key tissues. and the receptors are well conserved, being expressed mainly in stomach, and in the pituitary and brain, respectively. Twenty-one days of fasting resulted in a decrease in growth while Ghrelin plasma levels were elevated to decrease at 5 h post-prandial when pituitary expression was minimum. Gh in plasma increased during fasting and slowly felt upon refeeding, while plasma Igf-1 showed an inverse profile. Pituitary expression augmented during fasting reaching maximum levels at 1 day post-feeding while liver expression and that of its splice variants decreased to lowest levels. Liver Gh receptors expression was down-regulated during fasting and recovered after refeeding. This study demonstrates the important role of Ghrelin during fasting, its acute down-regulation in the post-prandial stage and its interaction with pituitary Ghsrs and Gh/Igf-1 axis.
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http://dx.doi.org/10.3389/fendo.2018.00399DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6077198PMC
July 2018

Temperature responsiveness of gilthead sea bream bone; an in vitro and in vivo approach.

Sci Rep 2018 07 25;8(1):11211. Epub 2018 Jul 25.

Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, 08028, Barcelona, Spain.

This study aimed to characterize the molecules involved in osteogenesis in seabream and establish using in vitro/in vivo approaches the responsiveness of selected key genes to temperature. The impact of a temperature drop from 23 to 13 °C was evaluated in juvenile fish thermally imprinted during embryogenesis. Both, in vitro/in vivo, Fib1a, appeared important in the first stages of bone formation, and Col1A1, ON and OP, in regulating matrix production and mineralization. OCN mRNA levels were up-regulated in the final larval stages when mineralization was more intense. Moreover, temperature-dependent differential gene expression was observed, with lower transcript levels in the larvae at 18 °C relative to those at 22 °C, suggesting bone formation was enhanced in the latter group. Results revealed that thermal imprinting affected the long-term regulation of osteogenesis. Specifically, juveniles under the low and low-to-high-temperature regimes had reduced levels of OCN when challenged, indicative of impaired bone development. In contrast, gene expression in fish from the high and high-to-low-temperature treatments was unchanged, suggesting imprinting may have a protective effect. Overall, the present study revealed that thermal imprinting modulates bone development in seabream larvae, and demonstrated the utility of the in vitro MSC culture as a reliable tool to investigate fish osteogenesis.
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http://dx.doi.org/10.1038/s41598-018-29570-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6060158PMC
July 2018

Proteolytic systems' expression during myogenesis and transcriptional regulation by amino acids in gilthead sea bream cultured muscle cells.

PLoS One 2017 20;12(12):e0187339. Epub 2017 Dec 20.

Departament de Biologia Cellular, Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, Barcelona, Spain.

Proteolytic systems exert an important role in vertebrate muscle controlling protein turnover, recycling of amino acids (AA) or its use for energy production, as well as other functions like myogenesis. In fish, proteolytic systems are crucial for the relatively high muscle somatic index they possess, and because protein is the most important dietary component. Thus in this study, the molecular profile of proteolytic markers (calpains, cathepsins and ubiquitin-proteasome system (UbP) members) were analyzed during gilthead sea bream (Sparus aurata) myogenesis in vitro and under different AA treatments. The gene expression of calpains (capn1, capn3 and capns1b) decreased progressively during myogenesis together with the proteasome member n3; whereas capn2, capns1a, capns1b and ubiquitin (ub) remained stable. Contrarily, the cathepsin D (ctsd) paralogs and E3 ubiquitin ligases mafbx and murf1, showed a significant peak in gene expression at day 8 of culture that slightly decreased afterwards. Moreover, the protein expression analyzed for selected molecules presented in general the same profile of the mRNA levels, which was confirmed by correlation analysis. These data suggest that calpains seem to be more important during proliferation, while cathepsins and the UbP system appear to be required for myogenic differentiation. Concerning the transcriptional regulation by AA, the recovery of their levels after a short starvation period did not show effects on cathepsins expression, whereas it down-regulated the expression of capn3, capns1b, mafbx, murf1 and up-regulated n3. With regards to AA deficiencies, the major changes occurred at day 2, when leucine limitation suppressed ctsb and ctsl expression. Besides at the same time, both leucine and lysine deficiencies increased the expression of mafbx and murf1 and decreased that of n3. Overall, the opposite nutritional regulation observed, especially for the UbP members, points out an efficient and complementary role of these factors that could be useful in gilthead sea bream diets optimization.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0187339PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5737955PMC
January 2018

Recombinant bovine growth hormone (rBGH) enhances somatic growth by regulating the GH-IGF axis in fingerlings of gilthead sea bream (Sparus aurata).

Gen Comp Endocrinol 2018 02 27;257:192-202. Epub 2017 Jun 27.

Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, 08028 Barcelona, Spain. Electronic address:

The growth hormone (GH)/insulin-like growth factors (IGFs) endocrine axis is the main growth-regulator system in vertebrates. Some authors have demonstrated the positive effects on growth of a sustained-release formulation of a recombinant bovine GH (rBGH) in different fish species. The aim of this work was to characterize the effects of a single injection of rBGH in fingerlings of gilthead sea bream on growth, GH-IGF axis, and both myogenic and osteogenic processes. Thus, body weight and specific growth rate were significantly increased in rBGH-treated fish respect to control fish at 6weeks post-injection, whereas the hepatosomatic index was decreased and the condition factor and mesenteric fat index were unchanged, altogether indicating enhanced somatic growth. Moreover, rBGH injection increased the plasma IGF-I levels in parallel with a rise of hepatic mRNA from total IGF-I, IGF-Ic and IGF-II, the binding proteins IGFBP-1a and IGFBP-2b, and also the receptors IGF-IRb, GHR-I and GHR-II. In skeletal muscle, the expression of IGF-Ib and GHR-I was significantly increased but that of IGF-IRb was reduced; the mRNA levels of myogenic regulatory factors, proliferation and differentiation markers (PCNA and MHC, respectively), or that of different molecules of the signaling pathway (TOR/AKT) were unaltered. Besides, the growth inhibitor myostatin (MSTN1 and MSTN2) and the hypertrophic marker (MLC2B) expression resulted significantly enhanced, suggesting altogether that the muscle is in a non-proliferative stage of development. Contrarily in bone, although the expression of most molecules of the GH/IGF axis was decreased, the mRNA levels of several osteogenic genes were increased. The histology analysis showed a GH induced lipolytic effect with a clear decrease in the subcutaneous fat layer. Overall, these results reveal that a better growth potential can be achieved on this species and supports the possibility to improve growth and quality through the optimization of its culture conditions.
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http://dx.doi.org/10.1016/j.ygcen.2017.06.019DOI Listing
February 2018

Caffeic acid and hydroxytyrosol have anti-obesogenic properties in zebrafish and rainbow trout models.

PLoS One 2017 1;12(6):e0178833. Epub 2017 Jun 1.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Barcelona, Spain.

Some natural products, known sources of bioactive compounds with a wide range of properties, may have therapeutic values in human health and diseases, as well as agronomic applications. The effect of three compounds of plant origin with well-known dietary antioxidant properties, astaxanthin (ATX), caffeic acid (CA) and hydroxytyrosol (HT), on zebrafish (Danio rerio) larval adiposity and rainbow trout (Onchorynchus mykiss) adipocytes was assessed. The zebrafish obesogenic test (ZOT) demonstrated the anti-obesogenic activity of CA and HT. These compounds were able to counteract the obesogenic effect produced by the peroxisome proliferator-activated receptor gamma (PPARγ) agonist, rosiglitazone (RGZ). CA and HT suppressed RGZ-increased PPARγ protein expression and lipid accumulation in primary-cultured rainbow trout adipocytes. HT also significantly reduced plasma triacylglycerol concentrations, as well as mRNA levels of the fasn adipogenic gene in the adipose tissue of HT-injected rainbow trout. In conclusion, in vitro and in vivo approaches demonstrated the anti-obesogenic potential of CA and HT on teleost fish models that may be relevant for studying their molecular mode of action. Further studies are required to evaluate the effect of these bioactive components as food supplements for modulating adiposity in farmed fish.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0178833PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5453583PMC
September 2017

Tributyltin and triphenyltin exposure promotes in vitro adipogenic differentiation but alters the adipocyte phenotype in rainbow trout.

Aquat Toxicol 2017 Jul 8;188:148-158. Epub 2017 May 8.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Barcelona 08028, Spain. Electronic address:

Numerous environmental pollutants have been identified as potential obesogenic compounds affecting endocrine signaling and lipid homeostasis. Among them, well-known organotins such as tributyltin (TBT) and triphenyltin (TPT), can be found in significant concentrations in aquatic environments. The aim of the present study was to investigate in vitro the effects of TBT and TPT on the development and lipid metabolism of rainbow trout (Onchorynchus mykiss) primary cultured adipocytes. Results showed that TBT and TPT induced lipid accumulation and slightly enhanced peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT enhancer binding protein alpha (C/EBPα) protein expression when compared to a control, both in the presence or absence of lipid mixture. However, the effects were higher when combined with lipid, and in the absence of it, the organotins did not cause complete mature adipocyte morphology. Regarding gene expression analyses, exposure to TBT and TPT caused an increase in fatty acid synthase (fasn) mRNA levels confirming the pro-adipogenic properties of these compounds. In addition, when added together with lipid, TBT and TPT significantly increased cebpa, tumor necrosis factor alpha (tnfa) and ATP-binding cassette transporter 1 (abca1) mRNA levels suggesting a synergistic effect. Overall, our data highlighted that TBT and TPT activate adipocyte differentiation in rainbow trout supporting an obesogenic role for these compounds, although by themselves they are not able to induce complete adipocyte development and maturation suggesting that these adipocytes might not be properly functional.
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http://dx.doi.org/10.1016/j.aquatox.2017.05.001DOI Listing
July 2017

Gene expression profile during proliferation and differentiation of rainbow trout adipocyte precursor cells.

BMC Genomics 2017 05 4;18(1):347. Epub 2017 May 4.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Av. Diagonal 643, 08028, Barcelona, Spain.

Background: Excessive accumulation of adipose tissue in cultured fish is an outstanding problem in aquaculture. To understand the development of adiposity, it is crucial to identify the genes which expression is associated with adipogenic differentiation. Therefore, the transcriptomic profile at different time points (days 3, 8, 15 and 21) along primary culture development of rainbow trout preadipocytes has been investigated using an Agilent trout oligo microarray.

Results: Our analysis identified 4026 genes differentially expressed (fold-change >3) that were divided into two major clusters corresponding to the main phases observed during the preadipocyte culture: proliferation and differentiation. Proliferation cluster comprised 1028 genes up-regulated from days 3 to 8 of culture meanwhile the differentiation cluster was characterized by 2140 induced genes from days 15 to 21. Proliferation was characterized by enrichment in genes involved in basic cellular and metabolic processes (transcription, ribosome biogenesis, translation and protein folding), cellular remodelling and autophagy. In addition, the implication of the eicosanoid signalling pathway was highlighted during this phase. On the other hand, the terminal differentiation phase was enriched with genes involved in energy production, lipid and carbohydrate metabolism. Moreover, during this phase an enrichment in genes involved in the formation of the lipid droplets was evidenced as well as the activation of the thyroid-receptor/retinoic X receptor (TR/RXR) and the peroxisome proliferator activated receptors (PPARs) signalling pathways. The whole adipogenic process was driven by a coordinated activation of transcription factors and epigenetic modulators.

Conclusions: Overall, our study demonstrates the coordinated expression of functionally related genes during proliferation and differentiation of rainbow trout adipocyte cells. Furthermore, the information generated will allow future investigations of specific genes involved in particular stages of fish adipogenesis.
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http://dx.doi.org/10.1186/s12864-017-3728-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5418865PMC
May 2017

Moderate and sustained exercise modulates muscle proteolytic and myogenic markers in gilthead sea bream ().

Am J Physiol Regul Integr Comp Physiol 2017 05 22;312(5):R643-R653. Epub 2017 Feb 22.

Departament de Biologia Cellular, Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, Barcelona, Spain

Swimming activity primarily accelerates growth in fish by increasing protein synthesis and energy efficiency. The role of muscle in this process is remarkable and especially important in teleosts, where muscle represents a high percentage of body weight and because many fish species present continuous growth. The aim of this work was to characterize the effects of 5 wk of moderate and sustained swimming in gene and protein expression of myogenic regulatory factors, proliferation markers, and proteolytic molecules in two muscle regions (anterior and caudal) of gilthead sea bream fingerlings. Western blot results showed an increase in the proliferation marker proliferating cell nuclear antigen (PCNA), proteolytic system members calpain 1 and cathepsin D, as well as vascular endothelial growth factor protein expression. Moreover, quantitative real-time PCR data showed that exercise increased the gene expression of proteases (calpains, cathepsins, and members of the ubiquitin-proteasome system in the anterior muscle region) and the gene expression of the proliferation marker PCNA and the myogenic factor MyoD in the caudal area compared with control fish. Overall, these data suggest a differential response of the two muscle regions during swimming adaptation, with tissue remodeling and new vessel formation occurring in the anterior muscle and enhanced cell proliferation and differentiation occurring in the caudal area. In summary, the present study contributes to improving the knowledge of the role of proteolytic molecules and other myogenic factors in the adaptation of muscle to moderate sustained swimming in gilthead sea bream.
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http://dx.doi.org/10.1152/ajpregu.00308.2016DOI Listing
May 2017

Adipogenic Gene Expression in Gilthead Sea Bream Mesenchymal Stem Cells from Different Origin.

Front Endocrinol (Lausanne) 2016 22;7:113. Epub 2016 Aug 22.

Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona , Barcelona , Spain.

During the last decades, adipogenesis has become an emerging field of study in aquaculture due to the relevance of the adipose tissue in many physiological processes and its connection with the endocrine system. In this sense, recent studies have translated into the establishment of preadipocyte culture models from several fish species, sometimes lacking information on the mRNA levels of adipogenic genes. Thus, the aim of this study was to determine the gene expression profile of gilthead sea bream (Sparus aurata) primary cultured mesenchymal stem cells (MSCs) from different origin (adipose tissue and vertebra bone) during adipogenesis. Both cell types differentiated into adipocyte-like cells, accumulating lipids inside their cytoplasm. Adipocyte differentiation of MSCs from adipose tissue resulted in downregulation of several adipocyte-related genes (such as lpl, hsl, pparα, pparγ and gapdh2) at day 4, gapdh1 at day 8, and fas and pparβ at day 12. In contrast, differences in lxrα mRNA expression were not observed, while g6pdh levels increased during adipocyte maturation. Gapdh and Pparγ protein levels were also detected in preadipocyte cultures; however, only the former increased its expression during adipogenesis. Moreover, differentiation of bone-derived cells into adipocytes also resulted in the downregulation of several adipocyte gene markers, such as fas and g6pdh at day 10 and hsl, pparβ, and lxrα at day 15. On the other hand, the osteogenic genes fib1a, mgp, and op remained stable, but an increase in runx2 expression at day 20 was observed. In summary, the present study demonstrates that gilthead sea bream MSCs, from both adipose tissue and bone, differentiate into adipocyte-like cells, although revealed some kind of species- and cell lineage-specific regulation with regards to gene expression. Present data also provide novel insights into some of the potential key genes controlling adipogenesis in gilthead sea bream that can help to better understand the regulation of lipid storage in fish.
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http://dx.doi.org/10.3389/fendo.2016.00113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4992700PMC
September 2016

Characterization data of gilthead sea bream (Sparus aurata) IGF-I receptors (IGF-IRa/Rb).

Data Brief 2016 Mar 6;6:507-13. Epub 2016 Jan 6.

Departament de Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, Avinguda Diagonal 643, 08028 Barcelona, Spain.

In this data article we describe the coding sequence of two IGF-IR paralogues (IGF-IRa and IGF-IRb) obtained from gilthead sea bream embryos. The putative protein architecture (domains and other important motifs) was determined and, amino acid sequences alignment and phylogenetic analysis of both receptors together with IGF-IR orthologues from different vertebrates was performed. Additionally, a semi-quantitative conventional PCR was done to analyze the mRNA expression of both receptors in different tissues of gilthead sea bream. These data will assist in further physiological studies in this species. In this sense, the expression of both receptors during ontogeny in muscle as well as the differential effects of IGF-I and IGF-II on their regulation during in vitro myogenesis has been recently studied (doi: 10.1016/j.ygcen.2015.11.011; [1]).
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http://dx.doi.org/10.1016/j.dib.2015.12.046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4724695PMC
March 2016

Lysine and Leucine Deficiencies Affect Myocytes Development and IGF Signaling in Gilthead Sea Bream (Sparus aurata).

PLoS One 2016 25;11(1):e0147618. Epub 2016 Jan 25.

Departament de Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, 08028, Barcelona, Spain.

Optimizing aquaculture production requires better knowledge of growth regulation and improvement in diet formulation. A great effort has been made to replace fish meal for plant protein sources in aquafeeds, making necessary the supplementation of such diets with crystalline amino acids (AA) to cover the nutritional requirements of each species. Lysine and Leucine are limiting essential AA in fish, and it has been demonstrated that supplementation with them improves growth in different species. However, the specific effects of AA deficiencies in myogenesis are completely unknown and have only been studied at the level of hepatic metabolism. It is well-known that the TOR pathway integrates the nutritional and hormonal signals to regulate protein synthesis and cell proliferation, to finally control muscle growth, a process also coordinated by the expression of myogenic regulatory factors (MRFs). This study aimed to provide new information on the impact of Lysine and Leucine deficiencies in gilthead sea bream cultured myocytes examining their development and the response of insulin-like growth factors (IGFs), MRFs, as well as key molecules involved in muscle growth regulation like TOR. Leucine deficiency did not cause significant differences in most of the molecules analyzed, whereas Lysine deficiency appeared crucial in IGFs regulation, decreasing significantly IGF-I, IGF-II and IGF-IRb mRNA levels. This treatment also down-regulated the gene expression of different MRFs, including Myf5, Myogenin and MyoD2. These changes were also corroborated by a significant decrease in proliferation and differentiation markers in the Lysine-deficient treatment. Moreover, both Lysine and Leucine limitation induced a significant down-regulation in FOXO3 gene expression, which deserves further investigation. We believe that these results will be relevant for the production of a species as appreciated for human consumption as it is gilthead sea bream and demonstrates the importance of an adequate level of Lysine in fishmeal diet formulation for optimum growth.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0147618PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4725776PMC
July 2016

Contribution of in vitro myocytes studies to understanding fish muscle physiology.

Comp Biochem Physiol B Biochem Mol Biol 2016 Sep 10;199:67-73. Epub 2015 Dec 10.

Departament de Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, 08028, Barcelona, Spain. Electronic address:

Research on the regulation of fish muscle physiology and growth was addressed originally by classical in vivo approaches; however, systemic interactions resulted in many questions that could be better considered through in vitro myocyte studies. The first paper published by our group in this field was with Tom Moon on brown trout cardiomyocytes, where the insulin and IGF-I receptors were characterized and the down-regulatory effects of an excess of peptides demonstrated. We followed the research on cultured skeletal muscle cells through the collaboration with INRA focused on the characterization of IGF-I receptors and its signaling pathways through in vitro development. Later on, we showed the important metabolic role of IGFs, although these studies were only the first stage of a prolific area of work that has offered a useful tool to advance in our knowledge of the endocrine and nutritional regulation of fish growth and metabolism. Obviously, the findings obtained in vitro serve the purpose to propose the scenario that will need confirmation in vivo, but this technique has made possible many different, easy, fast and better controlled studies. In this review, we have summarized the main advances that the use of cultured muscle cells has permitted, focusing mainly in the role of IGFs regulating fish metabolism and growth. Although many articles have already appeared using this model system in salmonids, gilthead sea bream or zebrafish, it is reasonable to expect new studies with cultured cells using innovative approaches that will help to understand fish physiology and its regulation.
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http://dx.doi.org/10.1016/j.cbpb.2015.12.003DOI Listing
September 2016

Effects of sustained exercise on GH-IGFs axis in gilthead sea bream (Sparus aurata).

Am J Physiol Regul Integr Comp Physiol 2016 Feb 9;310(4):R313-22. Epub 2015 Dec 9.

Departament de Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, Barcelona, Spain;

The endocrine system regulates growth mainly through the growth hormone (GH)/insulin-like growth factors (IGFs) axis and, although exercise promotes growth, little is known about its modulation of these factors. The aim of this work was to characterize the effects of 5 wk of moderate sustained swimming on the GH-IGFs axis in gilthead sea bream fingerlings. Plasma IGF-I/GH ratio and tissue gene expression of total IGF-I and three splice variants, IGF-II, three IGF binding proteins, two GH receptors, two IGF-I receptors, and the downstream molecules were analyzed. Fish under exercise (EX) grew more than control fish (CT), had a higher plasma IGF-I/GH ratio, and showed increased hepatic IGF-I expression (mainly IGF-Ia). Total IGF-I expression levels were similar in the anterior and caudal muscles; however, IGF-Ic expression increased with exercise, suggesting that this splice variant may be the most sensitive to mechanical action. Moreover, IGFBP-5b and IGF-II increased in the anterior and caudal muscles, respectively, supporting enhanced muscle growth. Furthermore, in EX fish, hepatic IGF-IRb was reduced together with both GHRs; GHR-II was also reduced in anterior muscle, while GHR-I showed higher expression in the two muscle regions, indicating tissue-dependent differences and responses to exercise. Exercise also increased gene and protein expression of target of rapamycin (TOR), suggesting enhanced muscle protein synthesis. Altogether, these data demonstrate that moderate sustained activity may be used to increase the plasma IGF-I/GH ratio and to potentiate growth in farmed gilthead sea bream, modulating the gene expression of different members of the GH-IGFs axis (i.e., IGF-Ic, IGF-II, IGFBP-5b, GHR-I, and TOR).
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http://dx.doi.org/10.1152/ajpregu.00230.2015DOI Listing
February 2016

IGF-I and IGF-II effects on local IGF system and signaling pathways in gilthead sea bream (Sparus aurata) cultured myocytes.

Gen Comp Endocrinol 2016 06 18;232:7-16. Epub 2015 Nov 18.

Departament de Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona, 08028 Barcelona, Spain. Electronic address:

The insulin-like growth factors (IGFs) have a fundamental role in a vast range of functions acting through a tyrosine-kinase receptor (IGF-IR). IGFs in muscle can affect the expression of components of the local IGF system, myogenic regulatory factors (MRFs), proliferating (proliferating cell nuclear antigen, PCNA) or differentiating molecules (myosin heavy chain, MHC) and, lead to the activation of different signaling pathways. The response of all these genes to IGFs incubation at two different times in day 4 cultured myocytes of gilthead sea bream was analyzed. Both IGFs increased the expression of IGF-I and IGFBP-5, but showed different effects on the receptors, with IGF-I suppressing the expression of both isoforms (IGF-IRa and IGF-IRb) and IGF-II up-regulating only IGF-IRb. Moreover, the protein levels of PCNA and target of rapamycin (TOR) increased after IGF-II incubation, although a decline in Myf5 and a rise in MHC gene expression was caused by IGF-I. Taken together, these results provide evidence for the importance of IGFs on controlling muscle development and growth in gilthead sea bream and suggest that each IGF may be preferentially acting through a specific IGF-IR. Moreover, the data support the hypothesis that IGF-II has a more important role during proliferation, whereas IGF-I seems to be relevant for the differentiation phase of myogenesis.
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http://dx.doi.org/10.1016/j.ygcen.2015.11.011DOI Listing
June 2016

Editorial: Control of Adipocyte Differentiation and Metabolism.

Front Endocrinol (Lausanne) 2015 28;6:132. Epub 2015 Aug 28.

Department of Physiology and Immunology, University of Barcelona , Barcelona , Spain.

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http://dx.doi.org/10.3389/fendo.2015.00132DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4551862PMC
September 2015

Roles of leptin and ghrelin in adipogenesis and lipid metabolism of rainbow trout adipocytes in vitro.

Comp Biochem Physiol A Mol Integr Physiol 2015 Oct 20;188:40-8. Epub 2015 Jun 20.

Department of Physiology and Immunology, Faculty of Biology, University of Barcelona, Barcelona 08028, Spain. Electronic address:

Leptin and ghrelin are important regulators of energy homeostasis in mammals, whereas their physiological roles in fish have not been fully elucidated. In the present study, the effects of leptin and ghrelin on adipogenesis, lipolysis and on expression of lipid metabolism-related genes were examined in rainbow trout adipocytes in vitro. Leptin expression and release increased from preadipocytes to mature adipocytes in culture, but did not affect the process of adipogenesis. While ghrelin and its receptor were identified in cultured differentiated adipocytes, ghrelin did not influence either preadipocyte proliferation or differentiation, indicating that it may have other adipose-related roles. Leptin and ghrelin increased lipolysis in mature freshly isolated adipocytes, but mRNA expression of lipolysis markers was not significantly modified. Leptin significantly suppressed the fatty acid transporter-1 expression, suggesting a decrease in fatty acid uptake and storage, but did not affect expression of any of the lipogenesis or β-oxidation genes studied. Ghrelin significantly increased the mRNA levels of lipoprotein lipase, fatty acid synthase and peroxisome proliferator-activated receptor-β, and thus appears to stimulate synthesis of triglycerides as well as their mobilization. Overall, the study indicates that ghrelin, but not leptin seems to be an enhancer of lipid turn-over in adipose tissue of rainbow trout, and this regulation may at least partly be mediated through autocrine/paracrine mechanisms. The mode of action of both hormones needs to be further explored to better understand their roles in regulating adiposity in fish.
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http://dx.doi.org/10.1016/j.cbpa.2015.06.017DOI Listing
October 2015

Characterisation and expression analysis of cathepsins and ubiquitin-proteasome genes in gilthead sea bream (Sparus aurata) skeletal muscle.

BMC Res Notes 2015 Apr 15;8:149. Epub 2015 Apr 15.

Department of Physiology and Immunology, Faculty of Biology, University of Barcelona, Av. Diagonal 643, Barcelona, 08028, Spain.

Background: The proteolytic enzymes involved in normal protein turnover in fish muscle are also responsible for post-mortem softening of the flesh and are therefore potential determinants of product quality. The main enzyme systems involved are calpains, cathepsins, and the ubiquitin-proteasome (UbP). In this study on Sparus aurata (Sa), the coding sequences of cathepsins (SaCTSB and SaCTSDb) and UbP family members (SaN3 and SaUb) were cloned from fast skeletal muscle, and their expression patterns were examined during ontogeny and in a fasting/re-feeding experiment.

Results: The amino acid sequences identified shared 66-100% overall identity with their orthologues in other vertebrates, with well conserved characteristic functional domains and catalytic residues. SaCTSDb showed phylogenetic, sequence and tissue distribution differences with respect to its paralogue SaCTSDa, previously identified in the ovary. Expression of gilthead sea bream cathepsins (B, L, Da, Db) and UbP members (N3, Ub, MuRF1 and MAFbx) in fast skeletal muscle was determined at three different life-history stages and in response to fasting and re-feeding in juveniles. Most of the proteolytic genes analysed were significantly up-regulated during fasting, and down-regulated with re-feeding and, between the fingerling (15 g) and juvenile/adult stages (~50/500 g), consistent with a decrease in muscle proteolysis in both later contexts. In contrast, SaCTSDa and SaMuRF1 expression was relatively stable with ontogeny and SaUb had higher expression in fingerlings and adults than juveniles.

Conclusions: The data obtained in the present study suggest that cathepsins and UbP genes in gilthead sea bream are co-ordinately regulated during ontogeny to control muscle growth, and indicate that feeding regimes can modulate their expression, providing a potential dietary method of influencing post-mortem fillet tenderisation, and hence, product quality.
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http://dx.doi.org/10.1186/s13104-015-1121-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4431372PMC
April 2015

Effects of nutritional status on plasma leptin levels and in vitro regulation of adipocyte leptin expression and secretion in rainbow trout.

Gen Comp Endocrinol 2015 Jan 8;210:114-23. Epub 2014 Nov 8.

Department of Physiology and Immunology, Faculty of Biology, University of Barcelona, Barcelona 08028, Spain. Electronic address:

As leptin has a key role on appetite, knowledge about leptin regulation is important in order to understand the control of energy balance. We aimed to explore the modulatory effects of adiposity on plasma leptin levels in vivo and the role of potential regulators on leptin expression and secretion in rainbow trout adipocytes in vitro. Fish were fed a regular diet twice daily ad libitum or a high-energy diet once daily at two ration levels; satiation (SA group) or restricted (RE group) to 25% of satiation, for 8weeks. RE fish had significantly reduced growth (p<0.001) and adipose tissue weight (p<0.001), and higher plasma leptin levels (p=0.022) compared with SA fish. Moreover, plasma leptin levels negatively correlated with mesenteric fat index (p=0.009). Adipocytes isolated from the different fish were treated with insulin, ghrelin, leucine, eicosapentaenoic acid or left untreated (control). In adipocytes from fish fed regular diet, insulin and ghrelin increased leptin secretion dose-dependently (p=0.002; p=0.033, respectively). Leptin secretion in control adipocytes was significantly higher in RE than in SA fish (p=0.022) in agreement with the in vivo findings, indicating that adipose tissue may contribute to the circulating leptin levels. No treatment effects were observed in adipocytes from the high-energy diet groups, neither in leptin expression nor secretion, except that leptin secretion was significantly reduced by leucine in RE fish adipocytes (p=0.025). Overall, these data show that the regulation of leptin in rainbow trout adipocytes by hormones and nutrients seems to be on secretion, rather than at the transcriptional level.
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http://dx.doi.org/10.1016/j.ygcen.2014.10.016DOI Listing
January 2015
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