RESUMO
Proteinograms, a semiquantitative analytical method that separates proteins into multiple bands, have not been explored in teleosts for diagnostic or prognostic purposes. This study aimed to establish reference values for proteinograms in the serum of gilthead seabream (Sparus aurata) and European sea bass (Dicentrarchus labrax), two important farmed fish species in the Mediterranean region. Serum proteins were studied using SDS-PAGE, electropherogram, and HPLC-mass spectrometry. SDS-PAGE analysis revealed four major bands of proteins around 11, 25, 70, and 100 kDa in the serum of gilthead seabream and European sea bass. Electropherogram results showed that a protein with a molecular weight of 76.8 kDa was the most abundant protein in the serum of gilthead seabream, while a peak of 75.5 kDa was the most abundant in European sea bass. HPLC-mass spectrometry detected 87 proteins and 119 proteins in the serum of gilthead seabream and European sea bass, respectively, including α1-globulins, α2-globulins, ß-globulins, and γ-globulins. Notably, the albumin sequence was not detected in either of the two species. These results help to characterize the serum protein profile and to establish reference proteinograms for these two fish species. They also provide a basis for the development of novel approaches for the rapid detection of loss of haemostasis due to stress, health disorders or disease in farmed fish.
Assuntos
Bass , Proteínas Sanguíneas , Proteínas de Peixes , Dourada , Animais , Bass/sangue , Dourada/sangue , Proteínas Sanguíneas/análise , Proteínas de Peixes/sangue , Proteínas de Peixes/química , Proteínas de Peixes/genética , Hemostasia , Eletroforese em Gel de Poliacrilamida/veterinária , Espectrometria de Massas/veterinária , Valores de Referência , Cromatografia Líquida de Alta Pressão/veterináriaRESUMO
This study assessed postprandial plasma aminoacidemia, glycemia, insulinemia and appetite responses to ingestion of a novel salmon-derived protein peptide (Salmon PP) compared with milk protein isolate (Milk PI). In a randomised, participant-blind crossover design, eleven healthy adults (M = 5, F = 6; mean ± sd age: 22 ± 3 years; BMI: 24 ± 3 kg/m2) ingested 0·3 g/kg/body mass of Salmon PP or Milk PI. Arterialised blood samples were collected whilst fasted and over a 240-min postprandial period. Appetite sensations were measured via visual analogue scales. An ad libitum buffet-style test meal was administered after each trial. The incremental AUC (iAUC) plasma essential amino acid (EAA) response was similar between Salmon PP and Milk PI. The iAUC plasma leucine response was significantly greater following Milk PI ingestion (P < 0·001), whereas temporal and iAUC plasma total amino acid (P = 0·001), non-essential amino acid (P = 0·002), glycine (P = 0·0025) and hydroxyproline (P < 0·001) responses were greater following Salmon PP ingestion. Plasma insulin increased similarly above post-absorptive values following Salmon PP and Milk PI ingestion, whilst plasma glucose was largely unaltered. Indices of appetite were similarly altered following Salmon PP and Milk PI ingestion, and total energy and macronutrient intake during the ad libitum meal was similar between Salmon PP and Milk PI. The postprandial plasma EAA, glycine, proline and hydroxyproline response to Salmon PP ingestion suggest this novel protein source could support muscle and possibly connective tissue adaptive remodelling, which warrants further investigation, particularly as the plasma leucine response to Salmon PP ingestion was inferior to Milk PI.
Assuntos
Aminoácidos , Apetite , Glicemia , Estudos Cross-Over , Insulina , Período Pós-Prandial , Salmão , Humanos , Feminino , Animais , Adulto Jovem , Apetite/efeitos dos fármacos , Apetite/fisiologia , Masculino , Aminoácidos/sangue , Adulto , Glicemia/metabolismo , Glicemia/análise , Insulina/sangue , Proteínas de Peixes/sangue , Proteínas do Leite/farmacologia , Peptídeos/sangue , Proteínas Alimentares/administração & dosagemRESUMO
Toll-like receptor 5 (TLR5) is responsible for bacterial flagellin recognition in vertebrates. In the present study, TLR5M was identified in the Nile tilapia Oreochromis niloticus (OnTLR5), containing a conserved LRR domain, a transmembrane region and a C-terminal TIR domain, similar to that of other fishes and mammals. OnTLR5 was broadly expressed in all the tissues examined, presenting the highest expression levels in the blood and the lowest in the kidney. OnTLR5 was detected from 2 d postfertilization (dpf) to 8 dpf during embryonic development. Moreover, expression levels of OnTLR5 were clearly altered in all five tissues examined in response to Streptococcus agalactiae infection in vivo. Overexpression of OnTLR5 in HEK293T cells revealed that OnTLR5 was distributed in the cytoplasm and significantly increased NF-κB activation. In response to cotransfection with OnMyd88, OnTLR5 significantly upregulated OnMyd88-induced NF-κB activation. Pulldown assays showed that OnTLR5 interacts with OnMyd88 and revealed an interaction between TLR5 and Aeromonas hydrophila flagellin. Taken together, these findings suggest that OnTLR5 plays important roles in TLR/IL-1R signalling pathways and the immune response to pathogen invasion.
Assuntos
Aeromonas hydrophila , Ciclídeos , Doenças dos Peixes , Fator 88 de Diferenciação Mieloide , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Peixes/sangue , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Proteínas de Peixes/metabolismo , Flagelina/farmacologia , Células HEK293 , Humanos , Fator 88 de Diferenciação Mieloide/metabolismo , NF-kappa B/metabolismo , Receptor 5 Toll-Like/biossíntese , Receptor 5 Toll-Like/sangue , Receptor 5 Toll-Like/genética , Receptor 5 Toll-Like/metabolismoRESUMO
Circulating insulin-like growth factor (IGF)-I has been proposed as a growth index in several teleosts, including salmonids, and its level in circulation is stabilized by multiple IGF-binding proteins (IGFBPs). Three IGFBPs, IGFBP-2b, -1a, and -1b, are consistently detected in salmonid blood and are suggested to be indices of positive or negative growth, although their applicability to rainbow trout (Oncorhynchus mykiss) is unclear. The present study examined the usefulness of IGFBPs along with IGF-I as a physiological indicator of growth rate in rainbow trout through a rearing experiment. Two groups of underyearling rainbow trout were pit-tagged and either fed or fasted for 33 days. A third group was fasted for 22 days, followed by refeeding for 11 days. Serum IGF-I levels were reduced after fasting for 22 days, but refeeding did not retore its levels to those of the fed control. Nevertheless, there was a positive relationship between serum IGF-I levels and individual growth rates over 33 days of experimentation, confirming its validity as a growth index. Ligand blotting using labeled human IGF-I revealed two IGFBP bands at 43 and 32 kDa, which corresponded to IGFBP-2b and an unidentified form, respectively. In contrast, bands corresponding to IGFBP-1a and -1b, which usually increase after fasting, were hardly detected, even in the fasted fish. The responses of circulating IGFBP-2b to fasting and refeeding were similar to those of circulating IGF-I and positively correlated with growth rate and IGF-I levels. The intensity of the serum 32-kDa IGFBP band was higher in constantly fed fish than in the fasted fish; however, its correlation with growth rate was weaker than those of IGF-I and IGFBP-2b. The present study shows that IGF-I and IGFBP-2b can be used as growth indices for rainbow trout. In contrast, circulating IGFBP-1a and -1b may not serve as negative growth indices in rainbow trout under regular aquaculture conditions because they are rarely detected by ligand blotting or respond to fasting/refeeding.
Assuntos
Proteínas de Peixes , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina , Fator de Crescimento Insulin-Like I , Oncorhynchus mykiss , Animais , Jejum , Proteínas de Peixes/sangue , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/sangue , Fator de Crescimento Insulin-Like I/análise , Oncorhynchus mykiss/metabolismoRESUMO
The study was designed to identify the types of mitogen-activated protein kinases (MAPKs) in erythrocytes and liver tissues of river lamprey Lampetra fluviatilis and monitor the changes in protein expression levels of found enzymes on the course of prespawning starvation (from November to the end of May). Immunoreactivity of the native and phosphorylated forms of ERK1/2, JNK and p38 was examined in the cytosolic and membrane cell fractions. Both lamprey erythrocytes and liver were found to highly express ERK1/2 and JNK, whereas only trace amounts of p38 were revealed in hepatic tissues. ERK1/2 was identified in cytosolic and membrane fractions, whereas JNK and p38 were predominantly cytosolic enzymes. Total cellular amounts of ERK1/2 and phospho-ERK1/2 in both erythrocytes and liver tissues appeared to be relatively stable on the course of prespawning starvation. However, before spawning ERK1/2 translocated from cytosol to membranes, with partial decline of its cytoplasmic expression being compensated by increases in membrane-bound pool. Immunoreactivity of cytoplasmic JNK, phospho-JNK and p38 were stable from November to March, but sharply decreased before spawning exhibiting almost negligible levels in May, which suggests the depletion of their cellular fractions. Most probably, ERK1/2 plays more important role in mediating adaptive responses of erythrocytes and liver tissues to conditions of natural starvation and maintenance of cell viability before spawning and death of animals in May.
Assuntos
Proteínas de Peixes/metabolismo , Lampreias/metabolismo , Fígado/enzimologia , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Animais , Eritrócitos/enzimologia , Feminino , Proteínas de Peixes/sangue , Lampreias/sangue , Masculino , Proteínas Quinases Ativadas por Mitógeno/sangue , Reprodução , Estações do Ano , Inanição/sangue , Inanição/enzimologia , Frações Subcelulares/enzimologiaRESUMO
Exosomes are associated with cancer progression, pregnancy, cardiovascular diseases, central nervous system-related diseases, immune responses and viral pathogenicity. However, study on the role of exosomes in the immune response of teleost fish, especially antiviral immunity, is limited. Herein, serum-derived exosomes from mandarin fish were used to investigate the antiviral effect on the exosomes of teleost fish. Exosomes isolated from mandarin fish serum by ultra-centrifugation were internalized by mandarin fish fry cells and were able to inhibit Infectious spleen and kidney necrosis virus (ISKNV) infection. To further investigate the underlying mechanisms of exosomes in inhibiting ISKNV infection, the protein composition of serum-derived exosomes was analyzed by mass spectrometry. It was found that myxovirus resistance 1 (Mx1) was incorporated by exosomes. Furthermore, the mandarin fish Mx1 protein was proven to be transferred into the recipient cells though exosomes. Our results showed that the serum-derived exosomes from mandarin fish could inhibit ISKNV replication, which suggested an underlying mechanism of the exosome antivirus in that it incorporates Mx1 protein and delivery into recipient cells. This study provided evidence for the important antiviral role of exosomes in the immune system of teleost fish.
Assuntos
Infecções por Vírus de DNA , Exossomos , Doenças dos Peixes , Proteínas de Peixes , Peixes , Iridoviridae , Proteínas de Resistência a Myxovirus , Animais , Linhagem Celular , Infecções por Vírus de DNA/sangue , Infecções por Vírus de DNA/imunologia , Infecções por Vírus de DNA/veterinária , Exossomos/imunologia , Exossomos/metabolismo , Doenças dos Peixes/sangue , Doenças dos Peixes/imunologia , Proteínas de Peixes/sangue , Proteínas de Peixes/imunologia , Peixes/sangue , Peixes/imunologia , Peixes/virologia , Iridoviridae/imunologia , Iridoviridae/metabolismo , Proteínas de Resistência a Myxovirus/sangue , Proteínas de Resistência a Myxovirus/imunologiaRESUMO
The effects of stocking density on growth performance, serum biochemistry, digestive enzymes, immune response, and muscle quality of largemouth bass (Micropterus salmoides) reared in nine in-pond raceway systems (IPRS, 22.0 m × 5.0 m × 2.0 m) were studied. M. salmoides with initial an body weight of 8.25 ± 0.51 g and body length of 6.99 ± 0.44 cm were reared at an initial stocking density of 90.91 ind./m3 (low stocking density, LSD), 113.63 ind./m3 (middle stocking density, MSD), and 136.36 ind./m3 (high stocking density, HSD) with triplication. After 300 days of culture, MSD recorded the highest final body weight, weight gain, specific growth rate, and yield, but the food conversion ratio in MSD was the lowest. The viscerosomatic index in LSD was significantly higher than other groups. The fish serum reared at HSD showed significantly lower total protein, higher total cholesterol, triglyceride, total bilirubin, glucose content, alanine transaminase, and aspartate transaminase activity. Significantly lower intestinal amylase, lipase, trypsin activities, hepatic superoxide dismutase (SOD) and catalase (CAT) activities, and higher malondialdehyde content were detected in HSD compared to others. The content of crude lipid, saturated fatty acid decreased, and total essential amino acid, delicious amino acid, and polyunsaturated fatty acid increased in muscle with stocking density increase. No significant difference was observed in muscle texture. Profitability analysis indicated the benefit-to-cost ratio varied between 1.10 and 1.68, of which MSD was significantly higher than others. The optimal stocking density for M. salmoides should be 113.63 ind./m3 in an IPRS farm.
Assuntos
Aquicultura/métodos , Bass , Alanina Transaminase/sangue , Aminoácidos/metabolismo , Amilases/metabolismo , Animais , Aspartato Aminotransferases/sangue , Bass/sangue , Bass/crescimento & desenvolvimento , Bass/imunologia , Bass/metabolismo , Catalase/metabolismo , Ácidos Graxos/metabolismo , Proteínas de Peixes/sangue , Imunidade , Intestinos/enzimologia , Lipase/metabolismo , Fígado/metabolismo , Músculos/química , Esteróis/sangue , Superóxido Dismutase/metabolismo , Triglicerídeos/sangue , Tripsina/metabolismoRESUMO
Kalliklectin is a unique fish-specific lectin that demonstrates sequence similarity to mammalian plasma kallikrein and coagulation factor XI, which are not lectins but proteases. Reported fish kalliklectins and these mammalian proteases comprise four characteristic "apple domains" (APDs). Bioinformatics analysis revealed that Siluriformes species possess anomalous kalliklectins comprising 6 to 16 APDs. Complementary DNA cloning showed that the full-length nucleotide sequence of Ictalurus punctatus consists of 2240 bp that encode 720 amino acid residues to produce a mature protein with a putative 18 amino acid N-terminus peptide sequence. This protein has a predicted molecular mass of 83,417.23 Da. Reverse transcription-polymerase chain reaction (RT-PCR) showed that this lectin gene expresses in the liver but not in any other tissues, including the mucosal tissues. This differential expression pattern makes this lectin unique compared to other lectins described in previous studies. We successfully detected an 85-kDa protein in the serum using western blotting analysis, suggesting that this lectin protein is produced by the liver and secreted into the bloodstream. We characterized a novel cDNA sequence encoding a new type of kalliklectin with eight APDs isolated from channel catfish, I. punctatus. Based on phylogenetic analysis, we speculated that there was a duplication of the third and fourth APD set in a common Siluriformes ancestor at some point after its separation from the common teleost ancestor and that these duplications then underwent independent repeats in different lineages resulting in the generation of the [APD1]-[APD2]-{[APD3]-[APD-4]} × n structure in modern catfishes.
Assuntos
Evolução Molecular , Proteínas de Peixes/química , Proteínas de Peixes/genética , Ictaluridae/genética , Lectinas/química , Lectinas/genética , Domínios Proteicos/genética , Sequência de Aminoácidos , Animais , Sequência de Bases/genética , Clonagem Molecular/métodos , DNA Complementar/genética , Proteínas de Peixes/sangue , Expressão Gênica , Ictaluridae/sangue , Lectinas/sangue , Filogenia , Alinhamento de Sequência/métodos , Homologia Estrutural de ProteínaRESUMO
Streptococcus agalactiae is one of the most important pathogens infecting tilapia worldwide and causes meningoencephalitis, septicemia and high mortalities with considerable losses. Various types of vaccines have been developed against S. agalactiae infection, such as inactivated vaccines, live attenuated vaccines and subunit vaccines. Bacterial ghosts (BGs) are nonliving, empty cell envelopes and have been reported as novel vaccine candidates. Therefore, the main aims of this study were to develop an S. agalactiae ghost vaccine (SAGV) and to evaluate the immune response and protective effect of SAGV against S. agalactiae with two novel adjuvants, Montanide™ ISA 763B VG and Montanide™ GEL02. Nile tilapia, mean weight 50 g, were divided into four groups as follows; 1) fish injected with PBS as control, 2) fish injected with the SAGV alone; 3) fish injected with the SAGV+Montanide™ ISA 763B VG; and 4) fish injected with SAGV+Montanide™ GEL02. Following vaccination, innate immunity parameters including serum lysozyme, myeloperoxidase, catalase, and bactericidal activity were all significantly enhanced. Moreover, specific serum IgM antibodies were induced and reached their highest level 2-8 weeks post vaccination. Importantly, the relative percent survival of tilapia vaccinated against the SAGV formulated with both adjuvants was 80-93%. Furthermore, the transcription of immune-related genes (IgM, TCRß, IL-1ß, IL-8 and TNFα) were up-regulated in tilapia after vaccination, indicating that both cellular and humoral immune responses were induced by these adjuvanted vaccines. In summary, Montanide™ ISA 763B VG and Montanide™ GEL02 can enhance immunoprotection induced by the SAGV vaccine against streptococcosis, demonstrating that both have value as potential adjuvants of fish vaccines.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Ciclídeos/imunologia , Doenças dos Peixes/prevenção & controle , Manitol/análogos & derivados , Manitol/administração & dosagem , Infecções Estreptocócicas/prevenção & controle , Vacinas Estreptocócicas/administração & dosagem , Streptococcus agalactiae/imunologia , Animais , Anticorpos Antibacterianos/sangue , Catalase/sangue , Ciclídeos/sangue , Doenças dos Peixes/sangue , Doenças dos Peixes/imunologia , Proteínas de Peixes/sangue , Fígado/imunologia , Muramidase/sangue , Peroxidase/sangue , Baço/imunologia , Infecções Estreptocócicas/sangue , Infecções Estreptocócicas/imunologiaRESUMO
In recent times, carbamazepine (CBZ) as an anticonvulsants drug has raised attention because of its safety concern in the aquatic environment. The present study aimed to evaluate the sub-lethal effects of CBZ (1%, 0.1 % and 0.01 % of 96 h LC50) on P. hypophthalmus for 60 days based on haematological, biochemical, and genotoxicity biomarkers. Chronic exposure of CBZ altered blood profiles (total erythrocyte count, packed cell volume, haemoglobin) and serum biomarkers such as alkaline phosphates, cholesterol, lactate dehydrogenase and transaminase enzymes. Oxidative stress biomarkers such as superoxide dismutase (SOD) and catalase (CAT) activity were also substantially affected in all treatments. Genotoxicity study revealed the formation of micronucleus in erythrocytes of exposed fish. Integrated Biomarker Response (IBR) study showed cholesterol, serum glutamic oxaloacetic transaminase (SGOT) in serum and SOD, CAT in liver tissue are the best organ-based enzyme biomarkers. The present report concludes that an environmentally realistic concentration of CBZ can pose a serious threat to aquatic organisms.
Assuntos
Anticonvulsivantes/toxicidade , Carbamazepina/toxicidade , Peixes-Gato , Poluentes Químicos da Água/toxicidade , Animais , Biomarcadores/sangue , Biomarcadores/metabolismo , Proteínas Sanguíneas/análise , Catalase/metabolismo , Peixes-Gato/sangue , Peixes-Gato/metabolismo , Contagem de Eritrócitos , Proteínas de Peixes/sangue , Proteínas de Peixes/metabolismo , Hemoglobinas/análise , Dose Letal Mediana , Fígado/efeitos dos fármacos , Fígado/metabolismo , Micronúcleos com Defeito Cromossômico/induzido quimicamente , Estresse Oxidativo/efeitos dos fármacos , Albumina Sérica/análise , Superóxido Dismutase/metabolismo , Testes de Toxicidade CrônicaRESUMO
Two IgM heavy (H) chain sub-isotypes (80 and 40 kDa) and two light (L) chain variants (25 and 30 kDa) were detected in the serum of giant grouper (Epinephelus lanceolatus), purified by ammonium sulphate precipitation followed by protein A affinity chromatography. This method yielded 5.6 mg/mL high purity IgM from grouper serum, with efficiency estimated at 39.5% recovery from crude serum. The H and L chains were identified by SDS-PAGE and mass spectrometry (MS). Nanopore long-read sequencing was used to generate a genomic contig (MW768935), containing Cµ, Cδ loci, VH regions, and a H chain Joining segment. cDNA sequencing of Cµ transcripts (MW768933 and MW768934) were used to polish the genomic contig and determine the exons and introns of the corresponding locus. MS peptide mapping revealed that the 80 kDa H chain consisted of CH1-4 domains while peptides from the 40 kDa H chain only mapped to CH1-2 domains. Our genomic contig showed the Cµ locus has a Cµ1-Cµ2-Cµ3-Cµ4 arrangement on the same strand as the other Ig loci identified in this genomic sequence. Our study corrects the NCBI annotations of the opposing Cµ loci (LOC117268697 and LOC117268550) in chromosome 16 (NC_047006). Further, we identified both κ and λ L chain isotypes in serum IgM. The molecular weight differences observed may result from different combinations of CL and VL genes. Putative IgM sub-isotypes have also been reported in Epinephelus itajara and Epinephelus coioides. The presence of IgM sub-isotypes may be a conserved trait among Epinephelus species.
Assuntos
Bass/genética , Proteínas de Peixes/sangue , Genoma , Cadeias Pesadas de Imunoglobulinas/sangue , Imunoglobulina M/sangue , Animais , Bass/imunologia , Cromatografia de Afinidade/veterinária , Espectrometria de Massas/veterinária , Análise de Sequência de DNA/veterináriaRESUMO
This study was done to evaluate the effects of Bisphenol A (BPA) on Siberian sturgeon (Acipenser baerii). As liver is the main organ in the homeostatic adjustments to stress, we used a proteomics method to address molecular response in this tissue. Also, we compared the levels of vitellogenin in plasma and mucus to propose that the last one be a non-invasive method to analyze this biomarker. The fish received 1, 10, and 100 µg g-1 week-1 BPA intraperitoneally for two weeks. The samples were taken on days 0, 7, and 14. Plasma vitellogenin level increased as the highest value was recorded in the group with 100 µg g-1 week-1 of BPA. Changes in the mucus and blood vitellogenin showed a similar pattern, suggesting that mucus could be used for evaluating the changes in blood vitellogenin. Comparative proteomics was used to determine the proteome of the liver of A. baerii in the highest dose of BPA in comparison with the control. Sixteen proteins were identified that their expression changed at least twice between the studied groups. The proteomic results showed that BPA increased the expression of proteins involved in the detoxification and metabolism, activated glycolysis, and produced necrosis in the liver.
Assuntos
Compostos Benzidrílicos/efeitos adversos , Disruptores Endócrinos/efeitos adversos , Poluentes Ambientais/efeitos adversos , Proteínas de Peixes/metabolismo , Peixes/metabolismo , Fenóis/efeitos adversos , Vitelogeninas/metabolismo , Animais , Proteínas de Peixes/análise , Proteínas de Peixes/sangue , Peixes/sangue , Fígado/efeitos dos fármacos , Fígado/metabolismo , Proteoma/análise , Proteoma/metabolismo , Vitelogeninas/análise , Vitelogeninas/sangueRESUMO
Extracellular vesicles (EVs) are lipid bilayer vesicles which are released from cells and play multifaceted roles in cellular communication in health and disease. EVs can be isolated from various body fluids, including serum and plasma, and are usable biomarkers as they can inform health status. Studies on EVs are an emerging research field in teleost fish, with accumulating evidence for important functions in immunity and homeostasis, but remain to be characterised in most fish species, including halibut. Protein deimination is a post-translational modification caused by a conserved family of enzymes, named peptidylarginine deiminases (PADs), and results in changes in protein folding and function via conversion of arginine to citrulline in target proteins. Protein deimination has been recently described in halibut ontogeny and halibut serum. Neither EV profiles, nor total protein or deiminated protein EV cargos have yet been assessed in halibut and are reported in the current study. Halibut serum EVs showed a poly-dispersed population in the size range of 50-600 nm, with modal size of EVs falling at 138 nm, and morphology was further confirmed by transmission electron microscopy. The assessment of EV total protein cargo revealed 124 protein hits and 37 deiminated protein hits, whereof 15 hits were particularly identified in deiminated form only. Protein interaction network analysis showed that deimination hits are involved in a range of gene regulatory, immune, metabolic and developmental processes. The same was found for total EV protein cargo, although a far wider range of pathways was found than for deimination hits only. The expression of complement component C3 and C4, as well as pentraxin-like protein, which were identified by proteomic analysis, was further verified in EVs by western blotting. This showed that C3 is exported in EVs at higher levels than C4 and deiminated C3 was furthermore confirmed to be at high levels in the deimination-enriched EV fractions, while, in comparison, C4 showed very low detection in deimination-enriched EV fractions. Pentraxin was exported in EVs, but not detected in the deimination-enriched fractions. Our findings provide novel insights into EV-mediated communication in halibut serum, via transport of protein cargo, including post-translationally deiminated proteins.
Assuntos
Citrulinação , Vesículas Extracelulares/metabolismo , Proteínas de Peixes/metabolismo , Proteoma/metabolismo , Animais , Proteínas do Sistema Complemento/metabolismo , Vesículas Extracelulares/ultraestrutura , Proteínas de Peixes/sangue , Linguado , Mapas de Interação de Proteínas , Desiminases de Arginina em Proteínas/metabolismoRESUMO
Two genotypes of the intestinal parasite Ceratonova shasta infect Oncorhynchus mykiss: genotype 0 results in a chronic infection with low mortality while genotype IIR causes disease with high mortality. We determined parasite load and the relative expression of six immune factors (IgT, IgM, IL-6, IL-8, IL-10, IFNG) in fish infected with either genotype over 29 days post-exposure. In genotype IIR infections the host responded with upregulation of inflammatory and regulatory cytokines. In contrast, genotype 0 infection did not elicit an inflammatory response and expression of IFNG and IL-10 was lower. Antibody expression was upregulated in both infections but appeared to have limited efficacy in the virulent genotype IIR infections. Histologically, in genotype 0 infections the parasite migrated through the tissue layers causing inflammation but minimal damage to the mucosal epithelium, which contrasts with the severe pathology found in genotype IIR infections.
Assuntos
Doenças dos Peixes/imunologia , Genótipo , Inflamação/imunologia , Mucosa/imunologia , Myxozoa/genética , Oncorhynchus mykiss/imunologia , Doenças Parasitárias em Animais/imunologia , Animais , Movimento Celular , Citocinas/genética , Citocinas/metabolismo , Proteínas de Peixes/sangue , Interações Hospedeiro-Parasita , Imunoglobulina M/sangue , Imunoglobulinas/sangue , Myxozoa/patogenicidade , Carga Parasitária , VirulênciaRESUMO
The natural antioxidants are well known for their antioxidative activity without side effects when compared to antibiotics. Hence, the present study aimed at evaluating p-Coumaric acid as an antioxidant additive on the blood and mRNA levels of antioxidant-related factors in common carp (Cyprinus carpio). Fish fed the basal diet supplemented with p-Coumaric at 0, 0.5, 1, and 1.5 g/kg for 56 days, then the serum, intestine, and liver samples were collected. The growth performance of fish fed with CA showed significantly (P < 0.05) improved FW, WG, and SGR compared to those of the control one. However, the feed conversion ratio was significantly (P < 0.05) reduced in fish fed 1 and 1.5 g/kg diet levels. SOD was not significantly differed among the groups fed with varied p-Coumaric acid (P > 0.05). Serum GPX and TAC were enhanced considerably by p-Coumaric acid regarding the control with the highest being in fish fed 1.5 g/kg diet (P < 0.05). Serum CAT was more elevated in fish provided p-Coumaric acid at 1 or 1.5 g/kg than the control while fish fed 0.5 g/kg did not display significant changes. MDA level significantly decreased by all p-Coumaric acid groups compared to the control one, and the lowest level was observed in 1.5 g/kg (P < 0.05). The mRNA level of CAT was significantly upregulated in the liver by p-Coumaric acid at 1 or 1.5 g/kg (P < 0.05), while the intestine CAT did not influence by p-Coumaric acid (P > 0.05). The measured SOD in the liver and intestine samples revealed no changes in common carp fed p-Coumaric acid (P > 0.05). GPX was significantly upregulated in the intestine by p-Coumaric acid at 1 or 1.5 g/kg (P < 0.05), whereas the liver GPX was upregulated by p-Coumaric acid at 1.5 g/kg. The mRNA level of the GST gene in the intestine of common carp was upregulated by p-Coumaric acid at 1.5 g/kg, whereas the liver displayed upregulated GST in fish fed 1 g/kg diet. The present study approved the application of p-Coumaric acid as a natural antioxidant for friendly, sustainable aquaculture.
Assuntos
Carpas/sangue , Carpas/genética , Ácidos Cumáricos/farmacologia , Suplementos Nutricionais , Animais , Dieta , Proteínas de Peixes/sangue , Proteínas de Peixes/genética , Glutationa Transferase/sangue , Glutationa Transferase/genética , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Oxirredutases/sangue , Oxirredutases/genética , RNA Mensageiro/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
In this study, changes in the blood tissue of rainbow trout (Oncorhynchus mykiss, Walbaum, 1792) caused by Fipronil (FP) insecticide were investigated using different biomarkers (Hematology parameters, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), malondialdehyde (MDA), paraoxonase (PON), arylesterase (ARE), myeleperoxidase (MPO), micronucleus (MN), 8-hydroxy-2-deoxyguanosine (8-OHdG)) level and caspase-3 activity. Statistically significant alterations in hematology parameters occurred with FP effect. In blood tissue, dose-dependent inhibition was determined in SOD-CAT-GPX-PON and ARE enzyme activities, but MDA and MPO were induced statistically significant. The results of MN assay were compared with the control group and it was obtained that genotoxicity of different dose groups was similar. The level of 8-OHdG and the activity and caspase-3 examined in blood tissue was increased depending on the dose. It was determined with different biomarkers that this insecticide caused physiological stress changes in the tissues examined.
Assuntos
Apoptose/efeitos dos fármacos , Dano ao DNA , Inseticidas/toxicidade , Micronúcleos com Defeito Cromossômico/induzido quimicamente , Oncorhynchus mykiss , Estresse Oxidativo/efeitos dos fármacos , Pirazóis/toxicidade , Poluentes Químicos da Água/toxicidade , 8-Hidroxi-2'-Desoxiguanosina/sangue , Animais , Biomarcadores/sangue , Caspase 3/sangue , Relação Dose-Resposta a Droga , Proteínas de Peixes/sangue , Oncorhynchus mykiss/sangue , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/metabolismoRESUMO
Cartilaginous fishes, comprising the chimeras, sharks, skates, and rays, split from the common ancestor with other jawed vertebrates approx. 450 million years ago. Being the oldest extant taxonomic group to possess an immunoglobulin (Ig)-based adaptive immune system, examination of this group has taught us much about the evolution of adaptive immunity, as well as the conserved and taxon-specific characteristics of Igs. Significant progress has been made analyzing sequences from numerous genomic and transcriptomic data sets. These findings have been supported by additional functional studies characterizing the Igs and humoral response of sharks and their relatives. This review will summarize what we have learned about the genomic organization, protein structure, and in vivo function of these Ig isotypes in cartilaginous fishes and highlight the areas where our knowledge is still lacking.
Assuntos
Elasmobrânquios/imunologia , Proteínas de Peixes/genética , Isotipos de Imunoglobulinas/genética , Imunidade Adaptativa/genética , Animais , Conjuntos de Dados como Assunto , Elasmobrânquios/sangue , Elasmobrânquios/genética , Proteínas de Peixes/sangue , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica , Isotipos de Imunoglobulinas/sangue , Isotipos de Imunoglobulinas/imunologiaRESUMO
In this study, we examined the effects of porcine growth hormone (GH) and cortisol on plasma insulin-like growth factor binding proteins (IGFBPs) in juveniles of three subspecies of Oncorhynchus masou (masu, amago, and Biwa salmon). Ligand blotting using digoxigenin-labeled human IGF-I was used to detect and semi-quantify three major circulating IGFBP bands at 41, 28, and 22 kDa, corresponding to IGFBP-2b, -1a, and -1b, respectively. GH increased plasma IGFBP-2b concentration in masu and Biwa salmon but suppressed it in amago salmon. Plasma IGFBP-2b levels were increased by cortisol in the three subspecies. Cortisol induced plasma IGFBP-1a in the three subspecies, whereas GH had a suppressive effect in masu and Biwa salmon. Sham and cortisol injections increased plasma IGFBP-1b levels after 1 day in masu and amago salmon, suggesting that IGFBP-1b is induced following exposure to stressors via cortisol. Increased IGFBP-1b levels were restored to basal levels when co-injected with GH in Biwa salmon, and the same trend was seen in masu and amago salmon. However, the suppressive effect of GH disappeared 2 days after injection in the three subspecies. Despite some differences among subspecies, the findings suggest that cortisol is a primary inducer of plasma IGFBP-1b; however, GH counteracts it in the short term. Therefore, GH has the potential to modulate the degree of increase in circulating IGFBP-1b levels during acute stress.
Assuntos
Proteínas de Peixes/sangue , Hormônio do Crescimento/farmacologia , Hidrocortisona/farmacologia , Proteína 1 de Ligação a Fator de Crescimento Semelhante à Insulina/sangue , Oncorhynchus/sangue , Animais , Western Blotting , Hormônio do Crescimento/administração & dosagem , Hidrocortisona/administração & dosagem , Fator de Crescimento Insulin-Like I/metabolismo , Oncorhynchus/classificação , Oncorhynchus/metabolismo , Isoformas de Proteínas/sangue , Especificidade da EspécieRESUMO
Temperature is a key environmental factor, and understanding how its fluctuations affect physiological and metabolic processes is critical for fish. The present study characterizes the energy response and fatty acid metabolism in Onychostoma macrolepis exposed to low temperature (10 °C). The results demonstrated that cold stress remarkably disrupted the energy homeostasis of O. macrolepis, then the AMP-activated protein kinase (AMPK) could strategically mobilize carbohydrates and lipids. In particular, when the O. macrolepis were faced with cold stress, the lipolysis was stimulated along with the enhanced fatty acid ß-oxidation for energy, while the fatty acid synthesis was supressed in the early stage. Additionally, the fatty acid composition analysis suggested that saturated fatty acid (SFA) might accumulate while monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) in storage lipids (mainly containing non-polar lipid, NPL) could be utilized to supply energy during cold acclimation. Altogether, this study may provide some meritorious for understanding the cold-tolerant mechanism of fish in the viewpoint of energy balance combined with fatty acid metabolism, and thus to contribute to this species rearing in fish farms in the future.
Assuntos
Resposta ao Choque Frio/fisiologia , Cyprinidae/metabolismo , Metabolismo Energético , Ácidos Graxos/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Nucleotídeos de Adenina/metabolismo , Tecido Adiposo/metabolismo , Animais , Colesterol/sangue , Resposta ao Choque Frio/genética , Proteínas de Peixes/sangue , Proteínas de Peixes/genética , Expressão Gênica , Glucose/metabolismo , Metabolismo dos Lipídeos , Fígado/metabolismo , Músculos/metabolismo , Temperatura , Triglicerídeos/metabolismoRESUMO
Vaccination plays a critical role in the protection of humans and other animals from infectious diseases. However, the same vaccine often confers different protection levels among individuals due to variation in genetics and/or immunological histories. While this represents a well-recognized issue in humans, it has received little attention in fish. Here we address this knowledge gap in a proteomic study of rainbow trout (Oncorhynchus mykiss, Walbaum), using non-lethal repeated blood sampling to establish the plasma protein response of individual fish following immunization. Six trout were immunized with adjuvanted hen egg-white lysozyme (HEL) and peripheral blood sampled at ten time points from day 0 to day 84 post-injection. We confirm that an antigen-specific antibody response to HEL was raised, showing differences in timing and magnitude among individuals. Using label-free liquid chromatography-mass spectrometry, we quantified the abundance of 278 plasma proteins across the timecourse. As part of the analysis, we show that this approach can distinguish many (but not all) duplicated plasma proteins encoded by paralogous genes retained from the salmonid-specific whole genome duplication event. Global variation in the plasma proteome was predominantly explained by individual differences among fish. However, sampling day explained a major component of variation in abundance for a statistically defined subset of 41 proteins, representing 15% of those detected. These proteins clustered into five groups showing distinct temporal responses to HEL immunization at the population level, and include classical immune (e.g. complement system members) and acute phase molecules (e.g. apolipoproteins, haptoglobins), several enzymes and other proteins supporting the immune response, in addition to evolutionarily conserved molecules that are as yet uncharacterized. Overall, this study improves our understanding of the fish plasma proteome, provides valuable marker proteins for different phases of the immune response, and has implications for vaccine development and the design of immune challenge experiments.