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1.
Anim Microbiome ; 4(1): 20, 2022 Mar 10.
Article in English | MEDLINE | ID: mdl-35272695

ABSTRACT

BACKGROUND: Mucosal surfaces of fish provide cardinal defense against environmental pathogens and toxins, yet these external mucosae are also responsible for maintaining and regulating beneficial microbiota. To better our understanding of interactions between host, diet, and microbiota in finfish and how those interactions may vary across mucosal tissue, we used an integrative approach to characterize and compare immune biomarkers and microbiota across three mucosal tissues (skin, gill, and gut) in Atlantic salmon receiving a control diet or diets supplemented with mannan-oligosaccharides, coconut oil, or both. Dietary impacts on mucosal immunity were further evaluated by experimental ectoparasitic sea lice (Lepeophtheirus salmonis) challenge. RESULTS: Fish grew to a final size of 646.5 g ± 35.8 during the 12-week trial, with no dietary effects on growth or sea lice resistance. Bacterial richness differed among the three tissues with the highest richness detected in the gill, followed by skin, then gut, although dietary effects on richness were only detected within skin and gill. Shannon diversity was reduced in the gut compared to skin and gill but was not influenced by diet. Microbiota communities clustered separately by tissue, with dietary impacts on phylogenetic composition only detected in the skin, although skin and gill communities showed greater overlap compared to the gut according to overall composition, differential abundance, and covariance networks. Inferred metagenomic functions revealed preliminary evidence for tissue-specific host-microbiota coadaptation, as putative microbiota functions showed ties to the physiology of each tissue. Immune gene expression profiles displayed tissue-specific signatures, yet dietary effects were also detected within each tissue and peripheral blood leukocytes. Procrustes analysis comparing sample-matched multivariate variation in microbiota composition to that of immune expression profiles indicated a highly significant correlation between datasets. CONCLUSIONS: Diets supplemented with functional ingredients, namely mannan-oligosaccharide, coconut oil, or a both, resulted in no difference in Atlantic salmon growth or resistance to sea lice infection. However, at the molecular level, functional ingredients caused physiologically relevant changes to mucosal microbiota and host immune expression. Putative tissue-specific metagenomic functions and the high correlation between expression profiles and microbiota composition suggest host and microbiota are interdependent and coadapted in a tissue-specific manner.

3.
J Acad Nutr Diet ; 113(2): 282-7, 2013 Feb.
Article in English | MEDLINE | ID: mdl-23351633

ABSTRACT

Enhanced n-3 fatty acid intake benefits cardiovascular disease (CVD) risk reduction. Increasing consumption at a population level may be better addressed by diet than through supplementation. However, limited data are available on the effect of the dose response to fish intake on plasma levels of n-3 fatty acids. To compare the effects of different doses of farmed Atlantic salmon on plasma phospholipid fatty acid proportions and CVD risk biomarkers (eg, glucose, insulin, homeostasis model of assessment-insulin resistance, high-sensitivity C-reactive protein, and interleukin-6) in healthy subjects we performed a randomized three-period crossover-designed trial (4-week treatment, 4- to 8-week washout) to compare the effects of twice per week consumption of farmed Atlantic salmon at doses of 90, 180, and 270 g in 19 apparently healthy men and women (mean age 40 to 65 years) and a body mass index between 25 and 34.9. All study visits were conducted at the US Department of Agriculture Agricultural Research Service Grand Forks Human Nutrition Research Center. Eicosapentaenoic acid and total n-3 concentrations were increased (P<0.05) by all treatments in a dose-response manner, with total n-3 of 8.03% ± 0.26% and 9.21% ± 0.26% for 180- and 270-g doses, respectively. Linoleic acid did not change in response to treatment, whereas arachidonic acid (P<0.05) and total n-6 fatty acids decreased dose dependently (<0.0001). The addition of farmed Atlantic salmon to the diet twice per week for 4 weeks at portions of 180 g and 270 g modifies phospholipid fatty acid proportions of n-3 and n-6 in a level associated with decreased risk for CVD.


Subject(s)
Cardiovascular Diseases/blood , Fatty Acids, Omega-3/administration & dosage , Fatty Acids, Omega-3/blood , Salmo salar , Adult , Aged , Animals , Biomarkers/blood , Body Mass Index , C-Reactive Protein/metabolism , Cardiovascular Diseases/epidemiology , Cardiovascular Diseases/prevention & control , Cross-Over Studies , Docosahexaenoic Acids/blood , Dose-Response Relationship, Drug , Eicosapentaenoic Acid/blood , Fatty Acids, Omega-6/blood , Female , Humans , Insulin/metabolism , Interleukin-6/blood , Linoleic Acid/blood , Male , Middle Aged , Seafood , Triglycerides/blood
4.
J Agric Food Chem ; 59(20): 11278-86, 2011 Oct 26.
Article in English | MEDLINE | ID: mdl-21919483

ABSTRACT

The consumption of seafood enriched in n-3 polyunsaturated fatty acids (PUFA) is associated with a decreased risk of cardiovascular disease. Several n-3 oxidation products from eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (22:6n-3) have known protective effects in the vasculature. It is not known whether the consumption of cooked seafood enriched in n-3 PUFA causes appreciable consumption of lipid oxidation products. We tested the hypothesis that baking Atlantic salmon (Salmo salar) increases the level of n-3 and n-6 PUFA oxidation products over raw salmon. We measured the contents of several monohydroxy-fatty acids (MHFA), prostanoids, and resolvins. Our data demonstrate that baking did not change the overall total levels of MHFA. However, baking resulted in selective regioisomeric loss of hydroxy fatty acids from arachidonic acid (20:4n-6) and EPA, while significantly increasing hydroxyl-linoleic acid levels. The contents of prostanoids and resolvins were reduced several-fold with baking. The inclusion of a coating on the salmon prior to baking reduced the loss of some MHFA but had no effect on prostanoid losses incurred by baking. Baking did not decrease n-3 PUFA contents, indicating that baking of salmon is an acceptable means of preparation that does not alter the potential health benefits of high n-3 seafood consumption. The extent to which the levels of MHFA, prostanoids, and resolvins in the raw or baked fish have physiologic consequence for humans needs to be determined.


Subject(s)
Docosahexaenoic Acids/analysis , Fatty Acids/analysis , Hot Temperature , Prostaglandins/analysis , Salmo salar , Seafood/analysis , Animals , Fatty Acids, Omega-3/analysis , Fatty Acids, Omega-6/analysis
5.
J Nutr ; 137(12): 2763-8, 2007 Dec.
Article in English | MEDLINE | ID: mdl-18029496

ABSTRACT

Supplementation of prebiotic compounds, including short-chain fructooligosaccharides (scFOS) has been shown to confer benefits on nutrient utilization, growth, and disease resistance of various animal species through improved gastrointestinal (GI) microbiota. However, potential uses of prebiotics for shrimp have not been defined. A 6-wk feeding trial was conducted in a recirculating system to determine the effects of scFOS supplementation on growth performance, immune functions, and GI microbiota composition of Pacific white shrimp (Litopenaeus vannamei). scFOS was supplemented in a nutritionally complete diet (35% crude protein) at 0.025, 0.0500, 0.075, 0.100, 0.200, 0.400, and 0.800% by weight. After 6 wk of feeding, shrimp fed 0, 0.1, and 0.8% scFOS were sampled for assays of immune function and GI microbiota. Dietary supplementation of scFOS did not improve weight gain, feed conversion ratio, or survival of shrimp. Denaturing gradient gel electrophoresis analysis suggested the intestinal tract microbial community from shrimp fed the basal diet was different from that of shrimp fed the scFOS diets [similarity coefficient (SC) = 74.9%)], although the intestinal tract microbial community from shrimp fed the scFOS-supplemented diets was very similar (SC = 92.3%). All the bacterial species contributing to the GI microbial differences were identified, although most of them are uncultured species. Both total hemocyte count and hemocyte respiratory burst increased (P < 0.05) by incremental dietary supplementation of scFOS (0-0.8%). This study is the first to our knowledge to show that dietary scFOS can selectively support growth of certain bacterial species in the GI tract of shrimp and enhance immunity, which may facilitate development of alternative strategies, including novel probiotics and synbiotics, for shrimp growth and health management.


Subject(s)
Aquaculture/methods , Gastrointestinal Tract/immunology , Gastrointestinal Tract/microbiology , Oligosaccharides/pharmacology , Penaeidae/immunology , Penaeidae/microbiology , Animal Feed , Animal Nutritional Physiological Phenomena , Animals , Diet , Dietary Supplements , Dose-Response Relationship, Drug , Gastrointestinal Tract/drug effects , Penaeidae/drug effects , Penaeidae/growth & development , Water
6.
Vis Neurosci ; 24(3): 363-75, 2007.
Article in English | MEDLINE | ID: mdl-17640446

ABSTRACT

Connexin 35/36 is the most widespread neuronal gap junction protein in the retina and central nervous system. Electrical and/or tracer coupling in a number of neuronal circuits that express this connexin are regulated by light adaptation. In many cases, the regulation of coupling depends on signaling pathways that activate protein kinases such as PKA, and Cx35 has been shown to be regulated by PKA phosphorylation in cell culture systems. To examine whether phosphorylation might regulate Cx35/36 in the retina we developed phospho-specific polyclonal antibodies against the two regulatory phosphorylation sites of Cx35 and examined the phosphorylation state of this connexin in the retina. Western blot analysis with hybrid bass retinal membrane preparations showed Cx35 to be phosphorylated at both the Ser110 and Ser276 sites, and this labeling was eliminated by alkaline phosphatase digestion. The homologous sites of mouse and rabbit Cx36 were also phosphorylated in retinal membrane preparations. Quantitative confocal immunofluorescence analysis showed gap junctions identified with a monoclonal anti-Cx35 antibody to have variable levels of phosphorylation at both the Ser110 and Ser276 sites. Unusual gap junctions that could be identified by their large size (up to 32 microm2) and location in the IPL showed a prominent shift in phosphorylation state from heavily phosphorylated in nighttime, dark-adapted retina to weakly phosphorylated in daytime, light-adapted retina. Both Ser110 and Ser276 sites showed significant changes in this manner. Under both lighting conditions, other gap junctions varied from non-phosphorylated to heavily phosphorylated. We predict that changes in the phosphorylation states of these sites correlate with changes in the degree of coupling through Cx35/36 gap junctions. This leads to the conclusion that connexin phosphorylation mediates changes in coupling in some retinal networks. However, these changes are not global and likely occur in a cell type-specific or possibly a gap junction-specific manner.


Subject(s)
Connexins/metabolism , Eye Proteins/metabolism , Retina/metabolism , Adaptation, Ocular/physiology , Amino Acid Sequence , Animals , Bass , Cell Membrane/metabolism , Cyclic AMP-Dependent Protein Kinases , Gene Expression Regulation/physiology , Gene Expression Regulation/radiation effects , In Vitro Techniques , Models, Biological , Phosphorylation , Photic Stimulation/methods , Retina/cytology , Serine/metabolism , Gap Junction delta-2 Protein
7.
Biochem Biophys Res Commun ; 335(4): 1191-8, 2005 Oct 07.
Article in English | MEDLINE | ID: mdl-16112650

ABSTRACT

We examined the interactions of calmodulin with neuronal gap junction proteins connexin35 (Cx35) from perch, its mouse homologue Cx36, and the related perch Cx34.7 using surface plasmon resonance. Calmodulin bound to the C-terminal domains of all three connexins with rapid kinetics in a concentration- and Ca2+-dependent manner. Dissociation was also very rapid. K(d)'s for calmodulin binding at a high-affinity site ranged from 11 to 72 nM, and K(1/2)'s for Ca2+ were between 3 and 5 microM. No binding to the intracellular loops was observed. Binding competition experiments with synthetic peptides mapped the calmodulin binding site to a 10-30 amino acid segment at the beginning of the C-terminal domain of Cx36. The micromolar K(1/2)'s and rapid on and off rates suggest that this interaction may change dynamically in neurons, and may occur transiently when Ca2+ is elevated to a level that would occur in the near vicinity of an activated synapse.


Subject(s)
Calcium/chemistry , Calmodulin/chemistry , Connexins/chemistry , Eye Proteins/chemistry , Fish Proteins/chemistry , Neurons/chemistry , Animals , Binding Sites , Mice , Perches , Protein Binding , Gap Junction delta-2 Protein
8.
Brain Res Mol Brain Res ; 135(1-2): 1-11, 2005 Apr 27.
Article in English | MEDLINE | ID: mdl-15857663

ABSTRACT

Connexin 35 (Cx35) is a major component of electrical synapses in the central nervous system. Many gap junctions containing Cx35 are regulated by dopamine receptor pathways that involve protein kinase A (PKA). To study the mechanism of PKA regulation, we analyzed direct phosphorylation of Cx35 by PKA in vitro and studied the regulation of neurobiotin tracer coupling in HeLa cells expressing Cx35 or Cx35 mutants that lack phosphorylation sites. In Cx35-transfected cells, application of the PKA activator Sp-8-cpt-cAMPS caused a significant decline in coupling, while a PKA inhibitor, Rp-8-cpt-cAMPS, significantly increased tracer coupling. In vitro phosphorylation and mutagenic analysis showed that PKA phosphorylates Cx35 directly at two major sites, Ser110 in the intracellular loop and Ser276 in the carboxyl terminus. In addition, a minor phosphorylation site in the C-terminus was identified by truncation of the last 7 amino acids at Ser298. The mutations Ser110Ala or Ser276Ala significantly reduced regulation of coupling by the PKA activator while a combination of the two eliminated regulation. Truncation at Ser298 reversed the regulation such that the PKA activator significantly increased and the PKA inhibitor significantly decreased coupling. The activation was eliminated in the S110A, S276A, S298ter triple mutant. We conclude that PKA regulates Cx35 coupling in a complex manner that requires both major phosphorylation sites. Furthermore, the tip of the C-terminus acts as a "switch" that determines whether phosphorylation will inhibit or enhance coupling. Reliance on the combined states of three sites provides fine control over the degree of coupling through Cx35 gap junctions.


Subject(s)
Connexins/metabolism , Cyclic AMP-Dependent Protein Kinases/metabolism , Cyclic AMP/analogs & derivatives , Gap Junctions/metabolism , Alanine/genetics , Amino Acid Sequence , Animals , Connexins/chemistry , Connexins/genetics , Cyclic AMP/pharmacology , Cyclic AMP-Dependent Protein Kinases/chemistry , Cyclic AMP-Dependent Protein Kinases/genetics , Enzyme Inhibitors/pharmacology , Gene Expression Regulation/drug effects , HeLa Cells , Humans , Immunohistochemistry/methods , Mice , Models, Biological , Mutagenesis/physiology , Mutation , Perches , Phosphorylation/drug effects , RNA, Messenger/biosynthesis , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/metabolism , Reverse Transcriptase Polymerase Chain Reaction/methods , Sequence Alignment , Serine/genetics , Serine/metabolism , Skates, Fish , Transfection/methods
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