Influences of subzero thermal acclimation on mitochondrial membrane composition of temperate zone marine bivalve mollusks

The phospholipid and phospholipid fatty acid composition of gill mitochondrial membranes from two temperate zone marine bivalve mollusks, the quahog, Mercenaria mercenaria, and the American oyster, Crassostrea virginica, were examined after acclimation to 12 and −1°C. Cardiolipin (CL) was the only phospholipid with proportions altered upon acclimation to −1°C, increasing 188% in the mitochondrial membranes of M. mercenaria. Although the ratio of bilayer stabilizing to destabilizing lipids is frequently associated with cold acclimation in ectothermic species, no change was found in this ratio in either of the species. Polyunsaturated fatty acids (PUFA) were found only to increase in C. virginica with cold acclimation, with total n-3 PUFA increasing in the phospholipid phosphatidylethanolamine, total n-6 PUFA increasing in CL, and total PUFA increasing in phosphatidylinositol. Monounsaturated fatty acids, not PUFA, were found to have increased in M. mercenaria, with 18:1n−9 increasing by 150% in CL, and 20∶1 increasing in both CL and phosphatidylcholine, by 146 and 192%, respectively. These manipulations of membrane phospholipid and fatty acid composition may represent an attempt by these species to help maintain membrane function at low temperatures.     

Effect of freezing on quality of sea bass and gilthead sea bream

Freezing is an efficient method of fish preservation. The aim of the present work was to examine the impact of freezing in fatty acid composition and in the in vitro inhibitory activity of sea bass (Dicentrarchus labrax) and gilthead sea bream (Sparus aurata) fillet lipids against platelet activating factor (PAF). The Bligh and Dyer extraction method and the counter‐current distribution method were used to obtain total, polar and neutral lipids. The fatty acid analysis conducted using the internal standard method and the biological assay on washed rabbit platelets took place calculating the in vitro inhibitory activity of fish lipids against 2.5 × 10^?11 M of PAF. No statistical changes (p<0.05) occurred in fatty acid content of fresh and thawed gilthead sea bream, while fatty acid amount in thawed sea bass was significantly higher (p<0.05) compared to fresh fish. Total lipids of both thawed fish species exhibited stronger anti‐thrombotic activity compared to fresh fish. Freezing preserved fish quality and reinforced the anti‐thrombotic properties of fish oils, since even after 6 months of freezing, fish oils preserve their nutritional value in terms of protecting against cardiovascular diseases. Practical applications: Fish fillets contain high amount of unsaturated lipids that may easily undergo lipid oxidation. Freezing and frozen storage prevent such oxidative changes so fish quality is retained. Fatty acids and PAF‐antagonists in fish are of major importance since they contribute to the nutritional value of fish. The practical application of this work lies on the evaluation of the nutritional value of fish in terms of cardio protection by examining the impact of freezing on the levels of fatty acids and PAF‐antagonists in aquacultured fish fillets.     

Cold acclimation or grapeseed oil feeding affects phospholipid composition and mitochondrial function in duckling skeletal muscle

The phospholipid fatty acid (FA) composition and functional properties of skeletal muscle and liver mitochondria were examined in cold-acclimated (CA, 4°C) ducklings. Phospholipid FA of isolated muscle mitochondria from CA birds were longer and more unsaturated than those from thermoneutral (TN, 25°C) reared ducklings. The rise in long-chain and polyunsaturated FA (PUFA, mainly 20∶4n-6) was associated with a higher State 4 respiration rate and a lower respiratory control ratio (RCR). Hepatic mitochondria, by contrast, were much less affected by cold acclimation. The cold-induced changes in phospholipid FA profile and functional properties of muscle mitochondria were reproduced by giving TN ducklings a diet enriched in grapeseed oil (GO, rich in n-6 FA), suggesting a causal relationship between the membrane structure and mitochondrial functional parameters. However, hepatic mitochondria from ducklings fed the GO diet also showed an enrichment in long-chain PUFA but opposite changes in their biochemical characteristics (lower State 4, higher RCR). It is suggested that the differential modulation of mitochondrial functional properties by membrane lipid composition between skeletal muscle and liver may depend on muscle-specific factors possibly interacting with long-chain PUFA and affecting the proton leakiness of mitochondrial membranes.     

Molecular characterization of three peroxisome proliferator-activated receptors from the sea bass (<Emphasis Type="Italic">Dicentrarchus labrax</Emphasis>)

Peroxisome proliferator-activated receptors (PPAR) are nuclear hormone receptors that control the expression of genes involved in lipid homeostasis in mammals. We searched for PPAR in sea bass, a marine fish of particular interest to aquaculture, after hypothesizing that the physiological and molecular processes that regulate lipid metabolism in fish are similar to those in mammals. Here, we report the identification of complementary DNA and corresponding genomic sequences that encode three distinct PPAR from sea bass. The sea bass PPAR are the structural homologs of the mammalian PPARα, β/δ and γ isotypes. As revealed by RNase protection, the tissue expression profile of the fish PPAR appears to be very similar to that of the mammalian PPAR homologs. Thus, PPARα is mainly expressed in the liver, PPARγ in adipose tissue, and PPARβ in all tissues tested, with its highest levels in the liver, where it is also the dominant isotype expressed. Like mammalian PPAR, the sea bass isotypes recognize and bind to PPAR response elements of both mammalian and piscine origin, as heterodimers with the 9-cis retinoic acid receptor. Through the coactivator-dependent receptor ligand assay, we also demonstrated that natural FA and synthetic hypolipidemic compounds can act as ligands of the sea bass PPARα and β isotypes. This suggests that the sea bass PPAR act through similar mechanisms and perform the same critical lipid metabolism functions as mammalian PPAR.     

Homeostatic control of membrane fatty acid composition in the rat after dietary lipid treatment

Diets in which both the lipid content and composition (polyunsaturated to saturated fatty acid ratio) were varied were fed to rats for 20 weeks, and the effects on the tissue lipid profiles were determined. The fatty acid profile of the plasma lipids, and the phospholipid fatty acids of the mitochondrial and microsomal fractions of liver, heart, kidney and brain, as well as erythrocyte membranes were determined. Despite large differences in the level and type of lipid present in the experimental diets and in the proportion of saturated fatty acids in the plasma lipids in response to the various diets, there was little effect on the proportion of saturated to unsaturated fatty acids in the phospholipids of the various membranes examined. The major effect of altering the dietary level of polyunsaturated to saturated fatty acids was on the ratio of the ω6/ω3 series of unsaturated fatty acids in the membrane lipids. This change occurred in all tissues except the brain, in which only a small response to altered dietary lipid intake was observed. The ω6/ω3 ratio was elevated upon feeding a diet rich in ω6 polyunsaturated fatty acids, but decreased when a diet rich in saturated fatty acids was fed. The failure to significantly alter membrane lipid saturation/unsaturation in the tissues examined would suggest that a homeostatic mechanism is operative in biological membranes and may act to buffer membranes from the effects of changes in the nature of the dietary lipid intake.     

Survey of qualitative traits of European sea bass cultivated in different rearing systems

The purpose of this study was to evaluate the main qualitative traits of European sea bass fattened in farms adopting different rearing techniques (offshore cages, inshore cages, land‐based basins) but fed the same diets in each production cycle, in a three year survey (2006, 2007, 2008). Three farms were chosen for each rearing technique and two sampling sessions of ten fish each were carried out for each rearing cycle. Proximate composition, fatty acid profile, and cholesterol content were evaluated in the fillet of marketable size fish as well as morpho‐biometric parameters and indices. Diets administered in 2008 had higher amounts of lipids, lower proportions of n–3 polyunsaturated fatty acids (PUFA) and higher proportions of n–6 PUFA compared to 2006 and 2007 diets. The chemical and fatty acid composition of the fish fillet was affected by the diet composition: fish fattened in 2008 exhibited statistically higher amounts of fat and cholesterol, lower proportions of n–3 PUFA and higher proportions of n–6 PUFA in comparison with fish cultivated in 2006 and 2007. Fish cultivated in cages, both offshore and inshore cages, exhibited leaner fillets, lower amounts of cholesterol and higher proportions of n–3 PUFA than fish cultivated in basins. Practical applications: In this study we have analyzed the combined effects of feeding and rearing systems of sea bass in a 3‐year survey for the first time. The results showed that feeding is the main factor affecting fish quality as regards both the chemical composition and the fatty acid profile. The partial substitution of fish meal and fish oil with vegetable sources, with the aim of saving the wild fish biomass as well as formulating less expensive diets, strongly alters the fillet composition. We also showed that the rearing system of fish in offshore cages makes it is possible to obtain better quality products.     

Incorporation ofcis- andtrans-octadecenoic acids into the membranes of rat liver mitochondria

The incorporation of dietary isomeric fatty acids into the membranes of liver mitochondria was investigated. Three groups of rats were fed diets containing 3% sunflower seed oil plus 15%, 20%, or 25% partially hydrogenated arachis oil. A fourth group was fed 25% partially hydrogenated arachis oil, but no sunflower seed oil. All diets were given for 3, 6, or 10 weeks. After 10 weeks, the content oftrans fatty acids in the lipids of the mitochondrial membranes was 15–19% of the total fatty acids. The composition of thetrans- and thecis-octadecenoic acids in the lipids of the mitochondrial membranes was similar for all groups supplemented with sunflower seed oil (SO), irrespective of time and dietary level of partially hydrogenated arachis oil (HAO). Thecis 18∶1 (n−8), which was a major isomer of the partially hydrogenated arachis oil, was almost excluded from the mitochondrial fatty acids. Likewise, the content oftrans 18∶1 (n−8) was considerably lower in the mitochondrial lipids than in the diet. On the contrary, the content oftrans 18∶1 (n−6) was higher in the mitochondrial lipids than in the diet. In the group fed without sunflower seed oil, isomers of linoleic acid and arachidonic acid were observed in the lipids of mitochondrial membranes. Presented in part at the ISF Congress, Marseille, September 1976.     

Dietary phospholipids are more efficient than neutral lipids for long-chain polyunsaturated fatty acid supply in European sea bass <Emphasis Type="Italic">Dicentrarchus labrax</Emphasis> larval development

We evaluated the effects of dietary lipid class (phospholipid vs. neutral lipid) and level of n−3 long-chain PUFA (LC-PUFA) on the growth, digestive enzymatic activity, and histological organization of the intestine and liver in European sea bass larvae. Fish were fed from the onset of exogenous feeding at 7 to 37 d post-hatch with five isoproteic and isolipidic compound diets with different levels of EPA and DHA. Diet names indicated the percentage of EPA and DHA contained in the phospholipids (PL) and neutral lipids (NL), as follows: PL5, PL3, PL1, NL1, and NL3. Histological observations showed different patterns of lipid absorption and accumulation in the intestinal mucosa depending on the level and nature of the dietary lipid fraction. Fish fed high levels of neutral lipids (11%, NL3 diet: 2.6% of EPA+DHA in the NL fraction) showed large intracellular and intercellular lipid deposits in the anterior intestine, but no such lipid accumulation was detected when larvae were fed with low and moderate levels of EPA and DHA in the phospholipid and neutral lipid fractions of the diet (PL and NL1 diets). PL were preferentially absorbed in the postvalvular intestine, and the accumulation of marine PL was inversely correlated to their dietary level. The postvalvular intestinal mucosa and liver showed signs of steatosis; large lipid vacuoles were observed in this region of the intestine and in the liver and were inversely correlated with the level of dietary neutral lipids. The best results in terms of growth, survival, and development (maturation of the digestive system and histological organization of the liver and intestinal mucosa) were obtained in the group fed with 2.3% of EPA and DHA in the PL fraction of the diet (PL3 diet), revealing that European sea bass larvae use the LC-PUFa contained in the PL fraction more efficiently than those from the NL fraction of the diet.     

Thermal adaptation affects the fatty acid composition of plasma phospholipids in trout

The adaptive changes in the fatty acid (FA) composition of plasma phospholipids (PL) in response to alterations in environmental water temperature were investigated in juvenile rainbow trout (Oncorhynchus mykiss). The changes observed during thermal adaptation from 22°C in summer to 8°C in winter were reproduced by laboratory cold acclimation (CA) at 6°C of 22°C-summer-acclimated animals. In cold-acclimated and winter-acclimated trout, the increase in the unsaturation of PL fatty acids was mainly due to an enrichment of approximately 7% in the total weight percentage of 22∶6n−3, while a concomitant significant decrease in the levels of 18∶0 and of the monoenoic n−9 FA was observed. A time course study revealed that the changes in PL fatty acids became significant after 10 d of CA and were complete after one month. These changes in the composition of the fatty acyl chains of plasma total PL indicate that the FA composition of plasma lipoprotein PL does not remain constant during thermal adaptation. This would suggest that plasma lipoproteins provide a rapid systemic supply of lipids containing more or less unsaturated FA during thermal adaptation of poikilothermic animals.     

The lipid composition of selected tissues from a Mediterranean monk seal,Monachus monachus

The lipid composition of blubber, brain, muscle and heart from a Mediterranean monk sealMonachus monachus (an endangered species) were examined to allow comparisons with more common species of seals. Only neutral lipids (mainly triacylglycerols) were detectable in the blubber lipids, whereas polar lipids predominated in the heart and in the brain. Neutral and polar lipids comprised almost equal proportions in both liver and muscle. Choline glycerophospholipids (CGP) were the major polar lipids, followed by ethanolamine glycerophospholipids (EGP) in the liver, heart and muscle. Cerebrosides accounted for 28.8% of the brain lipids. All lipid classes of the liver contained high levels (31–47%) of polyunsaturated fatty acids (PUFA), with the exception of phosphatidylserine. The total proportion of n−6 PUFA exceeded that of n−3 PUFA in all lipid classes of the liver, due mainly to the high levels of 20∶4n−6. The highest level of 20∶4n−6 occurred in phosphatidylinositol, where it comprised 32.4% of the total fatty acids. The CGP and EGP of the brain contained lower levels of PUFA than those of the liver, muscle and heart. Alkenyl ethers accounted for 35.8% of the total long-chain moieties in brain EGP. The fatty acid composition of blubber triacylglycerols differed from those of the lipid classes from other tissues in that it had a very low ratio of n−6 to n−3 PUFA (0.3) as a result of a lower content of 20∶4n−6.     

Mitochondrial membrane composition of two arctic marine bivalve mollusks, Serripes groenlandicus and Mya truncata

The phospholipid and fatty acid composition of gill mitochondria membranes from two Arctic marine bivalve mollusks, Mya truncata and Serripes groenlandicus, were examined. These animals were collected from the Arctic Ocean, where waters remain below 0°C throughout the year. In both species, the primary membrane phospholipids were phosphatidylcholine, and phosphatidylethanolamine. Although a low ratio of bilayer-stabilizing phospholipids to bilayer-destabilizing phospholipids is frequently associated with cold acclimation in temperate species, this ratio is very different between the two species. The monounsaturated fatty acid 20∶1 was abundant in the membranes of both Arctic species equaling 13.0% of the fatty acid composition in S. groenlandicus, and 17.7% in M. truncata. Polyunsaturated fatty acids were relatively low in the Arctic species, equaling 35.9% of total membrane fatty acids compared to that of temperate zone mollusks. It is suggested that monoenes are common in the tissues of Arctic species since they play a role in maintaining membrane function at subzero temperatures.     

Thermal adaptation in biological membranes: Functional significance of changes in phospholipid molecular species composition

Quantities of 1-palmitoyl 2-docosahexaenoyl phosphatidylcholine (16∶0/22∶6-PC or PDPC) increase from 24 to 40 weight percent as a consequence of cold acclimation in mitochondrial membranes of rainbow trout liver (J. Comp. Physiol. 156, 665–674, 1986). The present study was undertaken to assess the impact of such a large change in the proportions of a single molecular species on the fluidity, lateral packing (as sensed by phospholipase A₂), and permeability of biological membranes. These properties were examined in multilamellar liposomes prepared from binary mixtures of dipalmitoyl phosphatidylcholine (DPPC) and PDPC in proportions increasing from 10 to 40 mole% PDPC. Glucose permeability was positively correlated with both assay temperature and PDPC content. The temperature dependence of Na⁺ permeability declined steadily as the mole fraction of PDPC increased; consequently, sodium permeability was positively correlated with PDPC content at 5°C, but inversely correlated at 20°C. Phospholipase A₂ activity was independent of both assay temperature and vesicle composition. Vesicles of all compositions displayed a single transition in the temperature dependence of 1,6 diphenyl-1,3,5-hexatriene (DPH) fluorescence polarization, which shifted to lower temperature and broadened as proportions of PDPC increased. At temperatures belowthe transition, fluidity was positively correlated with the mole fraction of PDPC, but interfacial and deeper regions of the bilayer were affected differently by variations in PDPC content. Nonelectrolyte permeability was the only index of membrane structure or function to be significantly correlated with the fluidity of the bilayer interior. The tendencies of PDPC to both fluidize the membrane and to reduce the temperature sensitivity of electrolyte permeation may promote the adaptation of membrane function to low temperature.     

Temperature-dependent deacylation of molecular species of phosphatidylcholine by microsomal phospholipase A2 of thermally acclimated rainbow trout,Salmo gairdneri 1

Using the ratios of kinetic parameters, V/Km, the deacylation of different molecular species of 1-palmitoyl,2-acyl phosphatidylcholine via microsomal phospholipase A₂ (PLA₂) was studied in liver tissue of thermally acclimated rainbow trout (Salmo gairdneri). In general, PLA₂ from fish acclimated to cold temperatures showed an order of preference for the acyl moieties of 18∶1>18∶1>18∶2>18∶0. Trout acclimated to warm temperatures generally preferred 18∶0 PC, but the actual order of preference depended on the temperature of the assays and the presence of endogenous lipids in the enzyme preparation. At 5 C, the particulate (microsomal) enzyme preferred 18∶0>18∶2>18∶1, but a lipid-free preparation of the enzyme preferred 18∶2>18∶0>18∶1. At 20 C, particulate enzyme preferred 18∶1>18∶0>18∶2 but purified enzyme preferred 18∶0>18∶2>18∶1. Thus, assay temperature and the presence of microsomal lipids had a greater effect on PLA₂ from fish acclimated to warm temperatures than fish acclimated to cold temperatures. The substrate preference of PLA₂ is discussed with reference to the previously observed changes in membrane fatty acid composition that occur with thermal acclimation in rainbow trout.     

Sphingomyelin metabolism is linked to salt transport in the gills of euryhaline fish

Byin vivo andin vitro studies ofl-(3-³H)serine and [9,10(n)-³H]palmitic acid incorporation into phospholipids, we show a change in the renewal of the ceramide moiety of sphingomyelin in the gills of euryhaline fish (sea bass and eels) when the animals were subjected to abrupt alterations in environmental salinity.In vivo, decrease of the salinity from sea water (salinity 3.7%) to diluted sea water (salinity 1%) induced an increase of label incorporation into gill sphingomyelin. The same was true when gills from sea water-adapted sea bass or sea water-adapted eels were incubated in diluted sea water. A decrease in free ceramides synthesis was also observed in the gills of sea water-adapted sea bass when the salinity of the incubation medium was reduced. Direct inhibition of Na⁺/K⁺-ATPase activity with ouabain decreased the sphingomyelin synthesis in the gills of sea bass duringin vitro incubation in diluted sea water, whereas treatment with furosemide stimulated sphingomyelin synthesis in the same gills incubated in sea water. These findings indicate that changes in Na⁺ fluxes modify the sphingomyelin turnover and control the production of free ceramides and sphingosine in gill cells of euryhaline fish. In view of the well-known effects of these sphingomyelin degradation products on isolated tumor cell differentiation, we suggest that they play a very important role in modulating chloride cell distribution and metabolism of fish gills during abrupt changes in environmental salinity.     

Effect of Benzo(a)pyrene on Hepatic Biotransformation Activities: Time Course of Induction in Aquaculture European Sea Bass (Dicentrarchus labrax)

The responses of phase I and phase II enzymes to benzo(a)pyrene (BaP) exposure were characterized in liver of fish farm European sea bass (Dicentrarchus labrax). Treatment of sea bass with BaP by intraperitoneal injection produced a rapid putative MFO induction. Highest ethoxyresorufin-O-deethylase (EROD), ethoxycoumarin-O-deethylase (ECOD) and benzo(a)pyrene monooxygenase (BaPMO) activities were observed in microsomes 14–24 h after BaP injection. EROD was induced 14.5 fold, ECOD 3.9 and BaPMO 13 fold. Spectrophotometrically measured microsomal cytochrome P-450 and NADPH-cytochrome c reductase activity were not induced by BaP injection, whereas a low increase in cytochrome b₅ level was observed (induction factor 1.6 fold). Induction of phase II enzyme activities was lower than that of phase I enzymes: microsomal uridine diphospho-glucuronyl transferase (UDPGT) and cytosolic glutathion-S-transferase (GST) were induced 3.9 and 1.5 fold, respectively. The results indicate that sea bass is a sensitive species with respect to exposure to polycyclic aromatic hydrocarbons, exhibiting early marked responses of phase I and phase II biotransformation enzymes. The species offers potential as a tool for both laboratory studies and pollution monitoring. In the latter context it could be employed in the fish farms that occur alongside coastal regions.     

Growth Temperature and Salinity Impact Fatty Acid Composition and Degree of Unsaturation in Peanut-Nodulating Rhizobia

Growth and survival of bacteria depend on homeostasis of membrane lipids, and the capacity to adjust lipid composition to adapt to various environmental stresses. Membrane fluidity is regulated in part by the ratio of unsaturated to saturated fatty acids present in membrane lipids. Here, we studied the effects of high growth temperature and salinity (NaCl) stress, separately or in combination, on fatty acids composition and de novo synthesis in two peanut-nodulating Bradyrhizobium strains (fast-growing TAL1000 and slow-growing SEMIA6144). Both strains contained the fatty acids palmitic, stearic, and cis-vaccenic + oleic. TAL1000 also contained eicosatrienoic acid and cyclopropane fatty acid. The most striking change, in both strains, was a decreased percentage of cis-vaccenic + oleic (≥80% for TAL1000), and an associated increase in saturated fatty acids, under high growth temperature or combined conditions. Cyclopropane fatty acid was significantly increased in TAL1000 under the above conditions. De novo synthesis of fatty acids was shifted to the synthesis of a higher proportion of saturated fatty acids under all tested conditions, but to a lesser degree for SEMIA6144 compared to TAL1000. The major adaptive response of these rhizobial strains to increased temperature and salinity was an altered degree of fatty acid unsaturation, to maintain the normal physical state of membrane lipids.     

Molecular speciation of fish sperm phospholipids: Large amounts of dipolyunsaturated phosphatidylserine

The molecular species compositions of the main diacyl phosphoglyceride classes and ether-linked subclasses from sperm of three species of fish, sea bass Dicentrarchus labrax, Atlantic salmon Salmo salar and Chinook salmon Onchorhynchus tsawytscha, were determined. The phospholipids from sperm were highly unsaturated, dipolyunsaturated fatty acid (diPUFA) molecular species comprised 64.6 to 71.8% of phosphatidylserine (PS), 10.1 to 17.4% of phosphatidylethanolamine (PE), and 3.3 to 10.1% of phosphatidylcholine (PC). In sea bass sperm, di22∶6n-3 phospholipid was the predominant diPUFA molecular species, but in both salmon species 22∶5n-3/22∶6n-3 was also a major constituent of PS. Phospholipids containing 22∶6n-3 dominated in sea bass sperm with 16∶0/22∶6n-3 as a major component of PC and PE, and 18∶0/22∶6n-3 of PE and PS in addition to di22∶6n-3 in the latter two classes. In contrast, both salmon species contained much more 20∶5n-3 and less 22∶6n-3 so that saturated/20∶5n-3 and monounsaturated/20∶5n-3 molecular species were more abundant than the corresponding molecules containing 22∶6n-3. Ether-linked lipids comprised 11.3–36.3% of choline and ethanolamine phosphoglycerides in each fish species. Molecular species containing 22∶6n-3 were the major components of 1-O-alkyl-2-acyl-glycerophosphocholine, especially 16∶0a/22∶6n-3 in sea bass and 18∶1a/∶6n-3 in the two salmon species, while in 1-O-alk-1′-enyl-2-acyl-glycerophosphoethanolamine, 16∶0a/22∶6n-3 was the major component in both salmon and 18∶0a/22∶6n-3 in sea bass with 18∶1a/22∶6n-3 abundant in all three species. In Atlantic salmon 1-O-alkyl-2-acylglycerophosphoethanolamine comprised 24.6% of ethanolamine glycerophospholipids which were predominantly 16∶0a/22∶6n-3 and 18∶1a/22∶6n-3. Phosphatidylinositol from sperm was dominated by stearoyl/C₂₀ PUFA molecular species, in sea bass overwhelmingly 18∶0/20∶4n-6, while in both salmon species 18∶0/20∶4n-6 and 18∶0/20∶5n-3 were equally abundant.     

Recombinant Bovine Growth Hormone-Induced Metabolic Remodelling Enhances Growth of Gilthead Sea-Bream (Sparus aurata): Insights from Stable Isotopes Composition and Proteomics

Growth hormone and insulin-like growth factors (GH/IGF axis) regulate somatic growth in mammals and fish, although their action on metabolism is not fully understood in the latter. An intraperitoneal injection of extended-release recombinant bovine growth hormone (rbGH, Posilac^®) was used in gilthead sea bream fingerlings and juveniles to analyse the metabolic response of liver and red and white muscles by enzymatic, isotopic and proteomic analyses. GH-induced lipolysis and glycogenolysis were reflected in liver composition, and metabolic and redox enzymes reported higher lipid use and lower protein oxidation. In white and red muscle reserves, rBGH increased glycogen while reducing lipid. The isotopic analysis of muscles showed a decrease in the recycling of proteins and a greater recycling of lipids and glycogen in the rBGH groups, which favoured a protein sparing effect. The protein synthesis capacity (RNA/protein) of white muscle increased, while cytochrome-c-oxidase (COX) protein expression decreased in rBGH group. Proteomic analysis of white muscle revealed only downregulation of 8 proteins, related to carbohydrate metabolic processes. The global results corroborated that GH acted by saving dietary proteins for muscle growth mainly by promoting the use of lipids as energy in the muscles of the gilthead sea bream. There was a fuel switch from carbohydrates to lipids with compensatory changes in antioxidant pathways that overall resulted in enhanced somatic growth.     

Distributions of conjugated linoleic acid (CLA) isomers in tissue lipid classes of pigs fed a commercial CLA mixture determined by gas chromatography and silver ion-high-performance liquid chromatography

Pigs were fed a commercial conjugated linoleic acid (CLA) mixture, prepared by alkali isomerization of sunflower oil, at 2% of the basal diet, from 61.5 to 106 kg live weight, and were compared to pigs fed the same basal diet with 2% added sunflower oil. The total lipids from liver, heart, inner back fat, and omental fat of pigs fed the CLA diet were analyzed for the incorporation of CLA isomers into all the tissue lipid classes. A total of 10 lipid classes were isolated by three-directional thin-layer chromatography and analyzed by gas chromatography (GC) on long capillary columns and by silver-ion high-performance liquid chromatography (Ag⁺-HPLC); cholesterol was determined spectrophotometrically. Only trace amounts (<0.1%; by GC) of the 9,11–18∶2 cis/trans and trans, trans isomers were observed in pigs fed the control diet. Ten and twelve CLA isomers in the diet and in pig tissue lipids were sepatated by GC and Ag⁺-HPLC, respectively. The relative concentration of all the CLA isomers in the different lipid classes ranged from 1 to 6% of the total fatty acids. The four major cis/trans isomers (18.9% 11 cis, 13 trans-18∶2; 26.3% 10 trans, 12 cis-18∶2; 20.4% 9 cis, 11 trans-18∶2; and 16.1% 8 trans, 10 cis-18∶2) constituted 82% of the total CLA isomers in the dietary CLA mixture, and smaller amounts of the corresponding cis,cis (7.4%) and trans,trans (10.1%) isomers were present. The distribution of CLA isomers in inner back fat and in omental fat of the pigs was similar to that found in the diet. The liver triacylglycerols (TAG), free fatty acids (FFA), and cholesteryl esters showed a similar patterns to that found in the diet. The major liver phospholipids showed a marked increase of 9 cis,11 trans-18∶2, ranging from 36 to 54%, compared to that present in the diet. However, liver diphosphatidylglycerol (DPG) showed a high incorporation of the 11 cis,13 trans-18∶2 isomer (43%). All heart lipid classes, except TAG, showed a high content of 11 cis,13 trans-18∶2, which was in marked contrast to results in the liver. The relative proportion of 11 cis,13 trans-18∶2 ranged from 30% in the FFA to 77% in DPG. The second major isomer in all heart lipids was 9 cis,11 trans-18∶2. In both liver and heart lipids the relative proportions of both 10 trans,12 cis-18∶2 and 8 trans,10 cis-18∶2 were significantly lower compared to that found in the diet. The FFA in liver and heart showed the highest content of trans,trans isomers (31 to 36%) among all the lipid classes. The preferential accumulation of the 11 cis,13 trans-18∶2 into cardiac lipids, and in particular the major phospholipid in the inner mitochondrial membrane, DPG, in both heart and liver, appears unique and may be of concern. The levels of 11 cis,13 trans-18∶2 naturally found in foods have not been established.     

Fatty acid composition of wild and cultivated gilthead seabream (Sparus aurata) and sea bass (Dicentrarchus labrax)

Total lipid content and fatty acid composition were determined for wild and cultivated gilthead seabream and sea bass. Fatty acid analyses were carried out by gas chromatography‐ mass spectrometry. Respective total lipid content of flesh in cultivated gilthead seabream and sea bass were 1.7‐5.0‐times those of wild samples. Palmitic acid (C16:0) and oleic acid (C18:1n‐9) were the major fatty acids respectively among the saturated fatty acids and monounsaturated fatty acids of each fish species. It is noteworthy that both linoleic acid (C18:2n‐6) and arachidonic acid (C20:4n‐6) were predominant in total n‐6 polyunsaturated fatty acids in the respective cultivated and wild types. Eicosapentaenoic acid (C20:5n‐3) and docosahexaenoic acid (C22:6n‐3) amounts were significantly higher in flesh of cultivated fish than in wild fish.