Metabolic effects of diet containing blue mussel (Mytilus edulis) and blue mussel-fed salmon in a mouse model of obesity.

Alternative feed ingredients for farmed salmon are warranted due to increasing pressure on wild fish stocks. As locally farmed blue mussels may represent an environmentally sustainable substitute with a lower carbon footprint, we aimed to test the potential and safety of substituting fish meal with blue mussel meal in feed for Atlantic salmon. Salmon were fed diets in which fish meal was partially replaced with blue mussel meal in increments, accounting for up to 13.1 % of the ingredients. Fillets from the salmon were subsequently used to prepare obesity-promoting western diets for a 13-weeks mouse feeding trial. In a second mouse trial, we tested the effects of inclusion of up to 8% blue mussel meal directly in a meat-based western diet. Partial replacement of fish meal with blue mussel meal in fish feed preserved the n-3 polyunsaturated fatty acid (PUFA) content in salmon fillets. The observed blue mussel-induced changes in the fatty acid profiles in salmon fillets did not translate into similar changes in the livers of mice that consumed the salmon, and no clear dose-dependent responses were found. The relative levels of the marine n-3 fatty acids, EPA, and DHA were not reduced, and the n-3/n-6 PUFA ratios in livers from all salmon-fed mice were unchanged. The inclusion of blue mussel meal in a meat-based western diet led to a small, but dose-dependent increase in the n-3/n-6 PUFA ratios in mice livers. Diet-induced obesity, glucose intolerance, and hepatic steatosis were unaffected in both mice trials and no blue mussel-induced adverse effects were observed. In conclusion, our results suggest that replacing fish meal with blue mussel meal in salmon feed will not cause adverse effects in those who consume the salmon fillets.

Application of a Mytilus edulis-derived promoting calcium absorption peptide in calcium phosphate cements for bone.

The IEELEEELEAER peptide (PIE) identified from the protein hydrolysate of Mytilus edulis is reported to enhance osteoblast growth and differentiation, which also possesses a superior bone formation ability both in vitro and in vivo. Moreover, PIE bound to calcium spontaneously at the stoichiometry of 1:1, and there were amino nitrogen and carboxyl oxygen atoms in 2 glutamic acid residues at the calcium-binding sites in the PIE. The PIE-calcium complex facilitated calcium uptake through the Caco-2 cell monolayers. Incorporation of PIE into calcium phosphate cements enhanced calcium ion uptake and proliferation of osteoblasts and inhibit bacteria. This study suggest that calcium phosphate cements supplemented with PIE can serve as a potentially efficient material for bone graft used during spinal surgery.

A high fat-high sucrose diet enriched in blue mussels protects against systemic inflammation, metabolic dysregulation and weight gain in C57BL/6 mice.

High fat-high sucrose (HF-HS) diet, known as the western diet, has been shown to induce the onset of obesity via increasing metabolic inflammation, insulin resistance and adipose tissue dysfunction. Hyperleptinemia, hyperglycemia and dyslipidemia are also the primary observations of obesogenic diet induced obesity. We have previously reported anti-adipogenic and insulin sensitizing effects of blue mussels (BM) using 3T3-L1 cells. BM is a rich source of omega-3 polyunsaturated fatty acids, phytosterols and other micronutrients that has been shown to elicit benefits under obese conditions using in-vitro cell culture models. However, no studies to date have established the anti-obesity effects, safety and efficacy of BM in an in-vivo animal model. In the present study, we fed a HF-HS diet supplemented with different concentrations of BM freeze-dried powder (1.25, 2.5 and 5% w/w) to C57BL/6 mice for 12weeks. A HF-HS diet caused rapid weight gain, hyperglycemia, dyslipidemia, hyperleptinemia, and increased plasma levels of inflammatory cytokines; interleukin (IL)-6 and tumor necrosis factor (TNF)-α. Incorporating 2.5% BM in the HF-HS diet prevented weight gain, dyslipidemia, hyperglycemia and reduced the levels of inflammatory cytokines and leptin mRNA expression. Furthermore, plasma from 2.5% BM increased cholesterol efflux capacity of J774 macrophage cells, compared to plasma from HF-HS diet. There was no effect of 1.25% BM on any tested parameters, while 5% BM was not palatable after four weeks. In conclusion, our findings have established the efficacy and safety of BM using C57BL/6 mice, demonstrating that BM has the potential to target obesity and related complications.

Mussels as a tool for mitigation of nutrients in the marine environment.

Long-line mussel farming has been proposed as a mitigation tool for removal of excess nutrients in eutrophic coastal waters. A full-scale mussel farm optimized for cost efficient nutrient removal was established in the eutrophic Skive Fjord, Denmark where biological and economic parameters related to nutrient removal was monitored throughout a full production cycle (1 yr). The results showed that it was possible to obtain a high area specific biomass of 60 t WW ha(-1) eqvivalent to a nitrogen and phosphorus removal of 0.6-0.9 and 0.03-0.04 t ha(-1)yr, respectively. The analysis of the costs related to establishment, maintenance and harvest revealed that mussel production optimized for mitigation can be carried out at a lower cost compared to mussel production for (human) consumption. The costs for nutrient removal was 14.8 € kg(-1)N making mitigation mussel production a cost-efficient measure compared to the most expensive land-based measures.

Mussel oligopeptides ameliorate cognition deficit and attenuate brain senescence in D-galactose-induced aging mice.

Dietary supplementation exerts beneficial effects in reducing incidence of chronic neurodegenerative diseases. The purpose of this study was to examine protective effects of mussel (Mytilus edulis) oligopeptides supplementation on brain function in D-galactose induced aging mice. Sixty female 8-month-old mice were randomly divided into five groups: vehicle control, D-galactose, and D-galactose combined with 200, 500, 1000 mg/kg mussel oligopeptides. The results showed that mussel oligopeptides could improve cognitive learning and memory ability and protect the hippocampal neurons. In addition, GSH, SOD and GSH-pX activities were increased and MDA level was significantly decreased in mice fed with mussel oligopeptides. It was also found that mussel oligopeptides supplementation prevented D-galactose-induced elevations of iNOS activity and NO production and lactate acid levels in brain. Moreover, PI3K and Akt genes were up-regulated by mussel oligopeptides supplementation. These findings suggest that mussel oligopeptides are able to enhance exercise capacity and protect against oxidative damage caused by D-galactose in aging model mice through regulating oxidation metabolism and PI3K/Akt/NOS signal pathway. Therefore, mussel oligopeptides are good materials for future development of healthcare products to combat age-related brain dysfunction and to improve healthy life span.

Petroleum hydrocarbon contamination in Nelson Lagoon, Alaska, sampling three different matrices.

Polycyclic aromatic hydrocarbon (PAH) levels were measured in sediments, bivalves and semi-permeable membrane devices (SPMDs) in the relatively pristine marine environment of Nelson Lagoon, Alaska. Most PAH levels in Nelson Lagoon were low, and similar to global background concentrations. Sampling media type can significantly influence conclusions of PAH contamination in the environment. Concentration of a broad size range of PAHs was observed in the tissues of blue mussels (Mytilus edulis). SPMDs collected some two- to three-ring PAHs from the dissolved water phase, while sediments collected five- to six-ring PAHs that were likely adsorbed onto particulate matter. Benzo(a)pyrene, a potent carcinogen, was found in mussels at levels similar to more industrialized harbors in Alaska.

Mussel farming as a nutrient reduction measure in the Baltic Sea: consideration of nutrient biogeochemical cycles.

Nutrient loads from the land to the sea must be reduced to combat coastal eutrophication. It has been suggested that further mitigation efforts are needed in the brackish Baltic Sea to decrease nutrients, especially in eutrophic coastal areas. Mussel farming is a potential measure to remove nutrients directly from the sea. Mussels consume phytoplankton containing nitrogen (N) and phosphorus (P); when the mussels are harvested these nutrients are removed from the aquatic system. However, sedimentation of organic material in faeces and pseudo-faeces below a mussel farm consumes oxygen and can lead to hypoxic or even anoxic sediments causing an increased sediment release of ammonium and phosphate. Moreover, N losses from denitrification can be reduced due to low oxygen and reduced numbers of bioturbating organisms. To reveal if mussel farming is a cost-effective mitigation measure in the Baltic Sea the potential for enhanced sediment nutrient release must be assessed.

PAH body burden and biomarker responses in mussels (Mytilus edulis) exposed to produced water from a North Sea oil field: laboratory and field assessments.

In order to study the impact of produced water (PW) from a North Sea oil field on blue mussels (Mytilus edulis), chemical and biological markers were selected. A laboratory exposure (0.125%, 0.25% and 0.5% of PW) and a field study (6 stations 0.2-2 km from a PW discharge point) were conducted. In the laboratory study, PAH bioaccumulation increased in mussel soft tissue even at the lowest exposure dose. Micronuclei frequency demonstrated a dose-response pattern, whereas lysosomal membrane stability showed tendency towards a dose-response pattern. The same markers were assessed in the field study, biomarker analyses were consistent with the contamination level, as evaluated by mussel polycyclic aromatic hydrocarbons body burden. Overall, obtained results confirmed the value of an ecotoxicological approach for a scientifically sound characterisation of biological effects induced by offshore oilfield operational discharges.

Water column monitoring of the biological effects of produced water from the Ekofisk offshore oil installation from 2006 to 2009.

The Norwegian water column monitoring program investigates the biological effects of offshore oil and gas activities in Norwegian waters. In three separate surveys in 2006, 2008, and 2009, bioaccumulation and biomarker responses were measured in mussels (Mytilus edulis) and Atlantic cod (Gadus morhua) held in cages at known distances from the produced water (PW) discharge at the Ekofisk oil field. Identical monitoring studies performed in all three years have allowed the biological effects and bioaccumulation data to be compared, and in addition, enabled the potential environmental benefits of a PW treatment system (CTour), implemented in 2008, to be evaluated. The results of the 2009 survey showed that caged animals were exposed to low levels of PW components, with highest tissue concentrations in mussels located closest to the PW discharge. Mussels located approximately 1-2 km away demonstrated only background concentrations of target compounds. Concentrations of polycyclic aromatic hydrocarbons (PAH) and alkyl phenol (AP) metabolites in the bile of caged cod were elevated at stations 200-250 m from the discharge. There was also a signal of exposure relative to discharge for the biomarkers CYP1A in fish and micronuclei in mussels. All other fish and mussel biomarkers showed no significant exposure effects in 2009. The mussel bioaccumulation data in 2009 indicated a lower exposure to the PW effluent than seen previously in 2008 and 2006, resulting in an associated general improvement in the health of the caged mussels. This was due to the reduction in overall discharge of PW components (measured as oil in water) into the area in 2009 compared to previous years as a result of the improved PW treatment system.

Selenium in water enhances antioxidant defenses and protects against copper-induced DNA damage in the blue mussel Mytilus edulis.

Selenium and copper are naturally occurring elements in the environment that have important roles in cellular function. Selenium is known for its role in antioxidant defense, whereas copper is a redox-active metal capable of acting as a pro-oxidant. We investigated the effects of short term selenium (Na(2)SeO(3)) supplementation (4 µg/L for 3 days) on antioxidant parameters of the blue mussel, Mytilus edulis, and its possible protective effects against a subsequent copper (CuSO(4)) exposure (56 µg/L for 3 days). Selenium supplementation caused a 4-fold increase in glutathione levels in gills. The activity of selenium-dependent glutathione peroxidase was modulated by selenium in gills (2-fold increase) and also in cell-free haemolymph (40% increase). Copper exposure produced decreases in protein thiol levels (35%) and in thioredoxin reductase activity (60%) in gills and induced an increase in DNA damage in haemocytes (70% increase in % tail DNA observed using the comet assay). The decrease in thioredoxin reductase activity may constitute a mechanism of copper toxicity in bivalves, warranting further investigation. Pre-treatment with selenium largely prevented these deleterious effects of copper on protein thiols, thioredoxin reductase activity and DNA damage. The results suggest that induction of key antioxidant defenses such as glutathione and selenium-dependent glutathione peroxidase, as a result of selenium supplementation, may play an important role in protection of aquatic organisms against oxidative stress.

Complications with remediation strategies involving the biodegradation and detoxification of recalcitrant contaminant aromatic hydrocarbons.

Environmentally persistent aromatic hydrocarbons known as unresolved complex mixtures (UCMs) derived from crude oil can be accumulated by, and elicit toxicological responses in, marine organisms (e.g. mussels, Mytilus edulis). Comprehensive two-dimensional gas chromatography time-of-flight mass-spectrometry (GCxGC-ToF-MS) previously revealed that these UCMs included highly branched alkylated aromatic hydrocarbons. Here, the effects of biodegradation on the toxicity and chemical composition of an aromatic UCM hydrocarbon fraction isolated from Tia Juana Pesado (TJP) crude oil were examined. 48h exposure of mussels to the aromatic hydrocarbon fraction (F2) resulted in tissue concentrations of 900microgg(-1) (dry wt.) and approximately 45% decrease in clearance rate. Over 90% of the hydrocarbon burden corresponded to an UCM. Following a 5day recovery period, GCxGC-ToF-MS analysis of the tissues indicated depuration of most accumulated hydrocarbons and clearance rates returned to those observed in controls. To assess the potential of biodegradation to reduce UCM toxicity, TJP F2 was exposed to bacteria isolated from Whitley Bay, UK, for 46days. Mussels exposed to the undegraded TJP F2 from the abiotic control exhibited a reduction in clearance rate comparable with values for the pure crude oil TJP F2. Clearance rates of mussels exposed to biodegraded TJP F2 were statistically similar to seawater controls, suggesting biodegradation had reduced the TJP F2 toxicity. GCxGC-ToF-MS analysis revealed the same compound groups in the tissue of mussels exposed to pure TJP F2, undegraded TJP F2 and biodegraded TJP F2 samples; however >300 fewer compounds were observed in the biodegraded (954 compounds) compared to the undegraded TJP F2 (1261). The compound distributions were markedly different, possibly accounting for the decrease in toxicity. Extraction and analysis of pelleted bacterial cell material revealed that a significant proportion of the TJP F2 had adsorbed onto the cells. Thus extreme care must be taken in interpreting biodegradation data from recalcitrant UCM hydrocarbons.

Physiological-biochemical properties of blue mussel Mytilus edulis adaptation to oil contamination.

Bivalves have a known ability to accumulate different contaminants from ambient water and can therefore serve as bioindicators. The paper analyses certain biochemical and physiological parameters of blue mussels in response to varying oil product concentrations. The heart rate (HR) of blue mussels from the sublittoral zone exposed to different levels of oil products was investigated in a long-term experiment using non-invasive monitoring. A sharp rise in HR was observed at oil concentrations of 8.0 and 38.0 mg/l. A decreasing in mussel HR under the effect of lower concentrations (0.4 and 1.9 mg/l) was significant on the fourth day. Strong fluctuations of the cardiac activity were noted under all concentrations. After 6 days of oil treatment, tissues of the mussels were sampled to determine the total lipid composition. Low concentrations of oil products produced no reliable changes in the lipid composition whereas high concentrations induced significant changes in the ratio of lipid components (cholesterol and phospholipids).

Regional differences in mRNA responses in blue mussels within the Baltic proper.

Mussels (Mytilus sp.) from two regions along the permanent salinity gradient within the Baltic proper were exposed to copper (35 ppb) or petrol (0.3 mL/L) for 10 days and analyzed for mRNA expressions in gill tissue. Expression of mRNAs for the heat shock proteins HSP70 and HSP90 was significantly induced by copper, but not by petrol. For the metallothioneins MT10 and MT20, regional differences in mRNA expressions could be seen. In mussels from the northern Baltic proper, MT20 expression increased 2.8 and 3.4 times, after exposure to copper and petrol, respectively. In contrast, no change could be seen in MT20 expression for mussels from the southern Baltic proper. MT10 showed a peculiar expression not previously described. For some mussels, no expression at all was detected, some showed a weak expression and for some individuals a strong expression could be seen. For the mussels from the southern Baltic proper, the number of individuals with a strong expression of MT10 increased from 1 out of 18 (control), to 7 and 8, after exposure to copper and petrol, respectively. The results clearly show that responses vary between different regions within the Baltic proper, which emphasises the importance to study interactions between contaminants, populations and regions.

The metal-binding features of the recombinant mussel Mytilus edulis MT-10-IV metallothionein.

In contrast with the paradigmatic mammalian metallothioneins (MTs), mollusc MT systems consist at least of a high-cadmium induced form, possibly involved in detoxification, and another isoform either constitutive or regulated by essential metals and probably associated with housekeeping metabolism. With the aim of providing a deeper characterization of the coordination features of a molluscan MT peptide of the latter kind, we have analyzed here the metal-binding abilities of the recombinant MeMT-10-IV isoform of Mytilus edulis (MeMT). Also, comparison with other MTs of this type has been undertaken. A synthetic complementary DNA was constructed, cloned and expressed into two Escherichia coli systems. Upon zinc coordination, MeMT folds in vivo into highly chiral and stable Zn(7) complexes, with an exceptional reluctance to fully substitute cadmium(II) and/or copper(I) for zinc(II). In vivo cadmium binding leads to homometallic Cd(7) complexes that structurally differ from any of the in vitro prepared Cd(7) complexes. Homometallic Cu-MeMT can only be obtained in vitro from Zn(7)-MeMT after a great molar excess of copper(I) has been added. In vivo, two different heterometallic Zn,Cu-MeMT complexes are recovered, which nicely correspond to two distinct stages of the in vitro zinc/copper replacement. These MeMT metal-binding features are consistent with a physiological role related to basal/housekeeping metal, mainly zinc, metabolism, and confirm the correspondence between the MeMT gene response pattern and the functional properties of the encoded protein.

TENS stimulates constitutive nitric oxide release via opiate signaling in invertebrate neural tissues.

BACKGROUND: There is a major societal concern relating to the addictive properties of analgesic drugs such as morphine with regard to alleviating pain. Because of this, alternative methods of pain relief are, and have been, actively pursued. An extremely promising method for treatment of low to moderate levels of chronic pain in humans is transcutaneous electrical nerve stimulation (TENS). MATERIAL/METHODS: All experiments utilized the invertebrate marine bivalve mollusc Mytilus edulis pedal ganglia. TENS was achieved using a stimulation apparatus developed by Professor Han of Peking University. TENS experiments employed 2 stimulation protocols: 1) low 2 Hz frequency at 5 mA current, 2) alternating low and high frequencies at 2 and 100 Hz, respectively at 5 mA current. Real-time measurements of nitric oxide (NO), using an amperometric probe, measured NO released into the tissue bath subsequent to TENS. RESULTS: Pooled M. edulis pedal ganglia exposed to TENS demonstrate that stimulation at 2 Hz and 5 mA current promotes time-dependent release of NO. In another experiment, pooled ganglia were stimulated at alternating frequencies of 2 Hz and 100 Hz and 5 mA, which also released NO in a time-dependent manner. Unstimulated control ganglia did not release significant amounts of NO. NO release was antagonized by naloxone and L-NAME exposure, demonstrating that it was receptor and nitric oxide synthase mediated, respectively. CONCLUSIONS: It would appear that TENS stimulates endogenous morphine release since NO release was blocked by naloxone and opioid peptides do not release NO. The present study is highly suggestive of the occurrence of this same mechanism in mammalian neural systems since all biochemical and signaling components are present. Furthermore, it would appear that this process has evolutionary survival value since it occurs in an animal that evolved 500 million years ago.

The potential of ecotoxicoproteomics in environmental monitoring: biomarker profiling in mussel plasma using ProteinChip array technology.

New global technologies, allowing simultaneous analysis of thousands of genes, proteins, and metabolites (so-called "omics" technologies), are being adopted rapidly by industry, academia, and regulatory agencies. This study evaluated the potential of proteomics in ecotoxicological research (i.e., ecotoxicoproteomics). Filter-feeding mussels (Mytilus edulis) were exposed continuously for 3 wk to oil, or oil spiked with alkylphenols and extra polycyclic aromatic hydrocarbons. The influence of chronic exposure on mussel plasma protein expression was investigated utilizing ProteinChip array technology in combination with surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI TOF MS). Results indicated that exposure to spiked oil had a more significant effect on protein expression in mussels than oil alone. In total, 83 mass peaks (intact or modified proteins/peptides) were significantly altered by spiked oil, while 49 were altered by oil. In exposed organisms, the majority of peaks were upregulated compared to controls (i.e., 69% in oil and 71% in spiked oil). Some peaks (32 in total) were affected by both treatments; however, the degree of response was higher in the spiked oil group for 25 of the 32 commonly affected features. Additionally, certain peaks revealed exposure- or gender-specific responses. Multivariate analysis with regression tree-based methods detected protein patterns associated with exposure that correctly classified masked samples with 90-95% accuracy. Similarly, 92% of females and 85% of males were correctly classified (independent of exposure). Results indicate that proteomics have the potential to make a valuable contribution to environmental monitoring and risk assessment.

Structural characterization of the major extrapallial fluid protein of the mollusc Mytilus edulis: implications for function.

The major protein component of the extrapallial fluid of the mollusc Mytilus edulis has been previously isolated and partially characterized. It was postulated to play a role in shell mineralization because of its intriguing property of Ca(2+)-binding-induced self-assembling. However, it also binds other divalent ions, including Cd(2+), Cu(2+), Mn(2+), and Mg(2+). Herein is the initial report on the characterization of the primary structure of the extrapallial (EP) protein by RT-PCR and cDNA sequencing methods and by de novo peptide sequencing with mass spectrometry. The EP protein is comprised of 213 amino acids postcleavage of a signal peptide of 23 amino acids. The protein is rich in His, Glu, and Asp residues. The site of N-glycosylation, "NHTE", at amino acid positions 54-57 and the intramolecular disulfide bond between Cys 139 and Cys 171 of the protein have been characterized also. Sequence comparisons reveal that the EP protein possesses little homology to any presently known matrix proteins previously isolated from mollusc shells but rather it highly resembles a heavy metal binding protein and a histidine-rich glycoprotein, both from the hemolymph of M. edulis. The predicted domain profile and amino acid composition suggest that its N-terminus may be involved in calcium binding. The abundance of histidine residues of the protein may account for its heavy metal binding properties. Thus, the EP protein perhaps has multiple functions, serving as a Ca(2+)-transport protein, a shell matrix protein, and a heavy metal detoxification protein.