Human skeletal muscle metabolic responses to 6 days of high-fat overfeeding are associated with dietary n-3PUFA content and muscle oxidative capacity.

Understanding human physiological responses to high-fat energy excess (HFEE) may help combat the development of metabolic disease. We aimed to investigate the impact of manipulating the n-3PUFA content of HFEE diets on whole-body and skeletal muscle markers of insulin sensitivity. Twenty healthy males were overfed (150% energy, 60% fat, 25% carbohydrate, 15% protein) for 6 d. One group (n = 10) received 10% of fat intake as n-3PUFA rich fish oil (HF-FO), and the other group consumed a mix of fats (HF-C). Oral glucose tolerance tests with stable isotope tracer infusions were conducted before, and following, HFEE, with muscle biopsies obtained in basal and insulin-stimulated states for measurement of membrane phospholipids, ceramides, mitochondrial enzyme activities, and PKB and AMPKα2 activity. Insulin sensitivity and glucose disposal did not change following HFEE, irrespective of group. Skeletal muscle ceramide content increased following HFEE (8.5 ± 1.2 to 12.1 ± 1.7 nmol/mg, p = .03), irrespective of group. No change in mitochondrial enzyme activity was observed following HFEE, but citrate synthase activity was inversely associated with the increase in the ceramide content (r=-0.52, p = .048). A time by group interaction was observed for PKB activity (p = .003), with increased activity following HFEE in HF-C (4.5 ± 13.0mU/mg) and decreased activity in HF-FO (-10.1 ± 20.7 mU/mg) following HFEE. Basal AMPKα2 activity increased in HF-FO (4.1 ± 0.6 to 5.3 ± 0.7mU/mg, p = .049), but did not change in HF-C (4.6 ± 0.7 to 3.8 ± 0.9mU/mg) following HFEE. We conclude that early skeletal muscle signaling responses to HFEE appear to be modified by dietary n-3PUFA content, but the potential impact on future development of metabolic disease needs exploring.

Influence of Fish Oil-Derived n-3 Fatty Acid Supplementation on Changes in Body Composition and Muscle Strength During Short-Term Weight Loss in Resistance-Trained Men.

Background: A detrimental consequence of diet-induced weight loss, common in athletes who participate in weight cutting sports, is muscle loss. Dietary omega-3 polyunsaturated fatty acids (n-3PUFA) exhibit a protective effect on the loss of muscle tissue during catabolic situations such as injury-simulated leg immobilization. This study aimed to investigate the influence of dietary n-3PUFA supplementation on changes in body composition and muscle strength following short-term diet-induced weight loss in resistance-trained men. Methods: Twenty resistance-trained young (23 ± 1 years) men were randomly assigned to a fish oil group that supplemented their diet with 4 g n-3PUFA, 18 g carbohydrate, and 5 g protein (FO) or placebo group containing an equivalent carbohydrate and protein content (CON) over a 6 week period. During weeks 1-3, participants continued their habitual diet. During week 4, participants received all food items to control energy balance and a macronutrient composition of 50% carbohydrate, 35% fat, and 15% protein. During weeks 5 and 6, participants were fed an energy-restricted diet equivalent to 60% habitual energy intake. Body composition and strength were measured during weeks 1, 4, and 6. Results: The decline in total body mass (FO = -3.0 ± 0.3 kg, CON = -2.6 ± 0.3 kg), fat free mass (FO = -1.4 ± 0.3 kg, CON = -1.2 ± 0.3 kg) and fat mass (FO = -1.4 ± 0.2 kg, CON = -1.3 ± 0.3 kg) following energy restriction was similar between groups (all p > 0.05; d: 0.16-0.39). Non-dominant leg extension 1 RM increased (6.1 ± 3.4%) following energy restriction in FO (p < 0.05, d = 0.29), with no changes observed in CON (p > 0.05, d = 0.05). Dominant leg extension 1 RM tended to increase following energy restriction in FO (p = 0.09, d = 0.29), with no changes in CON (p > 0.05, d = 0.06). Changes in leg press 1 RM, maximum voluntary contraction and muscular endurance following energy restriction were similar between groups (p > 0.05, d = 0.05). Conclusion: Any possible improvements in muscle strength during short-term weight loss with n-3PUFA supplementation are not related to the modulation of FFM in resistance-trained men.

Adding Fish Oil to Whey Protein, Leucine, and Carbohydrate Over a Six-Week Supplementation Period Attenuates Muscle Soreness Following Eccentric Exercise in Competitive Soccer Players.

Soccer players often experience eccentric exercise-induced muscle damage given the physical demands of soccer match-play. Since long chain n-3 polyunsaturated fatty acids (n-3PUFA) enhance muscle sensitivity to protein supplementation, dietary supplementation with a combination of fish oil-derived n-3PUFA, protein, and carbohydrate may promote exercise recovery. This study examined the influence of adding n-3PUFA to a whey protein, leucine, and carbohydrate containing beverage over a six-week supplementation period on physiological markers of recovery measured over three days following eccentric exercise. Competitive soccer players were assigned to one of three conditions (2 × 200 mL): a fish oil supplement beverage (FO; n = 10) that contained n-3PUFA (1100 mg DHA/EPA-approximately 550 mg DHA, 550 mg EPA), whey protein (15 g), leucine (1.8 g), and carbohydrate (20 g); a protein supplement beverage (PRO; n = 10) that contained whey protein (15 g), leucine (1.8 g), and carbohydrate (20 g); and a carbohydrate supplement beverage (CHO; n = 10) that contained carbohydrate (24 g). Eccentric exercise consisted of unilateral knee extension/flexion contractions on both legs separately. Maximal force production was impaired by 22% during the 72-hour recovery period following eccentric exercise (p < 0.05). Muscle soreness, expressed as area under the curve (AUC) during 72-hour recovery, was less in FO (1948 ± 1091 mm × 72 h) than PRO (4640 ± 2654 mm × 72 h, p < 0.05) and CHO (4495 ± 1853 mm × 72 h, p = 0.10). Blood concentrations of creatine kinase, expressed as AUC, were ~60% lower in FO compared to CHO (p < 0.05) and tended to be lower (~39%, p = 0.07) than PRO. No differences in muscle function, soccer performance, or blood c-reactive protein concentrations were observed between groups. In conclusion, the addition of n-3PUFA to a beverage containing whey protein, leucine, and carbohydrate ameliorates the increase in muscle soreness and blood concentrations of creatine kinase following eccentric exercise in competitive soccer players.

Lipid Composition of Oil Extracted from Wasted Norway Lobster (Nephrops norvegicus) Heads and Comparison with Oil Extracted from Antarctic Krill (Euphasia superba).

In the UK, the Norway lobster (Nephrops norvegicus) supports its most important shellfish fishery. Nephrops are sold either whole, or as "tails-only" for the scampi trade. In the "tailing" process, the "head" (cephalothorax) is discarded as waste. A smaller crustacean species, the Antarctic krill Euphasia superba, represents an economically valuable industry, as its extractable oil is sold as a human dietary supplement. The aim of this study was to determine the amount and composition of the oil contained in discarded Nephrops heads and to compare its composition to the oil extracted from krill. Differences due to Geographical variation and seasonal patterns in the amount and composition of lipid were also noted. Results indicated that Nephrops head waste samples collected from more southern locations in Scotland (Clyde Sea area) contained higher levels of oil when compared to samples collected from northern locations in Iceland. Moreover, seasonal differences within the Clyde Sea area in Scotland were also observed, with oil extracted from Nephrops head waste peaking at around 11.5% during the summer months when larger and more mature females were caught by trawl. At this time of the year, the valuable fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) accounted for around 23% of the total fatty acid content in oil extracted from Nephrops head waste. A seasonal effect on EPA content was found, with higher levels obtained in the summer, while no trend was found in DHA percentages. Finally, oil from Nephrops head waste contained a higher proportion of EPA and DHA than krill oil but these fatty acids were more abundantly linked to the neutral lipids rather to than polar lipids. The characterization of lipid that could be extracted from Nephrops head waste should be seen as a first step for the commercial use of a valuable resource currently wasted. This approach is extremely relevant given the current limited supply of EPA and DHA and changes in the Common Fisheries Policy.

Fish oil supplementation suppresses resistance exercise and feeding-induced increases in anabolic signaling without affecting myofibrillar protein synthesis in young men.

Fish oil (FO) supplementation potentiates muscle protein synthesis (MPS) in response to a hyperaminoacidemic-hyperinsulinemic infusion. Whether FO supplementation potentiates MPS in response to protein ingestion or when protein ingestion is combined with resistance exercise (RE) remains unknown. In a randomized, parallel group design, 20 healthy males were randomized to receive 5 g/day of either FO or coconut oil control (CO) for 8 weeks. After supplementation, participants performed a bout of unilateral RE followed by ingestion of 30 g of whey protein. Skeletal muscle biopsies were obtained before and after supplementation for assessment of muscle lipid composition and relevant protein kinase activities. Infusion of L-[ring-(13)C6] phenylalanine was used to measure basal myofibrillar MP Sat rest (REST), in a nonexercised leg following protein ingestion (FED) and following RE and protein ingestion (FEDEX).MPS was significantly elevated above REST during FEDEX in both the FO and CO groups, but there was no effect of supplementation. There was a significant increase in MPS in both groups above REST during FED but no effect of supplementation. Supplementation significantly decreased pan PKB activity at RESTin the FO group but not the CO group. There was a significant increase from REST at post-RE for PKB and AMPKα2 activity in the CO group but not in the FO group. In FEDEX, there was a significant increase in p70S6K1 activity from REST at 3 h in the CO group only. These data highlight that 8 weeks of FO supplementation alters kinase signaling activity in response to RE plus protein ingestion without influencing MPS.

A novel SOD1-ALS mutation separates central and peripheral effects of mutant SOD1 toxicity.

Transgenic mouse models expressing mutant superoxide dismutase 1 (SOD1) have been critical in furthering our understanding of amyotrophic lateral sclerosis (ALS). However, such models generally overexpress the mutant protein, which may give rise to phenotypes not directly relevant to the disorder. Here, we have analysed a novel mouse model that has a point mutation in the endogenous mouse Sod1 gene; this mutation is identical to a pathological change in human familial ALS (fALS) which results in a D83G change in SOD1 protein. Homozgous Sod1(D83G/D83G) mice develop progressive degeneration of lower (LMN) and upper motor neurons, likely due to the same unknown toxic gain of function as occurs in human fALS cases, but intriguingly LMN cell death appears to stop in early adulthood and the mice do not become paralyzed. The D83 residue coordinates zinc binding, and the D83G mutation results in loss of dismutase activity and SOD1 protein instability. As a result, Sod1(D83G/D83G) mice also phenocopy the distal axonopathy and hepatocellular carcinoma found in Sod1 null mice (Sod1(-/-)). These unique mice allow us to further our understanding of ALS by separating the central motor neuron body degeneration and the peripheral effects from a fALS mutation expressed at endogenous levels.

Temporal changes in human skeletal muscle and blood lipid composition with fish oil supplementation.

The aim of this study was to examine changes in the lipid profile of red blood cells and muscle tissue along with the expression of anabolic signalling proteins in human skeletal muscle. Following a 2-week control period, 10 healthy male participants consumed 5 g d(-1) of fish oil (FO) for 4 weeks. Muscle biopsies and venous blood samples were collected in the fasted state 2 weeks prior (W-2) and immediately before (W0) the initiation of FO supplementation for internal control. Muscle biopsies and venous blood samples were again obtained at week 1 (W1), 2 (W2) and 4 (W4) during FO supplementation for assessment of changes in lipid composition and expression of anabolic signalling proteins. There was no change in the composition of any lipid class between W-2 and W0 confirming control. Following FO supplementation n-3 polyunsaturated fatty acid (n-3 PUFA) muscle lipid composition was increased from W0 to W2 and continued to rise at W4. n-3 PUFA blood lipid composition was increased from W0 to W1 and remained elevated for the remaining time points. Total protein content of focal adhesion kinase (FAK) increased from W0 to W4 whereas total mechanistic target of rapamycin (mTOR) was increased from W0 at W1 with no further significant increases at W2 and W4. These data show that FO supplementation results in discordant changes in the n-3 PUFA composition of skeletal muscle compared to blood that is associated with increases in total FAK content.

Influence of the dietary protein:lipid ratio and fish oil substitution on fatty acid composition and metabolism of Atlantic salmon (Salmo salar) reared at high water temperatures.

A factorial, two-way, experimental design was used for this 10-week nutritional trial, aiming to elucidate the interactive effects of decreasing dietary protein:lipid level and substitution of fish oil (FO) with rapeseed oil (RO) on tissue fatty acid (FA) composition and metabolism of large Atlantic salmon (Salmo salar L.) reared at high water temperatures (sub-optimal, summer temperatures: 11·6°C). The six experimental diets were isoenergetic and formulated to include either FO or RO (60 % of the added oil) at three dietary protein:lipid levels, specifically (1) 350 g/kg protein and 350 g/kg lipid, (2) 330 g/kg protein and 360 g/kg lipid, (3) 290 g/kg protein and 380 g/kg lipid. Final weight, specific growth rate and thermal growth coefficient were positively affected by the dietary RO inclusion at the expense of FO, while no significant effects were seen on growth due to the decreasing protein level. The oil source had a significant effect on muscle and liver FA composition. However, the changes in muscle and liver FA indicate selective utilisation or retention of individual FA and moderate reductions in tissue EPA and DHA. Pyloric caeca phospholipid FA composition was significantly affected by the two factors and, in some cases, significant interactions were also revealed. Liver and red muscle ß-oxidation capacities were significantly increased due to RO inclusion, while an interactive effect of protein level and oil source was shown for white muscle ß-oxidation capacity. The results could explain, at least partially, the better performance that was shown for the RO groups and the enhanced protein-sparing effect.

Effects of decontaminated fish oil or a fish and vegetable oil blend on persistent organic pollutant and fatty acid compositions in diet and flesh of Atlantic salmon ( Salmo salar).

The health benefits of seafood are well documented and based on the unique supply of n-3 highly unsaturated fatty acids (HUFA). Aquaculture now contributes about 50 % of food-grade seafood globally and Atlantic salmon (Salmo salar) is a rich source of n-3 HUFA. However, salmon and other oily fish can accumulate lipophilic persistent organic pollutants (POP), including dioxins (PCDD/F), polychlorinated biphenyls (PCB) and polybrominated diphenyl ethers (PBDE), derived largely from feed. In the present study, triplicate groups of salmon, of initial weight 0.78 kg, were fed one of three experimental diets for 11 weeks. The diets were coated with either a northern fish oil (FO) with a high POP content (cNFO), the same oil that had been decontaminated (deNFO) or a blend of southern fish oil, rapeseed and soyabean oils (SFO/RO/SO). Dietary PCDD/F+dioxin-like PCB (DL-PCB) concentrations were 17.36, 0.45 and 0.53 ng toxic equivalents (TEQ)/kg, respectively. After 11 weeks, the flesh concentrations in fish fed the cNFO, deNFO and SFO/RO/SO diets were 6.42, 0.34 and 0.41 ng TEQ/kg, respectively. There were no differences in flesh EPA and DHA between fish fed the cNFO or deNFO diets although EPA and DHA were reduced by 50 and 30 %, respectively, in fish fed the SFO/RO/SO diet. Thus, decontaminated FO can be used to produce salmon high in n-3 HUFA and low in POP. Salmon produced using deNFO would be of high nutritional value and very low in POP and would utilise valuable fish oils that would otherwise be destroyed due to their high pollutant concentrations.

The fatty acid compositions of erythrocyte and plasma polar lipids in children with autism, developmental delay or typically developing controls and the effect of fish oil intake.

The erythrocyte and plasma fatty acid compositions of children with autism were compared in a case-control study with typically developing (TD) children and with children showing developmental delay (DD). Forty-five autism subjects were age-matched with TD controls and thirty-eight with DD controls. Fatty acid data were compared using paired t tests. In addition, blood fatty acids from treatment-naive autism subjects were compared with autism subjects who had consumed fish oil supplements by two-sample t tests. Relatively few differences were seen between erythrocyte fatty acids in autism and TD subjects although the former had an increased arachidonic acid (ARA):EPA ratio. This ratio was also increased in plasma samples from the same children. No changes in n-3 fatty acids or ARA:EPA ratio were seen when comparing autism with DD subjects but some SFA and MUFA were decreased in the DD subjects, most notably 24 : 0 and 24 : 1, which are essential components of axonal myelin sheaths. However, if multiple comparisons are taken into account, and a stricter level of significance applied, most of these values would not be significant. Autism subjects consuming fish oil showed reduced erythrocyte ARA, 22 : 4n-6, 22 : 5n-6 and total n-6 fatty acids and increased EPA, 22 : 5n-3, 22 : 6n-3 and total n-3 fatty acids along with reduced n-6:n-3 and ARA:EPA ratios. Collectively, the autism subjects did not have an underlying phospholipid disorder, based on erythrocyte fatty acid compositions, although the increased ARA:EPA ratio observed suggested that an imbalance of essential highly unsaturated fatty acids may be present in a cohort of autism subjects.

Detection of common carotid artery stenosis using duplex ultrasonography: a validation study with computed tomographic angiography.

BACKGROUND: Severe stenosis of the common carotid artery (CCA), while uncommon, is associated with increased risk of transient ischemic attack and stroke. To date, no validated duplex ultrasound criteria have been established for grading the severity of CCA stenosis. The goal of this study was to use receiver-operating curve (ROC) analysis with computed tomographic angiography as the reference standard to establish duplex ultrasound criteria for diagnosing >or=50% CCA stenosis. METHODS: The study cohort included 64 patients (42 men, 22 women) with a mean age of 65 +/- 12 years (range, 16-89 years) who had CCA peak systolic velocity (PSV) >or=150 cm/sec and underwent computed tomographic angiography (CTA) of the cervical and intracerebral vessels within 1 month of the duplex examination. One study was excluded because the CTA was technically inadequate, whereas another was excluded because the patient underwent bilateral CCA stenting. The CCA ipsilateral to any of the following was excluded from the analysis: innominate artery occlusion (n = 1), previous stenting of the ICA or CCA (n = 7), carotid endarterectomy (n = 1), or carotid-to-carotid bypass (n = 1). Thus, the data set included 62 patients and 115 vessels. Bland-Altman analysis was used to examine the agreement between two measures of luminal reduction measured by CTA: percent diameter stenosis and percent area stenosis. Receiver operating characteristic (ROC) analysis was used to determine optimal PSV and EDV thresholds for diagnosing >or=50% CCA stenosis. RESULTS: Severity of CCA stenosis was <50% in 76 vessels, 50%-59% in eight, 60%-69% in eight, 70%-79% in nine, 80%-89% in three, 90%-99% in five, and occluded in six. Duplex ultrasonography identified six of six (100%) patients with 100% CCA occlusion by CTA. Bland-Altman analysis showed poor agreement between percent stenosis determined by vessel diameter compared with percent stenosis determined by reduction in lumen area. Therefore, subsequent analysis was performed using percent stenosis by area. ROC analysis of different PSV thresholds for detecting stenosis >or=50% showed that >182 cm/sec was the most accurate with a sensitivity of 64% and specificity of 88% (P < .0001). Sensitivity, specificity, and accuracy of carotid duplex were higher when the stenosis was located in the mid or distal aspects of the CCA (sensitivity 76%, specificity 89%, area under curve 0.84, P < .001) than in the intrathoracic and proximal segment of the artery (P = NS). ROC analysis of different EDV thresholds for detecting CCA stenosis >or=50% showed that >30 cm/sec was the most accurate with a sensitivity of 54% and a specificity of 74% (P < .0239). CONCLUSIONS: Duplex ultrasonography is highly sensitive, specific, and accurate for detecting CCA lesions in the mid and distal CCA. Use of peak systolic velocity may lead to improved detection of CCA disease and initiation of appropriate therapy to reduce the risk of stroke.

Absence of mycobacterium avium subsp. paratuberculosis in Crohn's patients.

BACKGROUND: Mycobacterium avium subspecies paratuberculosis (MAP) has been suspected of involvement in Crohn's disease (CD). We investigated this potential association by testing whole blood from CD patients and healthy controls for the presence of MAP by culture and molecular methods. In addition, each blood sample was analyzed for polymorphisms in the NOD2/CARD15 gene previously associated with CD. METHODS: Four 4-mL K(2)-EDTA tubes of whole blood were drawn from each subject (n = 260, 130 CD patients and 130 healthy controls). Two tubes of blood were cultured for MAP by the following methods: Mycobacterial Growth Indicator Tube, Herrold's Egg Yolk Agar, BACTEC 460, and Hungate. The remaining 2 tubes of blood were tested for MAP DNA and polymorphisms in the NOD2/CARD15 gene by polymerase chain reaction (PCR). RESULTS: One healthy control patient was positive for MAP via PCR; however, no viable MAP was cultured from this individual. All blood cultures were negative for MAP. One CD patient's blood was culture-positive for M. tuberculosis complex. CD patients exhibited a higher rate of polymorphism in the NOD2/CARD15 gene than healthy control patients. CONCLUSIONS: In this study MAP was not recovered from the blood of CD patients or healthy controls. However, CD patients showed higher mutation rates in the NOD2/CARD15 gene, compared with healthy controls, supporting the findings of other investigators. No correlation between these polymorphisms and MAP bacteremia in CD patients could be identified in this study.

Antimicrobial efficacy of riboflavin/UVA combination (365 nm) in vitro for bacterial and fungal isolates: a potential new treatment for infectious keratitis.

PURPOSE: To demonstrate the antimicrobial properties of riboflavin/UVA (365 nm) against common pathogens. METHODS: One group of bacteria (Pseudomonas aeruginosa [PA], Staphylococcus aureus [SA], and Staphylococcus epidermidis [SE]) was tested by using Kirby-Bauer discs with (1) empty disc (Control - C); (2) riboflavin 0.1% (B2); (3) riboflavin 0.1% previously activated by UVA (B2'); (4) UVA alone (UVA); (5) group 2+additional UVA exposure (UVA+B2); and (6) group 3+additional UVA exposure (UVA+B2'). In addition, another group of microbes was tested with the same approach: methicillin-resistant S. aureus (MRSA), multidrug-resistant P. aeruginosa (MDRPA), drug-resistant Streptococcus pneumoniae (DRSP), and Candida albicans (CA). The mean growth inhibition zone (GIZ) in square millimeters was measured around the discs. The mean standard deviation (MSD) was calculated to be 3.65 when alpha = 0.01. A mean deviation (MD) > MSD indicates a significant difference. RESULTS: In the first group, the GIZ was significantly greater after UVA (MD = 14.30), UVA+B2 (MD = 39.61), and UVA+B2' (MD = 40.45) when compared with C, B2, and B2'. UVA alone was less effective than UVA+B2 (MD = 25.31) and UVA+B2' (MD = 26.15). The second group demonstrated increased GIZ in UVA (MD = 6.98), UVA+B2 (MD = 17.80), and UVA+B2' (MD = 21.15) when compared with C, B2, and B2'. UVA alone was less effective against the second group of bacteria than was UVA+B2 (MD = 10.82) and UVA+B2' (MD = 14.17). CA did not show any GIZ after treatment. CONCLUSIONS: Riboflavin/UVA was effective against SA, SE, PA, MRSA, MDRPA, and DRSP, but was ineffective on CA and has the potential for use in treatment of microbial keratitis in the future.

A molecular analysis of prokaryotic and viral DNA sequences in prostate tissue from patients with prostate cancer indicates the presence of multiple and diverse microorganisms.

BACKGROUND: Inflammation, both acute and chronic, is a common feature of prostate histology. While inflammation has been proposed to play an important role in both benign and malignant growth of the prostate, the stimuli for this inflammation remain poorly characterized. Infectious pathogens are potential stimuli for prostatic inflammation. METHODS: Universal eubacterial PCR was used to test 170 prostate tissue core samples from 30 cancer patients for 16S rDNA gene sequences. Positive PCR products (n=64, 37%) were cloned and sequenced. For comparison, tissue samples from 30 patients were cultured using standard clinical microbiological techniques. DNA samples from 200 additional patients were tested by organism-specific PCR for the presence of Chlamydia trachomatis, Propionibacterium acnes, Trichomonas vaginalis, BK virus, Epstein-Barr virus, human cytomegalovirus, human papillomavirus, and xenotropic murine leukemia-related virus. RESULTS: 16S sequencing results indicated the presence of 83 distinct microorganisms. Microbiological culture isolated markedly fewer species. In general, organism-specific PCR failed to detect multiple organisms previously reported as common in the prostate. There was no significant association between the presence of particular species of bacteria and histologic evidence of acute or chronic inflammation. CONCLUSIONS: Most prostates from men undergoing prostatectomy (87%) contain bacterial DNA from one or more species. However, the majority of individual tissue core samples were negative, suggesting regional heterogeneity in the presence of bacteria and a lack of a generalized or ubiquitous prostatic flora. Culture results suggest either the "unculturable" nature of species present in the prostate or that 16S rDNA sequences were derived from non-viable bacteria.

Influence of dietary oil content and conjugated linoleic acid (CLA) on lipid metabolism enzyme activities and gene expression in tissues of Atlantic salmon (Salmo salar L.).

The overall objective is to test the hypothesis that conjugated linoleic acid (CLA) has beneficial effects in Atlantic salmon as a result of affecting lipid and FA metabolism. The specific aims of the present study were to determine the effects of CLA on some key pathways of FA metabolism, including FA oxidation and highly unsaturated FA (HUFA) synthesis. Salmon smolts were fed diets containing two levels of fish oil (low, approximately 17%, and high, approximately 34%) containing three levels of CLA (a 1:1 mixture of cis-9,trans-11 and trans-10,cis-12 at 0, 1, and 2% of diet) for 3 mon. The effects of dietary CLA on HUFA synthesis and beta-oxidation were measured, and the expression of key genes in the FA oxidation and HUFA synthesis pathways, and the potentially important transcription factors peroxisome proliferators activated receptors (PPAR), were determined in selected tissues. Liver HUFA synthesis and desaturase gene expression was increased by dietary CLA and decreased by high dietary oil content. Carnitine palmitoyltransferase-I (CPT-I) activity and gene expression were generally increased by CLA in muscle tissues although they were relatively unaffected by dietary oil content. In general CPT-I activity or gene expression was not correlated with P-oxidation. Dietary CLA tended to increase PPARalpha and beta gene expression in both liver and muscle tissues, and PPARgamma in liver. In summary, gene expression and activity of the FA pathways were altered in response to dietary CLA and/or oil content, with data suggesting that PPAR are also regulated in response to CLA. Correlations were observed between dietary CLA, liver HUFA synthesis and desaturase gene expression, and liver PPARalpha expression, and also between dietary CLA, CPT-I expression and activity, and PPARalpha expression in muscle tissues. In conclusion, this study suggests that dietary CLA has effects on FA metabolism in Atlantic salmon and on PPAR transcription factors. However, further work is required to assess the potential of CLA as a dietary supplement, and the role of PPAR in the regulation of lipid metabolism in fish.

Aqueous penetration and biological activity of moxifloxacin 0.5% ophthalmic solution and gatifloxacin 0.3% solution in cataract surgery patients.

PURPOSE: To measure the achievable perioperative aqueous concentration of the commercially available topically administered fourth generation fluoroquinolones, moxifloxacin 0.5% ophthalmic solution, and gatifloxacin 0.3% ophthalmic solution, and to correlate this concentration with the agents' biological efficacy in the aqueous humor of patients undergoing routine cataract surgery. DESIGN: Prospective, randomized, parallel, double-masked, clinical trial. PARTICIPANTS: Fifty patients undergoing cataract extraction. METHODS: Patients (n = 25) were given perioperative topical moxifloxacin 0.5% or topical gatifloxacin 0.3% (n = 25). One drop of antibiotic was administered every 10 minutes for 4 doses beginning 1 hour prior to surgery. Aqueous humor was sampled via paracentesis and antibiotic concentrations were determined using validated high performance liquid chromatography (HPLC) procedures. Dilution analyses were performed to determine the biological efficacy of the agents in the aqueous against Staphylococcus epidermidis, the most common cause of postcataract endophthalmitis. MAIN OUTCOME MEASURES: Aqueous humor antibiotic concentrations were measured using HPLC and microdilution bioassay techniques. Biological activity was measured as minimal inhibitory dilution and minimal bactericidal dilution. RESULTS: Aqueous humor concentrations for moxifloxacin via HPLC analysis were 1.80 (+/-1.21) microg/ml, whereas those for gatifloxacin were 0.48 (+/-0.34) microg/ml. This 3.8-fold difference in aqueous humor antibiotic concentrations was statistically significant (P = 0.00003). Similarly, the biological dilution analysis of the aqueous humor samples showed that moxifloxacin attained an estimated activity of 2.1 microg/ml, whereas the gatifloxacin activity was approximately 0.4 mug/ml, which represented a 4.9-fold difference. CONCLUSIONS: This study demonstrated that after topically administered perioperative antibiotics with cataract surgery, moxifloxacin 0.5% ophthalmic solution achieved a statistically significantly higher concentration in aqueous humor compared with gatifloxacin (P = 0.00003). Results from the broth dilution analysis showed that moxifloxacin 0.5% was biologically more active against S. epidermidis than gatifloxacin 0.3% in aqueous humor after topical application. There were no adverse events reported, and incision wounds healed quickly and as expected.

Fatty acid metabolism in Atlantic salmon (Salmo salar L.) hepatocytes and influence of dietary vegetable oil.

Isolated hepatocytes from Atlantic salmon (Salmo salar), fed diets containing either 100% fish oil or a vegetable oil blend replacing 75% of the fish oil, were incubated with a range of seven (14)C-labelled fatty acids. The fatty acids were [1-(14)C]16:0, [1-(14)C]18:1n-9, 91-(14)C]18:2n-6, [1-(14)C]18:3n-3, [1-(14)C]20:4n-6, [1-(14)C]20:5n-3, and [1-(14)C]22:6n-3. After 2 h of incubation, the hepatocytes and medium were analysed for acid soluble products, incorporation into lipid classes, and hepatocytes for desaturation and elongation. Uptake into hepatocytes was highest with [1-(14)C]18:2n-6 and [1-(14)C]20:5n-3 and lowest with [1-(14)C]16:0. The highest recovery of radioactivity in the cells was found in triacylglycerols. Of the phospholipids, the highest recovery was found in phosphatidylcholine, with [1-(14)C]16:0 and [1-(14)C]22:6n-3 being the most prominent fatty acids. The rates of beta-oxidation were as follows: 20:4n-6>18:2n-6=16:0>18:1n-9>22:6n-3=18:3n-3=20:5n-3. Of the fatty acids taken up by the hepatocytes, [1-(14)C]16:0 and [1-(14)C]18:1n-9 were subsequently exported the most, with the majority of radioactivity recovered in phospholipids and triacylglycerols, respectively. The major products from desaturation and elongation were generally one cycle of elongation of the fatty acids. Diet had a clear effect on the overall lipid metabolism, with replacing 75% of the fish oil with vegetable oil resulting in decreased uptake of all fatty acids and reduced incorporation of fatty acids into cellular lipids, but increased beta-oxidation activity and higher recovery in products of desaturation and elongation of [1-(14)C]18:2n-6 and [1-(14)C]18:3n-3.

A mutation in dynein rescues axonal transport defects and extends the life span of ALS mice.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative condition characterized by motoneuron degeneration and muscle paralysis. Although the precise pathogenesis of ALS remains unclear, mutations in Cu/Zn superoxide dismutase (SOD1) account for approximately 20-25% of familial ALS cases, and transgenic mice overexpressing human mutant SOD1 develop an ALS-like phenotype. Evidence suggests that defects in axonal transport play an important role in neurodegeneration. In Legs at odd angles (Loa) mice, mutations in the motor protein dynein are associated with axonal transport defects and motoneuron degeneration. Here, we show that retrograde axonal transport defects are already present in motoneurons of SOD1(G93A) mice during embryonic development. Surprisingly, crossing SOD1(G93A) mice with Loa/+ mice delays disease progression and significantly increases life span in Loa/SOD1(G93A) mice. Moreover, there is a complete recovery in axonal transport deficits in motoneurons of these mice, which may be responsible for the amelioration of disease. We propose that impaired axonal transport is a prime cause of neuronal death in neurodegenerative disorders such as ALS.

Molecular cloning and functional characterization of fatty acyl desaturase and elongase cDNAs involved in the production of eicosapentaenoic and docosahexaenoic acids from alpha-linolenic acid in Atlantic salmon (Salmo salar).

Fish are the only major dietary source for humans of omega-3 highly unsaturated fatty acids (HUFAs) and with declining fisheries farmed fish such as Atlantic salmon (Salmo salar) constitute an increasing proportion of the fish in the human diet. However, the current high use of fish oils, derived from wild capture marine fisheries, in aquaculture feeds is not sustainable in the longer term and will constrain continuing growth of aquaculture activities. Greater understanding of how fish metabolize and biosynthesize HUFA may lead to more sustainable aquaculture diets. The study described here contributes to an effort to determine the molecular genetics of the HUFA biosynthetic pathway in salmon, with the overall aim being to determine mechanisms for optimizing the use of vegetable oils in Atlantic salmon culture. In this paper we describe the cloning and functional characterization of 2 genes from salmon involved in the biosynthesis of HUFA. A salmon desaturase complementary DNA, SalDes, was isolated that include an open reading frame of 1362 bp specifying a protein of 454 amino acids. The protein sequence includes all the characteristics of microsomal fatty acid desaturases, including 3 histidine boxes, 2 transmembrane regions, and an N-terminal cytochrome b(5) domain containing a heme-binding motif similar to that of other fatty acid desaturases. Functional expression in the yeast Saccharomyces cerevisiae showed SalDes is predominantly an omega-3 delta5 desaturase, a key enzyme in the synthesis of eicosapentaenoic acid (20:5n-3) from alpha-linolenic acid (18:3n-3). The desaturase showed only low levels of delta6 activity toward C(18) polyunsaturated fatty acids. In addition, a fatty acid elongase cDNA, SalElo, was isolated that included an open reading frame of 888 bp, specifying a protein of 295 amino acids. The protein sequence of SalElo included characteristics of microsomal fatty acid elongases, including a histidine box and a transmembrane region. Upon expression in yeast SalElo showed broad substrate specificity for polyunsaturated fatty acids with a range of chain lengths, with the rank order being C(18) > C(20) > C(22). Thus this one polypeptide product displays all fatty acid elongase activities required for the biosynthesis of docosahexaenoic acid (22:6n-3) from 18:3n-3.

Effect of n-octanesulphonylacetamide (OSA) on ATP and protein expression in Mycobacterium bovis BCG.

OBJECTIVE: To determine the effect on BCG of n-octanesulphonylacetamide (OSA), a novel compound of the class beta-sulphonylcarboxamides, which has potent in vitro activity against pathogenic mycobacteria. METHODS AND RESULTS: The effect of OSA in BCG was examined using two-dimensional protein electrophoresis. Treatment of BCG with OSA resulted in overexpression of two proteins identified as the b-subunit of ATP synthase (Rv1306) and a 17 kDa heat shock protein (Rv0251c). [35S]Methionine pulse-labelling revealed that overexpression occurred within as little as 3.5 h post-exposure. These results were confirmed by RT-PCR. ATP levels decreased in OSA-treated BCG at 5 min, and 1, 3 and 24 h, with a 64%, 45%, 54% and 73% reduction in ATP, respectively. Only dicyclohexylcarbodiimide (DCCD), a known ATP synthase inhibitor, had a similar effect. No appreciable difference in ATP level was observed in BCG treated with standard antimycobacterial drugs, additional respiratory chain inhibitors or a fatty acid synthase inhibitor at a comparable time-point. Protein synthesis decreased within 5 min of exposure to OSA (56%), DCCD (74%) and thenoyltrifluoroacetone (TTFA) (77%). Ethanol (2.3%) potentiated the activity of OSA. In contrast, no synergic effect was observed with streptomycin and ethanol. Mycolic acid levels decreased 79% with DCCD, 46% with TTFA, a complex II inhibitor, and 43% with OSA compared with untreated controls. CONCLUSIONS: Our results suggest that OSA may interfere directly or indirectly with ATP synthase and possibly other components of the mycobacterial respiratory chain. These effects may hinder energy production, leading to interruption in the synthesis of large macromolecules including proteins and mycolic acids.