Hydroxyoctadecadienoic acids regulate apoptosis in human THP-1 cells in a PPARγ-dependent manner

Macrophage apoptosis, a key process in atherogenesis, is regulated by oxidation products, including hydroxyoctadecadienoic acids (HODEs). These stable oxidation products of linoleic acid (LA) are abundant in atherosclerotic plaque and activate PPARγ and GPR132. We investigated the mechanisms through which HODEs regulate apoptosis. The effect of HODEs on THP-1 monocytes and adherent THP-1 cells were compared with other C18 fatty acids, LA and α-linolenic acid (ALA). The number of cells was reduced within 24 hours following treatment with 9-HODE (p < 0.01, 30 μM) and 13 HODE (p < 0.01, 30 μM), and the equivalent cell viability was also decreased (p < 0.001). Both 9-HODE and 13-HODE (but not LA or ALA) markedly increased caspase-3/7 activity (p < 0.001) in both monocytes and adherent THP-1 cells, with 9-HODE the more potent. In addition, 9-HODE and 13-HODE both increased Annexin-V labelling of cells (p < 0.001). There was no effect of LA, ALA, or the PPARγ agonist rosiglitazone (1μM), but the effect of HODEs was replicated with apoptosis-inducer camptothecin (10μM). Only 9-HODE increased DNA fragmentation. The pro-apoptotic effect of HODEs was blocked by the caspase inhibitor DEVD-CHO. The PPARγ antagonist T0070907 further increased apoptosis, suggestive of the PPARγ-regulated apoptotic effects induced by 9-HODE. The use of siRNA for GPR132 showed no evidence that the effect of HODEs was mediated through this receptor. 9-HODE and 13-HODE are potent—and specific—regulators of apoptosis in THP-1 cells. Their action is PPARγ-dependent and independent of GPR132. Further studies to identify the signalling pathways through which HODEs increase apoptosis in macrophages may reveal novel therapeutic targets for atherosclerosis.

Randomized controlled trial examining the effects of fish oil and multivitamin supplementation on the incorporation of n-3 and n-6 fatty acids into red blood cells

The present randomized, placebo-controlled, double-blind, parallel-groups clinical trial examined the effects of fish oil and multivitamin supplementation on the incorporation of n-3 and n-6 fatty acids into red blood cells. Healthy adult humans (n = 160) were randomized to receive 6 g of fish oil, 6 g of fish oil plus a multivitamin, 3 g of fish oil plus a multivitamin or a placebo daily for 16 weeks. Treatment with 6 g of fish oil, with or without a daily multivitamin, led to higher eicosapentaenoic acid (EPA) composition at endpoint. Docosahexaenoic acid (DHA) composition was unchanged following treatment. The long chain LC n-3 PUFA index was only higher, compared to placebo, in the group receiving the combination of 6 g of fish oil and the multivitamin. Analysis by gender revealed that all treatments increased EPA incorporation in females while, in males, EPA was only significantly increased by the 6 g fish oil multivitamin combination. There was considerable individual variability in the red blood cell incorporation of EPA and DHA at endpoint. Gender contributed to a large proportion of this variability with females generally showing higher LC n-3 PUFA composition at endpoint. In conclusion, the incorporation of LC n-3 PUFA into red blood cells was influenced by dosage, the concurrent intake of vitamin/minerals and gender.

Predictors of treatment response in young people at ultra-high risk for psychosis who received long-chain omega-3 fatty acids.

Previous efforts in the prospective evaluation of individuals who experience attenuated psychotic symptoms have attempted to isolate mechanisms underlying the onset of full-threshold psychotic illness. In contrast, there has been little research investigating specific predictors of positive outcomes. In this study, we sought to determine biological and clinical factors associated with treatment response, here indexed by functional improvement in a pre-post examination of a 12-week randomized controlled intervention in individuals at ultra-high risk (UHR) for psychosis. Participants received either long-chain omega-3 (ω-3) polyunsaturated fatty acids (PUFAs) or placebo. To allow the determination of factors specifically relevant to each intervention, and to be able to contrast them, both treatment groups were investigated in parallel. Univariate linear regression analysis indicated that higher levels of erythrocyte membrane α-linolenic acid (ALA; the parent fatty acid of the ω-3 family) and more severe negative symptoms at baseline predicted subsequent functional improvement in the treatment group, whereas less severe positive symptoms and lower functioning at baseline were predictive in the placebo group. A multivariate machine learning analysis, known as Gaussian Process Classification (GPC), confirmed that baseline fatty acids predicted response to treatment in the ω-3 PUFA group with high levels of sensitivity, specificity and accuracy. In addition, GPC revealed that baseline fatty acids were predictive in the placebo group. In conclusion, our investigation indicates that UHR patients with higher levels of ALA may specifically benefit from ω-3 PUFA supplementation. In addition, multivariate machine learning analysis suggests that fatty acids could potentially be used to inform prognostic evaluations and treatment decisions at the level of the individual. Notably, multiple statistical analyses were conducted in a relatively small sample, limiting the conclusions that can be drawn from what we believe to be a first-of-its-kind study. Additional studies with larger samples are therefore needed to evaluate the generalizability of these findings.

Fishy business : effect of omega-3 fatty acids on zinc transporters and free zinc availability in human neuronal cells

Omega-3 (ω-3) fatty acids are one of the two main families of long chain polyunsaturated fatty acids (PUFA). The main omega-3 fatty acids in the mammalian body are α-linolenic acid (ALA), docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). Central nervous tissues of vertebrates are characterized by a high concentration of omega-3 fatty acids. Moreover, in the human brain, DHA is considered as the main structural omega-3 fatty acid, which comprises about 40% of the PUFAs in total. DHA deficiency may be the cause of many disorders such as depression, inability to concentrate, excessive mood swings, anxiety, cardiovascular disease, type 2 diabetes, dry skin and so on. On the other hand, zinc is the most abundant trace metal in the human brain. There are many scientific studies linking zinc, especially excess amounts of free zinc, to cellular death. Neurodegenerative diseases, such as Alzheimer's disease, are characterized by altered zinc metabolism. Both animal model studies and human cell culture studies have shown a possible link between omega-3 fatty acids, zinc transporter levels and free zinc availability at cellular levels. Many other studies have also suggested a possible omega-3 and zinc effect on neurodegeneration and cellular death. Therefore, in this review, we will examine the effect of omega-3 fatty acids on zinc transporters and the importance of free zinc for human neuronal cells. Moreover, we will evaluate the collective understanding of mechanism(s) for the interaction of these elements in neuronal research and their significance for the diagnosis and treatment of neurodegeneration.

Short-chain fatty acids increase expression and secretion of stromal cell-derived factor-1 in mouse and human pre-adipocytes

Objective: Stromal cell-derived factor-1 (SDF-1) is expressed in pre-adipocytes but its role is unknown. We investigated butyrate (a histone deacetylase inhibitor - HDACi) and other short-chain fatty acids (SCFA) in the regulation of SDF-1. We further investigated whether effects of SCFA were signalled through G protein-coupled receptors FFA2 and FFA3. Design and Results: SDF-1 mRNA expression and protein secretion were studied in 3T3-L1 cells and human pre-adipocytes. SDF-1 was abundant, with mRNA and protein levels increased by butyrate. This was replicated with acetate and propionate, but not with trichostatin or valproate. Trichostatin inhibited SDF-1 secretion. Pertussis toxin blocked stimulation by butyrate. The order of potency of SCFA in stimulating SDF-1 (C3 > C4 > C2) is consistent with action through FFA3. Silencing the FFA3 gene abolished butyrate-stimulated SDF-1 expression and secretion. FFA3 was expressed in both pre-adipocytes and adipocytes, while FFA2 was expressed in adipocytes only. SDF-1 expression was low in murine macrophage J774.2 cells, while the SDF-1 receptor CXCR4 was absent from 3T3-L1 cells but abundant in J774.2 macrophages. In human pre-adipocytes, FFA3 was also expressed and SCFA increased SDF-1 secretion. Conclusions: SDF-1 and CXCR4 may mediate the interaction between adipose stromal cells and macrophages. Effects of SCFA are mediated through FFA3, but not histone deacetylase inhibition.

Experimental reduction in dietary omega-3 polyunsaturated fatty acids depresses sperm competitiveness

 The health benefits of diets containing rich sources of long-chain omega-3 polyunsaturated fatty acids (n-3 LC-PUFA) are well documented and include reductions in the risk of several diseases typical of Western societies. The dietary intake of n-3 LC-PUFA has also been linked to fertility, and there is abundant evidence that a range of ejaculate traits linked to fertility in humans, livestock and other animals depend on an adequate intake of n-3 LC-PUFA from dietary sources. However, relatively few studies have explored how n-3 LC-PUFA influence reproductive fitness, particularly in the context of sexual selection. Here, we show that experimental reduction in the level of n-3 LC-PUFA in the diet of guppies (Poecilia reticulata) depresses a male’s share of paternity when sperm compete for fertilization, confirming that the currently observed trend for reduced n-3 LC-PUFA in western diets has important implications for individual reproductive fitness.

Dietary n-6/n-3 LC-PUFA ratio, temperature and time interactions on nutrients and fatty acids digestibility in Atlantic salmon

The objective of the present study was to evaluate the effects of altered dietary n-3/n-6 LC-PUFA ratio, adaptation to diet over time, different water temperatures, and their interactions on nutrients and fatty acids digestibility in juvenile Atlantic salmon. Three experimental diets were formulated to be identical, with the only exception of the ratio of eicosapentaenoic acid (EPA, 20:5n-3) to arachidonic acid (ARA, 20:4n-6), and fed to triplicate groups of juvenile Atlantic salmon (Salmo salar) of 55. g initial body weight. Fish were reared in a fully controlled recirculating aquaculture system, fed to apparent satiety twice daily and kept at 10. °C and for an initial period of 100. days, and faeces were collected for digestibility estimation. Then, half of the fish of each experimental tank were moved to a separate system, where the water temperature was gradually increased up to 20. °C. Fish were maintained in the two systems for an additional period of 50. days, and faeces were collected for digestibility estimation from both groups of fish at the two water temperatures. This study concluded that dietary treatments and time had only minor effects, whereas environmental temperature resulted in modified digestibility values, with increased nutrient digestibility with increasing temperature. Varying EPA/ARA ratio in the diet had only minor direct effects on digestibility, with no direct effect on overall nutrients digestibility, and fundamentally only statistically significant effects in the fatty acid digestibility of EPA and ARA themselves. Because of current increasing pressure for more efficient fish oil replacement strategies, increasing interest in dietary ARA in aquafeed and increasing relevance and occurrence of sub-optimal rearing temperature in commercial aquaculture, this study can be considered to be important as it provided a series of fundamental information, which are envisaged to be useful towards addressing these constraints and possible nutritional remedial strategies.

Improving quantification using curtain flow chromatography columns in the analysis of labile compounds: a study on amino acids.

The performance of curtain flow chromatography column technology with MS detection was evaluated for the analysis of labile compounds. The curtain flow column design allows for separations that are faster and/or more sensitive than conventional columns, depending on how exactly the curtain flow column is configured. For example, when mass spectral detection is employed, the curtain flow column can yield separations that are 5-times faster than conventional columns when the curtain flow and the conventional columns have the same internal diameter. Or when the internal diameter of the conventional column is reduced in order to yield the same analytical through-put as the curtain flow column, the sensitivity on the curtain flow column can be as much as 66-fold higher than the conventional column. As a consequence of the higher analytical through-put less standardization is required in the analysis of labile compounds because less sample degradation is apparent. Consequently the sample integrity is preserved yielding data of a higher quality.

What makes a good graphene-binding peptide? Adsorption of amino acids and peptides at aqueous graphene interfaces

Investigation of the non-covalent interaction of biomolecules with aqueous graphene interfaces is a rapidly expanding area. However, reliable exploitation of these interfaces in many applications requires that the links between the sequence and binding of the adsorbed peptide structures be clearly established. Molecular dynamics (MD) simulations can play a key role in elucidating the conformational ensemble of peptides adsorbed at graphene interfaces, helping to elucidate these rules in partnership with experimental characterisation. We apply our recently-developed polarisable force-field for biomolecule-graphene interfaces, GRAPPA, in partnership with advanced simulation approaches, to probe the adsorption behaviour of peptides at aqueous graphene. First we determine the free energy of adsorption of all twenty naturally occurring amino acids (AAs) via metadynamics simulations, providing a benchmark for interpreting peptide-graphene adsorption studies. From these free energies, we find that strong-binding amino acids have flat and/or compact side chain groups, and we relate this behaviour to the interfacial solvent structuring. Second, we apply replica exchange with solute tempering simulations to efficiently and widely sample the conformational ensemble of two experimentally-characterised peptide sequences, P1 and its alanine mutant P1A3, in solution and adsorbed on graphene. For P1 we find a significant minority of the conformational ensemble possesses a helical structure, both in solution and when adsorbed, while P1A3 features mostly extended, random-coil conformations. In solution this helical P1 configuration is stabilised through favourable intra-peptide interactions, while the adsorbed structure is stabilised via interaction of four strongly-binding residues, identified from our metadynamics simulations, with the aqueous graphene interface. Our findings rationalise the performance of the P1 sequence as a known graphene binder.

Propyl gallate and butylated hydroxytoluene influence the accumulation of saturated fatty acids, omega-3 fatty acid and carotenoids in thraustochytrids

Schizochytrium sp. S31 was shown to have potential for production of the functional food ingredients docosahexaenoic acid (DHA) and astaxanthin, with coproduction of biodiesel. Biomass and lipid levels were greater with glycerol than with glucose as carbon source. Addition of propyl gallate or butylated hydroxytoluene to the media resulted in increased biomass and lipid levels, with propyl gallate being the more effective of the two antioxidants. Medium supplementation with propyl gallate at 0.03% and glycerol as the carbon source resulted in enhanced biomass productivity (28.50 g L^-1), lipid accumulation (24.87 g L^-1) and astaxanthin levels (452.26 μg L^-1).

What is the most effective way of increasing the bioavailability of dietary long chain omega-3 fatty acids-daily vs. weekly administration of fish oil?

The recommendations on the intake of long chain omega-3 polyunsaturated fatty acids (n-3 LC-PUFA) vary from eating oily fish ("once to twice per week") to consuming specified daily amounts of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) ("250-500 mg per day"). It is not known if there is a difference in the uptake/bioavailability between regular daily consumption of supplementsvs. consuming fish once or twice per week. In this study, the bioavailability of a daily dose of n-3 LC-PUFA (Constant treatment), representing supplements, vs. a large weekly dose of n-3 LC-PUFA (Spike treatment), representing consuming once or twice per week, was assessed. Six-week old healthy male Sprague-Dawley rats were fed either a Constant treatment, a Spike treatment or Control treatment (no n-3 LC-PUFA), for six weeks. The whole body, tissues and faeces were analysed for fatty acid content. The results showed that the major metabolic fate of the n-3 LC-PUFA (EPA+docosapentaenoic acid (DPA) + DHA) was towards catabolism (β-oxidation) accounting for over 70% of total dietary intake, whereas deposition accounted less than 25% of total dietary intake. It was found that significantly more n-3 LC-PUFA were β-oxidised when originating from the Constant treatment (84% of dose), compared with the Spike treatment (75% of dose). Conversely, it was found that significantly more n-3 LC-PUFA were deposited when originating from the Spike treatment (23% of dose), than from the Constant treatment (15% of dose). These unexpected findings show that a large dose of n-3 LC-PUFA once per week is more effective in increasing whole body n-3 LC-PUFA content in rats compared with a smaller dose delivered daily.

Metabolic fate (absorption, β-oxidation and deposition) of long-chain n-3 fatty acids is affected by sex and by the oil source (krill oil or fish oil) in the rat

The effects of krill oil as an alternative source of n-3 long-chain PUFA have been investigated recently. There are conflicting results from the few available studies comparing fish oil and krill oil. The aim of this study was to compare the bioavailability and metabolic fate (absorption, β-oxidation and tissue deposition) of n-3 fatty acids originating from krill oil (phospholipid-rich) or fish oil (TAG-rich) in rats of both sexes using the whole-body fatty acid balance method. Sprague-Dawley rats (thirty-six male, thirty-six female) were randomly assigned to be fed either a krill oil diet (EPA+DHA+DPA=1·38 mg/g of diet) or a fish oil diet (EPA+DHA+DPA=1·61 mg/g of diet) to constant ration for 6 weeks. The faeces, whole body and individual tissues were analysed for fatty acid content. Absorption of fatty acids was significantly greater in female rats and was only minimally affected by the oil type. It was estimated that most of EPA (>90 %) and more than half of DHA (>60 %) were β-oxidised in both diet groups. Most of the DPA was β-oxidised (57 and 67 % for female and male rats, respectively) in the fish oil group; however, for the krill oil group, the majority of DPA was deposited (82-83 %). There was a significantly greater deposition of DPA and DHA in rats fed krill oil compared with those fed fish oil, not due to a difference in bioavailability (absorption) but rather due to a difference in metabolic fate (anabolism v. catabolism).