Preliminary data from studies in rodents suggests time-dependent associations between dietary LC n-3 PUFA and hepatic levels of the enzyme soluble epoxide hydrolase (sEH), which regulates the metabolism and availability of epoxyeicosatrienoic acids (EET).  EET are cytochrome P450 epoxygenase products of arachidonic acid associated with  lower blood pressure, decreased inflammatory response and inhibition of blood coagulation. To further investigate the association between LC n-3 PUFA and sEH, ApoE^-/^- mice were fed a high-fat high-cholesterol diet supplemented with either fish oil (EPA + DHA) or DHA or HOSF (all 2% w/w) for 10 weeks and livers and aortic roots were collected on day 2 and weeks 1, 2, 4 and 10.  Proteomics analysis showed an overall decreasing effect of fish oil (but not DHA) supplementation on hepatic protein levels of sEH compared to the control throughout the intervention period (<i>P</i> &lt; 0.05).  Neither fish oil nor DHA intervention affected atherosclerotic plaque size in the aortic root. We also examined how dietary supplementation with 1 g/day EPA or 1 g/day DHA for 10 days affects platelet sEH levels and platelet aggregation compared to 1 g/day HOSF (control) in healthy volunteers in a double-blind, placebo-controlled, cross-over trial.  We found that DHA decreased platelet aggregation by 10% (<i>P =</i> 0.04) and EPA also inhibited ADP (5 μM)-induced platelet aggregation by 14% compared to the control group but this effect did not reach statistical significance due to high variability between subjects.  EPA decreased platelet sEH levels by 25% (not significant), whereas DHA had no effect.  We also attempted to optimize a method for measuring EET in plasma and platelets.  However, the rapid conversion of EET to other compounds and their low concentration in tissues prevented us from optimizing such a method within the time limits of the project.

The endocannabinoid (EC) system has a role in metabolic homeostasis. The purpose of this study was to determine the effect of ECs and the fatty acids they are derived from on lipid metabolism and mitochondrial function in adipocytes. 3T3-L1 adipocytes on day 8 of differentiation were treated with ECs and fatty acids for 48 hours in the absence or presence of insulin and various inhibitors. Lysates were analyzed via Western immunoblotting, a lipolysis assay and Seahorse XF Analyzer for changes in protein levels, phosphorylation state, lipolysis, and oxygen consumption rate. Results showed that ECs (2-arachidonoyl glycerol) stimulated lipolysis via a novel AMPK-dependent pathway, while fatty acids had varying effects on insulin signaling and mitochondrial function . These data suggest adipose tissue EC receptors may be a suitable target for anti-obesity therapy. Further research is needed to understand how the dietary fatty acid profile may influence synthesis of ECs. / May 2016

Les MICI (maladies inflammatoires chroniques de l'intestin) invalident 200 000 personnes en France. La région Nord-Pas-de-Calais est particulièrement touchée par ces affections et les traitements disponibles pour ces pathologies demeurent coûteux et peu nombreux. Des études récentes ont suggéré que le système endocannabinoïde, exprimé au seing du tractus gastro-intestinal, est une cible thérapeutique prometteuse pour le traitement des MICI. Ce système se compose des récepteurs aux cannabinoïdes CB1 et CB2, des ligands endogènes de ces récepteurs, notamment l'anandamide et le 2-arachidonoylglycérol et des protéines impliquées dans l'anabolisme et le catabolisme des ligands. L'anandamide a présenté des capacités à prévenir la colite induite par le TNBS à des rongeurs. Toutefois, in vivo ce composé possède un temps de demi-vie court et est rapidement dégradé par une amidase à sérine, la FAAH (Fatty Acid Amide Hydrolase). Nous avons ainsi envisagé la conception, la synthèse et l'évaluation pharmacologique de nouveau inhibiteurs de la FAAH. L'une de nos molécules, le composé 95, présentant une CI50 sur l'enzyme de 88 nM a ensuite été injectée par voie intrapéritonéale à des souris dont la colite a été induite trois jours plus tard par l'injection intrarectale de TNBS. L'évaluation des scores macroscopiques et microscopiques des dommages causés sur le côlon par l'agent irritant a ensuite été effectuée. L'inflammation du côlon a été significativement réduite chez le groupe de souris ayant été traité par le composé 95, montrant que l'inhibition de la FAAH est une stratégie thérapeutique efficace dans le traitement des MICI. / IBD (Inflammatory Bowel Diseases) invalidate 200 000 persons in France. Nord-Pas-de-Calais region is particularly touched by these diseases and the available treatments for these pathologies are few and expensive. Recent studies have suggested that endocannabinoid system expressed in the gastrointestinal tract, was a promising therapeutic target for the IBD treatment. This system is made up of cannabinoids receptors CB1 and CB2, endogenous ligands of these receptors, notably anandamide and 2-arachidonoylglycerol, and proteins involved in ligands metabolism. Anandamide has shown properties to prevent TNBS-induced colitis in mice. However, in vivo, anandamide possesses a short life time and is rapidly hydrolyzed by a serine amidase, the FAAH (Fatty Acid Amide Hydrolase). In this context, we have considered the design, the synthesis and the pharmacological evaluation of new FAAH inhibitors. One of our molecules, compound 95, inhibiting the enzyme with an IC50 value of 88 nM has been injected intraperitonally to mice which the colitis was induced three days later by intrarectal TNBS-administration. The assessment of macroscopic and microscopic scores of colonic damages was undergone. Colonic inflammation was significatively reduced in the group of mice which has been treated by 95, showing evidence that FAAH inhibition was an effective therapeutic target for the treatment of IBD.

To evaluate the niche of coastal fish populations in the Beaufort Sea, stable isotopes (SI) and fatty acids (FA) were used to characterize species-specific niches, niche overlaps and resource partitioning (nicheROVER) of the Shingle Point fish populations. Fishes were grouped into three isotopic groups: marine, coastal, and freshwater (Ward’s clustering analysis), and five dietary groupings (using FA), where benthic feeding strategies were prevalent (correspondence analysis). Niche metrics were used to evaluate if total mercury (THg) could contribute complementary trophic information (residual permutation procedure (RPP)). Three THg groups (high, intermediate, low) were identified (boxplot analysis). High THg was identified in high trophic and benthic feeders, high THg ranges were observed in species with large niche sizes, high trophic feeding, and freshwater influences (RPP). The bioavailability of freshwater introduced THg to marine biota was assessed, however further research needs to be performed. Combining dietary indicators SI, FA, and THg, allowed for the characterization of the diet and habitat use of coastal fish populations, better understanding of the niches of these species, and developed baseline information for future monitoring in an MPA, as climate change continues to effect the Beaufort coastal environment. / February 2017

Introduction: An increased gluteofemoral adipose tissue (AT) mass is associated with a protective cardiovascular and metabolic risk profile, and effective fatty acid retention in femoral AT has been proposed as a possible mechanism. Catecholamines are important regulators of AT lipolysis and blood flow (ATBF). The aim of the thesis was to investigate regional differences in the adrenergic regulation of fatty acid release and ATBF between abdominal and femoral AT in vivo. Furthermore, in vivo regional fatty acid trafficking was studied in a physiological setting over 24 h. Methods: Regional fatty acid trafficking, along with the measurement of ATBF, was studied with the arterio-venous difference technique and stable isotope tracers in healthy volunteers. Adrenergic agonists (isoprenaline, adrenaline) were infused either locally by microinfusion, or systemically. Local microinfusion of adrenoreceptor antagonists (propranolol, phentolamine) was used to characterize specific adrenoreceptor subtype effects. The trafficking of dietary fatty acids was studied over a 24 h period involving three meals containing stable isotope-labelled fatty acids along with intravenous infusions of another labelled fatty acid. Results: Femoral ATBF and lipolysis was less responsive to adrenergic stimulation with adrenaline compared to abdominal AT. This was due to increased femoral α-adrenoreceptor responsiveness. When studied over 24 h, femoral AT showed a lower lipolysis rate compared to abdominal AT, while dietary fatty acids were extracted more avidly by abdominal AT. Uptake of non-dietary fatty acids (derived from very-low-density lipoproteins or unbound non-esterified fatty acids) was comparable between abdominal and femoral AT. Conclusion: There are fundamental differences in response to adrenergic stimuli between abdominal and gluteofemoral tissues and the ability of femoral AT to trap non-dietary fatty acids may provide protection of other tissues from ectopic fatty acid deposition.

N-Acylphosphatidylethanoiamine (NAPE) is synthesized in the microsomes of cotton seedlings by a mechanism that is possibly unique to plants, the ATP-, Ca2+-, and CoA-independent acylation ofphosphatidylethanolamine (PE) with unesterified free fatty acids (FFAs), catalyzed by NAPE synthase. A photoreactive free fatty acid analogue, 12-[(4- azidosalicyl)amino]dodecanoic acid (ASD), and its 125I-labeled derivative acted as substrates for the NAPE synthase enzyme.

One purpose of this investigation is to determine the fatty acid and lipid content of typical Vibrio cholerae cells. The comparison of cholera lipid constituents with those of closely-related bacteria might be of taxonomic value. Furthermore, chemical characterization of the cholera vibrio could provide useful criteria for identification of these disease-producing microorganisms.

NTRODUCTION: Colonic bacteria produce short-chain fatty acids (SCFAs) by fermentation of dietary carbohydrates and fiber. The production of SCFAs is greatest in proximal colon where propulsion is likely to be highly dependent on chemical/nutrient stimuli. Unabsorbed SCFAs entering the distal colon are likely to modify peristalsis initiated by fecal pellet-induced distension. AIM: To determine the effect of individual SCFAs on propulsive contractions in guinea pig proximal colon and on pellet propulsion in distal colon. METHODS: Proximal colon was excised, cannulated and placed in Krebs buffer in an organ bath. After equilibration, the colon was distended with 1ml of Krebs buffer alone or containing sodium salts of acetate, butyrate, or propionate at 10-100mM. Motility was video recorded, spatiotemporal maps generated, and the number of full-length propulsive contractions during a 5 min period was determined. The distal colon was removed from guinea pig and placed in an organ bath containing Krebs buffer. Following equilibration, video-tracking software was used to measure the velocity of propulsion of a clay pellet placed in the orad end of a segment. Krebs buffer alone or containing individual SCFAs at 30mM was perfused caudad to the pellet at 0.1 ml/min. RESULTS: The basal rate of propulsive contractions in Krebs buffer was 2.9 ± 0.7 per 5 min. Butyrate and propionate had concentration-dependent and opposing effects on propulsive contractions; acetate had no effect. Butyrate significantly increased propulsive contractions (maximal increase of 207% (p ≤ 0.05) at 30 mM) whereas propionate abolished propulsive contractions (91-100% inhibition at 10-30mM). Control Krebs buffer containing 10- 100mM NaCl had no effect. The mean rate of pellet propulsion during perfusion of Krebs buffer alone was 1.5 ± 0.2 mm/sec. Addition of sodium butyrate (30 mM) to the perfusate increased the velocity of pellet propulsion by 40 ± 4% (p<0.05) whereas addition of 30mM sodium propionate decreased velocity of pellet propulsion by 75 ± 8%. Acetate had no significant effect in either proximal or distal colon. Mixtures of SCFAs showed a general decrease in overall motility. CONCLUSION: SCFAs have differential effects on propulsive contractions in the proximal and distal colon of the guinea pig, with butyrate being excitatory and propionate being inhibitory. These studies suggest that the movement of feces in the colon depends not only on distension but the chemical (i.e. dietary) composition of the feces.

O objetivo deste estudo foi avaliar o efeito da competição de triathlon na função e morte de neutrófilos de atletas de elite e investigar o possível envolvimento dos ácidos graxos livres (AGs) neste processo. Os neutrófilos foram obtidos do sangue coletado de 11 sedentários e de 12 triatletas em repouso e após competição de triathlon (Half Ironman, 2 Km de natação, 80 Km de ciclismo e 20 Km de corrida). A competição de triathlon aumentou a capacidade dos neutrófilos de migrar e de realizar burst oxidativo, porém inibiu a fagociose realizada por estas células. Além disso, induziu aumento da fragmentação de DNA e externalização de fosfatidilserina. A competição de triathlon aumentou a concentração de AGs no plasma dos triatletas e esta foi correlacionada positivamente com a proporção de neutrófilos com DNA fragmentado e fosfatidilserina externalizada. A elevação da concentração plasmática dos ácidos oléico, linoléico e esteárico induzida pela competição parece estar envolvida nas alterações funcionais e na apoptose verificadas após a competição de triathlon. / The objective of this study was to evaluate the effect of triathlon competition on function and death of neutrophils from elite athletes and to investigate the involvement of fatty acids in this process. Neutrophils were obtained from blood collected from eleven sedentary volunteers and twelve triathletes under rest and after a Half Ironman triathlon competition (2 Km swimming, 80 Km cycling and 20 Km running). The triathlon competition increased the migration and reactive oxygen species production in neutrophils, however it reduced the phagocytosis activity. Moreover, it induced neutrophils death possibly by apoptosis as indicated by DNA fragmentation and phosphatidylserine externalization. The increase in plasma levels of oleic, linoleic and stearic acids induced by the competition may be involved in modulation on neutrophil function and death.

University of Macau / Institute of Chinese Medical Sciences