Isotope and fatty acid trends along continental shelf depth gradients: Inshore versus offshore hydrological influences on benthic trophic functioning

Anthropogenic activities and land-based inputs into the sea may influence the trophic structure and functioning of coastal and continental shelf ecosystems, despite the numerous opportunities and services the latter offer to humans and wildlife. In addition, hydrological structures and physical dynamics potentially influence the sources of organic matter (e.g., terrestrial versus marine, or fresh material versus detrital material) entering marine food webs. Understanding the significance of the processes that influence marine food webs and ecosystems (e.g., terrestrial inputs, physical dynamics) is crucially important because trophic dynamics are a vital part of ecosystem integrity. This can be achieved by identifying organic matter sources that enter food webs along inshore–offshore transects. We hypothesised that regional hydrological structures over wide continental shelves directly control the benthic trophic functioning across the shelf. We investigated this issue along two transects in the northern ecosystem of the Bay of Biscay (north-eastern Atlantic). Carbon and nitrogen stable isotope analysis (SIA) and fatty acid analysis (FAA) were conducted on different complementary ecosystem compartments that include suspended particulate organic matter (POM), sedimentary organic matter (SOM), and benthic consumers such as bivalves, large crustaceans and demersal fish. Samples were collected from inshore shallow waters (at ∼1 m in depth) to more than 200 m in depth on the offshore shelf break. Results indicated strong discrepancies in stable isotope (SI) and fatty acid (FA) compositions in the sampled compartments between inshore and offshore areas, although nitrogen SI (δ15N) and FA trends were similar along both transects. Offshore the influence of a permanently stratified area (described previously as a “cold pool”) was evident in both transects. The influence of this hydrological structure on benthic trophic functioning (i.e., on the food sources available for consumers) was especially apparent across the northern transect, due to unusual carbon isotope compositions (δ13C) in the compartments. At stations under the cold pool, SI and FA organism compositions indicated benthic trophic functioning based on a microbial food web, including a significant contribution of heterotrophic planktonic organisms and/or of SOM, notably in stations under the cold pool. On the contrary, inshore and shelf break areas were characterised by a microalgae-based food web (at least in part for the shelf break area, due to slope current and upwelling that can favour fresh primary production sinking on site). SIA and FAA were relevant and complementary tools, and consumers better medium- to long-term system integrators than POM samples, for depicting the trophic functioning and dynamics along inshore–offshore transects over continental shelves.

Ad-atoms and their enhancement effects on Pt activity for formic acid oxidation

Formic acid oxidation has been widely studied at Pt as a model reaction to understand fundamental aspects of electrocatalytic reactions in fuel cells. Electrocatalytic oxidation of formic acid takes place through two parallel pathways (direct and indirect). The indirect pathway proceeds via CO as an intermediate, which is known to be responsible for the poisoning of Pt and its consequent decrease in activity. Surface modification of Pt with ad-atoms is known to hinder this poisoning and promote the direct pathway. The incorporation of polymers (polyaniline, polycarbazole, polyindole) as supports also increases activity. Irreversibly adsorbed Sb and Bi on Pt are known to show high electrocatalytic activity for formic acid oxidation. This work presents the dependence of Sb and Bi irreversible adsorption on immersion time, metal solution concentration and pH. The activity of Sb and Bi modified Pt was correlated against immersion time and percent coverage of Pt by ad-atoms. Polyaniline support effects in combination with a Bi modified Pt catalyst showed enhancement in oxidation current compared to Pt-Bi.

Analgesic Effects of Fatty Acid Amide Hydrolase Inhibition in a Rat Model of Neuropathic Pain

Cannabinoid-based medicines have therapeutic potential for the treatment of pain. Augmentation of levels of endocannabinoids with inhibitors of fatty acid amide hydrolase (FAAH) is analgesic in models of acute and inflammatory pain states. The aim of this study was to determine whether local inhibition of FAAH alters nociceptive responses of spinal neurons in the spinal nerve ligation model of neuropathic pain. Electrophysiological studies were performed 14-18 days after spinal nerve ligation or sham surgery, and the effects of the FAAHinhibitor cyclohexylcarbamic acid 3-carbamoyl biphenyl-3-yl ester (URB597) on mechanically evoked responses of spinal neurons and levels of endocannabinoids were determined. Intraplantar URB597 (25 _g in 50 _l) significantly ( p _ 0.01) attenuated mechanically evoked responses of spinal neurons in sham-operated rats. Effects of URB597 were blocked by the cannabinoid 1 receptor (CB1 ) antagonist AM251 [N-1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-1-piperidinyl-1H-pyrazole-3-carboxamide] (30_g in50_l) and the opioid receptor antagonist naloxone. URB597 treatment increased levels of anandamide, 2-arachidonyl glycerol, and oleoyl ethanolamide in the ipsilateral hindpaw of shamoperated rats. Intraplantar URB597 (25 _g in 50 _l) did not, however, alter mechanically evoked responses of spinal neurons in spinal nerve ligated (SNL) rats or hindpaw levels of endocannabinoids. Intraplantar injection of a higher dose of URB597 (100 _g in 50 _l) significantly ( p_0.05) attenuated evoked responses of spinal neurons in SNL rats but did not alter hindpaw levels of endocannabinoids. Spinal administration of URB597 attenuated evoked responses of spinal neurons and elevated levels of endocannabinoids in shamoperated and SNL rats. These data suggest that peripheral FAAH activity may be altered or that alternative pathways of metabolism have greater importance in SNL rats.

Comparison of prophylactic and therapeutic use of short-chain fatty acid enemas in diversion colitis: a study in Wistar rats

OBJECTIVES: To study the effect of short-chain fatty-acids on atrophy and inflammation of excluded colonic segments before and after the development of diversion colitis. INTRODUCTION: Diversion colitis is a chronic inflammatory process affecting the dysfunctional colon, possibly evolving with mucous and blood discharge. The most favored hypotheses to explain its development is short-chain fatty-acid deficiency in the colon lumen. METHODS: Wistar rats were submitted to colostomy with distal colon exclusion. Two control groups (A1 and B1) received rectally administered physiological saline, whereas two experimental groups (A2 and B2) received rectally administered short-chain fatty-acids. The A groups were prophylactically treated (5th to 40th days postoperatively), whereas the B groups were therapeutically treated (after post-operative day 40). The mucosal thickness of the excluded colon was measured histologically. The inflammatory reaction of the mucosal lamina propria and the lymphoid tissue response were quantified through established scores. RESULTS: There was a significant thickness recovery of the colonic mucosa in group B2 animals (p = 0.0001), which also exhibited a significant reduction in the number of eosinophilic polymorphonuclear cells in the lamina propria (p = 0.0126) and in the intestinal lumen (p = 0.0256). Group A2 showed no mucosal thickness recovery and significant increases in the numbers of lymphocytes (p = 0.0006) and eosinophilic polymorphonuclear cells in the lamina propria of the mucosa (p = 0.0022). CONCLUSION: Therapeutic use of short-chain fatty-acids significantly reduced eosinophilic polymorphonuclear cell numbers in the intestinal wall and in the colonic lumen; it also reversed the atrophy of the colonic mucosa. Prophylactic use did not impede the development of mucosal atrophy

Development of a colorimetric indicator label for food oxidation based on hexanal detection

In this work, a colorimetric indicator for food oxidation based on the detection of hexanal in gas-phase, has been developed. In fact, in recent years, the food packaging industry has evolved towards new generation of packaging, like active and intelligent. According to literature (Pangloli P. et al. 2002), hexanal is the main product of a fatty acid oxidation: the linoleic acid. So, it was chosen to analyse two kinds of potato chips, fried in two different oils with high concentration of linoleic acid: olive oil and sunflower oil. Five different formulas were prepared and their colour change when exposed to hexanal in gas phase was evaluated. The formulas evaluations were first conducted on filter paper labels. The next step was to select the thickener to add to the formula, in order to coat a polypropylene film, more appropriate than the filter paper for a production at industrial scale. Three kinds of thickeners were tested: a cellulose derivative, an ethylene vinyl-alcohol and a polyvinyl alcohol. To obtain the final labels with the autoadhesive layer, the polypropylene film with the selected formula and thickener was coat with a water based adhesive. For both filter paper and polypropylene labels, with and without autoadhesive layer, the detection limit and the detection time were measured. For the selected formula on filter paper labels, the stability was evaluated, when conserved on the dark or on the light, in order to determine the storage time. Both potato chips samples, stocked at the same conditions, were analysed using an optimised Headspace-Solid Phase Microextraction-Gas Chromatography-Mass Spectrometry (HS-SPME-GC-MS) method, in order to determine the concentration of volatilized hexanal. With the aim to establish if the hexanal can be considered as an indicator of the end of potato chips shelf life, sensory evaluation was conducted each day of HS-SPME-GC-MS analysis.

Mitochondrial energy metabolism and redox responses to hypertriglyceridemia

In this work we review recent findings that explain how mitochondrial bioenergetic functions and redox state respond to a hyperlipidemic in vivo environment and may contribute to the maintenance of a normal metabolic phenotype. The experimental model utilized to evidence these adaptive mechanisms is especially useful for these studies since it exhibits genetic hypertriglyceridemia and avoids complications introduced by high fat diets. Liver from hypertrigliceridemic (HTG) mice have a greater content of glycerolipids together with increased mitochondrial free fatty acid oxidation. HTG liver mitochondria have a higher resting respiration rate but normal oxidative phosphorylation efficiency. This is achieved by higher activity of the mitochondrial potassium channel sensitive to ATP (mitoK(ATP)). The mild uncoupling mediated by mitoK(ATP) accelerates respiration rates and reduces reactive oxygen species generation. Although this response is not sufficient to inhibit lipid induced extra-mitochondrial oxidative stress in whole liver cells it avoids amplification of this redox imbalance. Furthermore, higher mitoK(ATP) activity increases liver, brain and whole body metabolic rates. These mitochondrial adaptations may explain why these HTG mice do not develop insulin resistance and obesity even under a severe hyperlipidemic state. On the contrary, when long term high fat diets are employed, insulin resistance, fatty liver and obesity develop and mitochondrial adaptations are inefficient to counteract energy and redox imbalances.

The peroxisome proliferator-activated receptor beta/delta agonist, GW501516, regulates the expression of genes involved in lipid catabolism and energy uncoupling in skeletal muscle cells

Lipid homeostasis is controlled by the peroxisome proliferator-activated receptors (PPARalpha, -beta/delta, and -gamma) that function as fatty acid-dependent DNA-binding proteins that regulate lipid metabolism. In vitro and in vivo genetic and pharmacological studies have demonstrated PPARalpha regulates lipid catabolism. In contrast, PPARgamma regulates the conflicting process of lipid storage. However, relatively little is known about PPARbeta/delta in the context of target tissues, target genes, lipid homeostasis, and functional overlap with PPARalpha and -gamma. PPARbeta/delta, a very low-density lipoprotein sensor, is abundantly expressed in skeletal muscle, a major mass peripheral tissue that accounts for approximately 40% of total body weight. Skeletal muscle is a metabolically active tissue, and a primary site of glucose metabolism, fatty acid oxidation, and cholesterol efflux. Consequently, it has a significant role in insulin sensitivity, the blood-lipid profile, and lipid homeostasis. Surprisingly, the role of PPARbeta/delta in skeletal muscle has not been investigated. We utilize selective PPARalpha, -beta/delta, -gamma, and liver X receptor agonists in skeletal muscle cells to understand the functional role of PPARbeta/delta, and the complementary and/or contrasting roles of PPARs in this major mass peripheral tissue. Activation of PPARbeta/delta by GW501516 in skeletal muscle cells induces the expression of genes involved in preferential lipid utilization, beta-oxidation, cholesterol efflux, and energy uncoupling. Furthermore, we show that treatment of muscle cells with GW501516 increases apolipoprotein-A1 specific efflux of intracellular cholesterol, thus identifying this tissue as an important target of PPARbeta/delta agonists. Interestingly, fenofibrate induces genes involved in fructose uptake, and glycogen formation. In contrast, rosiglitazone-mediated activation of PPARgamma induces gene expression associated with glucose uptake, fatty acid synthesis, and lipid storage. Furthermore, we show that the PPAR-dependent reporter in the muscle carnitine palmitoyltransferase-1 promoter is directly regulated by PPARbeta/delta, and not PPARalpha in skeletal muscle cells in a PPARgamma coactivator-1-dependent manner. This study demonstrates that PPARs have distinct roles in skeletal muscle cells with respect to the regulation of lipid, carbohydrate, and energy homeostasis. Moreover, we surmise that PPARgamma/delta agonists would increase fatty acid catabolism, cholesterol efflux, and energy expenditure in muscle, and speculate selective activators of PPARbeta/delta may have therapeutic utility in the treatment of hyperlipidemia, atherosclerosis, and obesity.

New mechanisms that regulate the expression of genes implicated in the process of ketogenesis

Dissertação para obtenção do Grau de Mestre em Biotecnologia

Obesity Preserves Myocardial Function During Blockade of the Glycolytic Pathway

Background: Obesity is defined by excessive accumulation of body fat relative to lean tissue. Studies during the last few years indicate that cardiac function in obese animals may be preserved, increased or diminished. Objective: Study the energy balance of the myocardium with the hypothesis that the increase in fatty acid oxidation and reduced glucose leads to cardiac dysfunction in obesity. Methods: 30-day-old male Wistar rats were fed standard and hypercaloric diet for 30 weeks. Cardiac function and morphology were assessed. In this paper was viewed the general characteristics and comorbities associated to obesity. The structure cardiac was determined by weights of the heart and left ventricle (LV). Myocardial function was evaluated by studying isolated papillary muscles from the LV, under the baseline condition and after inotropic and lusitropic maneuvers: myocardial stiffness; postrest contraction; increase in extracellular Ca2+ concentration; change in heart rate and inhibitor of glycolytic pathway. Results: Compared with control group, the obese rats had increased body fat and co-morbities associated with obesity. Functional assessment after blocking iodoacetate shows no difference in the linear regression of DT, however, the RT showed a statistically significant difference in behavior between the control and the obese group, most notable being the slope in group C. Conclusion: The energy imbalance on obesity did not cause cardiac dysfunction. On the contrary, the prioritization of fatty acids utilization provides protection to cardiac muscle during the inhibition of glycolysis, suggesting that this pathway is fewer used by obese cardiac muscle.

Double gene deletion reveals the lack of cooperation between PPARalpha and PPARbeta in skeletal muscle.

The peroxisome proliferator-activated receptors (PPARs) are involved in the regulation of most of the pathways linked to lipid metabolism. PPARalpha and PPARbeta isotypes are known to regulate muscle fatty acid oxidation and a reciprocal compensation of their function has been proposed. Herein, we investigated muscle contractile and metabolic phenotypes in PPARalpha-/-, PPARbeta-/-, and double PPARalpha-/- beta-/- mice. Heart and soleus muscle analyses show that the deletion of PPARalpha induces a decrease of the HAD activity (beta-oxidation) while soleus contractile phenotype remains unchanged. A PPARbeta deletion alone has no effect. However, these mild phenotypes are not due to a reciprocal compensation of PPARbeta and PPARalpha functions since double gene deletion PPARalpha-PPARbeta mostly reproduces the null PPARalpha-mediated reduced beta-oxidation, in addition to a shift from fast to slow fibers. In conclusion, PPARbeta is not required for maintaining skeletal muscle metabolic activity and does not compensate the lack of PPARalpha in PPARalpha null mice.

Do peroxisome proliferating compounds pose a hepatocarcinogenic hazard to humans?

The purpose of the workshop "Do Peroxisome Proliferating Compounds Pose a Hepatocarcinogenic Hazard to Humans?" was to provide a review of the current state of the science on the relationship between peroxisome proliferation and hepatocarcinogenesis. There has been much debate regarding the mechanism by which peroxisome proliferators may induce liver tumors in rats and mice and whether these events occur in humans. A primary goal of the workshop was to determine where consensus might be reached regarding the interpretation of these data relative to the assessment of potential human risks. A core set of biochemical and cellular events has been identified in the rodent strains that are susceptible to the hepatocarcinogenic effects of peroxisome proliferators, including peroxisome proliferation, increases in fatty acyl-CoA oxidase levels, microsomal fatty acid oxidation, excess production of hydrogen peroxide, increases in rates of cell proliferation, and expression and activation of the alpha subtype of the peroxisome proliferator-activated receptor (PPAR-alpha). Such effects have not been identified clinically in liver biopsies from humans exposed to peroxisome proliferators or in in vitro studies with human hepatocytes, although PPAR-alpha is expressed at a very low level in human liver. Consensus was reached regarding the significant intermediary roles of cell proliferation and PPAR-alpha receptor expression and activation in tumor formation. Information considered necessary for characterizing a compound as a peroxisome proliferating hepatocarcinogen include hepatomegaly, enhanced cell proliferation, and an increase in hepatic acyl-CoA oxidase and/or palmitoyl-CoA oxidation levels. Given the lack of genotoxic potential of most peroxisome proliferating agents, and since humans appear likely to be refractive or insensitive to the tumorigenic response, risk assessments based on tumor data may not be appropriate. However, nontumor data on intermediate endpoints would provide appropriate toxicological endpoints to determine a point of departure such as the LED10 or NOAEL which would be the basis for a margin-of-exposure (MOE) risk assessment approach. Pertinent factors to be considered in the MOE evaluation would include the slope of the dose-response curve at the point of departure, the background exposure levels, and variability in the human response.

The L162V polymorphism of the PPARA gene is associated with stage C of heart failure.

In human heart failure (HF) peroxisome proliferator-activated receptor alpha (PPAR alpha) is downregulated and consequently, the expression of genes involved in fatty acid oxidation repressed. The L162V (rs1800206) is a functional polymorphism of the human PPAR alpha gene (PPARA). In the present study we have investigated whether this polymorphism is associated with the development of stage C of HF.

Leucine supplementation protects from insulin resistance by regulating adiposity levels.

BACKGROUND Leucine supplementation might have therapeutic potential in preventing diet-induced obesity and improving insulin sensitivity. However, the underlying mechanisms are at present unclear. Additionally, it is unclear whether leucine supplementation might be equally efficacious once obesity has developed. METHODOLOGY/PRINCIPAL FINDINGS Male C57BL/6J mice were fed chow or a high-fat diet (HFD), supplemented or not with leucine for 17 weeks. Another group of HFD-fed mice (HFD-pairfat group) was food restricted in order to reach an adiposity level comparable to that of HFD-Leu mice. Finally, a third group of mice was exposed to HFD for 12 weeks before being chronically supplemented with leucine. Leucine supplementation in HFD-fed mice decreased body weight and fat mass by increasing energy expenditure, fatty acid oxidation and locomotor activity in vivo. The decreased adiposity in HFD-Leu mice was associated with increased expression of uncoupling protein 3 (UCP-3) in the brown adipose tissue, better insulin sensitivity, increased intestinal gluconeogenesis and preservation of islets of Langerhans histomorphology and function. HFD-pairfat mice had a comparable improvement in insulin sensitivity, without changes in islets physiology or intestinal gluconeogenesis. Remarkably, both HFD-Leu and HFD-pairfat mice had decreased hepatic lipid content, which likely helped improve insulin sensitivity. In contrast, when leucine was supplemented to already obese animals, no changes in body weight, body composition or glucose metabolism were observed. CONCLUSIONS/SIGNIFICANCE These findings suggest that leucine improves insulin sensitivity in HFD-fed mice by primarily decreasing adiposity, rather than directly acting on peripheral target organs. However, beneficial effects of leucine on intestinal gluconeogenesis and islets of Langerhans's physiology might help prevent type 2 diabetes development. Differently, metabolic benefit of leucine supplementation is lacking in already obese animals, a phenomenon possibly related to the extent of the obesity before starting the supplementation.

Oleoylethanolamide enhances β-adrenergic-mediated thermogenesis and white-to-brown adipocyte phenotype in epididymal white adipose tissue in rat.

β-adrenergic receptor activation promotes brown adipose tissue (BAT) β-oxidation and thermogenesis by burning fatty acids during uncoupling respiration. Oleoylethanolamide (OEA) can inhibit feeding and stimulate lipolysis by activating peroxisome proliferator-activating receptor-α (PPARα) in white adipose tissue (WAT). Here we explore whether PPARα activation potentiates the effect of β3-adrenergic stimulation on energy balance mediated by the respective agonists OEA and CL316243. The effect of this pharmacological association on feeding, thermogenesis, β-oxidation, and lipid and cholesterol metabolism in epididymal (e)WAT was monitored. CL316243 (1 mg/kg) and OEA (5 mg/kg) co-administration over 6 days enhanced the reduction of both food intake and body weight gain, increased the energy expenditure and reduced the respiratory quotient (VCO2/VO2). This negative energy balance agreed with decreased fat mass and increased BAT weight and temperature, as well as with lowered plasma levels of triglycerides, cholesterol, nonessential fatty acids (NEFAs), and the adipokines leptin and TNF-α. Regarding eWAT, CL316243 and OEA treatment elevated levels of the thermogenic factors PPARα and UCP1, reduced p38-MAPK phosphorylation, and promoted brown-like features in the white adipocytes: the mitochondrial (Cox4i1, Cox4i2) and BAT (Fgf21, Prdm16) genes were overexpressed in eWAT. The enhancement of the fatty-acid β-oxidation factors Cpt1b and Acox1 in eWAT was accompanied by an upregulation of de novo lipogenesis and reduced expression of the unsaturated-fatty-acid-synthesis enzyme gene, Scd1. We propose that the combination of β-adrenergic and PPARα receptor agonists promotes therapeutic adipocyte remodelling in eWAT, and therefore has a potential clinical utility in the treatment of obesity.

Differential effects of high-fat diet on myocardial lipid metabolism in failing and nonfailing hearts with angiotensin II-mediated cardiac remodeling in mice.

Normal myocardium adapts to increase of nutritional fatty acid supply by upregulation of regulatory proteins of the fatty acid oxidation pathway. Because advanced heart failure is associated with reduction of regulatory proteins of fatty acid oxidation, we hypothesized that failing myocardium may not be able to adapt to increased fatty acid intake and therefore undergo lipid accumulation, potentially aggravating myocardial dysfunction. We determined the effect of high-fat diet in transgenic mice with overexpression of angiotensinogen in the myocardium (TG1306/R1). TG1306/R1 mice develop ANG II-mediated left ventricular hypertrophy, and at one year of age approximately half of the mice present heart failure associated with reduced expression of regulatory proteins of fatty acid oxidation and reduced palmitate oxidation during ex vivo working heart perfusion. Hypertrophied hearts from TG1306/R1 mice without heart failure adapted to high-fat feeding, similarly to hearts from wild-type mice, with upregulation of regulatory proteins of fatty acid oxidation and enhancement of palmitate oxidation. There was no myocardial lipid accumulation or contractile dysfunction. In contrast, hearts from TG1306/R1 mice presenting heart failure were unable to respond to high-fat feeding by upregulation of fatty acid oxidation proteins and enhancement of palmitate oxidation. This resulted in accumulation of triglycerides and ceramide in the myocardium, and aggravation of contractile dysfunction. In conclusion, hearts with ANG II-induced contractile failure have lost the ability to enhance fatty acid oxidation in response to increased fatty acid supply. The ensuing accumulation of lipid compounds may play a role in the observed aggravation of contractile dysfunction.