Endogenous Fatty Acid Oxidation Research Articles

Identification of a weight loss-associated causal eQTL in MTIF3 and the effects of MTIF3 deficiency on human adipocyte function.

Genetic variation at the MTIF3 (Mitochondrial Translational Initiation Factor 3) locus has been robustly associated with obesity in humans, but the functional basis behind this association is not known. Here, we applied luciferase reporter assay to map potential functional variants in the haplotype block tagged by rs1885988 and used CRISPR-Cas9 to edit the potential functional variants to confirm the regulatory effects on MTIF3 expression. We further conducted functional studies on MTIF3-deficient differentiated human white adipocyte cell line (hWAs-iCas9), generated through inducible expression of CRISPR-Cas9 combined with delivery of synthetic MTIF3-targeting guide RNA. We demonstrate that rs67785913-centered DNA fragment (in LD with rs1885988, r2 > 0.8) enhances transcription in a luciferase reporter assay, and CRISPR-Cas9-edited rs67785913 CTCT cells show significantly higher MTIF3 expression than rs67785913 CT cells. Perturbed MTIF3 expression led to reduced mitochondrial respiration and endogenous fatty acid oxidation, as well as altered expression of mitochondrial DNA-encoded genes and proteins, and disturbed mitochondrial OXPHOS complex assembly. Furthermore, after glucose restriction, the MTIF3 knockout cells retained more triglycerides than control cells. This study demonstrates an adipocyte function-specific role of MTIF3, which originates in the maintenance of mitochondrial function, providing potential explanations for why MTIF3 genetic variation at rs67785913 is associated with body corpulence and response to weight loss interventions.

Hippo pathway inhibition promotes metabolic adaptability and antioxidant response in myoblasts

Metabolic plasticity in a hostile environment ensures cell survival. We investigated whether Hippo pathway inhibition contributed to cell adaptations under challenging conditions. We examined metabolic profiles and fuel substrate choices and preferences in C2C12 myoblasts after Hippo pathway inhibition via Salvador knockdown (SAV1 KD). SAV1 KD induced higher ATP production and a more energetic phenotype. Bioenergetic profiling showed enhanced key mitochondrial parameters including spare respiratory capacity. SAV1 KD cells showed markedly elevated glycolysis and glycolytic reserves; blocking other fuel-oxidation pathways enhanced mitochondrial flexibility of glucose oxidation. Under limited glucose, endogenous fatty acid oxidation increased to cope with bioenergetic stress. Gene expression patterns after SAV1 KD suggested transcriptional upregulation of key metabolic network regulators to promote energy production and free radical scavenging that may prevent impaired lipid and glucose metabolism. In SAV1 KD cells, sirtuin signaling was the top enriched canonical pathway linked with enhanced mitochondrial ATP production. Collectively, we demonstrated that Hippo pathway inhibition in SAV1 KD cells induces multiple metabolic properties, including enhancing mitochondrial spare respiratory capacity or glycolytic reserve to cope with stress and upregulating metabolic pathways supporting elevated ATP demand, bioenergetics, and glycolysis and counteracting oxidative stress. In response to metabolic challenges, SAV1 KD cells can increase fatty acid oxidation or glucose-coupled oxidative phosphorylation capacity to compensate for substrate limitations or alternative fuel oxidation pathway inhibition.

S100A4 enhances protumor macrophage polarization by control of PPAR-γ-dependent induction of fatty acid oxidation

BackgroundThe peroxisome proliferator-activated receptor γ (PPAR-γ)-dependent upregulation of fatty acid oxidation (FAO) mediates protumor (also known as M2-like) polarization of tumor-associated macrophages (TAMs). However, upstream factors determining PPAR-γ upregulation in...

FSMP-15. EVALUATING THE ROLE OF LONG-CHAIN FATTY ACID METABOLISM IN PROMOTING GLIOBLASTOMA GROWTH

Glioblastomas (GBM) or Stage IV gliomas, are the most aggressive of primary brain tumors and are associated with high mortality and morbidity. Patients diagnosed with this lethal cancer have a dismal survival rate of 14 months and a 5-year survival rate of 5.6% despite a multimodal therapeutic approach, including surgery, radiation therapy, and chemotherapy. Aberrant lipid metabolism, particularly abnormally active de novo fatty acid synthesis, is recognized to have a key role in tumor progression and chemoresistance in cancers. Previous studies have reported a high expression of fatty acid synthase (FASN) in patient tumors, leading to multiple investigations of FASN inhibition as a treatment strategy. However, none of these have developed as efficacious therapies. Furthermore, when we profiled FASN expression using The Cancer Genome Atlas (TCGA) we determined that high FASN expression in GBM patients did not confer a worse prognosis (HR: 1.06; p-value: 0.51) and was not overexpressed in GBM tumors compared to normal brain. Therefore, we need to reexamine the role of exogenous fatty acid uptake over de novofatty acid synthesis as a potential mechanism for tumor progression. Our study aims to measure and compare fatty acid oxidation (FAO) of endogenous and exogenous fatty acids between GBM patients and healthy controls. Using TCGA, we have identified the overexpression of multiple enzymes involved in mediating the transfer and activation of long-chain fatty acids (LCFA) in GBM tumors compared to normal brain tissue. We are currently conducting metabolic flux studies to (1) assess the biokinetics of LCFA degradation and (2) establish exogenous versus endogenous LCFA preferences between patient-derived primary GBM cells and healthy glial and immune cells during steady state and glucose-deprivation.

1945-P: The Mitochondrial Protein DRP1 Is Required for Endogenous Fatty Acid Oxidation

Introduction: Mitochondria are organelles that routinely undergo fission and fusion in a process known as mitochondrial dynamics. Increased mitochondrial fission is implicated in the pathogenesis of obesity, type 2 diabetes, and metabolic disease. DRP1 has been identified as a key regulator of mitochondrial fission. We and others have demonstrated that loss of DRP1 is associated with improvements in glucose homeostasis, however, the role of DRP1 in mitochondrial fatty acid oxidation is unknown. The purpose of this study was to measure endogenous fatty acid oxidation in DRP1-deficient skeletal muscle cells. Methods: C2C12 myoblasts were stably transfected with DRP1 shRNA (KO) or a scrambled control (CT). Myoblasts were then cultured in a substrate-limited DMEM (0.5mM glucose, 0.5mM carnitine, 1mM glutamine, and 1% FBS) overnight. 15 minutes prior to assay, etomoxir was added to inhibit endogenous fatty acid oxidation (FAO). Cells were then challenged with either bovine serum albumin (BSA) or palmitate (PA) and assessed for mitochondrial function using whole cell respirometry (Agilent XFe24). Additionally, cells were treated with 14C-labeled PA in the absence and presence of increasing glucose concentrations to assess metabolic flexibility and fatty acid oxidation. Results: Basal whole cell (P<0.05) and endogenous fatty acid (P<0.05) oxygen consumption were reduced in KO cells relative to CT (P<0.05). Furthermore, ATP linked fatty acid oxidation was blunted in the KO cells (P<0.05). Conclusion: Here, we show that DRP1 is required for FAO. After inhibition of FAO with etomoxir, basal and ATP-linked respiration were ablated in DRP1-deficient cells. These findings suggest that loss of DRP1 ablates the ability of cells to use endogenous fatty acids as substrate when carbohydrate is not available. These data suggest that impaired lipid metabolism, rather than inhibition of mitochondrial fission, may explain improvements in glucose homeostasis with the loss of DRP1. Disclosure W.T. King: None. C.L. Axelrod: None. G. Davuluri: None. R.C. Noland: None. J.P. Kirwan: None. Funding National Institutes of Health (R01DK108089, U54GM104940, R01DK103860)

Silencing of Supramammillary Nucleus (SuMN) Dopaminergic Neuronal Activity Induces the Obese Glucose Intolerant State

The “westernized” diet reduces brain dopamine synthesis and low brain striatal dopamine function has been associated with and implicated in potentiation of metabolic syndrome in animals and humans. However, hypothalamic dopamine function may be a more dominant regulator of peripheral fuel metabolism. The hypothalamic SuMN dopaminergic neurons project to the lateral septum, medial preoptic area, and suprachiasmatic nucleus, areas that prominently regulate both peripheral metabolism and striatal dopamine function. However, a causal role for such hypothalamic dopaminergic circuitry in modulating peripheral metabolism under normal feeding conditions has never been explored. This study therefore tested the impact of SuMN-selective knock-down of tyrosine hydroxylase (TH, the rate-limiting enzyme for dopamine synthesis) gene expression using AAV-mediated shRNA targeting TH (AAV-TH shRNA) on peripheral fuel metabolism in 14 weeks old rats fed standard low fat rodent chow (RC). Rats were micro-infused at the SuMN with either AAV-TH shRNA (N=12) or AAV-scrambled shRNA [control] (N=10) and maintained on RC ad lib for 77 days to assess changes in body weight (BW), glucose tolerance and fat metabolism. SuMN-selective TH knock down increased BW gain (54%, p=0.0005), total white adipose tissue mass (43%, p=0.012), feed efficiency (BW gain /g food) (25%, p= 0.048) and cumulative food consumption (15%, p= 0.0001) vs. control. SuMN-selective TH knock down increased glucose tolerance test glucose area under curve (AUC) by 27% (p=0.009) with no change in insulin AUC (p=0.99) vs. control. Such knock down also reduced adipose tissue mitochondrial fatty acid oxidation (FAO) rate (basal oxygen consumption rate 51%, p< 0.0001; endogenous FAO rate 58%, p< 0.0001; and exogenous FAO 54%, p=0.024). These findings indicate that reduced SuMN dopaminergic neuronal activity facilitates dysglycemia and obesity while reducing fat energy expenditure and increasing food consumption. Disclosure Y. Zhang: Employee; Self; VeroScience, LLC. T. Tsai: Employee; Self; VeroScience, LLC. M. Ezrokhi: Employee; Self; VeroScience, LLC. C. Stoelzel: Employee; Self; VeroScience, LLC. A. Cincotta: Employee; Self; VeroScience, LLC.. Stock/Shareholder; Self; VeroScience, LLC..

Anaplerotic Role of Glucose in the Oxidation of Endogenous Fatty Acids during Dengue Virus Infection.

Dengue virus (DENV) is among the most important human arboviruses and is clinically and experimentally associated with lipid metabolism disorders. Using high-resolution respirometry, we analyzed the metabolic switches induced by DENV in a human hepatic cell line. This experimental approach allowed us to determine the contribution of fatty acids, glutamine, glucose, and pyruvate to mitochondrial bioenergetics, shedding light on the mechanisms involved in DENV-induced metabolic alterations. We found that while infection strongly inhibits glutamine oxidation, it increases the cellular capacity of metabolizing glucose; remarkably, though, this substrate, instead being used as an energy source, performs an anaplerotic role in the oxidation of endogenous lipids. Fatty acids become the main energetic substrate in infected cell, and through the pharmacological modulation of β-oxidation we demonstrated that this pathway is essential for virus replication. Interestingly, infected cells were much less susceptible to the Crabtree effect, i.e., the glucose-mediated inhibition of mitochondrial oxygen consumption, suggesting that infection favors cellular respiration by increasing ADP availability. IMPORTANCE Dengue virus infection is a major cause of human arbovirosis, for which clinical and experimental evidence supports the idea that liver dysfunction and lipid metabolism disorders are characteristics of severe disease. Analyzing mitochondrial bioenergetics, here we show that infection of hepatic cells with dengue virus favors the cellular capacity of metabolizing glucose, impairing the normal metabolic flexibility that allows the oxidative machinery to switch among the main energetic substrates. However, instead of being used as an energy source, glucose performs an anaplerotic role in the oxidation of endogenous fatty acids, which become the main energetic substrate during infection. Taken together, the results shed light on metabolic mechanisms that may explain the profound alterations in lipid metabolism for severe dengue patients, contributing to the understanding of dengue physiopathology.

The Action of <i>p</i>-Synephrine on Lipid Metabolism in the Perfused Rat Liver

p-synephrine and p-octopamine were found to increase lipolysis in adipocytes. The present study approaches the question if these compounds, natural products of the bitter orange (Citrus aurantium fruit), increase lipolysis and fatty acid oxidation in the liver. Experiments were done in the perfused rat liver. Non-recirculating hemoglobin-free perfusion was done using the Krebs/ Henseleit-bicarbonate buffer (pH 7.4) as perfusion fluid. Both p-synephrine and p-octopamine, at the concentrations of 100 μM, were found to stimulate the hepatic triacylglycerol lipase by 40% and 51%, respectively. These seem to be the maximal stimulations possible in the liver. In the perfused liver, p-synephrine, when present at an initial concentration of 500 μM, was able to increase the non-esterified fatty acid release after one hour of recirculating perfusion. The effects of p-synephrine on the oxidation of exogenously supplied [1-14C]octanoate and [1-14C]oleate were minimal. Only oxygen uptake, already stimulated by octanoate or oleate, was additionally increased by the infusion of p-synephrine. These results contrast with those obtained in a previous study with p-octopamine, which increased the production of 14CO2 from both [1-14C]octanoate and [1-14C]oleate. Apparently only the oxidation of endogenous fatty acids is stimulated by p-synephrine. On the other hand, both p-synephrine and p-octopamine stimulate the hepatic triacylglycerol lipase to a much lesser extent than the adipocyte lipase. It can be concluded that p-synephrine affects much more carbohydrate metabolism in the liver than lipid metabolism.

Abstract 13015: Inhibition of Mitochondrial Calcium Uptake in the Heart Does Not Compromise Cardiac Energetics or Function Due to a Preferential Increase in Fatty Acid Oxidation During Increased Cardiac Work

The mitochondrial calcium (Ca 2+ ) uniporter (MCU) is primarily responsible for relaying cytosolic Ca 2+ transients to the mitochondria. Increasing cardiac workload by positive inotropes results in increased mitochondrial Ca 2+ , increasing glucose oxidation in comparison to fatty acid oxidation. This has been attributed to Ca 2+ stimulation of pyruvate dehydrogenase, the rate-limiting enzyme in glucose oxidation. We examined what effect increasing workload has on cardiac energetics and cardiac function if mitochondrial Ca 2+ levels are decreased in hearts from inducible cardio-specific deleted MCU (MCU -/- ) mice. Hearts from MCU -/- and MCU fl/fl control mice were isolated and perfused as working hearts with 5 mM glucose, 0.8 mM palmitate, 3% albumin, ± insulin (100 μU/ml) and isoproterenol (10 nM). Surprisingly, MCU -/- hearts showed increased cardiac work (p<0.05 vs. MCU fl/fl ). MCU -/- hearts were also not energy-starved, displaying basal rates of glucose and fatty acid oxidation comparable to controls, and even higher glucose oxidation in response to insulin (3027±213 vs. 2268±166 nmol . g dry wt -1. min -1 in MCU fl/fl , p <0.05). In response to isoproterenol treatment, MCU -/- hearts showed a similar increase in glucose oxidation, compared to controls. However, palmitate oxidation increased to a larger extent in MCU -/- hearts compared to controls (793±60 vs. 558±55 nmol . g dry wt -1. min -1 , p <0.05, respectively), resulting in a greater reliance on fatty acid oxidation for ATP production (40.1±1.9 % vs. 32.7±2.1% in MCU fl/fl , p <0.05). This high fatty acid oxidation supported the higher energy demand at increased workload and provided a higher energy reserve as evident by higher acetyl-CoA/ CoA ratios (0.36±0.03 vs. 0.26±0.02 in MCU fl/fl , p<0.05). The rise in fatty acid oxidation correlated with lower levels of malonyl CoA, an endogenous fatty acid oxidation inhibitor, (4.50±0.29 vs. 8.66±0.98 nmol . g dry wt -1 in MCU fl/fl , p< 0.05), and to an increase in acetylation of the fatty acid oxidation enzyme 3-hydroxyacyl CoA dehydrogenase (which stimulates activity). These results suggest that low mitochondrial Ca 2+ does not compromise cardiac energetics due to a compensatory stimulation of fatty acid oxidation.

Impaired energy metabolism of the taurine‑deficient heart.

Taurine is a β-amino acid found in high concentrations in excitable tissues, including the heart. A significant reduction in myocardial taurine content leads to the development of a unique dilated, atrophic cardiomyopathy. One of the major functions of taurine in the heart is the regulation of the respiratory chain. Hence, we tested the hypothesis that taurine deficiency-mediated defects in respiratory chain function lead to impaired energy metabolism and reduced ATP generation. We found that while the rate of glycolysis was significantly enhanced in the taurine-deficient heart, glucose oxidation was diminished. The major site of reduced glucose oxidation was pyruvate dehydrogenase, an enzyme whose activity is reduced by the increase in the NADH/NAD+ ratio and by decreased availability of pyruvate for oxidation to acetyl CoA and changes in [Mg2+]i. Also diminished in the taurine-deficient heart was the oxidation of two other precursors of acetyl CoA, endogenous fatty acids and exogenous acetate. In the taurine-deficient heart, impaired citric acid cycle activity decreased both acetate oxidation and endogenous fatty acid oxidation, but reductions in the activity of the mitochondrial transporter, carnitine palmitoyl transferase, appeared to also contribute to the reduction in fatty acid oxidation. These changes diminished the rate of ATP production, causing a decline in the phosphocreatine/ATP ratio, a sign of reduced energy status. The findings support the hypothesis that the taurine-deficient heart is energy starved primarily because of impaired respiratory chain function, an increase in the NADH/NAD+ ratio and diminished long chain fatty acid uptake by the mitochondria. The results suggest that improved energy metabolism contributes to the beneficial effect of taurine therapy in patients suffering from heart failure.

Adrenergic Metabolic and Hemodynamic Effects of Octopamine in the Liver

The fruit extracts of Citrus aurantium (bitter orange) are traditionally used as weight-loss products and as appetite suppressants. A component of these extracts is octopamine, which is an adrenergic agent. Weight-loss and adrenergic actions are always related to metabolic changes and this work was designed to investigate a possible action of octopamine on liver metabolism. The isolated perfused rat liver was used to measure catabolic and anabolic pathways and hemodynamics. Octopamine increased glycogenolysis, glycolysis, oxygen uptake, gluconeogenesis and the portal perfusion pressure. Octopamine also accelerated the oxidation of exogenous fatty acids (octanoate and oleate), as revealed by the increase in 14CO2 production derived from 14C labeled precursors. The changes in glycogenolysis, oxygen uptake and perfusion pressure were almost completely abolished by α1-adrenergic antagonists. The same changes were partly sensitive to the β-adrenergic antagonist propranolol. It can be concluded that octopamine accelerates both catabolic and anabolic processes in the liver via adrenergic stimulation. Acceleration of oxygen uptake under substrate-free perfusion conditions also means acceleration of the oxidation of endogenous fatty acids, which are derived from lipolysis. All these effects are compatible with an overall stimulating effect of octopamine on metabolism, which is compatible with its reported weight-loss effects in experimental animals.

Trimetazidine Reduces Endogenous Free Fatty Acid Oxidation and Improves Myocardial Efficiency in Obese Humans

The metabolic modulator trimetazidine (TMZ) has been suggested to induce a metabolic shift from myocardial fatty acid oxidation (FAO) to glucose utilization, but this mechanism remains unproven in humans. The oxidation of plasma derived FA is commonly measured in humans, whereas the contribution of FA from triglycerides stored in the myocardium has been poorly characterized. To verify the hypothesis that TMZ induces a metabolic shift, we combined positron emission tomography (PET) and magnetic resonance spectroscopy ((1)H-MRS) to measure myocardial FAO from plasma and intracellular lipids, and myocardial glucose metabolism. Nine obese subjects were studied before and after 1 month of TMZ treatment. Myocardial glucose and FA metabolism were assessed by PET with (18)F-fluorodeoxyglucose and (11)C-palmitate. (1)H-MRS was used to measure myocardial lipids, the latter being integrated into the PET data analysis to quantify myocardial triglyceride turnover. Myocardial FAO derived from intracellular lipids was at least equal to that of plasma FAs (P = NS). BMI and cardiac work were positively associated with the oxidation of plasma derived FA (P ≤ 0.01). TMZ halved total and triglyceride-derived myocardial FAO (32.7 ± 8.0 to 19.6 ± 4.0 μmol/min and 23.7 ± 7.5 to 10.3 ± 2.7 μmol/min, respectively; P ≤ 0.05). These changes were accompanied by increased cardiac efficiency since unchanged LV work (1.6 ± 0.2 to 1.6 ± 0.1 Watt/g × 10(2), NS) was associated with decreased work energy from the intramyocardial triglyceride oxidation (1.6 ± 0.5 to 0.4 ± 0.1 Watt/g × 10(2), P = 0.036). In obese subjects, we demonstrate that myocardial intracellular triglyceride oxidation significantly provides FA-derived energy for mechanical work. TMZ reduced the oxidation of triglyceride-derived myocardial FAs improving myocardial efficiency.

On the Control of Metabolic Remodeling in Mitochondria of the Failing Heart

Received June 5, 2009; accepted June 9, 2009. The metabolic phenotype of the failing heart may be defined as follows.1 Metabolism remodels in the failing heart, leading to a loss in energy reserve and the inability to increase ATP supply. Ultimately, this metabolic rigidity leads to a fall in ATP. The likely time line is decreased energy reserve via the phosphotransferase reactions (creatine kinase [CK] and adenylate kinase) leading to increases in ADP and AMP, triggering an increase in glycolysis. Although the contribution of glycolysis to overall ATP synthesis increases at least in the hypertrophied heart, glycolytic reserve is limited. Importantly, as heart failure evolves, ATP synthesis from oxidation of both endogenous and exogenous fatty acids by mitochondria, the major source of ATP in the heart, falls.2 Article see p 342 Remodeling of the failing myocardium is controlled by energy sensors such as AMP that lead to changes in phosphorylation state (as well as other chemical modifications) of many proteins for short-term preservation of ATP and by activation of transcription factors that coordinately control long-term remodeling of entire ATP synthesis and utilizing pathways. Given that the requirement for ATP for all metabolic processes and for cell viability is absolute, a renewed interest in metabolism has led to identification of the molecular links between physiological and metabolic stimuli and the regulation of gene expression in the heart. We not only have identified the metabolic targets of specific nuclear receptors and DNA-binding transcriptional activators but also are beginning to learn how their signals are amplified and sustained to remodel metabolism. Transcription is activated when transcriptional activators, including peroxisome proliferator-activated receptors, estrogen receptors, retinoid receptors, nuclear respiratory factors, and myocyte-enhancing factor-2 (MEF-2) form protein-protein complexes with the peroxisome proliferator-activated receptor-γ coactivators, namely PGC-1α and β, tethering PGC-1s to DNA (Figure). When …

Cell death–inducing DFF45-like effector C is reduced by caloric restriction and regulates adipocyte lipid metabolism

Members of the cell death–inducing DFF45-like effector (CIDE) gene family have been shown to regulate lipid metabolism. In this article, we report that the third member of the human CIDE family, CIDEC, is down-regulated in response to a reduced caloric intake. The down-regulation was demonstrated by microarray and real-time polymerase chain reaction analysis of subcutaneous adipose tissue in 2 independent studies on obese patients undergoing treatment with a very low calorie diet. By analysis of CIDEC expression in 65 human tissues, we conclude that human CIDEC is predominantly expressed in subcutaneous adipocytes. Together, these observations led us to investigate the effect of decreased CIDEC expression in cultured 3T3-L1 adipocytes. Small interfering RNA–mediated knockdown of CIDEC resulted in an increased basal release of nonesterified fatty acids, decreased responsiveness to adrenergic stimulation of lipolysis, and increased oxidation of endogenous fatty acids. Thus, we suggest that CIDEC is a regulator of adipocyte lipid metabolism and may be important for the adipocyte to adapt to changes in energy availability.

Aldosterone and Metabolic Syndrome

The classic role of aldosterone is to regulate water and electrolyte balance and, therefore, blood pressure homeostasis.1 Apart from that, experimental studies have demonstrated that aldosterone induces structural and functional alterations in the heart, kidneys, and vessels with effects such as myocardial fibrosis, nephrosclerosis, vascular inflammation and remodeling, and disturbed fibrinolysis.2,3 This damage seems to be aldosterone mediated, and aldosterone blockade with mineralocorticoid receptor (MR) antagonists, such as spironolactone, may prevent the onset of these effects.4,5 On the other hand, it cannot completely be ruled out that potassium and high blood pressure also play additional key roles in this damage.6,7 This evidence has impressively been supported by clinical studies, such as the Randomized Aldactone Evaluation Study and the Eplerenone Post-Acute Myocardial Infarction Survival and Efficacy Study.8,9 For example, increased mortality in patients with chronic heart failure has been associated with elevated aldosterone plasma levels,10 and high circulating plasma aldosterone levels predict the clinical outcome in patients after myocardial infarction.11 Furthermore, primary aldosteronism (PA) has been demonstrated to enhance the risk of cardiovascular events12 and kidney disease.13 In summary, aldosterone is considered a cardiovascular risk factor, promoting disease processes such as cardiac fibrosis, nephrosclerosis, and arteriosclerosis,2,3,14 all of which are increased in patients with obesity and the metabolic syndrome.15,16 The term “metabolic syndrome” (MSyn) has evolved various definitions in recent times; most of the studies introduced here use slight modifications. Nevertheless, all of the definitions used have a common denominator, which is reflected in a definition by the American Heart Association/National Heart, Lung, and Blood Institute.17 According to this definition, the MSyn is considered as a constellation of interrelated risk factors of metabolic origin, including arterial hypertension, dyslipidemias, alterations in glucose homeostasis with type 2 diabetes mellitus, and abdominal …

Life and death decisions of the pancreatic β-cell: the role of fatty acids

Both stimulatory and detrimental effects of NEFAs (non-esterified fatty acids) on pancreatic beta-cells have been recognized. Acute exposure of the pancreatic beta-cell to high glucose concentrations and/or saturated NEFAs results in a substantial increase in insulin release, whereas chronic exposure results in desensitization and suppression of secretion, followed by induction of apoptosis. Some unsaturated NEFAs also promote insulin release acutely, but they are less toxic to beta-cells during chronic exposure and can even exert positive protective effects. Therefore changes in the levels of NEFAs are likely to be important for the regulation of beta-cell function and viability under physiological conditions. In addition, the switching between endogenous fatty acid synthesis or oxidation in the beta-cell, together with alterations in neutral lipid accumulation, may have critical implications for beta-cell function and integrity. Long-chain acyl-CoA (formed from either endogenously synthesized or exogenous fatty acids) controls several aspects of beta-cell function, including activation of specific isoenzymes of PKC (protein kinase C), modulation of ion channels, protein acylation, ceramide formation and/or NO-mediated apoptosis, and transcription factor activity. In this review, we describe the effects of exogenous and endogenous fatty acids on beta-cell metabolism and gene and protein expression, and have explored the outcomes with respect to insulin secretion and beta-cell integrity.

The metabolic effects of thia fatty acids in rat liver depend on the position of the sulfur atom

The effects on oxidation and composition of fatty acids in rat liver were compared after administration of fatty acids with sulfur substituted in different positions. It has been hypothesized that drugs with hydrophobic backbone have lipid-lowering effects because they are not easily catabolized by mitochondrial β-oxidation. Thia fatty acids cannot be β-oxidized when sulfur is in 3-position, but β-oxidation is possible when sulfur is positioned further from the carboxyl group. To investigate whether catabolism of thia fatty acids would affect their ability to influence lipid metabolism, a series of thia fatty acids were synthesized and administered by oral gavage to male Wistar rats (300 mg/kg bodyweight/day for 7 days). Depending on the position of the sulfur atom and the chain length, the thia fatty acids were β-oxidized, desaturated and/or elongated, and the accumulated amounts were lower as the sulfur atom were positioned further from the carboxyl group. All thia fatty acids led to high peroxisomal β-oxidation of endogenous fatty acids, whereas the mitochondrial β-oxidation was high when sulfur was in 3-position, low when sulfur was in 4-position and similar to controls when sulfur was in 5- or 7-position. The changes in hepatic fatty acid composition were more pronounced when sulfur was positioned close to the carboxyl group. In conclusion, both the position of the sulfur atom and the chain length appear to determine the catabolic fate of thia fatty acids, and the non-β-oxidizable thia fatty acids were most potent in regulating oxidation and composition of endogenous fatty acids in rat liver.

Cytokine regulation of skeletal muscle fatty acid metabolism: effect of interleukin-6 and tumor necrosis factor-alpha.

IL-6 and TNF-alpha have been associated with insulin resistance and type 2 diabetes. Furthermore, abnormalities in muscle fatty acid (FA) metabolism are strongly associated with the development of insulin resistance. However, few studies have directly examined the effects of either IL-6 or TNF-alpha on skeletal muscle FA metabolism. Here, we used a pulse-chase technique to determine the effect of IL-6 (50-5,000 pg/ml) and TNF-alpha (50-5,000 pg/ml) on FA metabolism in isolated rat soleus muscle. IL-6 (5,000 pg/ml) increased exogenous and endogenous FA oxidation by approximately 50% (P < 0.05) but had no effect on FA uptake or incorporation of FA into endogenous lipid pools. In contrast, TNF-alpha had no effect on FA oxidation but increased FA incorporation into diacylglycerol (DAG) by 45% (P < 0.05). When both IL-6 (5,000 pg/ml) and insulin (10 mU/ml) were present, IL-6 attenuated insulin's suppressive effect on FA oxidation, increasing exogenous FA oxidation (+37%, P < 0.05). Furthermore, in the presence of insulin, IL-6 reduced the esterification of FA to triacylglycerol by 22% (P < 0.05). When added in combination with IL-6 or leptin (10 microg/ml), the TNF-alpha-induced increase in DAG synthesis was inhibited. In conclusion, the results demonstrate that IL-6 plays an important role in regulating fat metabolism in muscle, increasing rates of FA oxidation, and attenuating insulin's lipogenic effects. In contrast, TNF-alpha had no effect on FA oxidation but increased FA incorporation into DAG, which may be involved in the development of TNF-alpha-induced insulin resistance in skeletal muscle.

The stimulus–secretion coupling of amino acid-induced insulin release. Insulinotropic action of l-alanine

Available information on the fate and insulinotropic action of l-alanine in isolated pancreatic islets is restricted to data collected in obese hyperglycemic mice. Recent data, however, collected mostly in tumoral islet cells of either the RINm5F line or BRIN-BD11 line, have drawn attention to the possible role of Na + co-transport in the insulinotropic action of l-alanine. In the present study conducted in islets prepared from normal adult rats, l-alanine was found (i) to inhibit pyruvate kinase in islet homogenates, (ii) not to affect the oxidation of endogenous fatty acids in islets prelabelled with [U- 14C]palmitate, (iii) to stimulate 45Ca uptake in islets deprived of any other exogenous nutrient, and (iv) to augment insulin release evoked by either 2-ketoisocaproate or l-leucine, whilst failing to significantly affect glucose-induced insulin secretion. The oxidation of l-[U- 14C]alanine was unaffected by d-glucose, but inhibited by l-leucine. Inversely, l-alanine decreased the oxidation of d-[U- 14C]glucose, but failed to affect l-[U- 14C]leucine oxidation. It is concluded that the occurrence of a positive insulinotropic action of l-alanine is restricted to selected experimental conditions, the secretory data being compatible with the view that stimulation of insulin secretion by the tested nutrient(s) reflects, as a rule, their capacity to augment ATP generation in the islet B cells. However, the possible role of Na + co-transport in the secretory response to l-alanine cannot be ignored.

Concise review of the glutathione S-transferases and their significance to toxicology.

The glutathione S-transferases (GSTs; E.C. 2.5.1.18) are a multi-gene family of enzymes involved in the detoxification, and, in a few instances, activation of a wide variety of chemicals (Table 1). Since their discovery in the early 1970s, thousands of articles have been written on the structure, function, and toxicologic significance of GSTs, and it is obviously beyond the scope of this article to provide a comprehensive review of the field. This article will highlight the major features of this multi-gene family of enzymes, with a focus on human soluble (cytosolic) GSTs. A relatively comprehensive review of the GSTs was published in 1995, and provides an excellent source for detailed information and references about nearly all aspects of GSTs up to that time (Hayes and Pulford, 1995). (Because of space limitations, the present mini-review is not fully referenced, but it does include some newer references not covered in previous reviews.) GSTs catalyze the nucleophilic attack of glutathione (GSH) on electrophilic substrates, thereby decreasing their reactivity with cellular macromolecules (Armstrong, 1997). These proteins have been found in virtually every living species examined, including plants, animals, and bacteria (Hayes and Pulford, 1995). Most GSTs exist as soluble enzymes, although a small family of microsomal GSTs has been characterized (Andersson et al., 1994; Jakobsson et al., 1996), and a mitochondrial GST, referred to as GST Kappa, has also been identified (Pemble et al., 1996). In humans and in rodents, the soluble GSTs are collectively expressed in rather large amounts, constituting as much as 4% of total soluble protein in the liver. Although no definitive physiological role for GSTs has yet been identified, several reactive endogenous molecules, including ab-unsaturated keto prostaglandins (e.g., PGA2, PGJ2), and endogenous fatty acid oxidation products, such as 4-hydroxy-2-nonenal, serve as substrates for certain GSTs (Hayes and Pulford, 1995) (Table 2). One human GST (hGSTM2-2) has recently been found to exhibit unique and high catalytic activity toward reactive quinones of endogenous catecholamines (e.g., dopaminochrome, formed by oxidation of dopamine), and thus may play some protective role against endogenous oxidative tissue damage in the brain (Baez et al., 1997). The soluble forms exist as dimeric proteins, with subunit molecular weights of approximately 25 kDa (Table 1). Each subunit of the dimeric enzyme has an active site composed of 2 distinct functional regions: a hydrophilic G-site, which binds the physiological substrate glutathione, and an adjacent H-site which provides a hydrophobic environment for the binding of structurally diverse electrophilic substrates (Armstrong, 1997). The G-site is highly conserved between all GSTs due to its high specificity for GSH, whereas the H-site can be quite divergent between different GSTs, and exhibits broad and variable substrate binding specificity.