Elevated Free Fatty Acid Levels Research Articles

Letters to the Editor

Letters to the Editor

Association of Elevated Free Fatty Acids During Late Pregnancy With Preterm Delivery

To examine the association between moderately elevated maternal plasma free fatty acids (FFAs) during late pregnancy and preterm delivery. In a prospective observational cohort with 523 healthy pregnant women, fasting plasma FFAs were measured during the third trimester. Socioeconomic, demographic, and anthropometric measures were collected at entry to prenatal care, and pregnancy outcomes were abstracted from medical record at delivery. After control for confounders including prepregnant body mass index (multiple logistic regression analysis), women who had moderately elevated plasma FFAs (in the highest tertile), showed a greater than threefold increased risk of preterm delivery (adjusted odds ratio (AOR) 3.49, 95% (CI) 1.73-7.03, P<.001). The associations persisted in women who had spontaneous preterm delivery (AOR 2.35, 95% CI 1.05-5.28, P<.05) and after excluding women with gestational diabetes mellitus and preeclampsia (AOR 3.30, 95% CI 1.38-7.87, P<.01). Additional stratified analyses showed that the association of high maternal FFAs and increased risk of preterm delivery was independent of prepregnant obesity. Elevated fasting plasma FFA levels at 30 weeks of gestation were associated with an increased risk of preterm delivery. This effect was independent of prepregnant obesity and several other known risk factors for preterm delivery, including cigarette smoking, ethnicity, and prior preterm delivery. These data may have important clinical significance because they provide a possible link between preterm delivery and high lipid levels, a known risk factor for cardiovascular disease.

Elevated Toll-Like Receptor 4 Expression and Signaling in Muscle From Insulin-Resistant Subjects

OBJECTIVE— Tall-like receptor (TLR)4 has been implicated in the pathogenesis of free fatty acid (FFA)-induced insulin resistance by activating inflammatory pathways, including inhibitor of κB (IκB)/nuclear factor κB (NFκB). However, it is not known whether insulin-resistant subjects have abnormal TLR4 signaling. We examined whether insulin-resistant subjects have abnormal TLR4 expression and TLR4-driven (IκB/NFκB) signaling in skeletal muscle.RESEARCH DESIGN AND METHODS— TLR4 gene expression and protein content were measured in muscle biopsies in 7 lean, 8 obese, and 14 type 2 diabetic subjects. A primary human myotube culture system was used to examine whether FFAs stimulate IκB/NFκB via TLR4 and whether FFAs increase TLR4 expression/content in muscle.RESULTS— Obese and type 2 diabetic subjects had significantly elevated TLR4 gene expression and protein content in muscle. TLR4 muscle protein content correlated with the severity of insulin resistance. Obese and type 2 diabetic subjects also had lower IκBα content, an indication of elevated IκB/NFκB signaling. The increase in TLR4 and NFκB signaling was accompanied by elevated expression of the NFκB-regulated genes interleukin (IL)-6 and superoxide dismutase (SOD)2. In primary human myotubes, acute palmitate treatment stimulated IκB/NFκB, and blockade of TLR4 prevented the ability of palmitate to stimulate the IκB/NFκB pathway. Increased TLR4 content and gene expression observed in muscle from insulin-resistant subjects were reproduced by treating myotubes from lean, normal-glucose-tolerant subjects with palmitate. Palmitate also increased IL-6 and SOD2 gene expression, and this effect was prevented by inhibiting NFκB.CONCLUSIONS— Abnormal TLR4 expression and signaling, possibly caused by elevated plasma FFA levels, may contribute to the pathogenesis of insulin resistance in humans.

Deleterious action of FA metabolites on ATP synthesis: possible link between lipotoxicity, mitochondrial dysfunction, and insulin resistance

Insulin resistance is a characteristic feature of type 2 diabetes and obesity. Insulin-resistant individuals manifest multiple disturbances in free fatty acid (FFA) metabolism and have excessive lipid accumulation in insulin target tissues. Although much evidence supports a causal role for altered FFA metabolism in the development of insulin resistance, i.e., "lipotoxicity", the intracellular mechanisms by which elevated plasma FFA levels cause insulin resistance have yet to be completely elucidated. Recent studies have implicated a possible role for mitochondrial dysfunction in the pathogenesis of insulin resistance in skeletal muscle. We examined the effect of FFA metabolites [palmitoyl carnitine (PC), palmitoyl-coenzyme A (CoA), and oleoyl-CoA] on ATP synthesis in mitochondria isolated from mouse and human skeletal muscle. At concentrations ranging from 0.5 to 2 microM, these FFA metabolites stimulated ATP synthesis; however, above 5 microM, there was a dose-response inhibition of ATP synthesis. Furthermore, 10 microM PC inhibits ATP synthesis from pyruvate. Elevated PC concentrations (> or =10 microM) inhibit electron transport chain activity and decrease the mitochondrial inner membrane potential. These acquired mitochondrial defects, caused by a physiological increase in the concentration of FFA metabolites, provide a mechanistic link between lipotoxicity, mitochondrial dysfunction, and muscle insulin resistance.

CHANGES IN PLASMA FREE FATTY ACID LEVELS IN SEPTIC PATIENTS ARE ASSOCIATED WITH CARDIAC DAMAGE AND REDUCTION IN HEART RATE VARIABILITY

Free fatty acids (FFAs) have been shown to produce alteration of heart rate variability (HRV) in healthy and diabetic individuals. Changes in HRV have been described in septic patients and in those with hyperglycemia and elevated plasma FFA levels. We studied if sepsis-induced heart damage and HRV alteration are associated with plasma FFA levels in patients. Thirty-one patients with sepsis were included. The patients were divided into two groups: survivors(n = 12) and nonsurvivors (n = 19). The following associations were investigated: (a) troponin I elevation and HRV reduction and (b) clinical evolution and HRV index, plasma troponin, and plasma FFA levels. Initial measurements of C-reactive protein and gravity Acute Physiology and Chronic Health Evaluation scores were similar in both groups. Overall, an increase in plasma troponin level was related to increased mortality risk. From the first day of study, the nonsurvivor group presented a reduced left ventricular stroke work systolic index and a reduced low frequency (LF) that is one of HRV indexes. The correlation coefficient for LF values and troponin was r(2) = 0.75 (P < 0.05). All patients presented elevated plasma FFA levels on the first day of the study (5.11 +/- 0.53 mg/mL), and this elevation was even greater in the nonsurvivor group compared with the survivors (6.88 +/- 0.13 vs. 3.85 +/- 0.48 mg/mL, respectively; P < 0.05). Cardiac damage was confirmed by measurement of plasma troponin I and histological analysis. Heart dysfunction was determined by left ventricular stroke work systolic index and HRV index in nonsurvivor patients. A relationship was found between plasma FFA levels, LFnu index, troponin levels, and histological changes. Plasma FFA levels emerged as possible cause of heart damage in sepsis.

Increased mitochondrial uncoupling proteins, respiratory uncoupling and decreased efficiency in the chronically infarcted rat heart

Heart failure patients have abnormal cardiac high energy phosphate metabolism, the explanation for which is unknown. Patients with heart failure also have elevated plasma free fatty acid (FFA) concentrations. Elevated FFA levels are associated with increased cardiac mitochondrial uncoupling proteins (UCPs), which, in turn, are associated with decreased mitochondrial respiratory coupling and low cardiac efficiency. Here, we determined whether increased mitochondrial UCP levels contribute to decreased energetics in the failing heart by measuring UCPs and respiration in mitochondria isolated from the viable myocardium of chronically infarcted rat hearts and measuring efficiency (hydraulic work/O 2 consumption) in the isolated, working rat heart. Ten weeks after infarction, cardiac levels of UCP3 were increased by 53% in infarcted, failing hearts that had ejection fractions less than 45%. Cardiac UCP3 levels correlated positively with non-fasting plasma FFAs ( r = 0.81; p < 0.01). Mitochondria from failing hearts were less coupled than those from control hearts, as demonstrated by the lower ADP/O ratio of 1.9 ± 0.1 compared with 2.5 ± 0.2 in controls ( p < 0.05). The decreased ADP/O ratio was reflected in an efficiency of 14 ± 2% in the failing hearts when perfused with 1 mM palmitate, compared with 20 ± 1% in controls ( p < 0.05). We conclude that failing hearts have increased UCP3 levels that are associated with high circulating FFA concentrations, mitochondrial uncoupling, and decreased cardiac efficiency. Thus, respiratory uncoupling may underlie the abnormal energetics and low efficiency in the failing heart, although whether this is maladaptive or adaptive would require direct investigation.

Free fatty acids as mediators of adaptive compensatory responses to insulin resistance in dexamethasone‐treated rats

Chronic low-dose dexamethasone (DEX) treatment in rats is associated to insulin resistance with compensatory hyperinsulinaemia and reduction in food intake. We tested the hypothesis that the elevation in circulating free fatty acids (FFAs) induced by DEX is the common mediator of both insulin resistance and insulin hyperproduction. For this purpose, an anti-lipolytic agent was administered during DEX treatment to lower lipacidaemia for several hours prior to glucose and insulin tolerance tests. Leptin expression in adipose tissue (by Northern blot) and plasma leptin levels (by radioimmunoassay) were also investigated to verify whether a rise in circulating leptin could be responsible for the anorectic effect of DEX. Our data show that a transient pharmacological reduction of elevated plasma FFA levels abates the post-loading hyperinsulinaemia and counteracts the insulin resistance induced by DEX, supporting the hypothesis that the chronic elevation in FFAs is the common mediator of DEX-induced changes. Despite enhanced leptin expression in white adipose tissue, DEX-treated rats show no significant increase in plasma leptin levels. This suggests that the anorectic effect of DEX should be mediated, at least partially, by other factors, possibly related to the influence of concomitantly elevated plasma FFA and insulin levels on the hypothalamic centers regulating feeding. Our results sustain the idea that a prolonged increase in plasma FFA levels plays an important role in the adaptive regulation of glucose and energy homeostasis, not only by potentiating insulin secretion but also by providing a signal of 'nutrient abundance' capable of restraining food intake.

Differential activation of ER stress and apoptosis in response to chronically elevated free fatty acids in pancreatic β-cells

Chronic exposure to elevated saturated free fatty acid (FFA) levels has been shown to induce endoplasmic reticulum (ER) stress that may contribute to promoting pancreatic beta-cell apoptosis. Here, we compared the effects of FFAs on apoptosis and ER stress in human islets and two pancreatic beta-cell lines, rat INS-1 and mouse MIN6 cells. Isolated human islets cultured in vitro underwent apoptosis, and markers of ER stress pathways were elevated by chronic palmitate exposure. Palmitate also induced apoptosis in MIN6 and INS-1 cells, although the former were more resistant to both apoptosis and ER stress. MIN6 cells were found to express significantly higher levels of ER chaperone proteins than INS-1 cells, which likely accounts for the ER stress resistance. We attempted to determine the relative contribution that ER stress plays in palmitate-induced beta-cell apoptosis. Although overexpressing GRP78 in INS-1 cells partially reduced susceptibility to thapsigargin, this failed to reduce palmitate-induced ER stress or apoptosis. In INS-1 cells, palmitate induced apoptosis at concentrations that did not result in significant ER stress. Finally, MIN6 cells depleted of GRP78 were more susceptible to tunicamycin-induced apoptosis but not to palmitate-induced apoptosis compared with control cells. These results suggest that ER stress is likely not the main mechanism involved in palmitate-induced apoptosis in beta-cell lines. Human islets and MIN6 cells were found to express high levels of stearoyl-CoA desaturase-1 compared with INS-1 cells, which may account for the decreased susceptibility of these cells to the cytotoxic effects of palmitate.

Free fatty acids as a cardiovascular risk factor

Free fatty acids (FFAs) serve as physiologically important energy substrates and their release from the adipose tissue by lipolysis is regulated according to the energy demands of the body. FFAs are increased in obese patients and contribute to type 2 diabetes, hepatic steatosis and several cardiovascular diseases. In patients with heart failure and acute coronary syndromes, elevated FFA levels are a consequence of an increased lipolysis due to a surge in catecholamines and natriuretic peptides. FFAs contribute to myocardial dysfunction and are proarrhythmic, and their oxidation requires more oxygen than does glycolysis. Therapeutic approaches have already emerged that aim to reduce the uptake and/or oxidation of fatty acids in the myocardium. The routine use of FFAs as a diagnostic tool is limited by their high variability, this being strongly influenced by nutrition and the effects of several hormones. In addition, it remains to be clarified whether fasting or postprandial values or dynamic measurements such as changes in FFA concentrations induced by stress are better parameters for evaluating cardiovascular risk. In this review, we present an overview of the metabolism and role of FFAs as a cardiovascular risk factor and discuss the potential of FFAs in the diagnosis and treatment of cardiovascular diseases.

Rosiglitazone Prevents Free Fatty Acid-Induced Vascular Endothelial Dysfunction

Elevated plasma free fatty acid (FFA) levels contribute significantly to insulin resistance, while thiazolidinediones (TZDs) improve insulin resistance and lower plasma FFA levels in diabetic patients by acting as agonists of the peroxisome proliferator-activated receptor-γ (PPARγ). In particular, the TZD rosiglitazone improves endothelial function and increases nitric oxide release in patients with type 2 diabetes. It would be expected that reducing FFA by activating PPARγ would improve triglyceride/heparin-induced endothelial dysfunction. This randomized, double blind, placebo-controlled study examined the effects of 8 mg of rosiglitazone daily or placebo, given orally for 3 weeks to 16 healthy males. All subjects received 20% Intralipid, a triglyceride emulsion, and heparin by intravenous infusion to achieve FFA levels known to produce endothelial dysfunction and insulin resistance. Forearm blood flow (FBF) responses to acetylcholine, an endothelium-dependent vasodilator, and nitroglycerine, an endothelium-independent vasodilator, were measured in both arms by strain-gauge plethysmography. Circulating levels of C-reactive protein (CRP) and asymmetric dimethylarginine (ADMA), an endogenous nitric oxide synthase inhibitor, also were measured. Rosiglitazone administration did not alter FBF reactivity. Infusing triglyceride and heparin substantially increased plasma FFA concentrations, increased FBF in both study groups, and lowered endothelium-dependent vasodilation by 38%. Reduced reactivity to acetylcholine was observed in placebo recipients but not in those given rosiglitazone. In neither group did infusion of triglyceride/heparin influence endothelium-independent vasodilation in response to nitroglycerine. Levels of CRP remained unchanged, but insulin sensitivity and plasma ADMA levels decreased in the presence of elevated FFA levels in both study groups. In this trial rosiglitazone mitigated the increase in FFA levels following triglyceride/heparin infusion and also prevented FFA-induced endothelial dysfunction. These effects were observed in healthy individuals before any change in insulin sensitivity or plasma ADMA concentration.

In Vivo Effects of Insulin and Free Fatty Acids on Matrix Metalloproteinases in Rat Aorta

Obesity is associated with insulin resistance, hyperinsulinemia, elevated plasma free fatty acid (FFA), and increased risk for atherosclerotic vascular disease (ASVD). A part of this increased risk may be due to enhanced activation of matrix metalloproteinases (MMPs). Here, we have examined the effects of physiologically elevated levels of insulin and FFA on three MMPs and their inhibitors (tissue inhibitors of MMP [TIMPs]) in aortic tissue of male rats during euglycemic-hyperinsulinemic clamping. Four-hour euglycemic-hyperinsulinemic clamps with infusion of saline/glycerol, lipid/heparin, or insulin with or without lipid/heparin were performed in alert unrestrained male rats. Hyperinsulinemia increased MMP-2 ( approximately 6-fold), MMP-9 ( approximately 13-fold), membrane type 1-MMP (MT1-MMP; approximately 8-fold) (all Western blots), and gelatinolytic activity (zymography) of MMP-2 (2-fold), while not affecting TIMP-1 and TIMP-2. Insulin increased IRS-1-associated PI 3-kinase (PI3K), extracellular signal-regulated kinases 1/2 (ERK1/2), and c-jun NH(2)-terminal kinase (JNK) (by Western blots with phospho-specific antibodies). FFA augmented the insulin-mediated increases in MMP-2 (from approximately 6- to approximately 11-fold), MMP-9 (from approximately 3- to approximately 23-fold), MT1-MMP (from approximately 8- to approximately 20-fold), MMP-2 gelatinolytic activity (from 2- to 3-fold), and JNK and p38 mitogen-activated protein kinase (MAPK) activities but decreased insulin-mediated activation of PI3K and ERK1/2. Raising FFA without raising insulin affected neither MMPs nor TIMPs. FFA augmented insulin stimulation of the MMP/TIMP balance of three proatherogenic MMPs and increased activities of two MAPKs (JNK and p38 MAPK), both of which are known to stimulate the production of proinflammatory cytokines. This may, over time, increase degradation of extracellular matrix and together with inflammatory changes promote development of ASVD.

Mitochondria and heart failure

Energetic abnormalities in cardiac and skeletal muscle occur in heart failure and correlate with clinical symptoms and mortality. It is likely that the cellular mechanism leading to energetic failure involves mitochondrial dysfunction. Therefore, it is crucial to elucidate the causes of mitochondrial myopathy, in order to improve cardiac and skeletal muscle function, and hence quality of life, in heart failure patients. Recent studies identified several potential stresses that lead to mitochondrial dysfunction in heart failure. Chronically elevated plasma free fatty acid levels in heart failure are associated with decreased metabolic efficiency and cellular insulin resistance. Tissue hypoxia, resulting from low cardiac output and endothelial impairment, can lead to oxidative stress and mitochondrial DNA damage, which in turn causes dysfunction and loss of mitochondrial mass. Therapies aimed at protecting mitochondrial function have shown promise in patients and animal models with heart failure. Despite current therapies, which provide substantial benefit to patients, heart failure remains a relentlessly progressive disease, and new approaches to treatment are necessary. Novel pharmacological agents are needed that optimize substrate metabolism and maintain mitochondrial integrity, improve oxidative capacity in heart and skeletal muscle, and alleviate many of the clinical symptoms associated with heart failure.

Identification of genes that regulate multiple cellular processes/responses in the context of lipotoxicity to hepatoma cells

BackgroundIn order to devise efficient treatments for complex, multi-factorial diseases, it is important to identify the genes which regulate multiple cellular processes. Exposure to elevated levels of free fatty acids (FFAs) and tumor necrosis factor alpha (TNF-α) alters multiple cellular processes, causing lipotoxicity. Intracellular lipid accumulation has been shown to reduce the lipotoxicity of saturated FFA. We hypothesized that the genes which simultaneously regulate lipid accumulation as well as cytotoxicity may provide better targets to counter lipotoxicity of saturated FFA.ResultsAs a model system to test this hypothesis, human hepatoblastoma cells (HepG2) were exposed to elevated physiological levels of FFAs and TNF-α. Triglyceride (TG) accumulation, toxicity and the genomic responses to the treatments were measured. Here, we present a framework to identify such genes in the context of lipotoxicity. The aim of the current study is to identify the genes that could be altered to treat or ameliorate the cellular responses affected by a complex disease rather than to identify the causal genes. Genes that regulate the TG accumulation, cytotoxicity or both were identified by a modified genetic algorithm partial least squares (GA/PLS) analysis. The analyses identified NADH dehydrogenase and mitogen activated protein kinases (MAPKs) as important regulators of both cytotoxicity and lipid accumulation in response to FFA and TNF-α exposure. In agreement with the predictions, inhibiting NADH dehydrogenase and c-Jun N-terminal kinase (JNK) reduced cytotoxicity significantly and increased intracellular TG accumulation. Inhibiting another MAPK pathway, the extracellular signal regulated kinase (ERK), on the other hand, improved the cytotoxicity without changing TG accumulation. Much greater reduction in the toxicity was observed upon inhibiting the NADH dehydrogenase and MAPK (which were identified by the dual-response analysis), than for the stearoyl-CoA desaturase (SCD) activation (which was identified for the TG-alone analysis).ConclusionThese results demonstrate the applicability of GA/PLS in identifying the genes that regulate multiple cellular responses of interest and that genes regulating multiple cellular responses may be better candidates for countering complex diseases.

The importance of free fatty acids in the development of Type 2 diabetes

The recent increase in the prevalence of obesity has been associated with a coincident rise in the prevalence of Type 2 diabetes, whereas weight loss has been shown to decrease the risk of Type 2 diabetes. The pathophysiological mechanisms that have been proposed to explain this link are fundamentally concerned with insulin resistance and the decline in pancreatic B-cell function that accompanies an increase in visceral obesity. They involve the rise in the plasma concentrations of free fatty acids (FFAs) that are associated with an increase in fat mass. Elevated levels of FFAs can lead to insulin resistance, and evidence is growing that B-cell function is impaired through lipotoxicity. Factors such as tumour necrosis factor-alpha (TNF-alpha) and adiponectin, released from adipose tissue, can also modulate insulin resistance. Many interventions that are helpful in treating or preventing Type 2 diabetes, such as weight loss and certain pharmacological interventions, reduce circulating FFA concentrations to a greater or lesser extent. Recent study results suggest that peroxisome proliferator-activated receptor (PPAR)gamma agonists have an effect on the development of Type 2 diabetes. However, in light of concerns over the apparent increase in congestive heart failure with PPARgamma agonists, their place in the prevention of Type 2 diabetes remains to be determined.

This Month in Gastroenterology

This Month in Gastroenterology

Diabetes mellitus as a prothrombotic condition

Diabetes mellitus (DM) is characterized by fasting hyperglycaemia and a high risk of atherothrombotic disorders affecting the coronary, cerebral and peripheral arterial trees. The risk of myocardial infarction (MI) is 3-5 fold higher in Type 2 DM and a DM subject with no history of MI has the same risk as a non-DM subject with a past history of MI. In total around 70% of deaths are vascular with poorer outcomes to both acute events and cardiological interventions. It was proposed that clustering of vascular risk factors (hyperinsulinaemia, dysglycaemia, dyslipidaemia and hypertension) around insulin resistance (IR) accounted for the increase in risk with Type 2 DM. The importance of this became apparent with the recognition that risk clustering occurs in normoglycaemic and impaired glucose tolerance (IGT) subjects with IR, in total around 25% of the population in addition to long-standing Type 1 subjects with renal disease. Evidence indicates that thrombotic risk clustering also occurs in association with IR, suppression of fibrinolysis due to elevated concentrations of the fibrinolytic inhibitor, plasminogen activator inhibitor-1 (PAI-1) is invariable with IR and there is evidence that this is regulated by the effects of triglyceride on the PAI-1 gene promoter. Other studies indicated that prothrombotic risk (coagulation factors VII, XII and fibrinogen) also associates with the IR syndrome. The development of endothelial cell dysfunction with suppression of nitric oxide and prostacyclin synthesis, combined with platelet resistance to the anti-aggregatory effects of these hormones leads to loss of control over platelet activation. In addition, hyperglycaemia and glycation have marked effects on fibrin structure function, generating a clot which has a denser structure, resistant to fibrinolysis. The combination of increased circulating coagulation zymogens, inhibition of fibrinolysis, changes in fibrin structure/function and alterations in platelet reactivity creates a thrombotic risk clustering which underpins the development of cardiovascular disease.

Increased L-CPT-1 activity and altered gene expression in pancreatic islets of malnourished adult rats: a possible relationship between elevated free fatty acid levels and impaired insulin secretion

Intrauterine growth restriction is associated with chronically elevated levels of serum fatty acids and reduced glucose-stimulated insulin secretion. Lipid metabolism in pancreatic β cells is critical for the regulation of insulin secretion, and the chronic exposure to fatty acids results in higher palmitate oxidation rates and an altered insulin response to glucose. Using a rat model of isocaloric protein restriction, we examined whether pre- and postnatal protein malnutrition influences the properties of pancreatic islet carnitine palmitoyltransferase-1 (liver isoform, L-CPT-1), a rate-limiting enzyme that regulates fatty acid oxidation in mitochondria. The activity of L-CPT-1 in pancreatic islets increased in the low protein (LP), although the L-CPT-1 mRNA levels were unaffected by malnutrition. The susceptibility of enzyme to inhibition by malonyl-CoA was unaltered and the content of malonyl-CoA was reduced in LP cells. Because the mitochondrial oxidation of fatty acids is related to the altered expression of a number of genes encoding proteins involved in insulin secretion, the levels of expression of insulin and GLUT-2 mRNA were assessed. A reduced expression of both genes was observed in malnourished rats. These results provide further evidence that increased L-CPT-1 activity and changes in gene expression in pancreatic islets may be involved in the reduced insulin secretion seen in malnourished rats.

Sensitization of vascular smooth muscle cell to TNF-α-mediated death in the presence of palmitate

Saturated free fatty acids (FFAs), including palmitate, can activate the intrinsic death pathway in cells. However, the relationship between FFAs and receptor-mediated death pathway is still unknown. In this study, we have investigated whether FFAs are able to trigger receptor-mediated death. In addition, to clarify the mechanisms responsible for the activation, we examined the biochemical changes in dying vascular smooth muscle cell (VSMC) and the effects of various molecules to the receptor-mediated VSMC death. Tumor necrosis factor (TNF)-α-mediated VSMC death occurred in the presence of sub-cytotoxic concentration of palmitate as determined by assessing viability and DNA degradation, while the cytokine did not influence VSMC viability in the presence of oleate. The VSMC death was inhibited by the gene transfer of a dominant-negative Fas-associated death domain-containing protein and the baculovirus p35, but not by the bcl-xL or the c-Jun N-terminal kinase (JNK) binding domain of JNK-interacting protein-1, in tests utilizing recombinant adenoviruses. The VSMC death was also inhibited by a neutralizing anti-TNF receptor 1 antibody, the caspase inhibitor z-VAD, and the cathepsin B inhibitor CA074, a finding indicative of the role of both caspases and cathepsin B in this process. Consistent with this finding, caspase-3 activation and an increase in cytosolic cathepsin B activity were detected in the dying VSMC. Palmitate inhibited an increase of TNF-α-mediated nuclear factor kappa B (NF-κB) activity, the survival pathway activated by the cytokine, by hindering the translocation of the NF-κB subunit of p65 from the cytosol into the nucleus. The gene transfer of inhibitor of NF-κB predisposed VSMC to palmitate-induced cell death. To the best of our knowledge, this study is the first report to demonstrate the activation of TNF-α-mediated cell death in the presence of palmitate. The current study proposes that FFAs would take part in deleterious vascular consequences of such patients with elevated levels of FFAs as diabetics and obese individuals via the triggering of receptor-mediated death pathways of VSMC.

High-Fat- and Lipid-Induced Insulin Resistance in Rats: The Comparison of Glucose Metabolism, Plasma Resistin and Adiponectin Levels

Aims: In animal models, both an acute elevation in plasma free fatty acid (FFA) via intravenous infusion of a lipid emulsion and a chronic elevation in plasma FFA via high-fat feeding have been shown to induce skeletal muscle and liver insulin resistance. However, there have been very few studies comparing the effects of high-fat- and lipid-induced insulin resistance on glucose metabolism and adipocytokines. Methods: In the current study, we used lipid infusion and a high-fat feed in combination with the hyperinsulinemic-euglycemic clamp technique to assess the impact of acute and chronically elevated FFA levels on overall glucose metabolism and insulin action; two adipocytokines, resistin and adiponectin, were used. Results: At baseline, plasma FFA levels were significantly increased in the high-fat diet (HF) group compared to the control group (p < 0.05). During clamp steady-state, the FFA levels were reduced by ∼25% in the control and ∼48% in the HF groups. In contrast, there was a significant increase in the plasma FFA level in the lipid-infused group (from 0.82 ± 0.03 to 2.87 ± 0.18 mmol/l). The glucose infusion rates (GIRs) in the HF and lipid groups were obviously lower than in the control group (p < 0.01). Moreover, GIR was lower in the lipid group compared with the HF group (p < 0.05). The rate of glucose disappearance (G_Rd) was significantly lower in the lipid group compared with the control group. Hepatic glucose production in the control group was suppressed by ∼15% compared with the HF and lipid groups where it was suppressed by only ∼72 and ∼91%, respectively. The resistin level of muscle tissues in the lipid group was significantly higher compared with the control and HF groups (both p < 0.05). After the insulin clamp, the circulating adiponectin level was significantly decreased in the lipid group compared with the control and HF groups (p < 0.05). Conclusions: Lipid infusion, which was more effective than a high-fat diet, can induce peripheral and hepatic insulin resistance in rats. Insulin-induced resistance might be associated with elevated resistin and decreased adiponectin.

DIVERGENT EFFECTS OF WEIGHT LOSS AND MACRONUTRIENT COMPOSITION ON 24-HOUR LIPIDS WITH LOW-CALORIE/LOW-FAT VERSUS LOW-CARBOHYDRATE DIETS.

Introduction Diets with altered macronutrient composition have become a popular method of weight loss. Because a frequent claim made about low-carbohydrate diets relates to the beneficial effects on lipid and lipoprotein metabolism, ie lower triglycerides and higher HDL cholesterol, we sought to assess the impact of low-calorie/low-fat vs low-carbohydrate diets on fasting and 24-hour excursions of triglycerides (TGs) and free fatty acids (FFAs). Methods Thirty-five subjects were randomized to either a low-calorie/low-fat (LCAL; calorie restricted, 55% carbohydrate) or low-carbohydrate (LCARB; Results The average weight loss at 6 weeks was not significantly different by diet (−12.92 ± 7.3 and −14.47 ± 10.6 pounds) for the LCAL and LCARB groups, respectively. Twenty-four-hour TG excursions also did not differ between both groups at 6 weeks. In contrast, 24-hour FFAs were nearly twofold higher in the LCARB group (AUC, 9,864.3 ± 1711.9 vs 5,199.6 ± 1,119.1, p Conclusion Our data suggest that the effects of weight loss predominate over dietary macronutrient composition in the overall reduction of TGs. Conversely, the persistently elevated 24-hour FFA levels on the low-carbohydrate diet are likely a direct effect of differing dietary macronutrient compositions. The clinical implications of these divergent effects on lipids have not yet been defined.