Plasma Free Fatty Acid Availability Research Articles

GLUT4 localisation with the plasma membrane is unaffected by an increase in plasma free fatty acid availability

BackgroundInsulin-stimulated glucose uptake into skeletal muscle occurs via translocation of GLUT4 from intracellular storage vesicles to the plasma membrane. Elevated free fatty acid (FFA) availability via a lipid infusion reduces glucose disposal, but this occurs in the absence of impaired proximal insulin signalling. Whether GLUT4 localisation to the plasma membrane is subsequently affected by elevated FFA availability is not known.MethodsTrained (n = 11) and sedentary (n = 10) individuals, matched for age, sex and body mass index, received either a 6 h lipid or glycerol infusion in the setting of a concurrent hyperinsulinaemic-euglycaemic clamp. Sequential muscle biopsies (0, 2 and 6 h) were analysed for GLUT4 membrane localisation and microvesicle size and distribution using immunofluorescence microscopy.ResultsAt baseline, trained individuals had more small GLUT4 spots at the plasma membrane, whereas sedentary individuals had larger GLUT4 spots. GLUT4 localisation with the plasma membrane increased at 2 h (P = 0.04) of the hyperinsulinemic-euglycemic clamp, and remained elevated until 6 h, with no differences between groups or infusion type. The number of GLUT4 spots was unchanged at 2 h of infusion. However, from 2 to 6 h there was a decrease in the number of small GLUT4 spots at the plasma membrane (P = 0.047), with no differences between groups or infusion type.ConclusionGLUT4 localisation with the plasma membrane increases during a hyperinsulinemic-euglycemic clamp, but this is not altered by elevated FFA availability. GLUT4 appears to disperse from small GLUT4 clusters located at the plasma membrane to support glucose uptake during a hyperinsulinaemic-euglycaemic clamp.

Plasma free fatty acid concentration is closely tied to whole body peak fat oxidation rate during repeated exercise

Plasma free fatty acids (FFA) are a major contributor to whole body fat oxidation during exercise. However, the extent to which manipulating plasma FFA concentrations will influence whole body peak fat oxidation rate (PFO) during exercise remains elusive. In this study we aimed to increase plasma FFA concentrations through a combination of fasting and repeated exercise bouts. We hypothesized that an increase in plasma FFA concentration would increase PFO in a dose-dependent manner. Ten healthy young (31 ± 6 yr) (mean ± SD) well-trained (maximal oxygen uptake 65.9 ± 6.1 ml·min-1·kg-1) men performed four graded exercise tests (GXTs) on 1 day. The GXTs were interspersed by 4 h of bed rest. This was conducted either in a fasted state or with the consumption of a standardized carbohydrate-rich meal 3.5 h before each GXT. Fasting and previous GXTs resulted in a gradual increase in PFO from 0.63 ± 0.18 g/min after an overnight fast (10 h) to 0.93 ± 0.17 g/min after ∼22 h of fasting and three previous GXTs. This increase in PFO coincided with an increase in plasma FFA concentrations (r2 = 0.73, P < 0.0001). Ingestion of a carbohydrate-rich meal 3.5 h before each GXT resulted in unaltered PFO. This was also reflected in unchanged plasma FFA, glucose, and insulin concentrations. In this study we show that plasma FFA availability is closely tied to whole body PFO and that the length of fasting combined with previous exercise are robust stimuli toward increasing plasma FFA concentration, highlighting the importance for preexercise standardization when conducting GXTs measuring substrate oxidation. NEW & NOTEWORTHY We show that peak fat oxidation is increased in close relationship with plasma free fatty acid availability after combined fasting and repeated incremental exercise tests in healthy highly trained men. Therefore it may be argued that whole body fat oxidation rate measured in most cases after an overnight fast indeed does not represent whole body maximal fat oxidation rate but a whole body peak fat oxidation rate within the context of the preexercise standardization obtained in the study design.

Diurnal Variation in PDK4 Expression Is Associated With Plasma Free Fatty Acid Availability in People.

Many biological pathways involved in regulating substrate metabolism display rhythmic oscillation patterns. In rodents, clock genes regulate circadian rhythms of metabolic genes and substrate metabolism. However, the interrelationships among substrate metabolism, metabolic genes, and clock genes have not been fully explored in people. We tested the hypothesis that the diurnal expression pattern of pyruvate dehydrogenase kinase 4 (PDK4), a key metabolic enzyme involved in fuel switching between glucose and free fatty acids (FFAs), is associated with plasma FFA concentration and clock genes. We analyzed peripheral blood mononuclear cells (PBMCs), subcutaneous adipose tissue, and plasma samples obtained serially during 24 hours from metabolically healthy women (n = 10) and evaluated the interrelationships among PDK4, plasma FFA, and clock genes. We also determined the potential mechanisms responsible for PDK4 transcriptional regulation by using primary human PBMCs and adipocytes. We found that PDK4 diurnal expression patterns were similar in PBMCs and adipose tissue (ρ = 0.84, P < 0.001). The diurnal variation in PBMC PDK4 expression correlated more strongly with plasma FFA and insulin (ρ = 0.86 and 0.63, respectively, both P < 0.001) concentrations than clock genes. Data obtained from primary culture experiments demonstrated that FFAs directly induced PDK4 gene expression, at least in part through activation of peroxisome proliferator-activated receptor α. Our results suggest that plasma FFA availability is an important regulator of diurnal expression patterns of PDK4, and we identify a novel interaction between plasma FFA and cellular diurnal rhythms in regulating substrate metabolism.

One day of overfeeding impairs nocturnal glucose but not fatty acid homeostasis in overweight men

ObjectiveOverfeeding is associated with insulin resistance. Studies on animals suggest this is likely due to disruption of fatty acid metabolism and increased plasma free fatty acid (FFA) availability during the night. We tested the hypothesis that overfeeding induces insulin resistance and increases nocturnal but not daytime plasma FFA availability in human subjects.Design and MethodsWe measured plasma glucose, insulin, and FFA concentrations hourly for 24 hours during a day of isocaloric feeding and a day of hypercaloric feeding (30% calorie excess) in 8 overweight and obese, non-diabetic men (age: 38±3 years; body mass index: 34±2 kg/m2).ResultsOverfeeding had no effect on daytime plasma glucose, insulin, and FFA concentrations compared to isocaloric feeding, but increased nocturnal glucose (P=0.007) and insulin (P=0.003) concentrations and decreased nocturnal FFA concentration (P=0.006). The homeostasis model assessment of insulin resistance score was ~30% greater the morning after hypercaloric than isocaloric feeding (P=0.040).ConclusionsOne day of overfeeding has no effect on daytime plasma glucose and FFA concentrations but increases nocturnal plasma glucose and insulin concentrations, whereas nocturnal plasma FFA availability is reduced. The acute overfeeding-induced development of insulin resistant glucose metabolism therefore does not appear to be directly mediated by plasma FFA availability.

Skeletal muscle fat metabolism after exercise in humans: influence of fat availability

The mechanisms facilitating increased skeletal muscle fat oxidation following prolonged, strenuous exercise remain poorly defined. The aim of this study was to examine the influence of plasma free fatty acid (FFA) availability on intramuscular malonyl-CoA concentration and the regulation of whole-body fat metabolism during a 6-h postexercise recovery period. Eight endurance-trained men performed three trials, consisting of 1.5 h high-intensity and exhaustive exercise, followed by infusion of saline, saline + nicotinic acid (NA; low FFA), or Intralipid and heparin [high FFA (HFA)]. Muscle biopsies were obtained at the end of exercise (0 h) and at 3 and 6 h in recovery. Ingestion of NA suppressed the postexercise plasma FFA concentration throughout recovery (P < 0.01), except at 4 h. The alteration of the availability of plasma FFA during recovery induced a significant increase in whole-body fat oxidation during the 6-h period for HFA (52.2 ± 4.8 g) relative to NA (38.4 ± 3.1 g; P < 0.05); however, this response was unrelated to changes in skeletal muscle malonyl-CoA and acetyl-CoA carboxylase (ACC)β phosphorylation, suggesting mechanisms other than phosphorylation-mediated changes in ACC activity may have a role in regulating fat metabolism in human skeletal muscle during postexercise recovery. Despite marked changes in plasma FFA availability, no significant changes in intramuscular triglyceride concentrations were detected. These data suggest that the regulation of postexercise skeletal muscle fat oxidation in humans involves factors other than the 5'AMP-activated protein kinase-ACCβ-malonyl-CoA signaling pathway, although malonyl-CoA-mediated regulation cannot be excluded completely in the acute recovery period.

Free fatty acid kinetics in the late phase of postexercise recovery: importance of resting fatty acid metabolism and exercise-induced energy deficit

Free fatty acid (FFA) availability increases several-fold during exercise and remains significantly elevated for at least 3 to 6 hours after exercise cessation. Little, however, is known regarding the duration of the postexercise rise in FFA flux. In the present study, we used stable isotope–labeled palmitate infusion to examine fatty acid metabolism in 27 healthy untrained men and women (age, 29 ± 7 years; body mass index, 25 ± 4 kg/m2) between 13 to 16 hours and 21 to 24 hours after a single bout of moderate-intensity endurance exercise (1-2 hours at 60% of peak oxygen consumption), performed in the evening, and after a time-matched resting trial. Postabsorptive FFA rate of appearance (Ra) and FFA concentration in plasma were significantly greater after exercise than rest throughout the recovery period (P < .015), but the exercise-induced increases declined from approximately 40% at 13 to 16 hours to approximately 10% at 21 to 24 hours postexercise (P = .001). The magnitude of the exercise-induced increase in plasma FFA concentration was proportional to the increase in FFA Ra. Correlation analysis demonstrated that exercise-induced changes in plasma FFA Ra at 13 to 16 hours are (1) negatively associated with resting plasma FFA Ra and (2) positively associated with the net energy expenditure of exercise and the exercise-induced changes in whole-body fat oxidation rate (all P values < .05). In multivariate stepwise linear regression analysis, baseline plasma FFA Ra (P ≤ .008) and net energy expenditure of exercise (P ≤ .005) independently predicted the exercise-induced change in plasma FFA Ra at 13 to 16 hours. We conclude that the exercise-induced increase in FFA mobilization is (1) long-lived, persisting for 12 to 24 hours after exercise, with a progressive decline with time; (2) greater in subjects with low than high resting plasma FFA availability; and (3) greater after exercise with high than low energy demand.

Pioglitazone induces lipid accumulation in the rat heart despite concomitant reduction in plasma free fatty acid availability

Thiazolidinediones are insulin-sensitizing drugs which have been proved to be effective in the treatment of type 2 diabetes. However, the action of thiazolidinediones on myocardial metabolism is only poorly recognized. Therefore, the aim of our study was to investigate the effects of two-week pioglitazone treatment (3 mg/kg/d) on lipid and carbohydrate metabolism in the heart of rats fed on a standard chow or on a high-fat diet (HFD) for three weeks. High-fat feeding increased myocardial protein expression of all peroxisome proliferator-activated receptor (PPAR) isoforms. The greatest response was, however, noted in the case of PPARγ. Surprisingly, administration of pioglitazone induced accumulation of free fatty acids (FFA) and diacylglycerol in the heart in both groups, despite concomitant reduction in plasma FFA concentration. The content of triacylglycerol was increased only in the HFD group. Pioglitazone treatment also shifted myocardial substrate utilization towards greater contribution of glucose in both groups, as evidenced by decreased rate of palmitate oxidation and higher 2-deoxyglucose uptake and elevated glycogen content. This could induce a mismatch between the rate of myocardial fatty acid uptake and oxidation leading to increased intracellular availability of fatty acids for non-oxidative metabolic pathways like synthesis of acylglycerols. Our data suggests that thiazolidinediones improve cardiac insulin sensitivity by mechanisms other than reduction in intramyocardial lipid content.

Determinants of intramyocellular triglyceride turnover: implications for insulin sensitivity

Increased intramyocellular triglyceride (IMTG) content is found in both insulin-sensitive endurance-trained subjects and insulin-resistant obese/type 2 diabetic subjects. A high turnover rate of the IMTG pool in athletes is proposed to reduce accumulation of lipotoxic intermediates interfering with insulin signaling. IMTG turnover is a composite measure of the dynamic balance between lipolysis and lipid synthesis; both are influenced by mitochondrial fat oxidation and plasma free fatty acid availability. Therefore, more attention should be given to the factors controlling the rate of turnover of IMTG. In this review, particular attention has been given to muscle oxidative capacity, plasma free fatty acid availability, and IMTG hydrolysis (lipolysis) and synthesis. A higher oxidative, lipolytic, and lipid storage capacity in the muscle of endurance-trained subjects reflects a higher fractional turnover of the IMTG pool. Thus the co-localization of intermyofibrillar lipid droplets and mitochondria allows for a fine coupling of lipolysis of the IMTG pool to mitochondrial beta-oxidation. Conversely, reduced oxidative capacity and a mismatch between IMTG lipolysis and beta-oxidation might be detrimental to insulin sensitivity by generating several lipotoxic intermediates in sedentary populations including obese/type 2 diabetic subjects. Further studies are clearly required to better understand the relationship between the rate of turnover of IMTG and the accumulation of lipotoxic intermediates in the pathophysiology of insulin resistance.

Reduced plasma free fatty acid availability during exercise: effect on gene expression

Endurance exercise transiently increases the mRNA of key regulatory proteins involved in skeletal muscle metabolism. During prolonged exercise and subsequent recovery, circulating plasma fatty acid (FA) concentrations are elevated. The present study therefore aimed to determine the sensitivity of key metabolic genes to FA exposure, assessed in vitro using L6 myocytes and secondly, to measure the expression of these same set of genes in vivo, following a single exercise bout when the post-exercise rise in plasma FA is abolished by acipimox. Initial studies using L6 myotubes demonstrated dose responsive sensitivity for both PDK4 and PGC-1alpha mRNA to acute FA exposure in vitro. Nine active males performed two trials consisting of 2 h exercise, followed by 2 h of recovery. In one trial, plasma FA availability was reduced by the administration of acipimox (LFA), a pharmacological inhibitor of adipose tissue lipolysis, and in the second trial a placebo was provided (CON). During the exercise bout and during recovery, the rise in plasma FA and glycerol was abolished by acipimox treatment. Following exercise the mRNA abundance of PDK4 and PGC-1alpha were elevated and unaffected by either acipimox or placebo. Further analysis of skeletal muscle gene expression demonstrated that the CPT I gene was suppressed in both trials, whilst UCP-3 gene was only modestly regulated by exercise alone. Acipimox ingestion did not alter the response for both CPT I and UCP-3. Thus, this study demonstrates that the normal increase in circulating concentrations of FA during the later stages of exercise and subsequent recovery is not required to induce skeletal muscle mRNA expression of several proteins involved in regulating substrate metabolism.

Short‐term suppression of plasma free fatty acids fails to improve insulin sensitivity when intramyocellular lipid is elevated

Metabolic responses to manipulation of the plasma free fatty acid (FFA) concentration were assessed in six healthy men via cross-over design to determine whether FFAs independently influence insulin sensitivity. Intramyocellular lipid (IMCL) was measured by proton magnetic resonance spectroscopy and insulin sensitivity via frequently sampled intravenous glucose tolerance test (IVGTT) after 67 h of two identical low carbohydrate/high fat (LC) diets which were used to elevate IMCL and plasma FFAs. To uncouple the influence of FFAs and IMCL on insulin sensitivity, FFAs were suppressed 30 min prior to and during IVGTT in one treatment [LC + nicotinic acid (NA)] by NA ingestion. Vastus lateralis IMCL was significantly elevated in LC (13.3 +/- 1.1 x 10(-3)) and LC + NA (13.5 +/- 1.1 x 10(-3)) (P < 0.01 for both), but was not different between conditions (P > 0.05). Plasma FFAs were raised in LC (0.79 +/- 0.08 mmol/l) and LC + NA (0.80 +/- 0.11 mmol/l) (P < 0.01 for both) and were significantly reduced by NA ingestion prior to (0.36 +/- 0.05 mmol/l, P < 0.01) and during IVGTT (P < 0.05) in LC + NA. Despite marked differences in plasma FFA availability, insulin sensitivity and glucose tolerance were not different between LC and LC + NA (P > 0.05 for both). Plasma FFAs appear to exert no immediate effect on insulin sensitivity/glucose tolerance independent of their action on intracellular lipid moieties. Further research is required to elucidate the duration of FFA suppression required to restore insulin sensitivity following lipid-induced insulin resistance.

Increased intramuscular lipid storage in the insulin-resistant and endurance-trained state

Numerous studies have reported a strong correlation between intramuscular triacylglycerol (IMTG) content and insulin resistance. However, the proposed relationship between IMTG accumulation and skeletal muscle insulin resistance is not unambiguous, as trained athletes have been shown to be markedly insulin sensitive despite an elevated IMTG storage. Though the latter has often been attributed to differences in muscle fibre type composition and/or structural characteristics of the intramyocellular lipid deposits, recent studies have failed to provide such evidence. The greater insulin sensitivity despite an elevated IMTG deposition in the endurance-trained state has often been described as a metabolic paradox. However, divergent metabolic events are responsible for the greater IMTG content in the endurance-trained versus insulin-resistant states. The greater IMTG storage in the trained athlete represents an adaptive response to endurance training, allowing a greater contribution of the IMTG pool as a substrate source during exercise. In contrast, elevated IMTG stores in the obese and/or type 2 diabetes patient seem to be secondary to a structural imbalance between plasma free fatty acid availability, fatty acid (FA) storage and oxidation. Therefore, the reported correlation between IMTG content and insulin resistance does not represent a functional relationship, as it is strongly influenced by training status and/or habitual physical activity. It can be argued that the ratio between IMTG content and muscle oxidative capacity represents a more accurate marker of insulin resistance. Interventions to augment mitochondrial density and/or function are likely to improve the balance between FA uptake and oxidation and should be applied to prevent and/or treat insulin resistance.

Inhibition of adipose tissue lipolysis increases intramuscular lipid and glycogen use in vivo in humans

This study investigates the consequences of inhibition of adipose tissue lipolysis on skeletal muscle substrate use. Ten subjects were studied at rest and during exercise and subsequent recovery under normal, fasting conditions (control trial, CON) and following administration of a nicotinic acid analog (low plasma free fatty acid trial, LFA). Continuous [U-13C]palmitate and [6,6-2H2]glucose infusions were applied to quantify plasma free fatty acid (FFA) and glucose oxidation rates and to estimate intramuscular triacylglycerol (IMTG) and glycogen use. Muscle biopsies were collected to measure 1) fiber type-specific IMTG content; 2) allosteric regulators of hormone-sensitive lipase (HSL), glycogen phosphorylase, and pyruvate dehydrogenase; and 3) the phosphorylation status of HSL at Ser563 and Ser565. Administration of a nicotinic acid analog (acipimox) substantially reduced plasma FFA rate of appearance and subsequent plasma FFA concentrations (P < 0.0001). At rest, this substantially reduced plasma FFA oxidation rates, which was compensated by an increase in the estimated IMTG use (P < 0.05). During exercise, the progressive increase in FFA rate of appearance, uptake, and oxidation was prevented in the LFA trial and matched by greater IMTG and glycogen use. Differential phosphorylation of HSL or relief of its allosteric inhibition by long-chain fatty acyl-CoA could not explain the increase in muscle TG use, but there was evidence to support the contention that regulation may reside at the level of the glucose-fatty acid cycle. This study confirms the hypothesis that plasma FFA availability regulates both intramuscular lipid and glycogen use in vivo in humans.

Inhibition Of Adipose Tissue Lipolysis Increases Intramuscular Lipid Use In Type 2 Diabetes Patients

Numerous studies have reported the association between elevated plasma free fatty acid (FFA) concentrations, intramyocellular triacylglycerol (IMTG) accumulation and the development of insulin resistance. Excess IMTG accretion is the result of an imbalance between FFA uptake, IMTG deposition and its subsequent mobilisation and/or oxidation. Recent data indicate that this imbalance, and not the actual size or distribution of the IMTG pool, is responsible for the development of insulin resistance. Therefore, the turnover of the IMTG pool should form a major therapeutic target in the prevention and treatment of skeletal muscle insulin resistance. PURPOSE To investigate the consequences of adipose tissue lipolytic inhibition on skeletal muscle substrate use. METHODS Ten type 2 diabetes patients were studied at rest, during exercise and subsequent recovery under normal, fasting conditions (CON) and following administration of a nicotinic acid analogue (LFA). Continuous [U-13C]palmitate and [6,6-2H2]glucose infusions were applied to quantify plasma FFA and glucose oxidation rates, and to estimate intramyocellular lipid and glycogen use. Muscle biopsies were collected before and after exercise to determine net changes in fibre type specific lipid and glycogen content. RESULTS In the LFA trial, plasma FFA rate of appearance was effectively reduced, resulting in 2–5 fold lower plasma FFA concentrations (P<0.001). Plasma FFA oxidation rates were substantially reduced at rest, during exercise and recovery in the LFA trial, which was compensated by an increase in both IMTG and endogenous carbohydrate use. Plasma glucose disposal rates did not differ between trials. During exercise, IMTG use averaged 0.18 ± 0.05 vs 0.08 ± 0.03 g/min in the LFA and CON trial, respectively (P<0.01). In accordance to these tracer data, a greater net decline in type I and type II muscle fiber lipid content was observed following exercise in the LFA vs the CON trial (P<0.05). CONCLUSION This study shows that plasma FFA availability regulates IMTG use and introduces adipose tissue lipolytic inhibition as a means to augment IMTG use in type 2 diabetes patients. The latter seems to represent an effective strategy to improve whole-body insulin sensitivity.

Inhibition Of Adipose Tissue Lipolysis Increases Intramuscular Lipid Use In Type 2 Diabetes Patients

Numerous studies have reported the association between elevated plasma free fatty acid (FFA) concentrations, intramyocellular triacylglycerol (IMTG) accumulation and the development of insulin resistance. Excess IMTG accretion is the result of an imbalance between FFA uptake, IMTG deposition and its subsequent mobilisation and/or oxidation. Recent data indicate that this imbalance, and not the actual size or distribution of the IMTG pool, is responsible for the development of insulin resistance. Therefore, the turnover of the IMTG pool should form a major therapeutic target in the prevention and treatment of skeletal muscle insulin resistance. PURPOSE To investigate the consequences of adipose tissue lipolytic inhibition on skeletal muscle substrate use. METHODS Ten type 2 diabetes patients were studied at rest, during exercise and subsequent recovery under normal, fasting conditions (CON) and following administration of a nicotinic acid analogue (LFA). Continuous [U-13C]palmitate and [6,6-2H2]glucose infusions were applied to quantify plasma FFA and glucose oxidation rates, and to estimate intramyocellular lipid and glycogen use. Muscle biopsies were collected before and after exercise to determine net changes in fibre type specific lipid and glycogen content. RESULTS In the LFA trial, plasma FFA rate of appearance was effectively reduced, resulting in 2–5 fold lower plasma FFA concentrations (P<0.001). Plasma FFA oxidation rates were substantially reduced at rest, during exercise and recovery in the LFA trial, which was compensated by an increase in both IMTG and endogenous carbohydrate use. Plasma glucose disposal rates did not differ between trials. During exercise, IMTG use averaged 0.18 ± 0.05 vs 0.08 ± 0.03 g/min in the LFA and CON trial, respectively (P<0.01). In accordance to these tracer data, a greater net decline in type I and type II muscle fiber lipid content was observed following exercise in the LFA vs the CON trial (P<0.05). CONCLUSION This study shows that plasma FFA availability regulates IMTG use and introduces adipose tissue lipolytic inhibition as a means to augment IMTG use in type 2 diabetes patients. The latter seems to represent an effective strategy to improve whole-body insulin sensitivity.

Role of lipids in the early developmental stages of experimental immune diabetes induced by multiple low-dose streptozotocin

The present work examines the role of lipids in the development of the Type 1 diabetes induced by the administration of multiple low doses of streptozotocin (STZ) in C57BL/6J mice. The study was performed before and after the onset of clear hyperglycemia, and the results were as follows. First, 6 days after the first dose of STZ, while plasma glucose and insulin levels remained similar to those observed in the control mice, plasma free fatty acid (FFA) levels were significantly increased (P < 0.05). At that time, a marked increase of triglyceride content in gastronemius muscle was accompanied by a diminished activity of pyruvate dehydrogenase complex, suggesting an impaired glucose oxidation. Furthermore, a decrease of both triglyceride content and lipoprotein lipase activity was observed in the epididymal fat tissue. Second, 12 days after the first injection of STZ, hyperglycemia was accompanied by hypertriglyceridemia, a more pronounced increase of plasma FFA, and a significant (P < 0.05) reduction of insulinemia. At this time, both the adipose tissue and the gastrocnemius muscle showed a further deterioration of all parameters mentioned after 6 days. Moreover, in the gastrocnemius muscle, an impaired nonoxidative pathway of glucose metabolism was observed [significant reduction (P < 0.05) of glycogen mass, glucose-6-phosphate content, and glycogen synthase activities] at this time point. Finally, the data suggest for the first time that, in mice, Type 1 diabetes induced by multiple low doses of STZ and enhanced lipolysis of fat pads leads to an increase in the availability of plasma FFA, which seems to play a role in the early steps of diabetes evolution.

Suppression of plasma free fatty acids upregulates peroxisome proliferator-activated receptor (PPAR) α and δ and PPAR coactivator 1α in human skeletal muscle, but not lipid regulatory genes

Fatty acids are an important ligand for peroxisome proliferator-activated receptor (PPAR) activation and transcriptional regulation of metabolic genes. To examine whether reduced plasma free fatty acid (FFA) availability affects the mRNA content of proteins involved in fuel metabolism in vivo, the skeletal muscle mRNA content of various transcription factors, transcriptional coactivators and genes encoding for lipid regulatory proteins were examined before and after 3 h of cycle exercise with (NA) and without (CON) pre-exercise ingestion of nicotinic acid (NA). NA resulted in a marked (3- to 6-fold) increase (P<0.05) in PPARalpha, PPARdelta and PPAR coactivator 1alpha (PGC1alpha) mRNA, but was without effect on nuclear respiratory factor-1 and Forkhead transcription factor, fatty acid transcolase/CD36, carnitine palmitoyl transferase 1, hormone sensitive lipase (HSL) and pyruvate dehydrogenase kinase 4. Exercise in CON was associated with increased (P<0.05) PPARalpha, PPARdelta and PGC1alpha mRNA, which was similar in magnitude to levels observed with NA at rest. Exercise was generally without effect on the mRNA content of lipid regulatory proteins in CON and did not affect the mRNA content of the measured subset of transcription factors, transcriptional co-activators and lipid regulatory proteins during NA. To determine the possible mechanisms by which NA might affect PGC1alpha expression, we measured p38 MAP kinase (MAPK) and plasma epinephrine. Phosphorylation of p38 MAPK was increased (P<0.05) by NA treatment at rest, and this correlated (r2=0.84, P<0.01) with increased PGC1alpha. Despite this close relationship, increasing p38 MAPK in human primary myotubes was without effect on PGC1alpha mRNA content. Plasma epinephrine was elevated (P<0.05) by NA at rest (CON: 0.27+/-0.06, NA: 0.72+/-0.11 nM) and throughout exercise. Incubating human primary myotubes with epinephrine increased PGC1alpha independently of changes in p38 MAPK phosphorylation. Hence, despite the fact that NA ingestion decreased FFA availability, it promoted the induction of PPARalpha/delta and PGC1alpha gene expression to a similar degree as prolonged exercise. We suggest that the increase in PGC1alpha may be due to the elevated plasma epinephrine levels. Despite these changes in transcription factors/coactivators, the mRNA content of lipid regulatory proteins was generally unaffected by plasma FFA availability.

Effects of reduced free fatty acid availability on hormone-sensitive lipase activity in human skeletal muscle during aerobic exercise.

Hormone-sensitive lipase (HSL) catalyzes the hydrolysis of intramuscular triacylglycerol (IMTG); however, its regulation in skeletal muscle is poorly understood. To examine the effects of reduced free fatty acid (FFA) availability on HSL activity in skeletal muscle during aerobic exercise, 11 trained men exercised at 55% maximal O2 uptake for 40 min after the ingestion of nicotinic acid (NA) or nothing (control). Muscle biopsies were taken at rest and 5, 20, and 40 min of exercise. Plasma FFA were suppressed (P < 0.05) in NA during exercise ( approximately 0.40 +/- 0.04 vs. approximately 0.07 +/- 0.01 mM). The respiratory exchange ratio (RER) was increased throughout exercise (0.020 + 0.008) after NA ingestion. However, the provision of energy from fat oxidation only decreased from 33% of the total in the control trial to 26% in the NA trial, suggesting increased IMTG oxidation in the NA trial. Mean HSL activity was 2.25 + 0.15 mmol x kg dry mass(-1) x min(-1) at rest and increased (P < 0.05) to 2.94 +/- 0.20 mmol x kg dry mass(-1) x min(-1) at 5 min in control. Contrary to the hypothesis, mean HSL was not activated to a greater extent in the NA trial during exercise (2.20 + 0.28 at rest to 2.88 + 0.21 mmol x kg dry mass(-1) x min(-1) at 5 min). No further HSL increases were observed at 20 or 40 min in both trials. There was variability in the response to NA ingestion, as some subjects experienced a large increase in RER and decrease in fat oxidation, whereas other subjects experienced no shift in RER and maintained fat oxidation despite the reduced FFA availability in the NA trial. However, even in these subjects, HSL activity was not further increased during the NA trial. In conclusion, reduced plasma FFA availability accompanied by increased epinephrine concentration did not further activate HSL beyond exercise alone.

Hyperinsulinemia and triglyceride-rich lipoproteins.

Hypertriglyceridemia is the most frequent form of hyperlipidemia seen in diabetes. Because hypertriglyceridemia and hyperinsulinemia often coexist in the general population and because patients with NIDDM generally are hyperinsulinemic, we have undertaken a series of in vivo studies to examine the effects of hyperinsulinemia on VLDL production. These studies showed that chronic hyperinsulinemia is accompanied by increased VLDL production and that this occurs even when plasma free fatty acid (FFA) levels have fallen. By contrast, acute hyperinsulinemia is accompanied by a reduction in VLDL production, and this reduction is, at least in part, mediated by an associated reduction in the availability of plasma FFAs as a substrate for VLDL-triglyceride (TG). The studies also raise the possibility that the difference in the dependence of VLDL production on plasma FFAs in acute versus chronic hyperinsulinemia results from an increase in hepatic lipogenic enzymes and from the availability of an alternate substrate such as fructose. The overall effect of hyperinsulinemia on VLDL production is postulated to reflect both the effect of insulin on apolipoprotein B production and the hepatic synthesis of TG from either plasma FFAs or newly made fatty acids.

EFFECT OF ELEVATED [FFA] ON GLUCOSE UPTAKE AND LACTATE EFFLUX DURING SUBMAXIMAL EXERCISE IN MAN 9

The present investigation examined the effect of enhanced plasma free fatty acid availability on leg glucose uptake and lactate efflux during prolonged submaximal cycling exercise. Seven males cycled for 70 min (10 min at 40% and 60 min at 65% VO2max) while infused with either Intralipid (100 ml) and heparin (2000 U) (INTRA) or saline (CON). At the onset of exercise, plasma arterial [FFA] was 0.70 ± 0.03 mM (INTRA) vs. 0.22 ± 0.01 mM(CON). Arterial and femoral venous catheters were inserted and blood samples and expired gases were collected simultaneously at rest and at 8, 18, 34, 50 and 68 min of exercise. Blood flow was estimated from pulmonary O2 uptake and the leg a-v (O2) difference. No difference existed in VO2 or blood flow between trials, while average pulmonary RER was higher in CON (0.94 ± 0.01) compared to INTRA (0.91 ± 0.01)(p<0.05). Mean leg FFA uptake was higher in INTRA (0.16 ± 0.03 mmol/min) vs. CON (0.04 ± 0.01 mmol/min) during exercise (p<0.01). There was a significant decrease in lactate efflux during INTRA (1.55 ± 0.36 mmol/min) compared to CON (3.07 ± 0.47 mmol/min) (p<0.05). Leg glucose uptake was not different at any time point between trials. In conclusion, enhanced FFA availability leads to a decrease in lactate efflux and no change in glucose uptake. Therefore, if carbohydrate sparing occurs at these workloads, it must be a function of reduced glycogenolysis.

Effects of estrogen on free fatty acid metabolism in humans

To determine whether estrogen directly affects effective adipose lipolysis, palmitate rate of appearances ([14C]palmitate) was measured in 15 postmenopausal women. Each volunteer was studied after > or = 2 mo of estrogen treatment and again after > or = 2 mo of estrogen deficiency. Plasma hormone concentrations were controlled and identical on the 2 study days with use of the pancreatic clamp technique, and the lipolytic response to epinephrine and epinephrine + phentolamine was assessed. Results showed that overall palmitate flux was greater (10-20%, P < 0.05) during the estrogen-deficient than during the estrogen-replete study. Adrenergic stimulation of lipolysis was not specifically influenced by estrogen treatment, and control of plasma hormone concentrations did not eliminate the difference in palmitate flux between the estrogen-deficient and estrogen-replete study days. We conclude that estrogen deficiency is associated with increased plasma free fatty acid availability and that estrogen likely has direct, albeit small, effects on adipose tissue lipolysis.