Greater reliance on glycolysis is associated with lower mitochondrial substrate oxidation and insulin sensitivity in infant myogenic MSCs.

Individuals with insulin resistance and obesity display higher skeletal muscle production of nonoxidized glycolytic products (i.e., lactate), and lower complete mitochondrial substrate oxidation to CO2. These findings have also been observed in individuals without obesity and are associated with an increased risk for metabolic disease. The purpose of this study was to determine if substrate preference is evident at the earliest stage of life (birth) and to provide a clinical blood marker (lactate) that could be indicative of a predisposition for metabolic disease later. We used radiolabeled tracers to assess substrate oxidation and insulin sensitivity of myogenically differentiated mesenchymal stem cells (MSCs), a proxy of infant skeletal muscle tissue, derived from umbilical cords of full-term infants. We found that greater production of nonoxidized glycolytic products (lactate, pyruvate, alanine) is directly proportional to lower substrate oxidation and insulin sensitivity in MSCs. In addition, we found an inverse relationship between the ratio of complete glucose oxidation to CO2 and infant blood lactate at 1 mo of age. Collectively, considering that higher lactate was associated with lower MSC glucose oxidation and has been shown to be implicated with metabolic disease, it may be an early indicator of infant skeletal muscle phenotype.NEW & NOTEWORTHY In infant myogenically differentiated mesenchymal stem cells, greater production of nonoxidized glycolytic products was directly proportional to lower substrate oxidation and insulin resistance. Glucose oxidation was inversely correlated with infant blood lactate. This suggests that innate differences in infant substrate oxidation exist at birth and could be associated with the development of metabolic disease later in life. Clinical assessment of infant blood lactate could be used as an early indicator of skeletal muscle phenotype.

Impaired glucose partitioning in primary myotubes from severely obese women with type 2 diabetes.

The purpose of this study was to determine whether intramyocellular glucose partitioning was altered in primary human myotubes derived from severely obese women with type 2 diabetes. Human skeletal muscle cells were obtained from lean nondiabetic and severely obese Caucasian females with type 2 diabetes [body mass index (BMI): 23.6 ± 2.6 vs. 48.8 ± 1.9 kg/m2, fasting glucose: 86.9 ± 1.6 vs. 135.6 ± 12.0 mg/dL, n = 9/group]. 1-[14C]-Glucose metabolism (glycogen synthesis, glucose oxidation, and nonoxidized glycolysis) and 1- and 2-[14C]-pyruvate oxidation were examined in fully differentiated myotubes under basal and insulin-stimulated conditions. Tricarboxylic acid cycle intermediates were determined via targeted metabolomics. Myotubes derived from severely obese individuals with type 2 diabetes exhibited impaired insulin-mediated glucose partitioning with reduced rates of glycogen synthesis and glucose oxidation and increased rates of nonoxidized glycolytic products, when compared with myotubes derived from the nondiabetic individuals (P < 0.05). Both 1- and 2-[14C]-pyruvate oxidation rates were significantly blunted in myotubes from severely obese women with type 2 diabetes compared with myotubes from the nondiabetic controls. Lastly, concentrations of tricarboxylic acid cycle intermediates, namely, citrate (P < 0.05), cis-aconitic acid (P = 0.07), and α-ketoglutarate (P < 0.05), were lower in myotubes from severely obese women with type 2 diabetes. These data suggest that intramyocellular insulin-mediated glucose partitioning is intrinsically altered in the skeletal muscle of severely obese women with type 2 diabetes in a manner that favors the production of glycolytic end products. Defects in pyruvate dehydrogenase and tricarboxylic acid cycle may be responsible for this metabolic derangement associated with type 2 diabetes.

Plasma Lactate as a Marker for Metabolic Health.

Blood lactate concentrations traditionally have been used as an index of exercise intensity or clinical hyperlactatemia. However, more recent data suggest that fasting plasma lactate can also be indicative of the risk for subsequent metabolic disease. The hypothesis presented is that fasting blood lactate accumulation reflects impaired mitochondrial substrate use, which in turn influences metabolic disease risk.

Altered tricarboxylic acid cycle flux in primary myotubes from severely obese humans.

BACKGROUND/OBJECTIVE: The partitioning of glucose toward glycolytic end products rather than glucose oxidation and glycogen storage is evident in skeletal muscle with severe obesity and type 2 diabetes. The purpose of the present study was to determine the possible mechanism by which severe obesity alters insulin-mediated glucose partitioning in human skeletal muscle. SUBJECTS/METHODS: Primary human skeletal muscle cells (HSkMC) were isolated from lean (BMI = 23.6 ± 2.6 kg/m2, n = 9) and severely obese (BMI = 48.8 ± 1.9 kg/m2, n = 8) female subjects. Glucose oxidation, glycogen synthesis, non-oxidized glycolysis, pyruvate oxidation, and targeted TCA cycle metabolomics were examined in differentiated myotubes under basal and insulin-stimulated conditions. RESULTS: Myotubes derived from severely obese subjects exhibited attenuated response of glycogen synthesis (20.3%; 95% CI [4.7, 28.8]; P = 0.017) and glucose oxidation (5.6%; 95% CI [0.3, 8.6]; P = 0.046) with a concomitant greater increase (23.8%; 95% CI [5.7, 47.8]; P = 0.004) in non-oxidized glycolytic end products with insulin stimulation in comparison to the lean group (34.2% [24.9, 45.1]; 13.1% [8.6, 16.4], and 2.9% [-4.1, 12.2], respectively). These obesity-related alterations in glucose partitioning appeared to be linked with reduced TCA cycle flux, as 2-[14C]-pyruvate oxidation (358.4 pmol/mg protein/min [303.7, 432.9] vs. lean 439.2 pmol/mg protein/min [393.6, 463.1]; P = 0.013) along with several TCA cycle intermediates, were suppressed in the skeletal muscle of severely obese individuals. CONCLUSIONS: These data suggest that with severe obesity the partitioning of glucose toward anaerobic glycolysis in response to insulin is a resilient characteristic of human skeletal muscle. This altered glucose partitioning appeared to be due, at least in part, to a reduction in TCA cycle flux.

Fasting plasma lactate as a possible early clinical marker for metabolic disease risk.

BACKGROUND AND AIM: Elevated fasting plasma lactate concentrations are evident in individuals with metabolic diseases. However, it has yet to be determined if these associations exist in a young, healthy population as a possible early marker for metabolic disease risk. The purpose of this study was to determine if indices of the metabolic syndrome are related to plasma lactate concentrations in this population. METHODS: Fifty (29 ± 7 yr) men (n = 19) and women (n = 31) classified as overweight (26.4 ± 1.8 kg/m2) participated in this observational study. Blood pressure and blood metabolites were measured after an overnight fast. Lactate was also measured before and after a three-day eucaloric high-fat (70 %) diet. The homeostatic model assessment for insulin resistance (HOMA-IR) was calculated as a measure of insulin resistance. Visceral adipose tissue mass was determined via dual X-ray absorptiometry. RESULTS: Triglycerides (r = 0.55, p=<0.0001), HOMA-IR (r = 0.53, p=<0.0001), and systolic and diastolic (both, r = 0.36, p = 0.01) blood pressures associated with fasting plasma lactate. No differences in visceral adipose tissue existed between the sexes (p = 0.41); however, the relationship between visceral adipose tissue and lactate existed only in females (r = 0.59, p = 0.02) but not in males (p = 0.53). Fasting lactate and HOMA-IR increased in males (p = 0.01 and p = 0.02, respectively), but not females, following a three-day high-fat diet. CONCLUSION: Indices of the metabolic syndrome associated with fasting plasma lactates in young relatively healthy individuals. Fasting lactate also increased in a sex-specific manner after a three-day high fat diet. Thus, lactate could become a clinical marker for metabolic disease risk.

Severe obesity: evidence for a deranged metabolic program in skeletal muscle?

Severe obesity is increasing at a disproportionate rate compared with milder grade obesity. Our research group has obtained evidence indicative of an "obesity metabolic program" in skeletal muscles of severely obese individuals, which may be determined genetically or epigenetically. We believe that this represents a paradigm shift in thinking about metabolic regulation in obesity.

Ockham's razor and the metabolic syndrome.

The broad effects of bariatric/metabolic surgery on virtually every tissue and organ system remain unexplained. Weight loss, although a major factor, does not fully account for the rapid, full, and durable remission of type 2 diabetes, return of islet function, reduction of the prevalence of cancers, increase in gray matter of the brain, and decrease in all-cause mortality. This review supports the thesis that the metabolic syndrome is not a group of separate diseases but rather multiple expressions of a shared defect in the utilization of carbohydrates and lipids. That error is probably caused by a dysmetabolic signal from the foregut, stimulated by food, that limits entry of 2-carbon fragments into the tricarboxylic acid cycle, the accumulation of lactate and, in turn, increases in glucose and insulin. Surgery limits that signal by reducing contact between food and foregut mucosa. Speciation of that signal(s) may offer a new pathway for drug development.

Type 2 Diabetes Modifies Skeletal Muscle Gene Expression Response to Gastric Bypass Surgery.

Introduction: Roux-en-Y gastric bypass (RYGB) is an effective treatment for type 2 diabetes mellitus (T2DM) that can result in remission of clinical symptoms, yet mechanisms for improved skeletal muscle health are poorly understood. We sought to define the impact of existing T2DM on RYGB-induced muscle transcriptome changes. Methods: Vastus lateralis biopsy transcriptomes were generated pre- and 1-year post-RYGB in black adult females with (T2D; n = 5, age = 51 ± 6 years, BMI = 53.0 ± 5.8 kg/m2) and without (CON; n = 7, 43 ± 6 years, 51.0 ± 9.2 kg/m2) T2DM. Insulin, glucose, and HOMA-IR were measured in blood at the same time points. ANCOVA detected differentially expressed genes (p < 0.01, fold change < |1.2|), which were used to identify enriched biological pathways. Results: Pre-RYGB, 95 probes were downregulated with T2D including subunits of mitochondrial complex I. Post-RYGB, the T2D group had normalized gene expression when compared to their non-diabetic counterparts with only three probes remaining significantly different. In the T2D, we identified 52 probes upregulated from pre- to post-RYGB, including NDFUB7 and NDFUA1. Conclusion: Black females with T2DM show extensive downregulation of genes across aerobic metabolism pathways prior to RYGB, which resolves 1 year post-RYGB and is related to improvements in clinical markers. These data support efficacy of RYGB for improving skeletal muscle health, especially in patients with T2DM.

Elevated stearoyl-CoA desaturase-1 expression in skeletal muscle contributes to abnormal fatty acid partitioning in obese humans.

Obesity and type 2 diabetes are strongly associated with abnormal lipid metabolism and accumulation of intramyocellular triacylglycerol, but the underlying cause of these perturbations are yet unknown. Herein, we show that the lipogenic gene, stearoyl-CoA desaturase 1 (SCD1), is robustly up-regulated in skeletal muscle from extremely obese humans. High expression and activity of SCD1, an enzyme that catalyzes the synthesis of monounsaturated fatty acids, corresponded with low rates of fatty acid oxidation, increased triacylglycerol synthesis and increased monounsaturation of muscle lipids. Elevated SCD1 expression and abnormal lipid partitioning were retained in primary skeletal myocytes derived from obese compared to lean donors, implying that these traits might be driven by epigenetic and/or heritable mechanisms. Overexpression of human SCD1 in myotubes from lean subjects was sufficient to mimic the obese phenotype. These results suggest that elevated expression of SCD1 in skeletal muscle contributes to abnormal lipid metabolism and progression of obesity.

Lipid-induced insulin resistance is prevented in lean and obese myotubes by AICAR treatment.

The molecular mechanisms of obesity-associated insulin resistance are becoming increasingly clear, and the effects of various lipid molecules, such as diacylglycerol and ceramide, on the insulin signal are being actively explored. To better understand the divergent response to lipid exposure between lean and obese, we incubated primary human muscle cells from lean [body mass index (BMI) <25 kg/m(2)] and morbidly obese (BMI >40 kg/m(2)) subjects with the saturated fatty acid palmitate. Additionally, given that AMPK-activating drugs are widely prescribed for their insulin-sensitizing effects, we sought to determine whether 5-aminoimidazole-4-carboxamide 1-beta-D-ribofuranoside (AICAR)-stimulated AMPK activation could prevent or reverse the deleterious effects of lipid on insulin signaling. We found that a 1-h palmitate incubation in lean myotubes reduced (P < 0.05) insulin-stimulated phosphoprotein kinase B (Akt), Akt substrate 160 (AS160), and inhibitory factor kappaBalpha (IkappaBalpha) mass, all of which were prevented with AICAR inclusion. With a longer incubation, we observed that myotubes from morbidly obese individuals appear to be largely resistant to the detrimental effects of 16 h lipid exposure as was evident, in contrast to the lean, by the absence of a reduction in insulin-stimulated insulin receptor substrate (IRS)-1 Tyr phosphorylation, phospho-Akt, and phospho-AS160 (P < 0.05). Furthermore, 16 h lipid exposure significantly reduced IkappaBalpha levels and increased phosphorylation of c-Jun NH(2)-terminal kinase (JNK) and IRS1-Ser(312) in lean myotubes only (P < 0.05). Despite a divergent response to lipid between lean and obese myotubes, AICAR inclusion improved insulin signaling in all myotubes. These findings suggest an important role for regular exercise in addition to offering a potential mechanism of action for oral AMPK-activating agents, such as thiazolidinediones and metformin.

Metabolic profiling of muscle contraction in lean compared with obese rodents.

Interest in the pathophysiological relevance of intramuscular triacylglycerol (IMTG) accumulation has grown from numerous studies reporting that abnormally high glycerolipid levels in tissues of obese and diabetic subjects correlate negatively with glucose tolerance. Here, we used a hindlimb perfusion model to examine the impact of obesity and elevated IMTG levels on contraction-induced changes in skeletal muscle fuel metabolism. Comprehensive lipid profiling was performed on gastrocnemius muscles harvested from lean and obese Zucker rats immediately and 25 min after 15 min of one-legged electrically stimulated contraction compared with the contralateral control (rested) limbs. Predictably, IMTG content was grossly elevated in control muscles from obese rats compared with their lean counterparts. In muscles of obese (but not lean) rats, contraction resulted in marked hydrolysis of IMTG, which was then restored to near resting levels during 25 min of recovery. Despite dramatic phenotypical differences in contraction-induced IMTG turnover, muscle levels of diacylglycerol (DAG) and long-chain acyl-CoAs (LCACoA) were surprisingly similar between groups. Tissue profiles of acylcarnitine metabolites suggested that the surfeit of IMTG in obese rats fueled higher rates of fat oxidation relative to the lean group. Muscles of the obese rats had reduced lactate levels immediately following contraction and higher glycogen resynthesis during recovery, consistent with a lipid-associated glucose-sparing effect. Together, these findings suggest that contraction-induced mobilization of local lipid reserves in obese muscles promotes beta-oxidation, while discouraging glucose utilization. Further studies are necessary to determine whether persistent oxidation of IMTG-derived fatty acids contributes to systemic glucose intolerance in other physiological settings.

Plasma lactate as a marker of metabolic health: Implications of elevated lactate for impairment of aerobic metabolism in the metabolic syndrome.

BACKGROUND: Fasting lactate is elevated in metabolic diseases and could possibly be predictive of the risk of developing the metabolic syndrome. METHODS: Plasma samples were analyzed for fasting lactate to compare lean subjects, nondiabetic subjects with severe obesity, and metabolically impaired subjects. Subjects with severe obesity were studied 1 week before and 1 week to 9 months after gastric bypass surgery. Subjects with components of the metabolic syndrome were studied before and after 6 months of an exercise intervention. RESULTS: Metabolically impaired subjects had higher fasting lactate concentrations (P < .0001) and respond to a glucose or insulin challenge with higher lactates than non-obese subjects (P < .004). Lactate was significantly reduced a week after gastric bypass surgery (P < .05) and further reduced 1 to 9 months after surgery (0.95 ± 0.04 mM in non-obese, 1.26 ± 0.12 mM in subjects with severe obesity, and 0.68 ± 0.03 mM 1-3 months after gastric bypass). Six months of chronic exercise resulted in a 16% reduction (P = .028) in fasting lactate. CONCLUSION: Fasting plasma lactate was elevated in obese subjects with the metabolic syndrome compared with healthy lean individuals. Lactate was reduced by exercise and bariatric surgery, interventions that improve metabolic health and risk for subsequent disease. The results of this study and those previously published by our research group suggest that elevated lactate may be caused by an impairment in aerobic metabolism and may offer a metric assessing the severity of the metabolic syndrome.

Mechanism for improved insulin sensitivity after gastric bypass surgery.

CONTEXT: Surgical treatments of obesity have been shown to induce rapid and prolonged improvements in insulin sensitivity. OBJECTIVE: The aim of the study was to investigate the effects of gastric bypass surgery and the mechanisms that explain the improvement in insulin sensitivity. DESIGN: We performed a cross-sectional, nonrandomized, controlled study. SETTING: This study was conducted jointly between the Departments of Exercise Science and Physiology at East Carolina University in Greenville, North Carolina. SUBJECTS: Subjects were recruited into four groups: 1) lean [body mass index (BMI) < 25 kg/m(2); n = 93]; 2) weight-matched (BMI = 25 to 35 kg/m(2); n = 310); 3) morbidly obese (BMI > 35 kg/m(2); n = 43); and 4) postsurgery patients (BMI approximately 30 kg/m(2); n = 40). Postsurgery patients were weight stable 1 yr after surgery. MAIN OUTCOME MEASURES: Whole-body insulin sensitivity, muscle glucose transport, and muscle insulin signaling were assessed. RESULTS: Postsurgery subjects had insulin sensitivity index values that were similar to the lean and higher than morbidly obese and weight-matched control subjects. Glucose transport was higher in the postsurgery vs. morbidly obese and weight-matched groups. IRS1-pSer(312) in the postsurgery group was lower than morbidly obese and weight-matched groups. Inhibitor kappaBalpha was higher in the postsurgery vs. the morbidly obese and weight-matched controls, indicating reduced inhibitor of kappaB kinase beta activity. CONCLUSIONS: Insulin sensitivity and glucose transport are greater in the postsurgery patients than predicted from the weight-matched group, suggesting that improved insulin sensitivity after bypass is due to something other than, or in addition to, weight loss. Improved insulin sensitivity is related to reduced inhibitor of kappaB kinase beta activity and enhanced insulin signaling in muscle.

Time course metabolome of Roux-en-Y gastric bypass confirms correlation between leptin, body weight and the microbiome.

Roux-en-Y gastric bypass (RYGB) is an effective way to lose weight and reverse type 2 diabetes. We profiled the metabolome of 18 obese patients (nine euglycemic and nine diabetics) that underwent RYGB surgery and seven lean subjects. Plasma samples from the obese patients were collected before the surgery and one week and three months after the surgery. We analyzed the metabolome in association to five hormones (Adiponectin, Insulin, Ghrelin, Leptin, and Resistin), four peptide hormones (GIP, Glucagon, GLP1, and PYY), and two cytokines (IL-6 and TNF). PCA showed samples cluster by surgery time and many microbially driven metabolites (indoles in particular) correlated with the three months after the surgery. Network analysis of metabolites revealed a connection between carbohydrate (mannosamine and glucosamine) and glyoxylate and confirms glyoxylate association to diabetes. Only leptin and IL-6 had a significant association with the measured metabolites. Leptin decreased immediately after RYGB (before significant weight loss), whereas IL-6 showed no consistent response to RYGB. Moreover, leptin associated with tryptophan in support of the possible role of leptin in the regulation of serotonin synthesis pathways in the gut. These results suggest a potential link between gastric leptin and microbial-derived metabolites in the context of obesity and diabetes.

High Incomplete Skeletal Muscle Fatty Acid Oxidation Explains Low Muscle Insulin Sensitivity in Poorly Controlled T2D.

Context: Almost 50% of type 2 diabetic (T2D) patients are poorly controlled [glycated hemoglobin (HbA1c) ≥ 7%]; however, the mechanisms responsible for progressively worsening glycemic control are poorly understood. Lower skeletal muscle mitochondrial respiratory capacity is associated with low insulin sensitivity and the development of T2D. Objective: We investigated if skeletal muscle insulin sensitivity (SI) was different between well-controlled T2D (WCD) and poorly controlled T2D (PCD) and if the difference was associated with differences resulting from mitochondrial respiratory function. Design: Vastus lateralis muscle mitochondrial respiration, mitochondrial content, mitochondrial enzyme activity, and fatty acid oxidation (FAO) were measured. SI and the acute response to glucose (AIRg) were calculated by MINMOD analysis from glucose and insulin obtained during a modified, frequently sampled, intravenous glucose tolerance test. Results: SI and AIRg were lower in PCD than WCD. Muscle incomplete FAO was greater in PCD than WCD and greater incomplete FAO was associated with lower SI and higher HbA1c. Hydroxyacyl-coenzyme A dehydrogenase expression and activity were greater in PCD than WCD. There was no difference in maximal mitochondrial respiration or content between WCD and PCD. Conclusion: The current results suggest that greater skeletal muscle incomplete FAO in poorly controlled T2D is due to elevated ß oxidation and is associated with worsening muscle SI.

Lipid metabolism, exercise and insulin action.

Skeletal muscle constitutes 40% of body mass and takes up 80% of a glucose load. Therefore, impaired glucose removal from the circulation, such as that which occurs in obesity and type 2 diabetes, is attributable in large part to the insulin resistance in muscle. Recent research has shown that fatty acids, derived from adipose tissue, can interfere with insulin signalling in muscle. Hence, insulin-stimulated GLUT4 translocation to the cell surface is impaired, and therefore, the rate of glucose removal from the circulation into muscle is delayed. The mechanisms provoking lipid-mediated insulin resistance are not completely understood. In sedentary individuals, excess intramyocellular accumulation of triacylglycerols is only modestly associated with insulin resistance. In contrast, endurance athletes, despite accumulating large amounts of intramyocellular triacylglycerols, are highly insulin sensitive. Thus it appears that lipid metabolites, other than triacylglycerols, interfere with insulin signalling. These metabolites, however, are not expected to accumulate in athletic muscles, as endurance training increases the capacity for fatty acid oxidation by muscle. These observations, and others in severely obese individuals and type 2 diabetes patients, suggest that impaired rates of fatty acid oxidation are associated with insulin resistance. In addition, in obesity and type 2 diabetes, the rates of fatty acid transport into muscle are also increased. Thus, excess intracellular lipid metabolite accumulation, which interferes with insulin signalling, can occur as a result of impaired rates of fatty acid oxidation and/or increased rates of fatty acid transport into muscle. Accumulation of excess intramyocellular lipid can be avoided by exercise, which improves the capacity for fatty acid oxidation.

Differential acute and chronic responses in insulin action in cultured myotubes following from nondiabetic severely obese humans following gastric bypass surgery.

BACKGROUND: Roux-en-Y gastric bypass (RYGB) surgery has been shown to induce positive metabolic adaptations for individuals with severe obesity (body mass index ≥40 kg/m2), including improved peripheral insulin action. Although a major site of insulin action, the time course changes in skeletal muscle glucose metabolism following RYGB is unclear. OBJECTIVES: To investigate the acute and chronic effects of RYGB surgery on insulin-stimulated glucose metabolism in cultured human primary myotubes derived from nondiabetic severely obese humans. SETTING: East Carolina University Bariatric Surgery Center and East Carolina Diabetes and Obesity Institute. METHODS: Primary human skeletal muscle cells were isolated from biopsies obtained from 8 women with severe obesity before, 1 month, and 7 months following RYGB surgery. Glucose metabolism, glycogen content, and insulin signal transduction were determined in differentiated myotubes. RESULTS: Insulin-stimulated glycogen synthesis and glucose oxidation increased in human myotubes derived from patients with severe obesity at both 1 and 7 months post-RYGB. However, there were no alterations indicative of enhanced insulin signal transduction. At 1 month post-RYGB, muscle glycogen levels were lower (-23%) and phosphorylation of acetyl CoA carboxylase 2 (ACC2) was elevated (+16%); both returned to presurgery levels at 7 months after RYGB in myotubes derived from patients. At 7 months post-RYGB, there was an increase in peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC1α) protein content (+54%). CONCLUSION: These data indicate that insulin action intrinsically improves in cultured human primary myotubes derived from nondiabetic severely obese patients following RYGB surgery; however, the cellular alterations involved appear to consist of distinct acute and chronic components.

Circulating adipocyte-derived exosomal MicroRNAs associated with decreased insulin resistance after gastric bypass.

OBJECTIVE: Exosomes from obese adipose contain dysregulated microRNAs linked to insulin signaling, as compared with lean controls, providing a direct connection between adiposity and insulin resistance. This study tested the hypotheses that gastric bypass surgery and its subsequent weight loss would normalize adipocyte-derived exosomal microRNAs associated with insulin signaling and the associated metabolome related to glucose homeostasis. METHODS: African American female subjects with obesity (N = 6; age: 38.5 ± 6.8 years; BMI: 51.2 ± 8.8 kg/m2 ) were tested before and 1 year after surgery. Insulin resistance (HOMA), serum metabolomics, and global microRNA profiles of circulating adipocyte-derived exosomes were evaluated via ANCOVA and correlational analyses. RESULTS: One year postsurgery, patients showed decreased BMI (-18.6 ± 5.1 kg/m2 ; P < 0.001), ameliorated insulin resistance (HOMA: 1.94 ± 0.6 presurgery, 0.49 ± 0.1 postsurgery; P < 0.001), and altered metabolites including branched chain amino acids (BCAA). Biological pathway analysis of predicted mRNA targets of 168 surgery-responsive microRNAs (P < 0.05) identified the insulin signaling pathway (P = 1.27E-10; 52/138 elements), among others, in the data set. The insulin signaling pathway was also a target of 10 microRNAs correlated to changes in HOMA (P < 0.05; r > 0.4), and 48 microRNAs correlated to changes in BCAA levels. CONCLUSIONS: These data indicate that circulating adipocyte-derived exosomes are modified following gastric bypass surgery and correlate to improved postsurgery insulin resistance.

Defective signaling through Akt-2 and -3 but not Akt-1 in insulin-resistant human skeletal muscle: potential role in insulin resistance.

Recent evidence has shown that activation of phosphatidyinositol-3-kinase (PI3K) and Akt, necessary for insulin stimulation of glucose transport, is impaired in insulin resistance. It is unknown, however, which Akt isoform shows impaired activation in insulin resistance. Additionally, related growth factors (epidermal or platelet-derived vascular) also stimulate PI3K, but it is unknown whether production of 3,4,5 phosphatidyinositol is sufficient to stimulate glucose transport in insulin-resistant muscle. Moreover, these studies were performed in rodents, and little data exists from humans. Hence, we investigated the stimulation of PI3K and Akt-1, -2, and -3 by insulin and epidermal growth factors (EGFs) in skeletal muscles from lean and obese insulin-resistant humans. Insulin activated all Akt isoforms in lean muscles, whereas only Akt-1 was activated in obese muscles. Insulin receptor substrate (IRS)-1 was associated with PI3K activity, which is necessary for Akt activation by insulin, and was reduced in obese muscles, and this was accompanied by decreased IRS-1 expression. In contrast, insulin- or EGF-stimulated phosphotyrosine-associated PI3K activity was not different between lean and obese muscles. These results show that a defect in the ability of insulin to activate Akt-2 and -3 may explain the impaired insulin-stimulated glucose transport in insulin resistance. Additionally, these data also show that different upstream or downstream signals may regulate the activity of the various Akt isoforms.

Peroxisome proliferator-activated receptor-alpha regulates fatty acid utilization in primary human skeletal muscle cells.

In humans, skeletal muscle is a major site of peroxisome proliferator-activated receptor-alpha (PPAR-alpha) expression, but its function in this tissue is unclear. We investigated the role of hPPAR-alpha in regulating muscle lipid utilization by studying the effects of a highly selective PPAR-alpha agonist, GW7647, on [(14)C]oleate metabolism and gene expression in primary human skeletal muscle cells. Robust induction of PPAR-alpha protein expression occurred during muscle cell differentiation and corresponded with differentiation-dependent increases in oleate oxidation. In mature myotubes, 48-h treatment with 10-1,000 nmol/l GW7647 increased oleate oxidation dose-dependently, up to threefold. Additionally, GW7647 decreased oleate esterification into myotube triacylglycerol (TAG), up to 45%. This effect was not abolished by etomoxir, a potent inhibitor of beta-oxidation, indicating that PPAR-alpha-mediated TAG depletion does not depend on reciprocal changes in fatty acid catabolism. Consistent with its metabolic actions, GW7647 induced mRNA expression of mitochondrial enzymes that promote fatty acid catabolism; carnitine palmityltransferase 1 and malonyl-CoA decarboxylase increased approximately 2-fold, whereas pyruvate dehydrogenase kinase 4 increased 45-fold. Expression of several genes that regulate glycerolipid synthesis was not changed by GW7647 treatment, implicating involvement of other targets to explain the TAG-depleting effect of the compound. These results demonstrate a role for hPPAR-alpha in regulating muscle lipid homeostasis.