High fat diet-induced changes in mouse muscle mitochondrial phospholipids do not impair mitochondrial respiration despite insulin resistance

Background

Type 2 diabetes mellitus and muscle insulin resistance have been associated with reduced capacity of skeletal muscle mitochondria, possibly as a result of increased intake of dietary fat. Here, we examined the hypothesis that a prolonged high-fat diet consumption (HFD) increases the saturation of muscle mitochondrial membrane phospholipids causing impaired mitochondrial oxidative capacity and possibly insulin resistance.

Methodology

C57BL/6J mice were fed an 8-week or 20-week low fat diet (10 kcal%; LFD) or HFD (45 kcal%). Skeletal muscle mitochondria were isolated and fatty acid (FA) composition of skeletal muscle mitochondrial phospholipids was analyzed by thin-layer chromatography followed by GC. High-resolution respirometry was used to assess oxidation of pyruvate and fatty acids by mitochondria. Insulin sensitivity was estimated by HOMA-IR.

Principal Findings

At 8 weeks, mono-unsaturated FA (16∶1n7, 18∶1n7 and 18∶1n9) were decreased (−4.0%, p<0.001), whereas saturated FA (16∶0) were increased (+3.2%, p<0.001) in phospholipids of HFD vs. LFD mitochondria. Interestingly, 20 weeks of HFD descreased mono-unsaturated FA while n-6 poly-unsaturated FA (18∶2n6, 20∶4n6, 22∶5n6) showed a pronounced increase (+4.0%, p<0.001). Despite increased saturation of muscle mitochondrial phospholipids after the 8-week HFD, mitochondrial oxidation of both pyruvate and fatty acids were similar between LFD and HFD mice. After 20 weeks of HFD, the increase in n-6 poly-unsaturated FA was accompanied by enhanced maximal capacity of the electron transport chain (+49%, p = 0.002) and a tendency for increased ADP-stimulated respiration, but only when fuelled by a lipid-derived substrate. Insulin sensitivity in HFD mice was reduced at both 8 and 20 weeks.

Conclusions/Interpretation

Our findings do not support the concept that prolonged HF feeding leads to increased saturation of skeletal muscle mitochondrial phospholipids resulting in a decrease in mitochondrial fat oxidative capacity and (muscle) insulin resistance.     

Enhanced lipid-but not carbohydrate-supported mitochondrial respiration in skeletal muscle of PGC-1α overexpressing mice

Skeletal muscle mitochondrial dysfunction has been linked to several disease states as well as the process of aging. A possible factor involved is the peroxisome proliferator-activated receptor (PPAR) γ co-activator 1α (PGC-1α), a major player in the regulation of skeletal muscle mitochondrial metabolism. However, it is currently unknown whether PGC-1α, besides stimulating mitochondrial proliferation, also affects the functional capacity per mitochondrion. Therefore, we here tested whether PGC-1α overexpression, besides increasing mitochondrial content, also leads to intrinsic mitochondrial adaptations. Skeletal muscle mitochondria from 10 male, muscle-specific PGC-1α overexpressing mice (PGC-1αTg) and 8 wild-type (WT) mice were isolated. Equal mitochondrial quantities were then analyzed for their oxidative capacity by high-resolution respirometry, fuelled by a carbohydrate-derived (pyruvate) and a lipid (palmitoyl-CoA plus carnitine) substrate. Additionally, mitochondria were tested for reactive oxygen species (superoxide) production and fatty acid (FA)-induced uncoupling. PGC-1αTg mitochondria were characterized by an improved intrinsic mitochondrial fat oxidative capacity as evidenced by pronounced increase in ADP-stimulated respiration (P < 0.001) and maximal uncoupled respiration (P < 0.001) upon palmitoyl-CoA plus carnitine. Interestingly, intrinsic mitochondrial capacity on a carbohydrate-derived substrate tended to be reduced. Furthermore, the sensitivity to FA-induced uncoupling was diminished in PGC-1αTg mitochondria (P = 0.02) and this was accompanied by a blunted reduction in mitochondrial ROS production upon FAs in PGC-1αTg versus WT mitochondria (P = 0.04). Uncoupling protein 3 (UCP3) levels were markedly reduced in PGC-1αTg mitochondria (P < 0.001). Taken together, in addition to stimulating mitochondrial proliferation in skeletal muscle, we show here that overexpression of PGC-1α leads to intrinsic mitochondrial adaptations that seem restricted to fat metabolism.     

Activity-Based Protein Profiling Reveals Mitochondrial Oxidative Enzyme Impairment and Restoration in Diet-Induced Obese Mice

High-fat diet (HFD) induced obesity and concomitant development of insulin resistance (IR) and type 2 diabetes mellitus have been linked to mitochondrial dysfunction. However, it is not clear whether mitochondrial dysfunction is a direct effect of a HFD, or if mitochondrial function is reduced with increased HFD duration. We hypothesized that the function of mitochondrial oxidative and lipid metabolism functions in skeletal muscle mitochondria for HFD mice are similar, or elevated, relative to standard diet (SD) mice; thereby, IR is neither cause nor consequence of mitochondrial dysfunction. We applied a chemical probe approach to identify functionally reactive ATPases and nucleotide-binding proteins in mitochondria isolated from skeletal muscle of C57Bl/6J mice fed HFD or SD chow for 2-, 8-, or 16-weeks; feeding time points known to induce IR. A total of 293 probe-labeled proteins were identified by mass spectrometry-based proteomics, of which 54 differed in abundance between HFD and SD mice. We found proteins associated with the TCA cycle, oxidative phosphorylation (OXPHOS), and lipid metabolism were altered in function when comparing SD to HFD fed mice at 2-weeks, however by 16-weeks HFD mice had TCA cycle, β-oxidation, and respiratory chain function at levels similar to or higher than SD mice.     

High‐fat diet affects skeletal muscle mitochondria comparable to pressure overload‐induced heart failure

In heart failure, high‐fat diet (HFD) may exert beneficial effects on cardiac mitochondria and contractility. Skeletal muscle mitochondrial dysfunction in heart failure is associated with myopathy. However, it is not clear if HFD affects skeletal muscle mitochondria in heart failure as well. To induce heart failure, we used pressure overload (PO) in rats fed normal chow or HFD. Interfibrillar mitochondria (IFM) and subsarcolemmal mitochondria (SSM) from gastrocnemius were isolated and functionally characterized. With PO heart failure, maximal respiratory capacity was impaired in IFM but increased in SSM of gastrocnemius. Unexpectedly, HFD affected mitochondria comparably to PO. In combination, PO and HFD showed additive effects on mitochondrial subpopulations which were reflected by isolated complex activities. While PO impaired diastolic as well as systolic cardiac function and increased glucose tolerance, HFD did not affect cardiac function but decreased glucose tolerance. We conclude that HFD and PO heart failure have comparable effects leading to more severe impairment of IFM. Glucose tolerance seems not causally related to skeletal muscle mitochondrial dysfunction. The additive effects of HFD and PO may suggest accelerated skeletal muscle mitochondrial dysfunction when heart failure is accompanied with a diet containing high fat.     

Mitochondrial DNA Damage and Dysfunction,and Oxidative Stress Are Associated with Endoplasmic Reticulum Stress,Protein Degradation and Apoptosis in High Fat Diet-Induced Insulin Resistance Mice

Background

Recent studies showed a link between a high fat diet (HFD)-induced obesity and lipid accumulation in non-adipose tissues, such as skeletal muscle and liver, and insulin resistance (IR). Although the mechanisms responsible for IR in those tissues are different, oxidative stress and mitochondrial dysfunction have been implicated in the disease process. We tested the hypothesis that HFD induced mitochondrial DNA (mtDNA) damage and that this damage is associated with mitochondrial dysfunction, oxidative stress, and induction of markers of endoplasmic reticulum (ER) stress, protein degradation and apoptosis in skeletal muscle and liver in a mouse model of obesity-induced IR.

Methodology/Principal Findings

C57BL/6J male mice were fed either a HFD (60% fat) or normal chow (NC) (10% fat) for 16 weeks. We found that HFD-induced IR correlated with increased mtDNA damage, mitochondrial dysfunction and markers of oxidative stress in skeletal muscle and liver. Also, a HFD causes a change in the expression level of DNA repair enzymes in both nuclei and mitochondria in skeletal muscle and liver. Furthermore, a HFD leads to activation of ER stress, protein degradation and apoptosis in skeletal muscle and liver, and significantly reduced the content of two major proteins involved in insulin signaling, Akt and IRS-1 in skeletal muscle, and Akt in liver. Basal p-Akt level was not significantly influenced by HFD feeding in skeletal muscle and liver.

Conclusions/Significance

This study provides new evidence that HFD-induced mtDNA damage correlates with mitochondrial dysfunction and increased oxidative stress in skeletal muscle and liver, which is associated with the induction of markers of ER stress, protein degradation and apoptosis.     

Nox2 Mediates Skeletal Muscle Insulin Resistance Induced by a High Fat Diet

Inflammation and oxidative stress through the production of reactive oxygen species (ROS) are consistently associated with metabolic syndrome/type 2 diabetes. Although the role of Nox2, a major ROS-generating enzyme, is well described in host defense and inflammation, little is known about its potential role in insulin resistance in skeletal muscle. Insulin resistance induced by a high fat diet was mitigated in Nox2-null mice compared with wild-type mice after 3 or 9 months on the diet. High fat feeding increased Nox2 expression, superoxide production, and impaired insulin signaling in skeletal muscle tissue of wild-type mice but not in Nox2-null mice. Exposure of C2C12 cultured myotubes to either high glucose concentration, palmitate, or H₂O₂ decreases insulin-induced Akt phosphorylation and glucose uptake. Pretreatment with catalase abrogated these effects, indicating a key role for H₂O₂ in mediating insulin resistance. Down-regulation of Nox2 in C2C12 cells by shRNA prevented insulin resistance induced by high glucose or palmitate but not H₂O₂. These data indicate that increased production of ROS in insulin resistance induced by high glucose in skeletal muscle cells is a consequence of Nox2 activation. This is the first report to show that Nox2 is a key mediator of insulin resistance in skeletal muscle.     

Bioactives from bitter melon enhance insulin signaling and modulate acyl carnitine content in skeletal muscle in high-fat diet-fed mice

Bioactive components from bitter melon (BM) have been reported to improve glucose metabolism in vivo, but definitive studies on efficacy and mechanism of action are lacking. We sought to investigate the effects of BM bioactives on body weight, muscle lipid content and insulin signaling in mice fed a high-fat diet and on insulin signaling in L6 myotubes. Male C57BL/6J mice were randomly divided into low-fat diet control (LFD), high-fat diet (HFD) and HFD plus BM (BM) groups. Body weight, body composition, plasma glucose, leptin, insulin and muscle lipid profile were determined over 12 weeks. Insulin signaling was determined in the mouse muscle taken at end of study and in L6 myotubes exposed to the extract. Body weight, plasma glucose, insulin, leptin levels and HOMA-IR values were significantly lower in the BM-fed HFD group when compared to the HFD group. BM supplementation significantly increased IRS-2, IR β, PI 3K and GLUT4 protein abundance in skeletal muscle, as well as phosphorylation of IRS-1, Akt1 and Akt2 when compared with HFD (P<.05 and P<.01). BM also significantly reduced muscle lipid content in the HFD mice. BM extract greatly increased glucose uptake and enhanced insulin signaling in L6 myotubes. This study shows that BM bioactives reduced body weight, improved glucose metabolism and enhanced skeletal muscle insulin signaling. A contributing mechanism to the enhanced insulin signaling may be associated with the reduction in skeletal muscle lipid content. Nutritional supplementation with this extract, if validated for human studies, may offer an adjunctive therapy for diabetes.     

A Small Amount of Dietary Carbohydrate Can Promote the HFD-Induced Insulin Resistance to a Maximal Level

Both dietary fat and carbohydrates (Carbs) may play important roles in the development of insulin resistance. The main goal of this study was to further define the roles for fat and dietary carbs in insulin resistance. C57BL/6 mice were fed normal chow diet (CD) or HFD containing 0.1–25.5% carbs for 5 weeks, followed by evaluations of calorie consumption, body weight and fat gains, insulin sensitivity, intratissue insulin signaling, ectopic fat, and oxidative stress in liver and skeletal muscle. The role of hepatic gluconeogenesis in the HFD-induced insulin resistance was determined in mice. The role of fat in insulin resistance was also examined in cultured cells. HFD with little carbs (0.1%) induced severe insulin resistance. Addition of 5% carbs to HFD dramatically elevated insulin resistance and 10% carbs in HFD was sufficient to induce a maximal level of insulin resistance. HFD with little carbs induced ectopic fat accumulation and oxidative stress in liver and skeletal muscle and addition of carbs to HFD dramatically enhanced ectopic fat and oxidative stress. HFD increased hepatic expression of key gluconeogenic genes and the increase was most dramatic by HFD with little carbs, and inhibition of hepatic gluconeogenesis prevented the HFD-induced insulin resistance. In cultured cells, development of insulin resistance induced by a pathological level of insulin was prevented in the absence of fat. Together, fat is essential for development of insulin resistance and dietary carb is not necessary for HFD-induced insulin resistance due to the presence of hepatic gluconeogenesis but a very small amount of it can promote HFD-induced insulin resistance to a maximal level.     

Mitochondrial dysfunction and inhibition of myoblast differentiation in mice with high-fat-diet-induced pre-diabetes

Pre-diabetes is characterized by impaired glucose tolerance (IGT) and/or impaired fasting glucose. Impairment of skeletal muscle function is closely associated with the progression of diabetes. However, the entire pathological characteristics and mechanisms of pre-diabetes in skeletal muscle remain fully unknown. Here, we established a mouse model of pre-diabetes, in which 6-week-old male C57BL6/J mice were fed either normal diet or high-fat diet (HFD) for 8 or 16 weeks. Both non-fasting and fasting glucose levels and the results of glucose and insulin tolerance tests showed that mice fed an 8-week HFD developed pre-diabetes with IGT; whereas mice fed a 16-week HFD presented with impaired fasting glucose and impaired glucose tolerance (IFG-IGT). Mice at both stages of pre-diabetes displayed decreased numbers of mitochondria in skeletal muscle. Moreover, IFG-IGT mice exhibited decreased mitochondrial membrane potential and ATP production in skeletal muscle and muscle degeneration characterized by a shift in muscle fibers from predominantly oxidative type I to glycolytic type II. Western blotting and histological analysis confirmed that myoblast differentiation was only inhibited in IFG-IGT mice. For primary skeletal muscle satellite cells, inhibition of differentiation was observed in palmitic acid-induced insulin resistance model. Moreover, enhanced myoblast differentiation increased glucose uptake and insulin sensitivity. These findings indicate that pre-diabetes result in mitochondrial dysfunction and inhibition of myoblast differentiation in skeletal muscle. Therefore, interventions that enhance myoblast differentiation may improve insulin resistance of diabetes at the earlier stage.     

Lack of skeletal muscle liver kinase B1 alters gene expression,mitochondrial content,inflammation and oxidative stress without affecting high-fat diet-induced obesity or insulin resistance

Ad libitum high-fat diet (HFD) induces obesity and skeletal muscle metabolic dysfunction. Liver kinase B1 (LKB1) regulates skeletal muscle metabolism by controlling the AMP-activated protein kinase family, but its importance in regulating muscle gene expression and glucose tolerance in obese mice has not been established. The purpose of this study was to determine how the lack of LKB1 in skeletal muscle (KO) affects gene expression and glucose tolerance in HFD-fed, obese mice.KO and littermate control wild-type (WT) mice were fed a standard diet or HFD for 14 weeks. RNA sequencing, and subsequent analysis were performed to assess mitochondrial content and respiration, inflammatory status, glucose and insulin tolerance, and muscle anabolic signaling.KO did not affect body weight gain on HFD, but heavily impacted mitochondria-, oxidative stress-, and inflammation-related gene expression. Accordingly, mitochondrial protein content and respiration were suppressed while inflammatory signaling and markers of oxidative stress were elevated in obese KO muscles. KO did not affect glucose or insulin tolerance. However, fasting serum insulin and skeletal muscle insulin signaling were higher in the KO mice. Furthermore, decreased muscle fiber size in skmLKB1-KO mice was associated with increased general protein ubiquitination and increased expression of several ubiquitin ligases, but not muscle ring finger 1 or atrogin-1. Taken together, these data suggest that the lack of LKB1 in skeletal muscle does not exacerbate obesity or insulin resistance in mice on a HFD, despite impaired mitochondrial content and function and elevated inflammatory signaling and oxidative stress.     

Changes of skeletal muscle adiponectin content in diet-induced insulin resistant rats

The current study examined the relationship between skeletal muscle levels of adiponectin and parameters of insulin sensitivity. A high fat/sucrose diet (HFD) for 20 weeks resulted in significant increases in body weight, serum insulin, triglycerides (TG), and free fatty acids (FFA) (all p < 0.01). Interestingly, this diet leads to a slight increase in serum adiponectin, but significant decreases in gastrocnemius muscle and white adipose adiponectin (all p < 0.05). HFD for 4 weeks also resulted in a significant decrease in muscle adiponectin, which correlated with serum insulin, TG, and FFA (all p < 0.05). Treatment of the 4-week HFD rats with a PPARgamma agonist GI262570 ameliorated the diet-induced hyperinsulinemia and dyslipidemia, and effectively restored muscle adiponectin (all p < 0.05). This study demonstrated that HFD-induced hyperinsulinemia and dyslipidemia appeared without changes in serum adiponectin, but were associated with decreased tissue adiponectin. This provides the first evidence for a connection between tissue adiponectin and diet-induced hyperinsulinemia and dyslipidemia.     

Enhanced Lipid Oxidation and Maintenance of Muscle Insulin Sensitivity Despite Glucose Intolerance in a Diet-Induced Obesity Mouse Model

Background

Diet-induced obesity is a rising health concern which can lead to the development of glucose intolerance and muscle insulin resistance and, ultimately, type II diabetes mellitus. This research investigates the associations between glucose intolerance or muscle insulin resistance and tissue specific changes during the progression of diet-induced obesity.

Methodology

C57BL/6J mice were fed a normal or high-fat diet (HFD; 60% kcal fat) for 3 or 8 weeks. Disease progression was monitored by measurements of body/tissue mass changes, glucose and insulin tolerance tests, and ex vivo glucose uptake in intact muscles. Lipid metabolism was analyzed using metabolic chambers and ex vivo palmitate assays in intact muscles. Skeletal muscle, liver and adipose tissues were analyzed for changes in inflammatory gene expression. Plasma was analyzed for insulin levels and inflammatory proteins. Histological techniques were used on muscle and liver cryosections to assess metabolic and morphological changes.

Principal Findings/Conclusions

A rapid shift in whole body metabolism towards lipids was observed with HFD. Following 3 weeks of HFD, elevated total lipid oxidation and an oxidative fiber type shift had occurred in the skeletal muscle, which we propose was responsible for delaying intramyocellular lipid accumulation and maintaining muscle’s insulin sensitivity. Glucose intolerance was present after three weeks of HFD and was associated with an enlarged adipose tissue depot, adipose tissue inflammation and excess hepatic lipids, but not hepatic inflammation. Furthermore, HFD did not significantly increase systemic or muscle inflammation after 3 or 8 weeks of HFD suggesting that early diet-induced obesity does not cause inflammation throughout the whole body. Overall these findings indicate skeletal muscle did not contribute to the development of HFD-induced impairments in whole-body glucose tolerance following 3 weeks of HFD.     

Diet-Sensitive Sources of Reactive Oxygen Species in Liver Mitochondria: Role of Very Long Chain Acyl-CoA Dehydrogenases

High fat diets and accompanying hepatic steatosis are highly prevalent conditions. Previous work has shown that steatosis is accompanied by enhanced generation of reactive oxygen species (ROS), which may mediate further liver damage. Here we investigated mechanisms leading to enhanced ROS generation following high fat diets (HFD). We found that mitochondria from HFD livers present no differences in maximal respiratory rates and coupling, but generate more ROS specifically when fatty acids are used as substrates. Indeed, many acyl-CoA dehydrogenase isoforms were found to be more highly expressed in HFD livers, although only the very long chain acyl-CoA dehydrogenase (VLCAD) was more functionally active. Studies conducted with permeabilized mitochondria and different chain length acyl-CoA derivatives suggest that VLCAD is also a source of ROS production in mitochondria of HFD animals. This production is stimulated by the lack of NAD⁺. Overall, our studies uncover VLCAD as a novel, diet-sensitive, source of mitochondrial ROS.     

Effects of Dietary Eicosapentaenoic Acid (EPA) Supplementation in High-Fat Fed Mice on Lipid Metabolism and Apelin/APJ System in Skeletal Muscle

Various studies have shown that eicosapentaenoic acid (EPA) has beneficial effects on obesity and associated disorders. Apelin, the ligand of APJ receptor also exerts insulin-sensitizing effects especially by improving muscle metabolism. EPA has been shown to increase apelin production in adipose tissue but its effects in muscle have not been addressed. Thus, the effects of EPA supplementation (36 g/kg EPA) in high-fat diet (HFD) (45% fat, 20% protein, 35% carbohydrate) were studied in mice with focus on muscle lipid metabolism and apelin/APJ expression. Compared with HFD mice, HFD+EPA mice had significantly less weight gain, fat mass, lower blood glucose, insulinemia and hepatic steatosis after 10 weeks of diet. In addition, EPA prevented muscle metabolism alterations since intramuscular triglycerides were decreased and β-oxidation increased. In soleus muscles of HFD+EPA mice, apelin and APJ expression were significantly increased compared to HFD mice. However, plasma apelin concentrations in HFD and HFD+EPA mice were similar. EPA-induced apelin expression was confirmed in differentiated C2C12 myocytes but in this model, apelin secretion was also increased in response to EPA treatment. In conclusion, EPA supplementation in HFD prevents obesity and metabolic alterations in mice, especially in skeletal muscle. Since EPA increases apelin/APJ expression in muscle, apelin may act in a paracrine/autocrine manner to contribute to these benefical effects.     

Skeletal muscle reactive oxygen species: A target of good cop/bad cop for exercise and disease

Abstract

Metabolic stresses associated with disease, ageing, and exercise increase the levels of reactive oxygen species (ROS) in skeletal muscle. These ROS have been linked mechanistically to adaptations in skeletal muscle that can be favourable (i.e. in response to exercise) or detrimental (i.e. in response to disease). The magnitude, duration (acute versus chronic), and cellular origin of the ROS are important underlying factors in determining the metabolic perturbations associated with the ROS produced in skeletal muscle. In particular, insulin resistance has been linked to excess ROS production in skeletal muscle mitochondria. A chronic excess of mitochondrial ROS can impair normal insulin signalling pathways and glucose disposal in skeletal muscle. In contrast, ROS produced in skeletal muscle in response to exercise has been linked to beneficial metabolic adaptations including mitochondrial biogenesis and muscle hypertrophy. Moreover, unlike insulin resistance, exercise-induced ROS appears to be primarily of non-mitochondrial origin. The present review summarizes the diverse ROS-targeted metabolic outcomes associated with insulin resistance versus exercise in skeletal muscle, thus, presenting two contrasting perspectives of pathologically harmful versus physiologically beneficial ROS. Here, we discuss the key sites of ROS production during exercise and the effect of ROS in skeletal muscle of people with type 2 diabetes.     

Long-term programming of skeletal muscle and liver lipid and energy metabolism by resveratrol supplementation to suckling mice

Metabolic programming by dietary chemicals consumed in early life stages is receiving increasing attention. We here studied long-term effects of mild resveratrol (RSV) supplementation during lactation on muscular and hepatic lipid metabolism in adulthood. Newborn male mice received RSV or vehicle from day 2–20 of age, were weaned onto a chow diet on day 21, and were assigned to either a high-fat diet (HFD) or a normal-fat diet on day 90 of age for 10 weeks. RSV-treated mice showed in adulthood protection against HFD-induced triacylglycerol accumulation in skeletal muscle, enhanced muscular capacities for fat oxidation and mitochondria activity, signs of enhanced sirtuin 1 and AMP-dependent protein kinase signaling in muscle, and increased fat oxidation capacities and a decreased capacity for lipogenesis in liver compared with controls. Thus, RSV supplementation in early postnatal life may help preventing later diet-related disorders linked to ectopic lipid accumulation in muscle and liver tissues.     

High-fat feeding inhibits exercise-induced increase in mitochondrial respiratory flux in skeletal muscle

Twenty one healthy untrained male subjects were randomized to follow a high-fat diet (HFD; 55-60E% fat, 25-30E% carbohydrate, and 15E% protein) or a normal diet (ND; 25-35E% fat, 55-60E% carbohydrate, and 10-15E% protein) for 2(1/2) wk. Diets were isocaloric and tailored individually to match energy expenditure. At 2(1/2) wk of diet, one 60-min bout of bicycle exercise (70% of maximal oxygen uptake) was performed. Muscle biopsies were obtained before and after the diet, immediately after exercise, and after 3-h recovery. Insulin sensitivity (hyperinsulinemic-euglycemic clamp) and intramyocellular triacylglycerol content did not change with the intervention in either group. Indexes of mitochondrial density were similar across the groups and intervention. Mitochondrial respiratory rates, measured in permeabilized muscle fibers, showed a 31 ± 11 and 26 ± 9% exercise-induced increase (P < 0.05) in state 3 (glycolytic substrates) and uncoupled respiration, respectively. However, in HFD this increase was abolished. At recovery, no change from resting respiration was seen in either group. With a lipid substrate (octanoyl-carnitine with or without ADP), similar exercise-induced increases (31-62%) were seen in HFD and ND, but only in HFD was an elevated (P < 0.05) respiratory rate seen at recovery. With HFD complex I and IV protein expression decreased (P < 0.05 and P = 0.06, respectively). A fat-rich diet induces marked changes in the mitochondrial electron transport system protein content and in exercise-induced mitochondrial substrate oxidation rates, with the effects being present hours after the exercise. The effect of HFD is present even without effects on insulin sensitivity and intramyocellular lipid accumulation. An isocaloric high-fat diet does not cause insulin resistance.     

Globular adiponectin resistance develops independently of impaired insulin-stimulated glucose transport in soleus muscle from high-fat-fed rats

High-fat (HF) diets induce insulin resistance and alter lipid metabolism, although controversy exists regarding the impact of saturated vs. polyunsaturated fats. Adiponectin (Ad) stimulates fatty acid (FA) oxidation and improves insulin sensitivity in humans and rodents, due in part to the activation of AMP-activated protein kinase (AMPK) and subsequent deactivation of acetyl coenzyme A carboxylase (ACC). In genetically obese, diabetic mice, this acute stimulatory effect on AMPK in muscle is lost. The ability of a HF diet to induce skeletal muscle Ad resistance has not been examined. The purpose of this study was to determine whether Ad's effects on FA oxidation and AMPK/ACC would be reduced following different HF diets, and if this coincided with the development of impaired maximal insulin-stimulated glucose transport. Rats were fed a control (10% kcal fat, CON), high unsaturated fat (60% kcal safflower oil, SAFF), or high saturated fat diet (60% kcal lard, LARD) for 4 wk. Following the dietary intervention, glucose transport, lipid metabolism, and AMPK/ACC phosphorylation were measured in the presence and absence of globular Ad (gAd, 2.5 microg/ml) in isolated soleus muscle. LARD rats showed reduced rates of maximal insulin-stimulated glucose transport compared with CON and SAFF (+68 vs. +172 and +184%, P < or = 0.001). gAd increased pACC (+25%, P < or = 0.01) and FA oxidation (+28%, P < or = 0.05) in CON rats, but not in either HF group. Thus 4 wk of HF feeding results in the loss of gAd stimulatory effect on ACC phosphorylation and muscle FA oxidation, and this can occur independently of impaired maximal insulin-stimulated glucose transport.     

Altered estrogen receptor expression in skeletal muscle and adipose tissue of female rats fed a high-fat diet

Estrogen receptors (ERs) are expressed in adipose tissue and skeletal muscle, with potential implications for glucose metabolism and insulin signaling. Previous studies examining the role of ERs in glucose metabolism have primarily used knockout mouse models of ERα and ERβ, and it is unknown whether ER expression is altered in response to an obesity-inducing high-fat diet (HFD). The purpose of the current study was to determine whether modulation of glucose metabolism in response to a HFD in intact and ovariectomized (OVX) female rats is associated with alterations in ER expression. Our results demonstrate that a 6-wk HFD (60% calories from fat) in female rats induces whole body glucose intolerance with tissue-specific effects isolated to the adipose tissue, and no observed differences in insulin-stimulated glucose uptake, GLUT4, or ERα protein expression levels in skeletal muscle. In chow-fed rats, OVX resulted in decreased ERα with a trend toward decreased GLUT4 expression in adipose tissue. Sham-treated and OVX rats fed a HFD demonstrated a decrease in ERα and GLUT4 in adipose tissue. The HFD also increased activation of stress kinases (c-jun NH?-terminal kinase and inhibitor of κB kinase β) in the sham-treated rats and decreased expression of the protective heat shock protein 72 (HSP72) in both sham-treated and OVX rats. Our findings suggest that decreased glucose metabolism and increased inflammation in adipose tissue with a HFD in female rats could stem from a significant decrease in ERα expression.     

Disruption of CXC motif chemokine ligand-14 in mice ameliorates obesity-induced insulin resistance

In obese individuals, white adipose tissue (WAT) is infiltrated by large numbers of macrophages, resulting in enhanced inflammatory responses that contribute to insulin resistance. Here we show that expression of the CXC motif chemokine ligand-14 (CXCL14), which targets tissue macrophages, is elevated in WAT of obese mice fed a high fat diet (HFD) compared with lean mice fed a regular diet. We found that HFD-fed CXCL14-deficient mice have impaired WAT macrophage mobilization and improved insulin responsiveness. Insulin-stimulated phosphorylation of Akt kinase in skeletal muscle was severely attenuated in HFD-fed CXCL14+/- mice but not in HFD-fed CXCL14-/- mice. The insulin-sensitive phenotype of CXCL14-/- mice after HFD feeding was prominent in female mice but not in male mice. HFD-fed CXCL14-/- mice were protected from hyperglycemia, hyperinsulinemia, and hypoadiponectinemia and did not exhibit increased levels of circulating retinol-binding protein-4 and increased expression of interleukin-6 in WAT. Transgenic overexpression of CXCL14 in skeletal muscle restored obesity-induced insulin resistance in CXCL14-/- mice. CXCL14 attenuated insulin-stimulated glucose uptake in cultured myocytes and to a lesser extent in cultured adipocytes. These results demonstrate that CXCL14 is a critical chemoattractant of WAT macrophages and a novel regulator of glucose metabolism that functions mainly in skeletal muscle.