Genome-wide chromatin state transitions associated with developmental and environmental cues.

Differences in chromatin organization are key to the multiplicity of cell states that arise from a single genetic background, yet the landscapes of in vivo tissues remain largely uncharted. Here, we mapped chromatin genome-wide in a large and diverse collection of human tissues and stem cells. The maps yield unprecedented annotations of functional genomic elements and their regulation across developmental stages, lineages, and cellular environments. They also reveal global features of the epigenome, related to nuclear architecture, that also vary across cellular phenotypes. Specifically, developmental specification is accompanied by progressive chromatin restriction as the default state transitions from dynamic remodeling to generalized compaction. Exposure to serum in vitro triggers a distinct transition that involves de novo establishment of domains with features of constitutive heterochromatin. We describe how these global chromatin state transitions relate to chromosome and nuclear architecture, and discuss their implications for lineage fidelity, cellular senescence, and reprogramming.

Effect of resveratrol on insulin action in primary myotubes from lean individuals and individuals with severe obesity.

Resveratrol, a natural polyphenol compound contained in numerous plants, has been proposed as a treatment for obesity-related disease processes such as insulin resistance. However, in humans there are conflicting results concerning the efficacy of resveratrol in improving insulin action; the purpose of the present study was to determine whether obesity status (lean, severely obese) affects the response to resveratrol in human skeletal muscle. Primary skeletal muscle cells were derived from biopsies obtained from age-matched lean and insulin-resistant women with severe obesity and incubated with resveratrol (1 µM) for 24 h. Insulin-stimulated glucose oxidation and incorporation into glycogen, insulin signal transduction, and energy-sensitive protein targets [AMP-activated protein kinase (AMPK), Sirt1, and PGC1α] were analyzed. Insulin-stimulated glycogen synthesis, glucose oxidation, and AMPK phosphorylation increased with resveratrol incubation compared with the nonresveratrol conditions (main treatment effect for resveratrol). Resveratrol further increased IRS1, Akt, and TBC1D4 insulin-stimulated phosphorylation and SIRT1 content in myotubes from lean women, but not in women with severe obesity. Resveratrol improves insulin action in primary human skeletal myotubes derived from lean women and women with severe obesity. In women with obesity, these improvements may be associated with enhanced AMPK phosphorylation with resveratrol treatment.NEW & NOTEWORTHY A physiologically relevant dose of resveratrol increases insulin-stimulated glucose oxidation and glycogen synthesis in myotubes from individuals with severe obesity. Furthermore, resveratrol improved insulin signal transduction in myotubes from lean individuals but not from individuals with obesity. Activation of AMPK plays a role in resveratrol-induced improvements in glucose metabolism in individuals with severe obesity.

Muscle fiber phenotype: a culprit of abnormal metabolism and function in skeletal muscle of humans with obesity.

The proportion of the different types of fibers in a given skeletal muscle contributes to its overall metabolic and functional characteristics. Greater proportion of type I muscle fibers is associated with favorable oxidative metabolism and function of the muscle. Humans with obesity have a lower proportion of type I muscle fibers. We discuss how lower proportion of type I fibers in skeletal muscle of humans with obesity may explain metabolic and functional abnormalities reported in these individuals. These include lower muscle glucose disposal rate, mitochondrial content, protein synthesis, and quality/contractile function, as well as increased risk for heart disease, lower levels of physical activity, and propensity for weight gain/resistance to weight loss. We delineate future research directions and the need to examine hybrid muscle fiber populations, which are indicative of a transitory state of fiber phenotype within skeletal muscle. We also describe methodologies for precisely characterizing muscle fibers and gene expression at the single muscle fiber level to enhance our understanding of the regulation of muscle fiber phenotype in obesity. By contextualizing research in the field of muscle fiber type in obesity, we lay a foundation for future advancements and pave the way for translation of this knowledge to address impaired metabolism and function in obesity.

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.

Exercise during Pregnancy: Developmental Programming Effects and Future Directions in Humans.

Epidemiological studies show that low birth weight is associated with mortality from cardiovascular disease in adulthood, indicating that chronic diseases could be influenced by hormonal or metabolic insults encountered in utero. This concept, now known as the Developmental Origins of Health and Disease hypothesis, postulates that the intrauterine environment may alter the structure and function of the organs of the fetus as well as the expression of genes that impart an increased vulnerability to chronic diseases later in life. Lifestyle interventions initiated during the prenatal period are crucial as there is the potential to attenuate progression towards chronic diseases. However, how lifestyle interventions such as physical activity directly affect human offspring metabolism and the potential mechanisms involved in regulating metabolic balance at the cellular level are not known. The purpose of this review is to highlight the effects of exercise during pregnancy on offspring metabolic health and emphasize gaps in the current human literature and suggestions for future research.

Comparison of Activity and Participation, and Mental and Physical Functions of Immediate Post-Acute and Distant Post-Acute Adults with Chronic Cardiac Conditions.

This study compared activity participation, and mental and physical functions of two different groups of adults with chronic cardiac conditions. Eleven participants were assessed at the immediate post-acute stage and 26 participants were at the distant post-acute stage. Participants at distant post-acute were significantly more physically active (p < .001), more activity-limited in cognition-related activities (p = .035) and reported more depressive symptoms than immediate post-acute (p = .046). No significant difference in participation level was found. More attention to functional cognition and depressive symptoms at immediate post-acute and individualized approaches to remove participation barriers in complex activities at distant post-acute are likely needed for adults with chronic cardiac conditions.

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.

Roux-en-Y gastric bypass surgery restores insulin-mediated glucose partitioning and mitochondrial dynamics in primary myotubes from severely obese humans.

BACKGROUND/OBJECTIVES: Impaired insulin-mediated glucose partitioning is an intrinsic metabolic defect in skeletal muscle from severely obese humans (BMI ≥ 40 kg/m2). Roux-en-Y gastric bypass (RYGB) surgery has been shown to improve glucose metabolism in severely obese humans. The purpose of the study was to determine the effects of RYGB surgery on glucose partitioning, mitochondrial network morphology, and the markers of mitochondrial dynamics skeletal muscle from severely obese humans. SUBJECT/METHODS: Human skeletal muscle cells were isolated from muscle biopsies obtained from RYGB patients (BMI = 48.0 ± 2.1, n = 7) prior to, 1 month and 7 months following surgery and lean control subjects (BMI = 22.4 ± 1.1, n = 7). Complete glucose oxidation, non-oxidized glycolysis rates, mitochondrial respiratory capacity, mitochondrial network morphology, and the regulatory proteins of mitochondrial dynamics were determined in differentiated human myotubes. RESULTS: Myotubes derived from severely obese humans exhibited enhanced glucose oxidation (13.5%; 95% CI [7.6, 19.4], P = 0.043) and reduced non-oxidized glycolysis (-1.3%; 95% CI [-11.1, 8.6]) in response to insulin stimulation at 7 months after RYGB when compared with the presurgery state (-0.6%; 95% CI [-5.2, 4.0] and 19.5%; 95% CI [4.0, 35.0], P = 0.006), and were not different from the lean controls (16.7%; 95% CI [11.8, 21.5] and 1.9%; 95% CI [-1.6, 5.4], respectively). Further, the number of fragmented mitochondria and Drp1(Ser616) phosphorylation were trended to reduce/reduced (0.0104, 95% CI [0.0085, 0.0126], P = 0.091 and 0.0085, 95% CI [0.0068, 0.0102], P = 0.05) in myotubes derived from severely obese humans at 7 months after RYGB surgery in comparison with the presurgery state. Finally, Drp1(Ser616) phosphorylation was negatively correlated with insulin-stimulated glucose oxidation (r = -0.49, P = 0.037). CONCLUSION/INTERPRETATION: These data indicate that an intrinsic metabolic defect of glucose partitioning in skeletal muscle from severely obese humans is restored by RYGB surgery. The restoration of glucose partitioning may be regulated through reduced mitochondrial fission protein Drp1 phosphorylation.

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.

Electrical pulse stimulation induces differential responses in insulin action in myotubes from severely obese individuals.

KEY POINTS: Exercise/exercise training can enhance insulin sensitivity through adaptations in skeletal muscle, the primary site of insulin-mediated glucose disposal; however, in humans the range of improvement can vary substantially. The purpose of this study was to determine if obesity influences the magnitude of the exercise response in relation to improving insulin sensitivity in human skeletal muscle. Electrical pulse stimulation (EPS; 24 h) of primary human skeletal muscle myotubes improved insulin action in tissue from both lean and severely obese individuals, but responses to EPS were blunted with obesity. EPS improved insulin signal transduction in myotubes from lean but not severely obese subjects and increased AMP accumulation and AMPK Thr172 phosphorylation, but to a lesser degree in myotubes from the severely obese. These data reveal that myotubes of severely obese individuals enhance insulin action and stimulate exercise-responsive molecules with contraction, but in a manner and magnitude that differs from lean subjects. ABSTRACT: Exercise/muscle contraction can enhance whole-body insulin sensitivity; however, in humans the range of improvements can vary substantially. In order, to determine if obesity influences the magnitude of the exercise response, this study compared the effects of electrical pulse stimulation (EPS)-induced contractile activity upon primary myotubes derived from lean and severely obese (BMI ≥ 40 kg/m2 ) women. Prior to muscle contraction, insulin action was compromised in myotubes from the severely obese as was evident from reduced insulin-stimulated glycogen synthesis, glucose oxidation, glucose uptake, insulin signal transduction (IRS1, Akt, TBC1D4), and insulin-stimulated GLUT4 translocation. EPS (24 h) increased AMP, IMP, AMPK Thr172 phosphorylation, PGC1α content, and insulin action in myotubes of both the lean and severely obese subjects. However, despite normalizing indices of insulin action to levels seen in the lean control (non-EPS) condition, responses to EPS were blunted with obesity. EPS improved insulin signal transduction in myotubes from lean but not severely obese subjects and EPS increased AMP accumulation and AMPK Thr172 phosphorylation, but to a lesser degree in myotubes from the severely obese. These data reveal that myotubes of severely obese individuals enhance insulin action and stimulate exercise-responsive molecules with contraction, but in a manner and magnitude that differs from lean subjects.

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.

Roux-en-Y gastric bypass surgery enhances contraction-mediated glucose metabolism in primary human myotubes.

Contractile activity (e.g., exercise) evokes numerous metabolic adaptations in human skeletal muscle, including enhanced insulin action and substrate oxidation. However, there is intersubject variation in the physiological responses to exercise, which may be linked with factors such as the degree of obesity. Roux-en-Y gastric bypass (RYGB) surgery reduces body mass in severely obese (body mass index ≥ 40 kg/m2) individuals; however, it is uncertain whether RYGB can potentiate responses to contractile activity in this potentially exercise-resistant population. To examine possible interactions between RYGB and contractile activity, muscle biopsies were obtained from severely obese patients before and after RYGB, differentiated into myotubes, and electrically stimulated, after which changes in insulin action and glucose oxidation were determined. Before RYGB, myotubes were unresponsive to electrical stimulation, as indicated by no changes in insulin-stimulated glycogen synthesis and basal glucose oxidation. However, myotubes from the same patients at 1 mo after RYGB increased insulin-stimulated glycogen synthesis and basal glucose oxidation when subjected to contraction. While unresponsive before surgery, contraction improved insulin-stimulated phosphorylation of AS160 (Thr642, Ser704) after RYGB. These data suggest that RYGB surgery may enhance the ability of skeletal muscle from severely obese individuals to respond to contractile activity.

Overexpression of PGC-1α increases peroxisomal activity and mitochondrial fatty acid oxidation in human primary myotubes.

Peroxisomes are indispensable organelles for lipid metabolism in humans, and their biogenesis has been assumed to be under regulation by peroxisome proliferator-activated receptors (PPARs). However, recent studies in hepatocytes suggest that the mitochondrial proliferator PGC-1α (peroxisome proliferator-activated receptor gamma coactivator-1α) also acts as an upstream transcriptional regulator for enhancing peroxisomal abundance and associated activity. It is unknown whether the regulatory mechanism(s) for enhancing peroxisomal function is through the same node as mitochondrial biogenesis in human skeletal muscle (HSkM) and whether fatty acid oxidation (FAO) is affected. Primary myotubes from vastus lateralis biopsies from lean donors (BMI = 24.0 ± 0.6 kg/m2; n = 6) were exposed to adenovirus encoding human PGC-1α or GFP control. Peroxisomal biogenesis proteins (peroxins) and genes (PEXs) responsible for proliferation and functions were assessed by Western blotting and real-time qRT-PCR, respectively. [1-14C]palmitic acid and [1-14C]lignoceric acid (exclusive peroxisomal-specific substrate) were used to assess mitochondrial oxidation of peroxisomal-derived metabolites. After overexpression of PGC-1α, 1) peroxisomal membrane protein 70 kDa (PMP70), PEX19, and mitochondrial citrate synthetase protein content were significantly elevated (P < 0.05), 2) PGC-1α, PMP70, key PEXs, and peroxisomal ß-oxidation mRNA expression levels were significantly upregulated (P < 0.05), and 3) a concomitant increase in lignoceric acid oxidation by both peroxisomal and mitochondrial activity was observed (P < 0.05). These novel findings demonstrate that, in addition to the proliferative effect on mitochondria, PGC-1α can induce peroxisomal activity and accompanying elevations in long-chain and very-long-chain fatty acid oxidation by a peroxisomal-mitochondrial functional cooperation, as observed in HSkM cells.

Metabolic Inflexibility with Obesity and the Effects of Fenofibrate on Skeletal Muscle Fatty Acid Oxidation.

This study was designed to investigate mechanisms of lipid metabolic inflexibility in human obesity and the ability of fenofibrate (FENO) to increase skeletal muscle fatty acid oxidation (FAO) in primary human skeletal muscle cell cultures (HSkMC) exhibiting metabolic inflexibility. HSkMC from 10 lean and 10 obese, insulin resistant subjects were treated with excess fatty acid for 24 h (24hFA) to gauge lipid-related metabolic flexibility. Metabolically inflexible HSkMC from obese individuals were then treated with 24hFA in combination with FENO to determine effectiveness for increasing FAO. Mitochondrial enzyme activity and FAO were measured in skeletal muscle from subjects with prediabetes (n=11) before and after 10 weeks of fenofibrate in vivo. 24hFA increased FAO to a greater extent in HSkMC from lean versus obese subjects (+49% vs. +9%, for lean vs. obese, respectively; p<0.05) indicating metabolic inflexibility with obesity. Metabolic inflexibility was not observed for measures of cellular respiration in permeabilized cells using carbohydrate substrate. Fenofibrate co-incubation with 24hFA, increased FAO in a subset of HSkMC from metabolically inflexible, obese subjects (p<0.05), which was eliminated by PPARα antagonist. In vivo, fenofibrate treatment increased skeletal muscle FAO in a subset of subjects with prediabetes but did not affect gene transcription or mitochondrial enzyme activity. Lipid metabolic inflexibility observed in HSkMC from obese subjects is not due to differences in electron transport flux, but rather upstream decrements in lipid metabolism. Fenofibrate increases the capacity for FAO in human skeletal muscle cells, though its role in skeletal muscle metabolism in vivo remains unclear.

Age-related impairments in skeletal muscle PDH phosphorylation and plasma lactate are indicative of metabolic inflexibility and the effects of exercise training.

The purpose of this study was to determine whether plasma lactate and skeletal muscle glucose regulatory pathways, specifically PDH dephosphorylation, are impaired during hyperinsulinemic conditions in middle- to older-aged individuals and determine whether exercise training could improve key variables responsible for skeletal muscle PDH regulation. Eighteen young (19-29 yr; n = 9 males and 9 females) and 20 middle- to older-aged (57-82 yr; n = 10 males and 10 females) individuals underwent a 2-h euglycemic hyperinsulinemic clamp. Plasma samples were obtained at baseline and at 30, 50, 90, and 120 min for analysis of lactate, and skeletal muscle biopsies were performed at 60 min for analysis of protein associated with glucose metabolism. In response to insulin, plasma lactate was elevated in aged individuals when normalized to insulin action. Insulin-stimulated phosphorylation of skeletal muscle PDH on serine sites 232, 293, and 300 decreased in young individuals only. Changes in insulin-stimulated PDH phosphorylation were positively related to changes in plasma lactate. No age-related differences were observed in skeletal muscle phosphorylation of LDH, GSK-3α, or GSK-3ß in response to insulin or PDP1, PDP2, PDK2, PDK4, or MPC1 total protein. Twelve weeks of endurance- or strength-oriented exercise training improved insulin-stimulated PDH dephosphorylation, which was related to a reduced lactate response. These findings suggest that impairments in insulin-induced PDH regulation in a sedentary aging population contribute to impaired glucose metabolism and that exercise training is an effective intervention for treating metabolic inflexibility.

Plasma acylcarnitines during insulin stimulation in humans are reflective of age-related metabolic dysfunction.

The purpose of this study was to determine if plasma acylcarnitine (AC) profiling is altered under hyperinsulinemic conditions as part of the aging process. Fifteen young, lean (19-29 years) and fifteen middle-to older-aged (57-82 years) individuals underwent a 2-hr euglycemic-hyperinsulinemic clamp. Plasma samples were obtained at baseline, 20 min, 50 min, and 120 min for analysis of AC species and amino acids. Skeletal muscle biopsies were performed after 60 min of insulin-stimulation for analysis of acetyl-CoA carboxylase (ACC) phosphorylation. Insulin infusion decreased the majority of plasma short-, medium-, and long-chain (SC, MC, and LC, respectively) AC. However, during the initial 50 min, a number of MC and LC AC species (C10, C10:1, C12:1, C14, C16, C16:1, C18) remained elevated in aged individuals compared to their younger counterparts indicating a lag in responsiveness. Additionally, the insulin-induced decline in skeletal muscle ACC phosphorylation was blunted in the aged compared to young individuals (-24% vs. -56%, P < 0.05). These data suggest that a desensitization to insulin during aging, possibly at the level of skeletal muscle ACC phosphorylation, results in a diminished ability to transition to glucose oxidation indicative of metabolic inflexibility.

Examination of Lifestyle Behaviors and Cardiometabolic Risk Factors in University Students Enrolled in Kinesiology Degree Programs.

Preventing physical inactivity and weight gain during college is critical in decreasing lifelong obesity and associated disease risk. As such, we sought to compare cardiometabolic risk factors and lifestyle behaviors between college students enrolled in kinesiology and non-kinesiology degree programs to assess whether health and exercise degree programs may influence health behaviors and associated disease risk outcomes. Anthropometrics, fasting blood glucose, insulin, lipid profiles and HbA1c%, blood pressure, and peak oxygen consumption (V[Combining Dot Above]O2peak) were assessed in 247 healthy college students. The homeostasis model assessment of insulin sensitivity (HOMA) was calculated using glucose and insulin levels. Self-reported physical activity from the Paffenbarger questionnaire was collected to estimate the average caloric expenditure due to different types of physical activities. Despite no significant differences in body mass index or waist circumference between groups, kinesiology majors presented with ∼20% lower fasting insulin levels and HOMA (p = 0.01; p < 0.01, respectively) relative to nonmajors. Kinesiology majors reported increased weekly participation in vigorous-intensity sport and leisure activities and, on average, engaged in >300 metabolic equivalent-h·wk, whereas non-kinesiology majors engaged in <300 MET-h wk (p = 0.01). Our data suggest that students enrolled in kinesiology degree programs display improved healthy behaviors and associated outcomes (parameters of glucose homeostasis). Practical outcomes of this research indicate that implementing components of a comprehensive kinesiology curriculum encourages improved health behaviors and associated cardiometabolic risk factors.

Lipid exposure elicits differential responses in gene expression and DNA methylation in primary human skeletal muscle cells from severely obese women.

The skeletal muscle of obese individuals exhibits an impaired ability to increase the expression of genes linked with fatty acid oxidation (FAO) upon lipid exposure. The present study determined if this response could be attributed to differential DNA methylation signatures. RNA and DNA were isolated from primary human skeletal muscle cells (HSkMC) from lean and severely obese women following lipid incubation. mRNA expression and DNA methylation were quantified for genes that globally regulate FAO [PPARγ coactivator (PGC-1α), peroxisome proliferator-activated receptors (PPARs), nuclear respiratory factors (NRFs)]. With lipid oversupply, increases in NRF-1, NRF-2, PPARα, and PPARδ expression were dampened in skeletal muscle from severely obese compared with lean women. The expression of genes downstream of the PPARs and NRFs also exhibited a pattern of not increasing as robustly upon lipid exposure with obesity. Increases in CpG methylation near the transcription start site with lipid oversupply were positively related to PPARδ expression; increases in methylation with lipid were depressed in HSkMC from severely obese women. With severe obesity, there is an impaired ability to upregulate global transcriptional regulators of FAO in response to lipid exposure. Transient changes in DNA methylation patterns and differences in the methylation signature with severe obesity may play a role in the transcriptional regulation of PPARδ in response to lipid. The persistence of differential responses to lipid in HSkMC derived from lean and obese subjects supports the possibility of stable epigenetic programming of skeletal muscle cells by the respective environments.

Differential epigenetic and transcriptional response of the skeletal muscle carnitine palmitoyltransferase 1B (CPT1B) gene to lipid exposure with obesity.

The ability to increase fatty acid oxidation (FAO) in response to dietary lipid is impaired in the skeletal muscle of obese individuals, which is associated with a failure to coordinately upregulate genes involved with FAO. While the molecular mechanisms contributing to this metabolic inflexibility are not evident, a possible candidate is carnitine palmitoyltransferase-1B (CPT1B), which is a rate-limiting step in FAO. The present study was undertaken to determine if the differential response of skeletal muscle CPT1B gene transcription to lipid between lean and severely obese subjects is linked to epigenetic modifications (DNA methylation and histone acetylation) that impact transcriptional activation. In primary human skeletal muscle cultures the expression of CPT1B was blunted in severely obese women compared with their lean counterparts in response to lipid, which was accompanied by changes in CpG methylation, H3/H4 histone acetylation, and peroxisome proliferator-activated receptor-δ and hepatocyte nuclear factor 4α transcription factor occupancy at the CPT1B promoter. Methylation of specific CpG sites in the CPT1B promoter that correlated with CPT1B transcript level blocked the binding of the transcription factor upstream stimulatory factor, suggesting a potential causal mechanism. These findings indicate that epigenetic modifications may play important roles in the regulation of CPT1B in response to a physiologically relevant lipid mixture in human skeletal muscle, a major site of fatty acid catabolism, and that differential DNA methylation may underlie the depressed expression of CPT1B in response to lipid, contributing to the metabolic inflexibility associated with severe obesity.

SMAD3 augments FoxO3-induced MuRF-1 promoter activity in a DNA-binding-dependent manner.

Muscle-specific RING finger-1 (MuRF-1), a ubiquitin ligase and key regulator of proteasome-dependent protein degradation, is highly expressed during skeletal muscle atrophy. The transcription factor forkhead box O3 (FoxO3) induces MuRF-1 expression, but the direct role of other major atrophy-related transcription factors, such as SMAD3, is largely unknown. The goal of this study was to determine whether SMAD3 individually regulates, or with FoxO3 coordinately regulates, MuRF-1 expression. In cultured myotubes or human embryonic kidney cells, MuRF-1 mRNA content and promoter activity were increased by FoxO3 but not by SMAD3 overexpression. However, FoxO3 and SMAD3 coexpression synergistically increased MuRF-1 mRNA and promoter activity. Mutation of the SMAD-binding element (SBE) in the proximal MuRF-1 promoter or overexpression of a SMAD3 DNA-binding mutant attenuated FoxO3-dependent MuRF-1 promoter activation, showing that SMAD binding to DNA is required for optimal activation of FoxO3-induced transcription of MuRF-1. Using chromatin immunoprecipitation, SMAD3 DNA binding increased FoxO3 abundance and SBE mutation reduced FoxO3 abundance on the MuRF-1 promoter. Furthermore, SMAD3 overexpression dose-dependently increased FoxO3 protein content, and coexpression of FoxO3 and SMAD3 synergistically increased FoxO-dependent gene transcription [assessed with a FoxO response element (FRE)-driven reporter]. Collectively, these results show that SMAD3 regulates transcription of MuRF-1 by increasing FoxO3 binding at a conserved FRE-SBE motif within the proximal promoter region, and by increasing FoxO3 protein content and transcriptional activity. These data are the first to indicate that two major transcription factors regulating protein degradation, FoxO3 and SMAD3, converge to coordinately and directly regulate transcription of MuRF-1.