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Context: Humans with obesity and insulin resistance exhibit lipid accumulation in skeletal muscle, but the underlying biological mechanisms responsible for the accumulation of lipid in the muscle of these individuals remain unknown. Objective: We investigated how plasma insulin modulates the extraction of circulating triglycerides (TGs) and non-esterified fatty acids (NEFAs) from dietary and endogenous sources in the muscle of lean, insulin-sensitive humans (Lean-IS) and contrasted these responses to those in humans with obesity and insulin resistance (Obese-IR). Methods: The studies were performed in a postprandial state associated with steady-state plasma TG concentrations. The arterio-venous blood sampling technique was employed to determine the extraction of circulating lipids across the forearm muscle before and after insulin infusion. We distinguished kinetics of TGs and NEFAs from dietary sources across muscle from those from endogenous sources by incorporating stable isotope-labeled triolein in ingested fat. Results: Plasma insulin rapidly suppressed the extraction of plasma TGs from endogenous, but not dietary, sources in the Lean-IS, but same response was absent in the Obese-IR. Furthermore, in the muscle of Lean-IS, plasma insulin decreased the extraction of circulating NEFAs from both dietary and endogenous sources, but in Obese-IR subjects this response was absent for NEFAs from dietary sources. Conclusions: Partitioning of circulating lipids away from the skeletal muscle when plasma insulin increases, such as during the postprandial period, is impaired in humans with obesity and insulin resistance. Trial Registration: ClinicalTrials.gov ( NCT01860911 ).
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OBJECTIVE: This study tested the hypothesis that expression of insulin-like growth factor 1 (IGF-1) protein and mRNA splice variants is lower in skeletal muscle of humans with obesity who have a lower mixed-muscle protein fractional synthesis rate (MMP-FSR) when compared with individuals without obesity. METHODS: The study included nine participants with obesity (OB, mean [SD], BMI = 35 [3] kg/m2 , MMP-FSR = 0.06%/h [0.02%/h]) and nine participants without obesity (W-OB, BMI = 24 [3] kg/m2 , MMP-FSR = 0.08%/h [0.02%/h]; for both BMI and MMP-FSR p < 0.05). MMP-FSR and mitochondrial protein FSR were measured following an overnight fast. RESULTS: Along with lower MMP-FSR, OB participants displayed lower mitochondrial protein FSR (p = 0.03) compared with W-OB participants. Expression of IGF-1 (p = 0.04) and IGF-1 receptor (p < 0.01) proteins was lower in muscle of OB participants. In addition, OB participants had lower (p < 0.05) mRNA expression of IGF1 variants Eb and Ec. This study demonstrates that lower protein synthesis in muscle of humans with obesity occurs concurrently with lower expression of muscle IGF-1 and IGF-1 receptor proteins, as well as lower mRNA expression of the IGF1 splice variants. CONCLUSIONS: These findings indicate that lower protein synthesis observed in muscle of humans with obesity may result from diminished muscle IGF1 gene expression.
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Factor I del Crecimiento Similar a la Insulina , Proteínas Musculares , Humanos , Factor I del Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/metabolismo , Receptor IGF Tipo 1/genética , Receptor IGF Tipo 1/metabolismo , Músculo Esquelético/metabolismo , Obesidad/genética , Obesidad/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Mitocondriales/metabolismoRESUMEN
Obesity negatively impacts skeletal muscle protein metabolism, and also impairs skeletal muscle maintenance and regeneration. We analyzed muscle biopsy samples from humans with increased body mass index (BMI) (i.e. > 30 kg/m2) and controls (i.e., BMI < 25 kg/m2) for expression of syncytin-1, a fusogenic protein regulating skeletal muscle regeneration. When compared to controls, humans with increased BMI and concomitant reduction in muscle protein synthesis had higher expression of syncytin-1 in skeletal muscle (p < 0.05). Across human subjects, muscle protein synthesis correlated inversely (r = -0.51; p = 0.03) with syncytin-1 expression in muscle. Using a C2C12 cell line we found that expression of syncytin-A (i.e, corresponding protein in murine tissue) is increased by insulin, and that this response is impaired in the presence of fatty acids, whose metabolism is altered within the metabolic environment induced by increased BMI. In C2C12 cells, the response of the protein 4E-BP1, which signals increase in protein synthesis in muscle, resembled that of syncytin-A. These findings provide novel insights into the expression of syncytin-1 in skeletal muscle of humans with increased BMI, as well as its basic regulation by insulin and fatty acids in muscle. The findings signify the need for further research into the regulation of syncytin-1 in skeletal muscle of humans with increased BMI, as well as its biological implications for altering muscle protein metabolism and regeneration.
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Skeletal muscle is highly plastic and dynamically regulated by the body's physical demands. This study aimed to determine the plasticity of skeletal muscle DNA methylation in response to 8 weeks of supervised exercise training in volunteers with a range of insulin sensitivities. We studied 13 sedentary participants and performed euglycemic hyperinsulinemic clamps with basal vastus lateralis muscle biopsies and peak aerobic activity (VO2 peak) tests before and after training. We extracted DNA from the muscle biopsies and performed global methylation using Illumina's Methylation EPIC 850K BeadChip. Training significantly increased peak aerobic capacity and insulin-stimulated glucose disposal. Fasting serum insulin and insulin levels during the steady state of the clamp were significantly lower post-training. Insulin clearance rates during the clamp increased following the training. We identified 13 increased and 90 decreased differentially methylated cytosines (DMCs) in response to 8 weeks of training. Of the 13 increased DMCs, 2 were within the following genes, FSTL3, and RP11-624M8.1. Of the 90 decreased DMCs, 9 were within the genes CNGA1, FCGR2A, KIF21A, MEIS1, NT5DC1, OR4D1, PRPF4B, SLC26A7, and ZNF280C. Moreover, pathway analysis showed an enrichment in metabolic and actin-cytoskeleton pathways for the decreased DMCs, and for the increased DMCs, an enrichment in signal-dependent regulation of myogenesis, NOTCH2 activation and transmission, and SMAD2/3: SMAD4 transcriptional activity pathways. Our findings showed that 8 weeks of exercise training alters skeletal muscle DNA methylation of specific genes and pathways in people with varying degrees of insulin sensitivity.
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BACKGROUND: The mechanisms of weight loss and metabolic improvements following bariatric surgery in skeletal muscle are not well known; however, epigenetic modifications are likely to contribute. The aim of our study was to investigate skeletal muscle DNA methylation after weight loss induced by Roux-en-Y gastric bypass (RYGB) surgery. Muscle biopsies were obtained basally from seven insulin-resistant obese (BMI > 40 kg/m2) female subjects (45.1 ± 3.6 years) pre- and 3-month post-surgery with euglycemic hyperinsulinemic clamps to assess insulin sensitivity. Four lean (BMI < 25 kg/m2) females (38.5 ± 5.8 years) served as controls. We performed reduced representation bisulfite sequencing next generation methylation on DNA isolated from the vastus lateralis muscle biopsies. RESULTS: Global methylation was significantly higher in the pre- (32.97 ± 0.02%) and post-surgery (33.31 ± 0.02%) compared to the lean (30.46 ± 0.02%), P < 0.05. MethylSig analysis identified 117 differentially methylated cytosines (DMCs) that were significantly altered in the post- versus pre-surgery (Benjamini-Hochberg q < 0.05). In addition, 2978 DMCs were significantly altered in the pre-surgery obese versus the lean controls (Benjamini-Hochberg q < 0.05). For the post-surgery obese versus the lean controls, 2885 DMCs were altered (Benjamini-Hochberg q < 0.05). Seven post-surgery obese DMCs were normalized to levels similar to those observed in lean controls. Of these, 5 were within intergenic regions (chr11.68,968,018, chr16.73,100,688, chr5.174,115,531, chr5.1,831,958 and chr9.98,547,011) and the remaining two DMCs chr17.45,330,989 and chr14.105,353,824 were within in the integrin beta 3 (ITGB3) promoter and KIAA0284 exon, respectively. ITGB3 methylation was significantly decreased in the post-surgery (0.5 ± 0.5%) and lean controls (0 ± 0%) versus pre-surgery (13.6 ± 2.7%, P < 0.05). This decreased methylation post-surgery was associated with an increase in ITGB3 gene expression (fold change + 1.52, P = 0.0087). In addition, we showed that ITGB3 promoter methylation in vitro significantly suppressed transcriptional activity (P < 0.05). Transcription factor binding analysis for ITGB3 chr17.45,330,989 identified three putative transcription factor binding motifs; PAX-5, p53 and AP-2alphaA. CONCLUSIONS: These results demonstrate that weight loss after RYGB alters the epigenome through DNA methylation. In particular, this study highlights ITGB3 as a novel gene that may contribute to the metabolic improvements observed post-surgery. Future additional studies are warranted to address the exact mechanism of ITGB3 in skeletal muscle.
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Metilación de ADN/genética , Epigénesis Genética/genética , Derivación Gástrica/métodos , Músculo Esquelético/metabolismo , Obesidad/cirugía , Pérdida de Peso/genética , Femenino , Humanos , Persona de Mediana Edad , Obesidad/genéticaRESUMEN
Mitochondria from skeletal muscle of humans with obesity often display alterations with respect to their morphology, proteome, biogenesis, and function. These changes in muscle mitochondria are considered to contribute to metabolic abnormalities observed in humans with obesity. Most of the evidence describing alterations in muscle mitochondria in humans with obesity, however, lacks reference to a specific subcellular location. This is despite data over the years showing differences in the morphology and function of subsarcolemmal (found near the plasma membrane) and intermyofibrillar (nested between the myofibrils) mitochondria in skeletal muscle. Recent studies reveal that impairments in mitochondrial function in obesity with respect to the subcellular location of the mitochondria in muscle are more readily evident following exposure of the skeletal muscle to physiological stimuli. In this review, we highlight the need to understand skeletal muscle mitochondria metabolism in obesity in a subpopulation-specific manner and in the presence of physiological stimuli that modify mitochondrial function in vivo. Experimental approaches employed under these conditions will allow for more precise characterization of impairments in skeletal muscle mitochondria and their implications in inducing metabolic dysfunction in human obesity.
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Ejercicio Físico/fisiología , Mitocondrias Musculares/metabolismo , Músculo Esquelético/metabolismo , Fenómenos Fisiológicos de la Nutrición , Obesidad/metabolismo , Animales , HumanosRESUMEN
INTRODUCTION: Current evidence indicates mitochondrial dysfunction in humans with obesity. Acute exercise appears to enhance mitochondrial function in the muscle of nonobese humans, but its effects on mitochondrial function in muscle of humans with obesity are not known. We sought to determine whether acute aerobic exercise stimulates mitochondrial function in subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondria in humans with obesity. METHODS: We assessed maximal adenosine triphosphate production rate (MAPR) and citrate synthase (CS) activity in isolated SS and IMF mitochondria from subjects with body mass index < 27 kg·m (median age, 25 yr; interquartile range, 22-39 yr) and subjects with body mass index > 32 kg·m (median age, 29 yr; interquartile range, 20-39 yr) before and 3 h after a 45-min cycling exercise at an intensity corresponding to 65% HR reserve. The SS and IMF mitochondria were isolated from muscle biopsies using differential centrifugation. Maximal adenosine triphosphate production rate and CS activities were determined using luciferase-based and spectrophotometric enzyme-based assays, respectively. RESULTS: Exercise increased MAPR in IMF mitochondria in both nonobese subjects and subjects with obesity (P < 0.05), but CS-specific activity did not change in either group (P > 0.05). Exercise increased MAPR supported by complex II in SS mitochondria, in both groups (P < 0.05), but MAPR supported by complex I or palmitate did not increase by exercise in the subjects with obesity (P > 0.05). Citrate synthase-specific activity increased in SS mitochondria in response to exercise only in nonobese subjects (P < 0.05). CONCLUSIONS: In nonobese humans, acute aerobic exercise increases MAPR in both SS and IMF mitochondria. In humans with obesity, the exercise increases MAPR in IMF mitochondria, but this response is less evident in SS mitochondria.
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Adenosina Trifosfato/biosíntesis , Ejercicio Físico , Mitocondrias Musculares/metabolismo , Obesidad/metabolismo , Adulto , Glucemia/análisis , Citrato (si)-Sintasa/metabolismo , Femenino , Humanos , Resistencia a la Insulina , Masculino , Músculo Esquelético/metabolismo , Adulto JovenRESUMEN
BACKGROUND: Skeletal muscle mitochondrial content and function appear to be altered in obesity. Mitochondria in muscle are found in well-defined regions within cells, and they are arranged in a way that form distinct subpopulations of subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondria. We sought to investigate differences in the proteomes of SS and IMF mitochondria between lean subjects and subjects with obesity. METHODS: We performed comparative proteomic analyses on SS and IMF mitochondria isolated from muscle samples obtained from lean subjects and subjects with obesity. Mitochondria were isolated using differential centrifugation, and proteins were subjected to label-free quantitative tandem mass spectrometry analyses. Collected data were evaluated for abundance of mitochondrial proteins using spectral counting. The Reactome pathway database was used to determine metabolic pathways that are altered in obesity. RESULTS: Among proteins, 73 and 41 proteins showed different (mostly lower) expression in subjects with obesity in the SS and IMF mitochondria, respectively (false discovery rate-adjusted Pâ¯≤â¯0.05). We specifically found an increase in proteins forming the tricarboxylic acid cycle and electron transport chain (ETC) complex II, but a decrease in proteins forming protein complexes I and III of the ETC and adenosine triphosphate (ATP) synthase in subjects with obesity in the IMF, but not SS, mitochondria. Obesity was associated with differential effects on metabolic pathways linked to protein translation in the SS mitochondria and ATP formation in the IMF mitochondria. CONCLUSIONS: Obesity alters the expression of mitochondrial proteins regulating key metabolic processes in skeletal muscle, and these effects are distinct to mitochondrial subpopulations located in different regions of the muscle fibers. TRIAL REGISTRATION: ClinicalTrials.gov (NCT01824173).
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Mitocondrias Musculares/ultraestructura , Proteínas Mitocondriales/metabolismo , Obesidad/metabolismo , Complejos de ATP Sintetasa/metabolismo , Adulto , Femenino , Voluntarios Sanos , Humanos , Masculino , Redes y Vías Metabólicas , Mitocondrias Musculares/metabolismo , Mitocondrias Musculares/patología , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/ultraestructura , Obesidad/patología , Proteómica , Sarcolema/metabolismo , Sarcolema/ultraestructura , Fracciones Subcelulares/metabolismo , Fracciones Subcelulares/ultraestructura , Espectrometría de Masas en TándemRESUMEN
Context: Obesity is associated with mitochondrial dysfunction in skeletal muscle. Increasing the plasma amino acid (AA) concentrations stimulates mitochondrial adenosine triphosphate (ATP) production in lean individuals. Objective: To determine whether acute elevation in plasma AAs enhances muscle mitochondrial respiration and ATP production in subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondria in obese adults. Design: Assessment of SS and IMF mitochondrial function during saline (i.e., control) and AA infusions. Participants: Eligible participants were healthy lean (body mass index, <25 kg/m2; age, 37 ± 3 years; n = 10) and obese (body mass index >30 kg/m2; age 35 ± 3 years; n = 11) subjects. Intervention: Single trial of saline infusion followed by AA infusion. SS and IMF mitochondria were isolated from muscle biopsies collected at the end of the saline and AA infusions. Main Outcomes: Mitochondrial respiration and ATP production. Results: AA infusion increased adenosine 5'-diphosphate (ADP)-stimulated respiration and ATP production rates of SS mitochondria in the lean (P < 0.05), but not obese, subjects. Furthermore, AA infusion increased the uncoupled (i.e., non-ADP-stimulated) respiration of SS mitochondria in the lean subjects only (P < 0.05). AA infusion had no effect on any of these parameters in IMF mitochondria in either lean or obese subjects (P > 0.05). Conclusions: Increasing the plasma AA concentrations enhances the capacity for respiration and ATP production of muscle SS, but not IMF, mitochondria in lean individuals, in parallel with increases in uncoupled respiration. However, neither of these parameters increases in muscle SS or IMF mitochondria in obese individuals.
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Aminoácidos/sangre , Aminoácidos/farmacología , Mitocondrias Musculares/metabolismo , Obesidad/metabolismo , Consumo de Oxígeno/efectos de los fármacos , Sarcolema/metabolismo , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/biosíntesis , Adulto , Glucemia/metabolismo , Índice de Masa Corporal , Femenino , Hormonas/sangre , Humanos , Masculino , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/metabolismo , Miofibrillas/metabolismo , Adulto JovenRESUMEN
BACKGROUND: Obesity is a disease that is caused by genetic and environmental factors. However, epigenetic mechanisms of obesity are less well known. DNA methylation provides a mechanism whereby environmental factors can influence gene transcription. The aim of our study was to investigate skeletal muscle DNA methylation of sorbin and SH3 domain containing 3 (SORBS3) with weight loss induced by Roux-en-Y gastric bypass (RYGB). RESULTS: Previously, we had shown increased methylation (5.0 to 24.4%) and decreased gene expression (fold change - 1.9) of SORBS3 with obesity (BMI > 30 kg/m2) compared to lean controls. In the present study, basal muscle biopsies were obtained from seven morbidly obese (BMI > 40 kg/m2) female subjects pre- and 3 months post-RYGB surgery, in combination with euglycemic-hyperinsulinemic clamps to assess insulin sensitivity. We identified 30 significantly altered promoter and untranslated region methylation sites in SORBS3 using reduced representation bisulfite sequencing (RRBS). Twenty-nine of these sites were decreased (- 5.6 to - 24.2%) post-RYGB compared to pre-RYGB. We confirmed the methylation in 2 (Chr.8:22,423,690 and Chr.8:22,423,702) of the 29 decreased SORBS3 sites using pyrosequencing. This decreased methylation was associated with an increase in SORBS3 gene expression (fold change + 1.7) post-surgery. In addition, we demonstrated that SORBS3 promoter methylation in vitro significantly alters reporter gene expression (P < 0.0001). Two of the SORBS3 methylation sites (Chr.8:22,423,111 and Chr.8:22,423,205) were strongly correlated with fasting plasma glucose levels (r = 0.9, P = 0.00009 and r = 0.8, P = 0.0010). Changes in SORBS3 gene expression post-surgery were correlated with obesity measures and fasting insulin levels (r = 0.5 to 0.8; P < 0.05). CONCLUSIONS: These results demonstrate that SORBS3 methylation and gene expression are altered in obesity and restored to normal levels through weight loss induced by RYGB surgery.
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Proteínas Adaptadoras Transductoras de Señales/genética , Derivación Gástrica/métodos , Músculo Esquelético/química , Obesidad Mórbida/cirugía , Adulto , Biopsia , Metilación de ADN , Epigénesis Genética , Femenino , Regulación de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Proteínas Musculares , Músculo Esquelético/patología , Obesidad Mórbida/genética , Análisis de Secuencia de ADN , Resultado del TratamientoRESUMEN
Obesity can increase the risk of complex metabolic diseases, including insulin resistance. Moreover, obesity can be caused by environmental and genetic factors. However, the epigenetic mechanisms of obesity are not well defined. Therefore, the identification of novel epigenetic biomarkers of obesity allows for a more complete understanding of the disease and its underlying insulin resistance. The aim of our study was to identify DNA methylation changes in whole-blood that were strongly associated with obesity and insulin resistance. Whole-blood was obtained from lean (n = 10; BMI = 23.6 ± 0.7 kg/m2) and obese (n = 10; BMI = 34.4 ± 1.3 kg/m2) participants in combination with euglycemic hyperinsulinemic clamps to assess insulin sensitivity. We performed reduced representation bisulfite sequencing on genomic DNA isolated from the blood. We identified 49 differentially methylated cytosines (DMCs; q < 0.05) that were altered in obese compared with lean participants. We identified 2 sites (Chr.21:46,957,981 and Chr.21:46,957,915) in the 5' untranslated region of solute carrier family 19 member 1 (SLC19A1) with decreased methylation in obese participants (lean 0.73 ± 0.11 vs. obese 0.09 ± 0.05; lean 0.68 ± 0.10 vs. obese 0.09 ± 0.05, respectively). These 2 DMCs identified by obesity were also significantly predicted by insulin sensitivity (r = 0.68, P = 0.003; r = 0.66; P = 0.004). In addition, we performed a differentially methylated region (DMR) analysis and demonstrated a decrease in methylation of Chr.21:46,957,915-46,958,001 in SLC19A1 of -34.9% (70.4% lean vs. 35.5% obese). The decrease in whole-blood SLC19A1 methylation in our obese participants was similar to the change observed in skeletal muscle (Chr.21:46,957,981, lean 0.70 ± 0.09 vs. obese 0.31 ± 0.11 and Chr.21:46,957,915, lean 0.72 ± 0.11 vs. obese 0.31 ± 0.13). Pyrosequencing analysis further demonstrated a decrease in methylation at Chr.21:46,957,915 in both whole-blood (lean 0.71 ± 0.10 vs. obese 0.18 ± 0.06) and skeletal muscle (lean 0.71 ± 0.10 vs. obese 0.30 ± 0.11). Our findings demonstrate a new potential epigenetic biomarker, SLC19A1, for obesity and its underlying insulin resistance.
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Biomarcadores/sangre , Epigénesis Genética , Resistencia a la Insulina , Obesidad/genética , Adulto , Femenino , Humanos , Masculino , Obesidad/metabolismoRESUMEN
PURPOSE OF REVIEW: The purpose of this review is to provide an update of recent additions to our understanding of the prevalence of nutrient deficiencies and the potential role of preoperative weight loss in contributing to these deficiencies in obese individuals planning to undergo bariatric surgery. RECENT FINDINGS: Recent reports that have included bariatric surgery candidates from sites around the world have shown consistent deficiencies in a variety of nutrients. Although protein-energy malnutrition is uncommon preoperatively, micronutrient deficiencies occur commonly with multiple deficiencies often present in the same individual. No difference in the prevalence of deficiency between men and women is apparent, and a standard profile of susceptibility to deficiency has not been identified. In the only studies that have evaluated dietary intake of total energy, macronutrients and micronutrients preoperatively, despite an excess of calories ingested, micronutrient intake tends to be lower than recommended. SUMMARY: A high prevalence of micronutrient deficiencies, especially vitamin D, folate, B12 and iron, is present in obese individuals being considered for bariatric surgery. Despite high-caloric intake, the deficiencies present appear to be related to the poor quality of the diet and low micronutrient intake. These findings strengthen prior recommendations of routine preoperative nutritional screening. Because a standard profile of susceptibility to deficiency has not been identified, extensive nutritional screening, including micronutrient testing, should be considered in all patients in the preoperative setting. Finally, we recommend early supplementation of vitamins and minerals based on laboratory assessment and incorporation of a program to optimize eating behaviors prior to surgery.
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Cirugía Bariátrica , Enfermedades Carenciales/etiología , Micronutrientes/deficiencia , Obesidad/complicaciones , Enfermedades Carenciales/epidemiología , Femenino , Humanos , Masculino , Estado Nutricional , Obesidad/cirugía , Periodo Preoperatorio , PrevalenciaRESUMEN
Our previous studies show reduced abundance of the ß-subunit of mitochondrial H+-ATP synthase (ß-F1-ATPase) in skeletal muscle of obese individuals. The ß-F1-ATPase forms the catalytic core of the ATP synthase, and it is critical for ATP production in muscle. The mechanism(s) impairing ß-F1-ATPase metabolism in obesity, however, are not completely understood. First, we studied total muscle protein synthesis and the translation efficiency of ß-F1-ATPase in obese (BMI, 36±1 kg/m2) and lean (BMI, 22±1 kg/m2) subjects. Both total protein synthesis (0.044±0.006 vs 0.066±0.006%·h-1) and translation efficiency of ß-F1-ATPase (0.0031±0.0007 vs 0.0073±0.0004) were lower in muscle from the obese subjects when compared to the lean controls (P<0.05). We then evaluated these same responses in a primary cell culture model, and tested the specific hypothesis that circulating non-esterified fatty acids (NEFA) in obesity play a role in the responses observed in humans. The findings on total protein synthesis and translation efficiency of ß-F1-ATPase in primary myotubes cultured from a lean subject, and after exposure to NEFA extracted from serum of an obese subject, were similar to those obtained in humans. Among candidate microRNAs (i.e., non-coding RNAs regulating gene expression), we identified miR-127-5p in preventing the production of ß-F1-ATPase. Muscle expression of miR-127-5p negatively correlated with ß-F1-ATPase protein translation efficiency in humans (r = - 0.6744; P<0.01), and could be modeled in vitro by prolonged exposure of primary myotubes derived from the lean subject to NEFA extracted from the obese subject. On the other hand, locked nucleic acid inhibitor synthesized to target miR-127-5p significantly increased ß-F1-ATPase translation efficiency in myotubes (0.6±0.1 vs 1.3±0.3, in control vs exposure to 50 nM inhibitor; P<0.05). Our experiments implicate circulating NEFA in obesity in suppressing muscle protein metabolism, and establish impaired ß-F1-ATPase translation as an important consequence of obesity.
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Ácidos Grasos no Esterificados/metabolismo , ATPasas de Translocación de Protón Mitocondriales/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Obesidad/metabolismo , Adulto , Células Cultivadas , Grasas de la Dieta/administración & dosificación , Epigénesis Genética , Ácidos Grasos no Esterificados/sangre , Femenino , Humanos , Masculino , MicroARNs/genética , MicroARNs/metabolismo , Mitocondrias Musculares/enzimología , ATPasas de Translocación de Protón Mitocondriales/genética , Desarrollo de Músculos/genética , Fibras Musculares Esqueléticas/enzimología , Proteínas Musculares/biosíntesis , Proteínas Musculares/genética , Proteína MioD/genética , Miogenina/genética , Obesidad/sangre , Obesidad/genética , Delgadez/sangre , Delgadez/genética , Delgadez/metabolismoRESUMEN
BACKGROUND: Obesity is a metabolic disease caused by environmental and genetic factors. However, the epigenetic mechanisms of obesity are incompletely understood. The aim of our study was to investigate the role of skeletal muscle DNA methylation in combination with transcriptomic changes in obesity. RESULTS: Muscle biopsies were obtained basally from lean (n = 12; BMI = 23.4 ± 0.7 kg/m(2)) and obese (n = 10; BMI = 32.9 ± 0.7 kg/m(2)) participants in combination with euglycemic-hyperinsulinemic clamps to assess insulin sensitivity. We performed reduced representation bisulfite sequencing (RRBS) next-generation methylation and microarray analyses on DNA and RNA isolated from vastus lateralis muscle biopsies. There were 13,130 differentially methylated cytosines (DMC; uncorrected P < 0.05) that were altered in the promoter and untranslated (5' and 3'UTR) regions in the obese versus lean analysis. Microarray analysis revealed 99 probes that were significantly (corrected P < 0.05) altered. Of these, 12 genes (encompassing 22 methylation sites) demonstrated a negative relationship between gene expression and DNA methylation. Specifically, sorbin and SH3 domain containing 3 (SORBS3) which codes for the adapter protein vinexin was significantly decreased in gene expression (fold change -1.9) and had nine DMCs that were significantly increased in methylation in obesity (methylation differences ranged from 5.0 to 24.4 %). Moreover, differentially methylated region (DMR) analysis identified a region in the 5'UTR (Chr.8:22,423,530-22,423,569) of SORBS3 that was increased in methylation by 11.2 % in the obese group. The negative relationship observed between DNA methylation and gene expression for SORBS3 was validated by a site-specific sequencing approach, pyrosequencing, and qRT-PCR. Additionally, we performed transcription factor binding analysis and identified a number of transcription factors whose binding to the differentially methylated sites or region may contribute to obesity. CONCLUSIONS: These results demonstrate that obesity alters the epigenome through DNA methylation and highlights novel transcriptomic changes in SORBS3 in skeletal muscle.
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Proteínas Adaptadoras Transductoras de Señales/genética , Metilación de ADN , Perfilación de la Expresión Génica/métodos , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Obesidad/genética , Adulto , Epigénesis Genética , Femenino , Regulación de la Expresión Génica , Humanos , Masculino , Proteínas Musculares , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Regiones Promotoras Genéticas , Análisis de Secuencia de ADN/métodos , Análisis de Secuencia de ARN/métodosRESUMEN
The mechanisms of metabolic improvements after Roux-en-Y gastric bypass (RYGB) surgery are not entirely clear. Therefore, the aim of our study was to investigate the role of obesity and RYGB on the human skeletal muscle proteome. Basal muscle biopsies were obtained from seven obese (BMI >40 kg/m(2)) female subjects (45.1 ± 3.6 years) pre- and 3 months post-RYGB, and euglycemic-hyperinsulinemic clamps were used to assess insulin sensitivity. Four age-matched (48.5 ± 4.7 years) lean (BMI <25 kg/m(2)) females served as control subjects. We performed quantitative mass spectrometry and microarray analyses on protein and RNA isolated from the muscle biopsies. Significant improvements in fasting plasma glucose (104.2 ± 7.8 vs. 86.7 ± 3.1 mg/dL) and BMI (42.1 ± 2.2 vs. 35.3 ± 1.8 kg/m(2)) were demonstrated in the pre- versus post-RYGB, both P < 0.05. Proteomic analysis identified 2,877 quantifiable proteins. Of these, 395 proteins were significantly altered in obesity before surgery, and 280 proteins differed significantly post-RYGB. Post-RYGB, 49 proteins were returned to normal levels after surgery. KEGG pathway analysis revealed a decreased abundance in ribosomal and oxidative phosphorylation proteins in obesity, and a normalization of ribosomal proteins post-RYGB. The transcriptomic data confirmed the normalization of the ribosomal proteins. Our results provide evidence that obesity and RYGB have a dynamic effect on the skeletal muscle proteome.
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Derivación Gástrica , Músculo Esquelético/metabolismo , Proteoma/análisis , Proteómica/métodos , Glucemia/metabolismo , Ayuno/sangre , Femenino , Técnica de Clampeo de la Glucosa , Humanos , Técnicas In Vitro , Insulina/sangre , Masculino , Espectrometría de Masas , Análisis por MicromatricesRESUMEN
Loss of skeletal muscle in patients who have undergone gastric bypass is a consistent observation. Skeletal muscle constitutes the largest protein/amino acid pool in the body, and loss of skeletal muscle has important implications in health and disease. Sustaining a given level of muscle protein requires a balance between the rates of muscle protein synthesis and breakdown. Current evidence suggests that reduced rate of protein synthesis is implicated in the loss of muscle after gastric bypass. This is not surprising given a less than optimal dietary protein intake after the procedure and because, unlike other macronutrients, protein/amino acids are not stored in the body. Ingesting essential amino acids (EAAs), which cannot be synthesized de novo and have the primary role in the regulation of muscle protein synthesis, can potentially ameliorate loss of muscle protein after gastric bypass. At the same time, ingestion of EAAs provides a more efficient nutritional approach (i.e., greater stimulation of protein synthesis relative to the amount of amino acids ingested) to enhance muscle protein synthesis compared with the ingestion of intact protein. Changing current dietary practices toward increasing ingestion of EAAs provides an approach that can potentially prevent loss of lean body tissue and ultimately achieve a more sustained level of health in patients who have undergone gastric bypass.
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Aminoácidos Esenciales/uso terapéutico , Suplementos Dietéticos , Derivación Gástrica/efectos adversos , Proteínas Musculares/biosíntesis , Músculo Esquelético/efectos de los fármacos , Biosíntesis de Proteínas/efectos de los fármacos , Aminoácidos Esenciales/metabolismo , Proteínas en la Dieta/administración & dosificación , Proteínas en la Dieta/metabolismo , Humanos , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Obesidad Mórbida/cirugíaRESUMEN
Enrichment from the easily accessible blood amino acid pool is commonly used as precursor enrichment to calculate rates of muscle protein fractional synthesis in relevant human studies in lieu of the less accessible muscle fluid amino acid pool. However, the accuracy of this approach depends largely on the extent to which there is low discrepancy in free amino acid enrichment between blood and muscle. Steady-state gradient (i.e., ratio) of amino acid enrichment between blood and muscle fluid in the basal state and in response to amino acid infusion were determined in five healthy subjects, and in association with two separate tracers: d9-leucine, introduced endogenously by the metabolism of d10-leucine (i.e., l-[2,3,3,4,5,5,5,6,6,6-(2)H10]leucine) infused in blood, and (13)C6-phenylalanine introduced/infused in blood. The blood-to-muscle fluid amino acid enrichment ratio was lower (P < 0.05) for d9-leucine compared to (13)C6-phenylalanine both before (1.5 ± 0.1 vs. 2.5 ± 0.1) and during (1.1 ± 0.1 vs. 1.2 ± 0.1) amino acid infusion. Importantly, the decrease in this ratio in association with the amino acid infusion was considerably less for the d9-leucine than the (13)C6-phenylalanine (-0.38 ± 0.03 vs. -1.29 ± 0.07; P < 0.05). In conclusion, blood d9-leucine enrichment introduced endogenously by intravenous infusion of d10-leucine provides a closer estimate of the muscle fluid amino acid enrichment, and its associated changes, than blood phenylalanine enrichment to calculate rates of muscle protein synthesis in humans.
RESUMEN
Obesity is a growing worldwide epidemic, increasingly addressed through surgical options for weight loss. Benefits of these operations, such as weight loss and improvement or reversal of obesity-related comorbidities, are well established; however, postoperative complications do occur. This article will evaluate common causes for hospital admissions in the post-bariatric surgery population as they relate to the hospitalist who is often responsible for their care. Here we provide an overview of the most common bariatric procedures currently performed, early postoperative complications, late medical complications (ie, abdominal complaints, weight fluctuations, nutritional deficiencies, and metabolic bone disease), and late surgical complications that often affect these patients and result in hospital admissions. Special attention will be paid to radiologic pearls that can assist in the initial evaluation and diagnosis of these patients.
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Cirugía Bariátrica/efectos adversos , Médicos Hospitalarios/educación , Obesidad Mórbida/cirugía , Complicaciones Posoperatorias/prevención & control , Cirugía Bariátrica/métodos , Humanos , Obesidad Mórbida/complicaciones , Complicaciones Posoperatorias/etiología , Pérdida de PesoRESUMEN
BACKGROUND: Little is known about management of hyperglycemia in inpatients. OBJECTIVE: To gain insight into caring for hospitalized patients with hyperglycemia. DESIGN: Retrospective analysis. SETTING: Teaching hospital. PATIENTS: Data on all patients discharged between January 1, 2001, and December 31, 2004 with a diagnosis of diabetes or hyperglycemia were extracted and linked to laboratory and pharmacy databases. Only the data on patients who did not require intensive care and who were hospitalized for at least 3 days were analyzed. MEASUREMENTS: Average bedside glucose during the first and last 24 hours of hospital stay and for the entire length of stay; assessment of changes in insulin regimen and dose. RESULTS: The average age of patients included in the study (n = 2916) was 69 years. Fifty-seven percent of the patients were men, 90% were white, and average length of stay was 5.7 days. More than 20% of the patients had evidence of sustained hyperglycemia. Forty-two percent of the patients who showed poor control of glycemia (glucose > 200 mg/dL) during the first 24 hours were discharged in poor control. The frequency of hypoglycemia was low (only 2.2 of 100 measurements per person) compared with hyperglycemia (25.5 of 100 measurements per person). Most patients (72%) received insulin during hospitalization, but there was high use of short-acting insulin and less than optimal intensification of therapy (clinical inertia); many patients had insulin therapy decreased despite persistent hyperglycemia (negative therapeutic momentum). CONCLUSIONS: Glycemic control in the hospital was frequently poor, and there was suboptimal use of insulin, even among patients with sustained hyperglycemia. Educational programs directed at practitioners should focus on the importance of inpatient glucose control and provide guidelines on how and when to change therapy.
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Diabetes Mellitus/sangre , Hospitalización , Hiperglucemia/sangre , Anciano , Glucemia/metabolismo , Distribución de Chi-Cuadrado , Diabetes Mellitus/terapia , Manejo de la Enfermedad , Femenino , Índice Glucémico , Humanos , Hiperglucemia/terapia , Pacientes Internos , Insulina/administración & dosificación , Tiempo de Internación/estadística & datos numéricos , Masculino , Estudios Retrospectivos , Resultado del TratamientoRESUMEN
OBJECTIVE: To develop insight into resident physician attitudes about inpatient hyperglycemia and determine perceived barriers to optimal management. METHODS: As part of a planned educational program, a questionnaire was designed and administered to determine the opinions of residents about the importance of inpatient glucose control, their perceptions about what glucose ranges were desirable, and the problems they encountered when trying to manage hyperglycemia in hospitalized patients. RESULTS: Of 70 resident physicians from various services, 52 completed the survey (mean age, 31 years; 48% men; 37% in first year of residency training). Most respondents indicated that glucose control was "very important" in critically ill and perioperative patients but only "somewhat important" in non-critically ill patients. Most residents indicated that they would target a therapeutic glucose range within the recommended levels in published guidelines. Most residents also said they felt "somewhat comfortable" managing hyperglycemia and hypoglycemia and using subcutaneous insulin therapy, whereas most residents (48%) were "not at all comfortable" with use of intravenous administration of insulin. In general, respondents were not very familiar with existing institutional policies and preprinted order sets relating to glucose management. The most commonly reported barrier to management of inpatient hyperglycemia was lack of knowledge about appropriate insulin regimens and how to use them. Anxiety about hypoglycemia was only the third most frequent concern. CONCLUSION: Most residents acknowledged the importance of good glucose control in hospitalized patients and chose target glucose ranges consistent with existing guidelines. Lack of knowledge about insulin treatment options was the most commonly cited barrier to ideal management. Educational programs should emphasize inpatient treatment strategies for glycemic control.