Journal of applied physiology (Bethesda, Md. : 1985), Jan 25, 2015
We determined the effects of 'periodized nutrition' on skeletal muscle and whole-body res... more We determined the effects of 'periodized nutrition' on skeletal muscle and whole-body responses to a bout of prolonged exercise the following morning. Seven cyclists completed two trials receiving isoenergetic diets differing in the timing of ingestion: They consumed either 8 g•kg(-1) BM of CHO before undertaking an evening session of high-intensity training (HIT) and slept without eating (FASTED), or 4 g•kg(-1) BM of CHO before HIT then 4 g•kg(-1) BM of CHO before sleeping (FED). The next morning subjects completed 2 h cycling (120SS) while overnight fasted. Muscle biopsies were taken on day 1 (D1) before and 2 h after HIT and on Day 2 (D2) pre-, post-, and 4 h after 120SS. Muscle [glycogen] was higher in FED at all times post-HIT (P< 0.001). HIT increased PGC1α mRNA (P< 0.01) while PDK4 mRNA was elevated to a greater extent in FASTED (P< 0.05). Resting phosphorylation of AMPK(Thr172), p38MAPK(Thr180/Tyr182) and p-ACC(Ser79) (D2) was greater in FASTED (P< 0.05)....
Acute exercise increases glucose uptake in skeletal muscle by an insulin-independent mechanism. I... more Acute exercise increases glucose uptake in skeletal muscle by an insulin-independent mechanism. In the period after exercise insulin sensitivity to increase glucose uptake is enhanced. The molecular mechanisms underpinning this phenomenon are poorly understood, but appear to involve an increased cell surface abundance of GLUT4. While increased proximal insulin signaling does not seem to mediate this effect, elevated phosphorylation of TBC1D4, a downstream target of both insulin (Akt) and exercise (AMPK) signaling, appears to play a role. The main purpose of this study was to determine whether AMPK activation increases skeletal muscle insulin sensitivity. We found that prior AICAR stimulation of wild-type mouse muscle increases insulin sensitivity to stimulate glucose uptake. However, this was not observed in mice with reduced or ablated AMPK activity in skeletal muscle. Furthermore, prior AICAR stimulation enhanced insulin-stimulated phosphorylation of TBC1D4 at Thr(649) and Ser(711...
microRNAs (miRNAs) are short noncoding RNAs that regulate gene expression through posttranscripti... more microRNAs (miRNAs) are short noncoding RNAs that regulate gene expression through posttranscriptional repression of target genes. miRNAs exert a fundamental level of control over many developmental processes, but their role in the differentiation and development of skeletal muscle from myogenic progenitor cells in humans remains incompletely understood. Using primary cultures established from human skeletal muscle satellite cells, we performed microarray profiling of miRNA expression during differentiation of myoblasts (day 0) into myotubes at 48 h intervals (day 2, 4, 6, 8, and 10). Based on a time-course analysis, we identified 44 miRNAs with altered expression [false discovery rate (FDR) < 5%, fold change > ±1.2] during differentiation, including the marked upregulation of the canonical myogenic miRNAs miR-1, miR-133a, miR-133b, and miR-206. Microarray profiling of mRNA expression at day 0, 4, and 10 identified 842 and 949 genes differentially expressed (FDR < 10%) at da...
American journal of physiology. Cell physiology, 2014
Two-dimensional difference gel electrophoresis (2-D DIGE)-based proteome analysis has revealed in... more Two-dimensional difference gel electrophoresis (2-D DIGE)-based proteome analysis has revealed intrinsic insulin resistance in myotubes derived from type 2 diabetic patients. Using 2-D DIGE-based proteome analysis, we identified a subset of insulin-resistant proteins involved in protein turnover in skeletal muscle of type 2 diabetic patients, suggesting aberrant regulation of the protein homeostasis maintenance system underlying metabolic disease. We then validated the role of the ubiquitin-proteasome system (UPS) in myotubes to investigate whether impaired proteasome function may lead to metabolic arrest or insulin resistance. Myotubes derived from muscle biopsies obtained from people with normal glucose tolerance (NGT) or type 2 diabetes were exposed to the proteasome inhibitor bortezomib (BZ; Velcade) without or with insulin. BZ exposure increased protein carbonylation and lactate production yet impaired protein synthesis and UPS function in myotubes from type 2 diabetic patients...
In mature human skeletal muscle, insulin-stimulated glucose transport is mediated primarily via t... more In mature human skeletal muscle, insulin-stimulated glucose transport is mediated primarily via the GLUT4 glucose transporter. However, in contrast to mature skeletal muscle, cultured muscle expresses significant levels of the GLUT1 glucose transporter. To assess the relative contribution of these two glucose transporters, we used a novel photolabelling techniques to assess the cell surface abundance of GLUT1 and GLUT4 specifically in primary cultures of human skeletal muscle. We demonstrate that insulin-stimulated glucose transport in cultured human skeletal muscle is mediated by GLUT4, as no effect on GLUT1 appearance at the plasma membrane was noted. Furthermore, GLUT4 mRNA and protein increased twofold (p < 0.05), after differentiation, whereas GLUT1 mRNA and protein decreased 55% (p < 0.005). Incubation of differentiated human skeletal muscle cells with a non-peptide insulin mimetic significantly (p < 0.05) increased glucose uptake and glycogen synthesis. Thus, culture...
Arterial stiffness is an important cardiovascular risk marker, which can be measured noninvasivel... more Arterial stiffness is an important cardiovascular risk marker, which can be measured noninvasively with different techniques. To validate such techniques in healthy subjects, we compared the recently introduced oscillometric Arteriograph (AG) technique with the tonometric SphygmoCor (SC) method and their associations with carotid ultrasound measures and traditional risk indicators. Sixty-three healthy subjects aged 20-69 (mean 48 ± 15) years were included. We measured aortic pulse wave velocity (PWVao) and augmentation index (AIx) by AG and SC, and with SC also the PWVao standardized to 80% of the direct distance between carotid and femoral sites (St-PWVaoSC). The carotid strain, stiffness index and intima-media thickness (cIMTmean) were evaluated by ultrasound. PWVaoAG (8.00 ± 2.16 m s(-1)) was higher (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001) than PWVaoSC (6.87 ± 1.47 m s(-1)), but did not differ from St-PWVaoSC (7.68 ± 1.58 m s(-1)), and correlated (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001) with both (r = 0.54 and 0.59). St-PWVaoSC was significantly (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.01) higher than PWVaoAG for values below median (7.4 m s(-1)). PWVao by SC and AG differed significantly in females (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001), but not in males (P=0.40). AIxaoAG (27.5 ± 14.5%) was higher (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001) than AIxaoSC (20.5 ± 17.4%), but related closely (r=0.97, P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001). St-PWVaoSC, PWVao and AIxao by SC, and PWVao and AIxao by AG were all related to serum cholesterol and to cIMTmean (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001). Arterial stiffness indices by AG and SC correlate with vascular risk markers in healthy subjects. AIxao results by AG and SC are closely interrelated, but higher values are obtained by AG. In the lower range, PWVao values by AG and SC are similar, but differ for higher values. Our results imply the necessity to apply one and the same technique for repeated studies.
The identification of microRNAs (miRNAs) has established new mechanisms that control skeletal m... more The identification of microRNAs (miRNAs) has established new mechanisms that control skeletal muscle adaptation to exercise. The present study investigated the mRNA regulation of components of the miRNA biogenesis pathway (Drosha, Dicer and Exportin-5), muscle enriched miRNAs, (miR-1, -133a, -133b and -206), and several miRNAs dysregulated in muscle myopathies (miR-9, -23, -29, -31 and -181). Measurements were made in muscle biopsies from nine healthy untrained males at rest, 3 h following an acute bout of moderate-intensity endurance cycling and following 10 days of endurance training. Bioinformatics analysis was used to predict potential miRNA targets. In the 3 h period following the acute exercise bout, Drosha, Dicer and Exportin-5, as well as miR-1, -133a, -133-b and -181a were all increased. In contrast miR-9, -23a, -23b and -31 were decreased. Short-term training increased miR-1 and -29b, while miR-31 remained decreased. Negative correlations were observed between miR-9 and HDAC4 protein (r=-0.71; P=0.04), miR-31 and HDAC4 protein (r=-0.87; P=0.026) and miR-31 and NRF1 protein (r=-0.77; P=0.01) 3 h following exercise. miR-31 binding to the HDAC4 and NRF1 3 untranslated region (UTR) reduced luciferase reporter activity. Exercise rapidly and transiently regulates several miRNA species in muscle. Several of these miRNAs may be involved in the regulation of skeletal muscle regeneration, gene transcription and mitochondrial biogenesis. Identifying endurance exercise-mediated stress signals regulating skeletal muscle miRNAs, as well as validating their targets and regulatory pathways post exercise, will advance our understanding of their potential role/s in human health.
Skeletal muscle is a malleable organ that responds to a single acute exercise bout by inducing th... more Skeletal muscle is a malleable organ that responds to a single acute exercise bout by inducing the expression of genes involved in structural, metabolic and functional adaptations. Several epigenetic mechanisms including histone H4 deacetylation and loss of promoter methylation have been implicated in modifying exercise-responsive gene expression. These transient changes suggest that epigenetic mechanisms are not restricted to early stages of human development but are broad dynamic controllers of genomic plasticity in response to environmental factors.
Journal of applied physiology (Bethesda, Md. : 1985), Jan 25, 2015
We determined the effects of 'periodized nutrition' on skeletal muscle and whole-body res... more We determined the effects of 'periodized nutrition' on skeletal muscle and whole-body responses to a bout of prolonged exercise the following morning. Seven cyclists completed two trials receiving isoenergetic diets differing in the timing of ingestion: They consumed either 8 g•kg(-1) BM of CHO before undertaking an evening session of high-intensity training (HIT) and slept without eating (FASTED), or 4 g•kg(-1) BM of CHO before HIT then 4 g•kg(-1) BM of CHO before sleeping (FED). The next morning subjects completed 2 h cycling (120SS) while overnight fasted. Muscle biopsies were taken on day 1 (D1) before and 2 h after HIT and on Day 2 (D2) pre-, post-, and 4 h after 120SS. Muscle [glycogen] was higher in FED at all times post-HIT (P< 0.001). HIT increased PGC1α mRNA (P< 0.01) while PDK4 mRNA was elevated to a greater extent in FASTED (P< 0.05). Resting phosphorylation of AMPK(Thr172), p38MAPK(Thr180/Tyr182) and p-ACC(Ser79) (D2) was greater in FASTED (P< 0.05)....
Acute exercise increases glucose uptake in skeletal muscle by an insulin-independent mechanism. I... more Acute exercise increases glucose uptake in skeletal muscle by an insulin-independent mechanism. In the period after exercise insulin sensitivity to increase glucose uptake is enhanced. The molecular mechanisms underpinning this phenomenon are poorly understood, but appear to involve an increased cell surface abundance of GLUT4. While increased proximal insulin signaling does not seem to mediate this effect, elevated phosphorylation of TBC1D4, a downstream target of both insulin (Akt) and exercise (AMPK) signaling, appears to play a role. The main purpose of this study was to determine whether AMPK activation increases skeletal muscle insulin sensitivity. We found that prior AICAR stimulation of wild-type mouse muscle increases insulin sensitivity to stimulate glucose uptake. However, this was not observed in mice with reduced or ablated AMPK activity in skeletal muscle. Furthermore, prior AICAR stimulation enhanced insulin-stimulated phosphorylation of TBC1D4 at Thr(649) and Ser(711...
microRNAs (miRNAs) are short noncoding RNAs that regulate gene expression through posttranscripti... more microRNAs (miRNAs) are short noncoding RNAs that regulate gene expression through posttranscriptional repression of target genes. miRNAs exert a fundamental level of control over many developmental processes, but their role in the differentiation and development of skeletal muscle from myogenic progenitor cells in humans remains incompletely understood. Using primary cultures established from human skeletal muscle satellite cells, we performed microarray profiling of miRNA expression during differentiation of myoblasts (day 0) into myotubes at 48 h intervals (day 2, 4, 6, 8, and 10). Based on a time-course analysis, we identified 44 miRNAs with altered expression [false discovery rate (FDR) < 5%, fold change > ±1.2] during differentiation, including the marked upregulation of the canonical myogenic miRNAs miR-1, miR-133a, miR-133b, and miR-206. Microarray profiling of mRNA expression at day 0, 4, and 10 identified 842 and 949 genes differentially expressed (FDR < 10%) at da...
American journal of physiology. Cell physiology, 2014
Two-dimensional difference gel electrophoresis (2-D DIGE)-based proteome analysis has revealed in... more Two-dimensional difference gel electrophoresis (2-D DIGE)-based proteome analysis has revealed intrinsic insulin resistance in myotubes derived from type 2 diabetic patients. Using 2-D DIGE-based proteome analysis, we identified a subset of insulin-resistant proteins involved in protein turnover in skeletal muscle of type 2 diabetic patients, suggesting aberrant regulation of the protein homeostasis maintenance system underlying metabolic disease. We then validated the role of the ubiquitin-proteasome system (UPS) in myotubes to investigate whether impaired proteasome function may lead to metabolic arrest or insulin resistance. Myotubes derived from muscle biopsies obtained from people with normal glucose tolerance (NGT) or type 2 diabetes were exposed to the proteasome inhibitor bortezomib (BZ; Velcade) without or with insulin. BZ exposure increased protein carbonylation and lactate production yet impaired protein synthesis and UPS function in myotubes from type 2 diabetic patients...
In mature human skeletal muscle, insulin-stimulated glucose transport is mediated primarily via t... more In mature human skeletal muscle, insulin-stimulated glucose transport is mediated primarily via the GLUT4 glucose transporter. However, in contrast to mature skeletal muscle, cultured muscle expresses significant levels of the GLUT1 glucose transporter. To assess the relative contribution of these two glucose transporters, we used a novel photolabelling techniques to assess the cell surface abundance of GLUT1 and GLUT4 specifically in primary cultures of human skeletal muscle. We demonstrate that insulin-stimulated glucose transport in cultured human skeletal muscle is mediated by GLUT4, as no effect on GLUT1 appearance at the plasma membrane was noted. Furthermore, GLUT4 mRNA and protein increased twofold (p < 0.05), after differentiation, whereas GLUT1 mRNA and protein decreased 55% (p < 0.005). Incubation of differentiated human skeletal muscle cells with a non-peptide insulin mimetic significantly (p < 0.05) increased glucose uptake and glycogen synthesis. Thus, culture...
Arterial stiffness is an important cardiovascular risk marker, which can be measured noninvasivel... more Arterial stiffness is an important cardiovascular risk marker, which can be measured noninvasively with different techniques. To validate such techniques in healthy subjects, we compared the recently introduced oscillometric Arteriograph (AG) technique with the tonometric SphygmoCor (SC) method and their associations with carotid ultrasound measures and traditional risk indicators. Sixty-three healthy subjects aged 20-69 (mean 48 ± 15) years were included. We measured aortic pulse wave velocity (PWVao) and augmentation index (AIx) by AG and SC, and with SC also the PWVao standardized to 80% of the direct distance between carotid and femoral sites (St-PWVaoSC). The carotid strain, stiffness index and intima-media thickness (cIMTmean) were evaluated by ultrasound. PWVaoAG (8.00 ± 2.16 m s(-1)) was higher (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001) than PWVaoSC (6.87 ± 1.47 m s(-1)), but did not differ from St-PWVaoSC (7.68 ± 1.58 m s(-1)), and correlated (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001) with both (r = 0.54 and 0.59). St-PWVaoSC was significantly (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.01) higher than PWVaoAG for values below median (7.4 m s(-1)). PWVao by SC and AG differed significantly in females (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001), but not in males (P=0.40). AIxaoAG (27.5 ± 14.5%) was higher (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001) than AIxaoSC (20.5 ± 17.4%), but related closely (r=0.97, P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001). St-PWVaoSC, PWVao and AIxao by SC, and PWVao and AIxao by AG were all related to serum cholesterol and to cIMTmean (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001). Arterial stiffness indices by AG and SC correlate with vascular risk markers in healthy subjects. AIxao results by AG and SC are closely interrelated, but higher values are obtained by AG. In the lower range, PWVao values by AG and SC are similar, but differ for higher values. Our results imply the necessity to apply one and the same technique for repeated studies.
The identification of microRNAs (miRNAs) has established new mechanisms that control skeletal m... more The identification of microRNAs (miRNAs) has established new mechanisms that control skeletal muscle adaptation to exercise. The present study investigated the mRNA regulation of components of the miRNA biogenesis pathway (Drosha, Dicer and Exportin-5), muscle enriched miRNAs, (miR-1, -133a, -133b and -206), and several miRNAs dysregulated in muscle myopathies (miR-9, -23, -29, -31 and -181). Measurements were made in muscle biopsies from nine healthy untrained males at rest, 3 h following an acute bout of moderate-intensity endurance cycling and following 10 days of endurance training. Bioinformatics analysis was used to predict potential miRNA targets. In the 3 h period following the acute exercise bout, Drosha, Dicer and Exportin-5, as well as miR-1, -133a, -133-b and -181a were all increased. In contrast miR-9, -23a, -23b and -31 were decreased. Short-term training increased miR-1 and -29b, while miR-31 remained decreased. Negative correlations were observed between miR-9 and HDAC4 protein (r=-0.71; P=0.04), miR-31 and HDAC4 protein (r=-0.87; P=0.026) and miR-31 and NRF1 protein (r=-0.77; P=0.01) 3 h following exercise. miR-31 binding to the HDAC4 and NRF1 3 untranslated region (UTR) reduced luciferase reporter activity. Exercise rapidly and transiently regulates several miRNA species in muscle. Several of these miRNAs may be involved in the regulation of skeletal muscle regeneration, gene transcription and mitochondrial biogenesis. Identifying endurance exercise-mediated stress signals regulating skeletal muscle miRNAs, as well as validating their targets and regulatory pathways post exercise, will advance our understanding of their potential role/s in human health.
Skeletal muscle is a malleable organ that responds to a single acute exercise bout by inducing th... more Skeletal muscle is a malleable organ that responds to a single acute exercise bout by inducing the expression of genes involved in structural, metabolic and functional adaptations. Several epigenetic mechanisms including histone H4 deacetylation and loss of promoter methylation have been implicated in modifying exercise-responsive gene expression. These transient changes suggest that epigenetic mechanisms are not restricted to early stages of human development but are broad dynamic controllers of genomic plasticity in response to environmental factors.
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