Regulation of HSL serine phosphorylation in skeletal muscle and adipose tissue

Hormone-sensitive lipase (HSL) is important for the degradation of triacylglycerol in adipose and muscle tissue, but the tissue-specific regulation of this enzyme is not fully understood. We investigated the effects of adrenergic stimulation and AMPK activation in vitro and in circumstances where AMPK activity and catecholamines are physiologically elevated in humans in vivo (during physical exercise) on HSL activity and phosphorylation at Ser563 and Ser660, the PKA regulatory sites, and Ser565, the AMPK regulatory site. In human experiments, skeletal muscle, subcutaneous adipose and venous blood samples were obtained before, at 15 and 90 min during, and 120 min after exercise. Skeletal muscle HSL activity was increased by ∼80% at 15 min compared with rest and returned to resting rates at the cessation of and 120 min after exercise. Consistent with changes in plasma epinephrine, skeletal muscle HSL Ser563 and Ser660 phosphorylation were increased by 27% at 15 min ( P < 0.05), remained elevated at 90 min, and returned to preexercise values postexercise. Skeletal muscle HSL Ser565 phosphorylation and AMPK signaling were increased at 90 min during, and after, exercise. Phosphorylation of adipose tissue HSL paralleled changes in skeletal muscle in vivo, except HSL Ser660 was elevated 80% in adipose compared with 35% in skeletal muscle during exercise. Studies in L6 myotubes and 3T3-L1 adipocytes revealed important tissue differences in the regulation of HSL. AMPK inhibited epinephrine-induced HSL activity in L6 myotubes and was associated with reduced HSL Ser660 but not Ser563 phosphorylation. HSL activity was reduced in L6 myotubes expressing constitutively active AMPK, confirming the inhibitory effects of AMPK on HSL activity. Conversely, in 3T3-L1 adipocytes, AMPK activation after epinephrine stimulation did not prevent HSL activity or glycerol release, which coincided with maintenance of HSL Ser660 phosphorylation. Taken together, these data indicate that HSL activity is maintained in the face of AMPK activation as a result of elevated HSL Ser660 phosphorylation in adipose tissue but not skeletal muscle.

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CTRP9 transgenic mice are protected from diet-induced obesity and metabolic dysfunction

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00110.2013 ◽

2013 ◽

Vol 305 (5) ◽

pp. R522-R533 ◽

Cited By ~ 70

Author(s):

Jonathan M. Peterson ◽

Zhikui Wei ◽

Marcus M. Seldin ◽

Mardi S. Byerly ◽

Susan Aja ◽

...

Keyword(s):

Skeletal Muscle ◽

Adipose Tissue ◽

Transgenic Mice ◽

High Fat Diet ◽

Fat Oxidation ◽

Acid Oxidation ◽

Ampk Activation ◽

High Fat ◽

Diet Induced Obesity

CTRP9 is a secreted multimeric protein of the C1q family and the closest paralog of the insulin-sensitizing adipokine, adiponectin. The metabolic function of this adipose tissue-derived plasma protein remains largely unknown. Here, we show that the circulating levels of CTRP9 are downregulated in diet-induced obese mice and upregulated upon refeeding. Overexpressing CTRP9 resulted in lean mice that dramatically resisted weight gain induced by a high-fat diet, largely through decreased food intake and increased basal metabolism. Enhanced fat oxidation in CTRP9 transgenic mice resulted from increases in skeletal muscle mitochondrial content, expression of enzymes involved in fatty acid oxidation (LCAD and MCAD), and chronic AMPK activation. Hepatic and skeletal muscle triglyceride levels were substantially decreased in transgenic mice. Consequently, CTRP9 transgenic mice had a greatly improved metabolic profile with markedly reduced fasting insulin and glucose levels. The high-fat diet-induced obesity, insulin resistance, and hepatic steatosis observed in wild-type mice were prevented in transgenic mice. Consistent with the in vivo data, recombinant protein significantly enhanced fat oxidation in L6 myotubes via AMPK activation and reduced lipid accumulation in H4IIE hepatocytes. Collectively, these data establish CTRP9 as a novel metabolic regulator and a new component of the metabolic network that links adipose tissue to lipid metabolism in skeletal muscle and liver.

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Adrenergic regulation of HSL serine phosphorylation and activity in human skeletal muscle during the onset of exercise

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00130.2006 ◽

2006 ◽

Vol 291 (4) ◽

pp. R1094-R1099 ◽

Cited By ~ 12

Author(s):

Jason L. Talanian ◽

Rebecca J. Tunstall ◽

Matthew J. Watt ◽

Mylinh Duong ◽

Christopher G. R. Perry ◽

...

Keyword(s):

Skeletal Muscle ◽

Cyclic Amp ◽

High Intensity ◽

Human Skeletal Muscle ◽

High Intensity Exercise ◽

Serine Phosphorylation ◽

Hormone Sensitive Lipase ◽

Peak Oxygen Consumption ◽

Adrenergic Stimulation ◽

Arterial Blood

Skeletal muscle hormone-sensitive lipase (HSL) activity is increased by contractions and increases in blood epinephrine (EPI) concentrations and cyclic AMP activation of the adrenergic pathway during prolonged exercise. To determine the importance of hormonal stimulation of HSL activity during the onset of moderate- and high-intensity exercise, nine men [age 24.3 ± 1.2 yr, 80.8 ± 5.0 kg, peak oxygen consumption (V̇o2 peak) 43.9 ± 3.6 ml·kg−1·min−1] cycled for 1 min at ∼65% V̇o2 peak, rested for 60 min, and cycled at ∼90% V̇o2 peak for 1 min. Skeletal muscle biopsies were taken pre- and postexercise, and arterial blood was sampled throughout exercise. Arterial EPI increased ( P < 0.05) postexercise at 65% (0.45 ± 0.10 to 0.78 ± 0.27 nM) and 90% V̇o2 peak (0.57 ± 0.34 to 1.09 ± 0.50 nM). HSL activity increased ( P < 0.05) following 1 min of exercise at 65% V̇o2 peak [1.05 ± 0.39 to 1.78 ± 0.54 mmol·min−1·kg dry muscle (dm)−1] and 90% V̇o2 peak (1.07 ± 0.24 to 1.91 ± 0.62 mmol·min−1·kg dm−1). Cyclic AMP content also increased ( P < 0.05) at both exercise intensities (65%: 1.52 ± 0.67 to 2.75 ± 1.12, 90%: 1.85 ± 0.65 to 2.64 ± 0.93 μmol/kg dm). HSL Ser660 phosphorylation (∼55% increase) and ERK1/2 phosphorylation (∼33% increase) were augmented following exercise at both intensities, whereas HSL Ser563 and Ser565 phosphorylation were not different from rest. The results indicate that increases in arterial EPI concentration during the onset of moderate- and high-intensity exercise increase cyclic AMP content, which results in the phosphorylation of HSL Ser660. This adrenergic stimulation contributes to the increase in HSL activity that occurs in human skeletal muscle in the first minute of exercise at 65% and 90% V̇o2 peak.

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Hormone-Sensitive Lipase Serine Phosphorylation and Glycerol Exchange Across Skeletal Muscle in Lean and Obese Subjects: Effect of -Adrenergic Stimulation

Diabetes ◽

10.2337/db07-0857 ◽

2008 ◽

Vol 57 (7) ◽

pp. 1834-1841 ◽

Cited By ~ 32

Author(s):

J. W.E. Jocken ◽

C. Roepstorff ◽

G. H. Goossens ◽

P. van der Baan ◽

M. van Baak ◽

...

Keyword(s):

Skeletal Muscle ◽

Serine Phosphorylation ◽

Hormone Sensitive Lipase ◽

Adrenergic Stimulation ◽

Hormone Sensitive ◽

Obese Subjects

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Effects of an oral and intravenous fat load on adipose tissue and forearm lipid metabolism

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1999.276.2.e241 ◽

1999 ◽

Vol 276 (2) ◽

pp. E241-E248 ◽

Cited By ~ 17

Author(s):

Kevin Evans ◽

Mo L. Clark ◽

Keith N. Frayn

Keyword(s):

Skeletal Muscle ◽

Fatty Acids ◽

Adipose Tissue ◽

Fatty Acid ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Hormone Sensitive Lipase ◽

Nonesterified Fatty Acid ◽

Subcutaneous Adipose

We have studied the fate of lipoprotein lipase (LPL)-derived fatty acids by measuring arteriovenous differences across subcutaneous adipose tissue and skeletal muscle in vivo. Six subjects were fasted overnight and were then given 40 g of triacylglycerol either orally or as an intravenous infusion over 4 h. Intracellular lipolysis (hormone-sensitive lipase action; HSL) was suppressed after both oral and intravenous fat loads ( P < 0.001). Insulin, a major regulator of HSL activity, showed little change after either oral or intravenous fat load, suggesting that suppression of HSL action occurred independently of insulin. The rate of action of LPL (measured as triacylglycerol extraction) increased with both oral and intravenous fat loads in adipose tissue ( P = 0.002) and skeletal muscle ( P = 0.001). There was increased escape of LPL-derived fatty acids into the circulation from adipose tissue, shown by lack of reesterification of fatty acids. There was no release into the circulation of LPL-derived fatty acids from skeletal muscle. These results suggest that insulin is not essential for HSL suppression or increased triacylglycerol clearance but is important in reesterification of fatty acids in adipose tissue but not uptake by skeletal muscle, thus affecting fatty acid partitioning between adipose tissue and the circulation, postprandial nonesterified fatty acid concentrations, and hepatic very low density lipoprotein secretion.

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Triglyceride lipases alter fuel metabolism and mitochondrial gene expressionThis paper is one of a selection of papers published in this Special Issue, entitled 14th International Biochemistry of Exercise Conference – Muscles as Molecular and Metabolic Machines, and has undergone the Journal’s usual peer review process.

Applied Physiology Nutrition and Metabolism ◽

10.1139/h09-019 ◽

2009 ◽

Vol 34 (3) ◽

pp. 340-347 ◽

Cited By ~ 13

Author(s):

Matthew J. Watt

Keyword(s):

Skeletal Muscle ◽

Adipose Tissue ◽

Fatty Acid ◽

Peer Review Process ◽

Mitochondrial Gene ◽

Hormone Sensitive Lipase ◽

Moderate Intensity ◽

Moderate Intensity Exercise ◽

Important Regulator

Fatty acids derived from the hydrolysis of adipose tissue and skeletal muscle triacylglycerol (TG) are an important energy substrate at rest and during prolonged moderate-intensity exercise. Hormone sensitive lipase (HSL) was long considered to be the rate-limiting enzyme for adipocyte and skeletal muscle TG lipolysis. However, the understanding of TG lipolysis regulation was recently challenged by the finding that adipose TG lipase (ATGL) is the predominant TG lipase in adipose tissue and an important regulator of TG degradation in skeletal muscle. Thus, it is now proposed that ATGL and HSL regulate lipolysis in a serial manner, with ATGL cleaving the first fatty acid and HSL the second fatty acid of TG. Further to this biochemical evaluation, the generation and metabolic characterization of ATGL−/− and HSL−/− mice have revealed distinct phenotypes. ATGL−/− mice are obese, exhibit impaired thermogenesis, oxidize more carbohydrate, and die prematurely due to cardiac dysfunction. Studies in HSL−/− mice report defective β-adrenergic stimulated lipolysis, protection against high-fat diet-induced obesity, and possible impairments in insulin secretion. This review outlines the current understanding of the cellular regulation of TG lipases, lipolytic regulation, and the functional implications of manipulating ATGL and HSL in vivo.

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Nitric oxide increases GLUT4 expression and regulates AMPK signaling in skeletal muscle

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00045.2007 ◽

2007 ◽

Vol 293 (4) ◽

pp. E1062-E1068 ◽

Cited By ~ 113

Author(s):

Vitor A. Lira ◽

Quinlyn A. Soltow ◽

Jodi H. D. Long ◽

Jenna L. Betters ◽

Jeff E. Sellman ◽

...

Keyword(s):

Nitric Oxide ◽

Skeletal Muscle ◽

Glucose Transporter ◽

No Donor ◽

Ampk Signaling ◽

L6 Myotubes ◽

Glut4 Expression ◽

Compound C ◽

Cgmp Analog

Nitric oxide (NO) and 5′-AMP-activated protein kinase (AMPK) are involved in glucose transport and mitochondrial biogenesis in skeletal muscle. Here, we examined whether NO regulates the expression of the major glucose transporter in muscle (GLUT4) and whether it influences AMPK-induced upregulation of GLUT4. At low levels, the NO donor S-nitroso- N-penicillamine (SNAP, 1 and 10 μM) significantly increased GLUT4 mRNA (∼3-fold; P < 0.05) in L6 myotubes, and cotreatment with the AMPK inhibitor compound C ablated this effect. The cGMP analog 8-bromo-cGMP (8-Br-cGMP, 2 mM) increased GLUT4 mRNA by ∼50% ( P < 0.05). GLUT4 protein expression was elevated 40% by 2 days treatment with 8-Br-cGMP, whereas 6 days treatment with 10 μM SNAP increased GLUT4 expression by 65%. Cotreatment of cultures with the guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one prevented the SNAP-induced increase in GLUT4 protein. SNAP (10 μM) also induced significant phosphorylation of α-AMPK and acetyl-CoA carboxylase and translocation of phosphorylated α-AMPK to the nucleus. Furthermore, L6 myotubes exposed to 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) for 16 h presented an approximately ninefold increase in GLUT4 mRNA, whereas cotreatment with the non-isoform-specific NOS inhibitor NG-nitro-l-arginine methyl ester, prevented ∼70% of this effect. In vivo, GLUT4 mRNA was increased 1.8-fold in the rat plantaris muscle 12 h after AICAR injection, and this induction was reduced by ∼50% in animals cotreated with the neuronal and inducible nitric oxide synthases selective inhibitor 1-(2-trifluoromethyl-phenyl)-imidazole. We conclude that, in skeletal muscle, NO increases GLUT4 expression via a cGMP- and AMPK-dependent mechanism. The data are consistent with a role for NO in the regulation of AMPK, possibly via control of cellular activity of AMPK kinases and/or AMPK phosphatases.

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Reduced plasma FFA availability increases net triacylglycerol degradation, but not GPAT or HSL activity, in human skeletal muscle

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00542.2003 ◽

2004 ◽

Vol 287 (1) ◽

pp. E120-E127 ◽

Cited By ~ 64

Author(s):

Matthew J. Watt ◽

Anna G. Holmes ◽

Gregory R. Steinberg ◽

Jose L. Mesa ◽

Bruce E. Kemp ◽

...

Keyword(s):

Skeletal Muscle ◽

Adipose Tissue ◽

Fatty Acid ◽

Body Fat ◽

Plasma Free Fatty Acid ◽

Fat Oxidation ◽

Dry Mass ◽

Hormone Sensitive Lipase ◽

Whole Body ◽

Plasma Ffa

Intramuscular triacylglycerols (IMTG) are proposed to be an important metabolic substrate for contracting muscle, although this remains controversial. To test the hypothesis that reduced plasma free fatty acid (FFA) availability would increase IMTG degradation during exercise, seven active men cycled for 180 min at 60% peak pulmonary O2 uptake either without (CON) or with (NA) prior ingestion of nicotinic acid to suppress adipose tissue lipolysis. Skeletal muscle and adipose tissue biopsy samples were obtained before and at 90 and 180 min of exercise. NA ingestion decreased ( P < 0.05) plasma FFA at rest and completely suppressed the exercise-induced increase in plasma FFA (180 min: CON, 1.42 ± 0.07; NA, 0.10 ± 0.01 mM). The decreased plasma FFA during NA was associated with decreased ( P < 0.05) adipose tissue hormone-sensitive lipase (HSL) activity (CON: 13.9 ± 2.5, NA: 9.1 ± 3.0 nmol·min−1·mg protein−1). NA ingestion resulted in decreased whole body fat oxidation and increased carbohydrate oxidation. Despite the decreased whole body fat oxidation, net IMTG degradation was greater in NA compared with CON (net change: CON, 2.3 ± 0.8; NA, 6.3 ± 1.2 mmol/kg dry mass). The increased IMTG degradation did not appear to be due to reduced fatty acid esterification, because glycerol 3-phosphate activity was not different between trials and was unaffected by exercise (rest: 0.21 ± 0.07; 180 min: 0.17 ± 0.04 nmol·min−1·mg protein−1). HSL activity was not increased from resting rates during exercise in either trial despite elevated plasma epinephrine, decreased plasma insulin, and increased ERK1/2 phosphorylation. AMP-activated protein kinase (AMPK)α1 activity was not affected by exercise or NA, whereas AMPKα2 activity was increased ( P < 0.05) from rest during exercise in NA and was greater ( P < 0.05) than in CON at 180 min. These data suggest that plasma FFA availability is an important mediator of net IMTG degradation, and in the absence of plasma FFA, IMTG degradation cannot maintain total fat oxidation. These changes in IMTG degradation appear to disassociate, however, from the activity of the key enzymes responsible for synthesis and degradation of this substrate.

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Lack of adipose-specific hexose-6-phosphate dehydrogenase causes inactivation of adipose glucocorticoids and improves metabolic phenotype in mice

Clinical Science ◽

10.1042/cs20190679 ◽

2019 ◽

Vol 133 (21) ◽

pp. 2189-2202

Author(s):

Jian Wang ◽

Ying Wang ◽

Limei Liu ◽

Kabirullah Lutfy ◽

Theodore C. Friedman ◽

...

Keyword(s):

Adipose Tissue ◽

Fasting Blood Glucose ◽

Visceral Obesity ◽

Conditional Knockout ◽

Hormone Sensitive Lipase ◽

Metabolic Phenotype ◽

Mrna Levels ◽

Obesity And Diabetes ◽

Factor C

Abstract Excessive glucocorticoid (GC) production in adipose tissue promotes the development of visceral obesity and metabolic syndrome (MS). 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) is critical for controlling intracellular GC production, and this process is tightly regulated by hexose-6-phosphate dehydrogenase (H6PDH). To better understand the integrated molecular physiological effects of adipose H6PDH, we created a tissue-specific knockout of the H6PDH gene mouse model in adipocytes (adipocyte-specific conditional knockout of H6PDH (H6PDHAcKO) mice). H6PDHAcKO mice exhibited almost complete absence of H6PDH expression and decreased intra-adipose corticosterone production with a reduction in 11β-HSD1 activity in adipose tissue. These mice also had decreased abdominal fat mass, which was paralleled by decreased adipose lipogenic acetyl-CoA carboxylase (ACC) and ATP-citrate lyase (ACL) gene expression and reduction in their transcription factor C/EBPα mRNA levels. Moreover, H6PDHAcKO mice also had reduced fasting blood glucose levels, increased glucose tolerance, and increased insulin sensitivity. In addition, plasma free fatty acid (FFA) levels were decreased with a concomitant decrease in the expression of lipase adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) in adipose tissue. These results indicate that inactivation of adipocyte H6PDH expression is sufficient to cause intra-adipose GC inactivation that leads to a favorable pattern of metabolic phenotypes. These data suggest that H6PDHAcKO mice may provide a good model for studying the potential contributions of fat-specific H6PDH inhibition to improve the metabolic phenotype in vivo. Our study suggests that suppression or inactivation of H6PDH expression in adipocytes could be an effective intervention for treating obesity and diabetes.

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Myofibrillar Protein Abundance and Anabolic Signaling in Myotubes Depleted of E1 Alpha Subunit of Branched-Chain Ketoacid Dehydrogenase Complex

Current Developments in Nutrition ◽

10.1093/cdn/nzaa049_035 ◽

2020 ◽

Vol 4 (Supplement_2) ◽

pp. 642-642

Author(s):

Glory Madu ◽

Olasunkanmi Adegoke

Keyword(s):

Skeletal Muscle ◽

Amino Acids ◽

Protein Content ◽

Muscle Protein ◽

Essential Amino Acids ◽

Myofibrillar Protein ◽

L6 Myotubes ◽

Branched Chain ◽

Ketoacid Dehydrogenase

Abstract Objectives Branched-chain amino acids (BCAAs) are essential amino acids that are crucial for skeletal muscle anabolism. Thus, alterations in their levels are associated with muscle atrophic diseases such as cancer, chronic inflammatory and neurological disorders. Others have linked impairments in BCAA metabolism to the development of insulin resistance and its sequelae. Compared to the effects of theses amino acids, much less is known on how impairment in BCAA catabolism affects skeletal muscle. BCAA catabolism starts with the reversible transamination by the mitochondrial enzyme branched-chain aminotransferase 2 (BCAT2). This is followed by the irreversible carboxylation, catalyzed by branched-chain ketoacid dehydrogenase (BCKD) complex. We have shown that BCAT2 and BCKD are essential for the differentiation of skeletal myoblasts into myotubes. Here, we investigated the effect of depletion of BCAT2 or of E1a subunit of BCKD in differentiated myotubes. Methods On day 4 of differentiation, L6 myotubes were transfected with the following siRNA oligonucleotides: scrambled (control), BCAT2, or E1a subunit of BCKD. Results Forty-eight hours after transfection, compared to control or BCAT2 siRNA group, we observed improved myotube structure in BCKD-depleted cells. BCKD depletion augmented myofibrillar protein levels: myosin heavy chain (MHC, 2-fold) and tropomyosin (4-fold), P < 0.05, n = 3. To further analyze the increase in myofibrillar protein content, we examined signaling through mTORC1 (mechanistic target of rapamycin complex 1), a vital complex necessary for skeletal muscle anabolism. BCKD depletion increased the phosphorylation of mTORC1 upstream activator AKT (52%, P < 0.05, n = 3), and of mTORC1 downstream substrates by 25%-86%, consistent with the increase in myofibrillar proteins. Finally, in myotubes treated with the catabolic cytokine (tumor necrosis factor-a), BCKD depletion tended to increase the abundance of tropomyosin (a myofibrillar protein). Conclusions We showed that depletion of BCKD enhanced myofibrillar protein content and anabolic signaling. If these data are confirmed in vivo, development of dietary and other interventions that target BCKD abundance or functions may promote muscle protein anabolism in individuals with muscle wasting conditions. Funding Sources MHRC, NSERC York U.

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