Two weeks of metformin treatment induces AMPK-dependent enhancement of insulin-stimulated glucose uptake in mouse soleus muscle

Metformin-induced activation of the 5′-AMP-activated protein kinase (AMPK) has been associated with enhanced glucose uptake in skeletal muscle, but so far no direct causality has been examined. We hypothesized that an effect of in vivo metformin treatment on glucose uptake in mouse skeletal muscles is dependent on AMPK signaling. Oral doses of metformin or saline treatment were given to muscle-specific kinase dead (KD) AMPKα2 mice and wild-type (WT) littermates either once or chronically for 2 wk. Soleus and extensor digitorum longus muscles were used for measurements of glucose transport and Western blot analyses. Chronic treatment with metformin enhanced insulin-stimulated glucose uptake in soleus muscles of WT (∼45%, P < 0.01) but not of AMPK KD mice. Insulin signaling at the level of Akt protein expression or Thr308 and Ser473 phosphorylation was not changed by metformin treatment. Insulin signaling at the level of Akt and TBC1D4 protein expression as well as Akt Thr308/Ser473 and TBC1D4 Thr642/Ser711 phosphorylation were not changed by metformin treatment. Also, protein expressions of Rab4, GLUT4, and hexokinase II were unaltered after treatment. The acute metformin treatment did not affect glucose uptake in muscle of either of the genotypes. In conclusion, we provide novel evidence for a role of AMPK in potentiating the effect of insulin on glucose uptake in soleus muscle in response to chronic metformin treatment.

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Reciprocity between Skeletal Muscle AMPK Deletion and Insulin Action in Diet-Induced Obese Mice

10.2337/figshare.12328136 ◽

2020 ◽

Author(s):

Ada Admin ◽

Louise Lantier ◽

Ashley S. Williams ◽

Ian M.Williams ◽

Amanda Guerin ◽

...

Keyword(s):

Skeletal Muscle ◽

Insulin Signaling ◽

Insulin Action ◽

Energy State ◽

Energy Charge ◽

Ampk Signaling ◽

Diet Induced Obesity ◽

Energy Sensor ◽

Amp Activated Protein Kinase

Insulin resistance due to overnutrition places a burden on energy-producing pathways in skeletal muscle (SkM). Nevertheless, energy state is not compromised. The hypothesis that the energy sensor AMP-activated protein kinase (AMPK) is necessary to offset the metabolic burden of overnutrition was tested using chow-fed and high fat (HF)-fed SkM-specific AMPKa1a2 knockout (mdKO) mice and AMPKa1a2lox/lox littermates (WT). Lean mdKO and WT mice were phenotypically similar. HF-fed mice were equally obese and maintained lean mass regardless of genotype. Results did not support the hypothesis that AMPK is protective during overnutrition. Paradoxically, mdKO mice were more insulin sensitive. Insulin-stimulated SkM glucose uptake was ~two-fold greater in mdKO mice in vivo. Furthermore, insulin signaling, SkM GLUT4 translocation, hexokinase activity, and glycolysis were increased. AMPK and insulin signaling intersect at mTOR, a critical node for cell proliferation and survival. Basal mTOR activation was reduced by 50% in HF-fed mdKO mice, but was normalized by insulin-stimulation. Mitochondrial function was impaired in mdKO mice, but energy charge was preserved by AMP deamination. Results show a surprising reciprocity between SkM AMPK signaling and insulin action that manifests with diet-induced obesity, as insulin action is preserved to protect fundamental energetic processes in the muscle.

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Reciprocity between Skeletal Muscle AMPK Deletion and Insulin Action in Diet-Induced Obese Mice

10.2337/figshare.12328136.v1 ◽

2020 ◽

Author(s):

Ada Admin ◽

Louise Lantier ◽

Ashley S. Williams ◽

Ian M.Williams ◽

Amanda Guerin ◽

...

Keyword(s):

Skeletal Muscle ◽

Insulin Signaling ◽

Insulin Action ◽

Energy State ◽

Energy Charge ◽

Ampk Signaling ◽

Diet Induced Obesity ◽

Energy Sensor ◽

Amp Activated Protein Kinase

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Insulin Signaling and Glucose Uptake in the Soleus Muscle of 30-Month-Old Rats After Calorie Restriction With or Without Acute Exercise

The Journals of Gerontology Series A ◽

10.1093/gerona/glv142 ◽

2015 ◽

Vol 71 (3) ◽

pp. 323-332 ◽

Cited By ~ 10

Author(s):

Haiyan Wang ◽

Naveen Sharma ◽

Edward B. Arias ◽

Gregory D. Cartee

Keyword(s):

Glucose Uptake ◽

Calorie Restriction ◽

Soleus Muscle ◽

Insulin Signaling ◽

Acute Exercise ◽

Old Rats

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α-Hederin Inhibits the Growth of Lung Cancer A549 Cells in vitro and in vivo by Increasing SIRT6 Dependent Glycolysis

10.21203/rs.3.rs-36418/v1 ◽

2020 ◽

Author(s):

cong fang ◽

Yahui Liu ◽

Lanying Chen ◽

Yingying Luo ◽

Yaru Cui ◽

...

Keyword(s):

Lung Cancer ◽

Mouse Model ◽

Protein Expression ◽

Glucose Uptake ◽

A549 Cells ◽

Lactate Production ◽

Tumor Xenograft ◽

Xenograft Mouse Model ◽

Atp Generation

Abstract Background: α-hederin an effective component of Pulsatilla chinensis (Bunge) Regel, Studies showed that α-hederin exert many pharmacological activities, However, the effect of α-hederin on metabolism is still unclear. This study aimed to illuminate the role of α-hederin in glucose metabolism in lung cancer cells and investigate the molecular mechanism of α-hederin. Methods: CCK8 and colony formation assays were employed to assess the anti-proliferative effects induced by α-hederin. Glucose uptake, ATP generation, and reduced lactate production were measured using kits, and an A549 tumor xenograft mouse model of lung cancer was used to assess the in vivo antitumor effect of α-hederin (5, 10 mg/kg). Glycolytic-related key enzymes hexokinase 2 (HK2), glucose transporters 1 (GLUT1), pyruvate kinase M2 (PKM2), lactate dehydrogenase A (LDHA), monocarboxylate transporter (MCT4), c-Myc, Hypoxia inducible factor-1α (HIF-1α) and Sirtuin 6 (SIRT6) protein expression were detected by western blotting and immunohistochemical staining and SIRT6 inhibitors was verified in A549 cells. Results: Our results showed that cell proliferation was significantly inhibited by α-hederin in a dose-dependent manner and that α-hederin inhibited glucose uptake and ATP generation and reduced lactate production. Furthermore, α-hederin remarkably inhibited HK2, GLUT1, PKM2, LDHA, MCT4, c-Myc, HIF-1α and activated SIRT6 protein expression. Using inhibitors, we proved that α-hederin inhibits glycolysis by activating SIRT6. Moreover, a tumor xenograft mouse model of lung cancer further confirmed that α-hederin inhibits lung cancer growth via inhibiting glycolysis in vivo. Conclusions: α-hederin inhibits the growth of non-small cell lung cancer A549 cells by inhibiting glycolysis. The mechanism of glycolysis inhibition includes α-hederin activating the expression of the glycolytic related protein SIRT6.

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Abstract 344: FNDC5, a PGC1-a-Dependent Myokine, Increases Glucose Utilization and Fatty-Acid Oxidation by AMPK Signaling Pathway

Circulation Research ◽

10.1161/res.113.suppl_1.a344 ◽

2013 ◽

Vol 113 (suppl_1) ◽

Author(s):

Ling Tao ◽

Yi Liu ◽

Chao Xin ◽

Weidong Huang ◽

Lijian Zhang ◽

...

Keyword(s):

Fatty Acid ◽

Glucose Uptake ◽

Fatty Acid Oxidation ◽

Glucose Utilization ◽

C2c12 Cells ◽

Time Dependent ◽

Acid Oxidation ◽

Ampk Signaling

FNDC5 is a hormone secreted by myocytes that could reduce obesity and insulin resistance, However, the exact effect of FNDC5 on glucose and lipid metabolism remain poorly identified; More importantly, the signaling pathways that mediate the metabolic effects of FNDC5 is completely unknown. Here we showed that FNDC5 stimulates β-oxidation and glucose uptake in C2C12 cells in a dose- and time-dependent fashion in vitro (n=8, all P<0.01). In vivo study revealed that FNDC5 also enhanced glucose tolerance in diabetic mice and increased the glucose uptake evidenced by increased [18F] FDG accumulation in hearts by PET scan (n=6, all P<0.05). FNDC5 decreased the expression of gluconeogenesis related molecules (PEPCK and G6Pase) and increased the phosphorylation of ACC, a key modulator of fatty-acid oxidation, both in hepatocytes and C2C12 cells (n=3, all P<0.05). In parallel with its stimulation of β-oxidation and glucose uptake, FNDC5 increased the phosphorylation of AMPK both in hepatocytes and C2C12 cells in a dose- and time-dependent fashion in vitro and in vivo. More importantly, the β-oxidation and glucose uptake, the expression of PEPCK and G6Pase and the phosphorylation of ACC induced by FNDC5 were attenuated by AMPK inhibitor in hepatocytes and C2C12 cells (P<0.05). Most importantly, the FNDC5 induced glucose uptake and phosphorylation of ACC were attenuated in AMPK-DN mice (n=6, all P<0.05). The glucose-lowering effect of FNDC5 in diabetic mice was also attenuated by AMPK inhibitor. Our data presents the direct evidence that FNDC5 stimulates glucose utilization and fatty-acid oxidation by AMPK signaling pathway, suggesting that FNDC5 be a novel pharmacological approach for type 2 diabetes.

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Overexpression of SH2-Containing Inositol Phosphatase 2 Results in Negative Regulation of Insulin-Induced Metabolic Actions in 3T3-L1 Adipocytes via Its 5′-Phosphatase Catalytic Activity

Molecular and Cellular Biology ◽

10.1128/mcb.21.5.1633-1646.2001 ◽

2001 ◽

Vol 21 (5) ◽

pp. 1633-1646 ◽

Cited By ~ 134

Author(s):

Tsutomu Wada ◽

Toshiyasu Sasaoka ◽

Makoto Funaki ◽

Hiroyuki Hori ◽

Shihou Murakami ◽

...

Keyword(s):

Protein Kinase ◽

Glucose Uptake ◽

Phosphatase Activity ◽

Insulin Signaling ◽

Glycogen Synthase ◽

Glycogen Synthesis ◽

Dominant Negative ◽

Inositol Phosphatase

ABSTRACT Phosphatidylinositol (PI) 3-kinase plays an important role in various metabolic actions of insulin including glucose uptake and glycogen synthesis. Although PI 3-kinase primarily functions as a lipid kinase which preferentially phosphorylates the D-3 position of phospholipids, the effect of hydrolysis of the key PI 3-kinase product PI 3,4,5-triphosphate [PI(3,4,5)P3] on these biological responses is unknown. We recently cloned rat SH2-containing inositol phosphatase 2 (SHIP2) cDNA which possesses the 5′-phosphatase activity to hydrolyze PI(3,4,5)P3 to PI 3,4-bisphosphate [PI(3,4)P2] and which is mainly expressed in the target tissues of insulin. To study the role of SHIP2 in insulin signaling, wild-type SHIP2 (WT-SHIP2) and 5′-phosphatase-defective SHIP2 (ΔIP-SHIP2) were overexpressed in 3T3-L1 adipocytes by means of adenovirus-mediated gene transfer. Early events of insulin signaling including insulin-induced tyrosine phosphorylation of the insulin receptor β subunit and IRS-1, IRS-1 association with the p85 subunit, and PI 3-kinase activity were not affected by expression of either WT-SHIP2 or ΔIP-SHIP2. Because WT-SHIP2 possesses the 5′-phosphatase catalytic region, its overexpression marked by decreased insulin-induced PI(3,4,5)P3 production, as expected. In contrast, the amount of PI(3,4,5)P3 was increased by the expression of ΔIP-SHIP2, indicating that ΔIP-SHIP2 functions in a dominant-negative manner in 3T3-L1 adipocytes. Both PI(3,4,5)P3 and PI(3,4)P2 were known to possibly activate downstream targets Akt and protein kinase Cλ in vitro. Importantly, expression of WT-SHIP2 inhibited insulin-induced activation of Akt and protein kinase Cλ, whereas these activations were increased by expression of ΔIP-SHIP2 in vivo. Consistent with the regulation of downstream molecules of PI 3-kinase, insulin-induced 2-deoxyglucose uptake and Glut4 translocation were decreased by expression of WT-SHIP2 and increased by expression of ΔIP-SHIP2. In addition, insulin-induced phosphorylation of GSK-3β and activation of PP1 followed by activation of glycogen synthase and glycogen synthesis were decreased by expression of WT-SHIP2 and increased by the expression of ΔIP-SHIP2. These results indicate that SHIP2 negatively regulates metabolic signaling of insulin via the 5′-phosphatase activity and that PI(3,4,5)P3 rather than PI(3,4)P2 is important for in vivo regulation of insulin-induced activation of downstream molecules of PI 3-kinase leading to glucose uptake and glycogen synthesis.

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Maternal Obesity and Western-style Diet Impair Fetal and Juvenile Offspring Skeletal Muscle Insulin-Stimulated Glucose Transport in Nonhuman Primates

10.1101/864082 ◽

2019 ◽

Author(s):

William Campodonico-Burnett ◽

Byron Hetrick ◽

Stephanie R. Wesolowski ◽

Simon Schenk ◽

Diana L. Takahashi ◽

...

Keyword(s):

Glucose Uptake ◽

Insulin Signaling ◽

Maternal Obesity ◽

Metabolic Diseases ◽

Ex Vivo ◽

Primate Model ◽

Intravenous Insulin ◽

Pregnancy And Lactation ◽

Signaling Activation

AbstractInfants born to mothers with obesity have a greater risk for childhood obesity and metabolic diseases; however, the underlying biological mechanisms remain poorly understood. We used a nonhuman primate model to investigate whether maternal obesity combined with a western-style diet (WSD) impairs offspring muscle insulin action. Briefly, adult females were fed a control (CON) or WSD prior to and during pregnancy and lactation. Offspring were weaned to a CON or WSD. Muscle glucose uptake and insulin signaling were measured ex vivo in fetal and juvenile offspring. In vivo signaling was evaluated before and after an intravenous insulin bolus just prior to weaning. We find that fetal muscle exposed to maternal WSD had reduced insulin-stimulated glucose uptake and impaired insulin signaling. In juvenile offspring, insulin-stimulated glucose uptake was similarly reduced by both maternal and post-weaning WSD. Analysis of insulin signaling activation revealed distinct changes between fetal and post-weaning WSD exposure. We conclude that maternal WSD leads to a persistent decrease in insulin-stimulated glucose uptake in juvenile offspring even in the absence of increased offspring adiposity or markers of systemic insulin resistance. Switching offspring to a healthy diet did not ameliorate the effects of maternal WSD suggesting earlier interventions may be necessary.

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A conserved role for AMP-activated protein kinase in NGLY1 deficiency

PLoS Genetics ◽

10.1371/journal.pgen.1009258 ◽

2020 ◽

Vol 16 (12) ◽

pp. e1009258

Author(s):

Seung Yeop Han ◽

Ashutosh Pandey ◽

Tereza Moore ◽

Antonio Galeone ◽

Lita Duraine ◽

...

Keyword(s):

Energy Metabolism ◽

Protein Kinase ◽

Global Developmental Delay ◽

Genetic Studies ◽

Ampk Signaling ◽

Biochemical Assays ◽

Severe Reduction ◽

Amp Activated Protein Kinase ◽

Impaired Energy Metabolism

Mutations in human N-glycanase 1 (NGLY1) cause the first known congenital disorder of deglycosylation (CDDG). Patients with this rare disease, which is also known as NGLY1 deficiency, exhibit global developmental delay and other phenotypes including neuropathy, movement disorder, and constipation. NGLY1 is known to regulate proteasomal and mitophagy gene expression through activation of a transcription factor called "nuclear factor erythroid 2-like 1" (NFE2L1). Loss of NGLY1 has also been shown to impair energy metabolism, but the molecular basis for this phenotype and its in vivo consequences are not well understood. Using a combination of genetic studies, imaging, and biochemical assays, here we report that loss of NGLY1 in the visceral muscle of the Drosophila larval intestine results in a severe reduction in the level of AMP-activated protein kinase α (AMPKα), leading to energy metabolism defects, impaired gut peristalsis, failure to empty the gut, and animal lethality. Ngly1–/– mouse embryonic fibroblasts and NGLY1 deficiency patient fibroblasts also show reduced AMPKα levels. Moreover, pharmacological activation of AMPK signaling significantly suppressed the energy metabolism defects in these cells. Importantly, the reduced AMPKα level and impaired energy metabolism observed in NGLY1 deficiency models are not caused by the loss of NFE2L1 activity. Taken together, these observations identify reduced AMPK signaling as a conserved mediator of energy metabolism defects in NGLY1 deficiency and suggest AMPK signaling as a therapeutic target in this disease.

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AMP-activated protein kinase activates transcription of the UCP3 and HKII genes in rat skeletal muscle

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00278.2002 ◽

2002 ◽

Vol 283 (6) ◽

pp. E1239-E1248 ◽

Cited By ~ 64

Author(s):

James Stoppani ◽

Audrey L. Hildebrandt ◽

Kei Sakamoto ◽

David Cameron-Smith ◽

Laurie J. Goodyear ◽

...

Keyword(s):

Skeletal Muscle ◽

Blood Glucose ◽

Protein Kinase ◽

Energy Demand ◽

Uncoupling Protein ◽

Hexokinase Ii ◽

Arterial Infusion ◽

Ampk Signaling ◽

Lactate Levels ◽

Amp Activated Protein Kinase

AMP-activated protein kinase (AMPK) has recently emerged as a key signaling protein in skeletal muscle, coordinating the activation of both glucose and fatty acid metabolism in response to increased cellular energy demand. To determine whether AMPK signaling may also regulate gene transcription in muscle, rats were given a single subcutaneous injection (1 mg/g) of the AMP analog 5-aminoimidazole-4-carboxamide-1-β-d-ribonucleoside (AICAR). AICAR injection activated ( P < 0.05) AMPK-α2 (∼2.5-fold) and transcription of the uncoupling protein-3 (UCP3, ∼4-fold) and hexokinase II (HKII, ∼10-fold) genes in both red and white skeletal muscle. However, AICAR injection also elicited ( P < 0.05) an acute drop (60%) in blood glucose and a sustained (2-h) increase in blood lactate, prompting concern regarding the specificity of AICAR on transcription. To maximize AMPK activation in muscle while minimizing potential systemic counterregulatory responses, a single-leg arterial infusion technique was employed in fully conscious rats. Relative to saline-infused controls, single-leg arterial infusion of AICAR (0.125, 0.5, and 2.5 μg · g−1 · min−1for 60 min) induced a dose-dependent increase (2- to 4-fold, P < 0.05) in UCP3 and HKII transcription in both red and white skeletal muscle. Importantly, AICAR infusion activated transcription only in muscle from the infused leg and had no effect on blood glucose or lactate levels. These data provide evidence that AMPK signaling is linked to the transcriptional regulation of select metabolic genes in skeletal muscle.

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Local hindlimb antioxidant infusion does not affect muscle glucose uptake during in situ contractions in rat

Journal of Applied Physiology ◽

10.1152/japplphysiol.01335.2009 ◽

2010 ◽

Vol 108 (5) ◽

pp. 1275-1283 ◽

Cited By ~ 18

Author(s):

T. L. Merry ◽

R. M. Dywer ◽

E. A. Bradley ◽

S. Rattigan ◽

G. K. McConell

Keyword(s):

Skeletal Muscle ◽

Glucose Uptake ◽

Ex Vivo ◽

Moderate Intensity ◽

Ampk Signaling ◽

Epigastric Artery ◽

Hindlimb Muscle ◽

Skeletal Muscle Glucose Uptake ◽

Amp Activated Protein Kinase

There is evidence that reactive oxygen species (ROS) contribute to the regulation of skeletal muscle glucose uptake during highly fatiguing ex vivo contraction conditions via AMP-activated protein kinase (AMPK). In this study we investigated the role of ROS in the regulation of glucose uptake and AMPK signaling during low-moderate intensity in situ hindlimb muscle contractions in rats, which is a more physiological protocol and preparation. Male hooded Wistar rats were anesthetized, and then N-acetylcysteine (NAC) was infused into the epigastric artery (125 mg·kg−1·h−1) of one hindlimb (contracted leg) for 15 min before this leg was electrically stimulated (0.1-ms impulse at 2 Hz and 35 V) to contract at a low-moderate intensity for 15 min. The contralateral leg did not receive stimulation or local NAC infusion (rest leg). NAC infusion increased ( P < 0.05) plasma cysteine and cystine (by ∼360- and 1.4-fold, respectively) and muscle cysteine (by 1.5-fold, P = 0.001). Although contraction did not significantly alter muscle tyrosine nitration, reduced (GSH) or oxidized glutathione (GSSG) content, S-glutathionylation of protein bands at ∼250 and 150 kDa was increased ( P < 0.05) ∼1.7-fold by contraction, and this increase was prevented by NAC. Contraction increased ( P < 0.05) skeletal muscle glucose uptake 20-fold, AMPK phosphorylation 6-fold, ACCβ phosphorylation 10-fold, and p38 MAPK phosphorylation 60-fold, and the muscle fatigued by ∼30% during contraction and NAC infusion had no significant effect on any of these responses. This was despite NAC preventing increases in S-glutathionylation with contraction. In conclusion, unlike during highly fatiguing ex vivo contractions, local NAC infusion during in situ low-moderate intensity hindlimb contractions in rats, a more physiological preparation, does not attenuate increases in skeletal muscle glucose uptake or AMPK signaling.

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