Isoflurane activates AMP-activated protein kinase to inhibit proliferation, and promote apoptosis and autophagy in cervical carcinoma both in vitro and in vivo

The hypothesis was tested that hypoxia increases AMP-activated protein kinase (AMPK) activity independently of AMP concentration ([AMP]) in heart. In isolated perfused rat hearts, cytosolic [AMP] was changed from 0.2 to 16 μM using metabolic inhibitors during both normal oxygenation (95% O2-5% CO2, normoxia) and limited oxygenation (95% N2-5% CO2, hypoxia). Total AMPK activity measured in vitro ranged from 2 to 40 pmol·min−1·mg protein−1 in normoxic hearts and from 5 to 55 pmol·min−1·mg protein−1 in hypoxic hearts. The dependence of the in vitro total AMPK activity on the in vivo cytosolic [AMP] was determined by fitting the measurements from individual hearts to a hyperbolic equation. The [AMP] resulting in half-maximal total AMPK activity ( A0.5) was 3 ± 1 μM for hypoxic hearts and 28 ± 13 μM for normoxic hearts. The A0.5 for α2-isoform AMPK activity was 2 ± 1 μM for hypoxic hearts and 13 ± 8 μM for normoxic hearts. Total AMPK activity correlated with the phosphorylation of the Thr172 residue of the AMPK α-subunit. In potassium-arrested hearts perfused with variable O2 content, α-subunit Thr172 phosphorylation increased at O2 ≤ 21% even though [AMP] was <0.3 μM. Thus hypoxia or O2 ≤ 21% increased AMPK phosphorylation and activity independently of cytosolic [AMP]. The hypoxic increase in AMPK activity may result from either direct phosphorylation of Thr172 by an upstream kinase or reduction in the A0.5 for [AMP].

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5′-AMP-activated protein kinase is inactivated by adrenergic signalling in adult cardiac myocytes

Bioscience Reports ◽

10.1042/bsr20110076 ◽

2011 ◽

Vol 32 (2) ◽

pp. 197-209 ◽

Cited By ~ 10

Author(s):

Yugo Tsuchiya ◽

Fiona C. Denison ◽

Richard B. Heath ◽

David Carling ◽

David Saggerson

Keyword(s):

Protein Kinase ◽

Cardiac Myocytes ◽

Calcium Signalling ◽

Metabolic Stress ◽

Protein Kinase Activity ◽

Adult Rat ◽

Protein Kinase C Inhibitor ◽

Amp Activated Protein Kinase

In adult rat cardiac myocytes adrenaline decreased AMPK (AMP-activated protein kinase) activity with a half-time of approximately 4 min, decreased phosphorylation of AMPK (α-Thr172) and decreased phosphorylation of ACC (acetyl-CoA carboxylase). Inactivation of AMPK by adrenaline was through both α1- and β-ARs (adrenergic receptors), but did not involve cAMP or calcium signalling, was not blocked by the PKC (protein kinase C) inhibitor BIM I (bisindoylmaleimide I), by the ERK (extracellular-signal-regulated kinase) cascade inhibitor U0126 or by PTX (pertussis toxin). Adrenaline caused no measurable change in LKB1 activity. Adrenaline decreased AMPK activity through a process that was distinct from AMPK inactivation in response to insulin or PMA. Neither adrenaline nor PMA altered the myocyte AMP:ATP ratio although the adrenaline effect was attenuated by oligomycin and by AICAR (5-amino-4-imidazolecarboxamide-1-β-D-ribofuranoside), agents that mimic ‘metabolic stress’. Inactivation of AMPK by adrenaline was abolished by 1 μM okadaic acid suggesting that activation of PP2A (phosphoprotein phosphatase 2A) might mediate the adrenaline effect. However, no change in PP2A activity was detected in myocyte extracts. Adrenaline increased phosphorylation of the AMPK β-subunit in vitro but there was no detectable change in vivo in phosphorylation of previously identified AMPK sites (β-Ser24, β-Ser108 or β-Ser182) suggesting that another site(s) is targeted.

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Statins Activate AMP-Activated Protein Kinase In Vitro and In Vivo

Circulation ◽

10.1161/circulationaha.106.630194 ◽

2006 ◽

Vol 114 (24) ◽

pp. 2655-2662 ◽

Cited By ~ 183

Author(s):

Wei Sun ◽

Tzong-Shyuan Lee ◽

Minjia Zhu ◽

Chunang Gu ◽

Yinsheng Wang ◽

...

Keyword(s):

Protein Kinase ◽

Amp Activated Protein Kinase

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Adiponectin promotes VEGF-C-dependent lymphangiogenesis by inhibiting miR-27b through a CaMKII/AMPK/p38 signaling pathway in human chondrosarcoma cells

Clinical Science ◽

10.1042/cs20160117 ◽

2016 ◽

Vol 130 (17) ◽

pp. 1523-1533 ◽

Cited By ~ 20

Author(s):

Chun-Yin Huang ◽

An-Chen Chang ◽

Hsien-Te Chen ◽

Shih-Wei Wang ◽

Yuan-Shun Lo ◽

...

Keyword(s):

Protein Kinase ◽

Signaling Pathway ◽

Tube Formation ◽

Human Chondrosarcoma ◽

Dependent Protein ◽

And Migration ◽

Factor C ◽

Amp Activated Protein Kinase

Chondrosarcoma is the second most frequently occurring type of bone malignancy characterized by distant metastatic propensity. Vascular endothelial growth factor-C (VEGF-C) is the major lymphangiogenic factor, and makes crucial contributions to tumour lymphangiogenesis and lymphatic metastasis. Adiponectin is a protein hormone secreted predominantly by differentiated adipocytes. In recent years, adiponectin has also been indicated as facilitating tumorigenesis, angiogenesis and metastasis. However, the effect of adiponectin on VEGF-C regulation and lymphangiogenesis in chondrosarcoma has remained largely a mystery. In the present study, we have shown a clinical correlation between adiponectin and VEGF-C, as well as tumour stage, in human chondrosarcoma tissues. We further demonstrated that adiponectin promoted VEGF-C expression and secretion in human chondrosarcoma cells. The conditioned medium from adiponectin-treated cells significantly induced tube formation and migration of human lymphatic endothelial cells. In addition, adiponectin knock down inhibited lymphangiogenesis in vitro and in vivo. We also found that adiponectin-induced VEGF-C is mediated by the calmodulin-dependent protein kinase II (CaMKII), AMP-activated protein kinase (AMPK) and p38 signaling pathway. Furthermore, the expression of miR-27b was negatively regulated by adiponectin via the CaMKII, AMPK and p38 cascade. The present study is the first to describe the mechanism of adiponectin-promoted lymphangiogenesis by up-regulating VEGF-C expression in chondrosarcomas. Thus, adiponectin could serve as a therapeutic target in chondrosarcoma metastasis and lymphangiogenesis.

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AMP-activated protein kinase (AMPK) regulates in vitro bone formation and bone mass in vivo

Bone ◽

10.1016/j.bone.2010.04.079 ◽

2010 ◽

Vol 47 ◽

pp. S44

Author(s):

M. Shah⁎ ◽

A. Bataveljic ◽

T.R. Arnett ◽

B. Viollet ◽

L.K. Saxon ◽

...

Keyword(s):

Protein Kinase ◽

Bone Formation ◽

Bone Mass ◽

Amp Activated Protein Kinase

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Dissecting the Role of 5′-AMP for Allosteric Stimulation, Activation, and Deactivation of AMP-activated Protein Kinase

Journal of Biological Chemistry ◽

10.1074/jbc.m606357200 ◽

2006 ◽

Vol 281 (43) ◽

pp. 32207-32216 ◽

Cited By ~ 309

Author(s):

Marianne Suter ◽

Uwe Riek ◽

Roland Tuerk ◽

Uwe Schlattner ◽

Theo Wallimann ◽

...

Keyword(s):

Protein Kinase ◽

Protein Phosphatase ◽

Energy Homeostasis ◽

Adenine Nucleotides ◽

Enzyme Preparations ◽

Activity Measurements ◽

Ampk Activity ◽

Amp Activated Protein Kinase

AMP-activated protein kinase (AMPK) is a heterotrimeric protein kinase that is crucial for cellular energy homeostasis of eukaryotic cells and organisms. Here we report on the activation of AMPK α1β1γ1 and α2β2γ1 by their upstream kinases (Ca2+/calmodulin-dependent protein kinase kinase-β and LKB1-MO25α-STRADα), the deactivation by protein phosphatase 2Cα, and on the extent of stimulation of AMPK by its allosteric activator AMP, using purified recombinant enzyme preparations. An accurate high pressure liquid chromatography-based method for AMPK activity measurements was established, which allowed for direct quantitation of the unphosphorylated and phosphorylated artificial peptide substrate, as well as the adenine nucleotides. Our results show a 1000-fold activation of AMPK by the combined effects of upstream kinase and saturating concentrations of AMP. The two AMPK isoforms exhibit similar specific activities (6 μmol/min/mg) and do not differ significantly by their responsiveness to AMP. Due to the inherent instability of ATP and ADP, it proved impossible to assay AMPK activity in the absolute absence of AMP. However, the half-maximal stimulatory effect of AMP is reached below 2 μm. AMP does not appear to augment phosphorylation by upstream kinases in the purified in vitro system, but deactivation by dephosphorylation of AMPK α-subunits at Thr-172 by protein phosphatase 2Cα is attenuated by AMP. Furthermore, it is shown that neither purified NAD+ nor NADH alters the activity of AMPK in a concentration range of 0–300 μm, respectively. Finally, evidence is provided that ZMP, a compound formed in 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside-treated cells to activate AMPK in vivo, allosterically activates purified AMPK in vitro, but compared with AMP, maximal activity is not reached. These data shed new light on physiologically important aspects of AMPK regulation.

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Effects of AMP-Activated Protein Kinase in Cerebral Ischemia

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.2009.255 ◽

2009 ◽

Vol 30 (3) ◽

pp. 480-492 ◽

Cited By ~ 104

Author(s):

Jun Li ◽

Louise D McCullough

Keyword(s):

Protein Kinase ◽

In Vivo Studies ◽

Ampk Activation ◽

Vigorous Exercise ◽

Nitric Oxide Signaling ◽

Reductase Inhibitors ◽

Amp Activated Protein Kinase ◽

The Brain

AMP-activated protein kinase (AMPK) is a serine threonine kinase that is highly conserved through evolution. AMPK is found in most mammalian tissues including the brain. As a key metabolic and stress sensor/effector, AMPK is activated under conditions of nutrient deprivation, vigorous exercise, or heat shock. However, it is becoming increasingly recognized that changes in AMPK activation not only signal unmet metabolic needs, but also are involved in sensing and responding to ‘cell stress’, including ischemia. The downstream effect of AMPK activation is dependent on many factors, including the severity of the stressor as well as the tissue examined. This review discusses recent in vitro and in vivo studies performed in the brain/neuronal cells and vasculature that have contributed to our understanding of AMPK in stroke. Recent data on the potential role of AMPK in angiogenesis and neurogenesis and the interaction of AMPK with 3-hydroxy-3-methy-glutaryl-CoA reductase inhibitors (statins) agents are highlighted. The interaction between AMPK and nitric oxide signaling is also discussed.

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Docosahexaenoic Acid Lessens Hepatic Lipid Accumulation and Inflammation via the AMP-Activated Protein Kinase and Endoplasmic Reticulum Stress Signaling Pathway in Grass Carp (Ctenopharyngodon Idella)

10.21203/rs.3.rs-845760/v1 ◽

2021 ◽

Author(s):

Xiaocheng Huang ◽

Jian Sun ◽

Chenchen Bian ◽

Shanghong Ji ◽

Hong ji

Keyword(s):

Endoplasmic Reticulum ◽

Protein Kinase ◽

Hepatic Steatosis ◽

Grass Carp ◽

Lipid Accumulation ◽

Ctenopharyngodon Idella ◽

Hepatic Inflammation ◽

Amp Activated Protein Kinase

Abstract Background: The liver is the primary organ for frontline immune defense and lipid metabolism. Excessive lipid accumulation in the liver severely affects its metabolic homeostasis and causes metabolic diseases. Docosahexaenoic acid (DHA) is known for its beneficial effects on lipid metabolism and anti-inflammation, but its molecular mechanism remains unknown, especially in fish. In this study, we evaluated the protective effects of DHA on hepatic steatosis of grass carp (Ctenopharyngodon idella) in vivo and in vitro and mainly focused on lipogenesis and inflammation. Grass carp were fed with purified diets supplemented with 0%, 0.5% and 1% DHA for 8 weeks in vivo. Hepatocytes were treated with palmitic acid (PA) (200 μM) with or without DHA (50 or 100 μM) for 24 h in vitro. In addition, Compound C (CC, the inhibitor of AMP-activated protein kinase) was used to examine the mechanism of DHA on hepatic steatosis in vitro.Results: In this study, 1% DHA significantly decreased the liver triglyceride (TG), malondialdehyde (MDA), serum tumor necrosis factor α (TNFα) and nuclear factor kappa B (NFκB) contents. DHA (100 μM) effectively attenuated PA-induced lipid accumulation (P<0.05). Furthermore, DHA significantly inhibited endoplasmic reticulum (ER) stress and stimulated the expression of AMP-activated protein kinase (AMPK) and its downstream factors related to hepatic inflammation and lipogenesis in vivo and in vitro. However, the effects of DHA could be abrogated by CC in vitro.Conclusions: DHA exerted a protective effect on hepatic steatosis by inhibiting ER stress, improving antioxidant ability, relieving hepatic inflammation and inhibiting hepatic lipogenesis in an AMPK-dependent manner. Our findings give a theoretical foundation for further elucidation of the beneficial role of DHA in vertebrates.

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AMP-activated protein kinase phosphorylation of the R domain inhibits PKA stimulation of CFTR

AJP Cell Physiology ◽

10.1152/ajpcell.00677.2008 ◽

2009 ◽

Vol 297 (1) ◽

pp. C94-C101 ◽

Cited By ~ 51

Author(s):

J Darwin King ◽

Adam C. Fitch ◽

Jeffrey K. Lee ◽

Jill E. McCane ◽

Don-On Daniel Mak ◽

...

Keyword(s):

Protein Kinase ◽

Dominant Negative ◽

Ampk Phosphorylation ◽

Ampk Activity ◽

Amp Activated Protein Kinase ◽

Stimulation Of ◽

In Cells ◽

R Domain

The metabolic sensor AMP-activated protein kinase (AMPK) has emerged as an important link between cellular metabolic status and ion transport activity. We previously found that AMPK binds to and phosphorylates CFTR in vitro and inhibits PKA-dependent stimulation of CFTR channel gating in Calu-3 bronchial serous gland epithelial cells. To further characterize the mechanism of AMPK-dependent regulation of CFTR, whole cell patch-clamp measurements were performed with PKA activation in Calu-3 cells expressing either constitutively active or dominant-negative AMPK mutants (AMPK-CA or AMPK-DN). Baseline CFTR conductance in cells expressing AMPK-DN was substantially greater than controls, suggesting that tonic AMPK activity in these cells inhibits CFTR under basal conditions. Although baseline CFTR conductance in cells expressing AMPK-CA was comparable to that of controls, PKA stimulation of CFTR was completely blocked in AMPK-CA-expressing cells, suggesting that AMPK activation renders CFTR resistant to PKA activation in vivo. Phosphorylation studies of CFTR in human embryonic kidney-293 cells using tetracycline-inducible expression of AMPK-DN demonstrated AMPK-dependent phosphorylation of CFTR in vivo. However, AMPK activity modulation had no effect on CFTR in vivo phosphorylation in response to graded doses of PKA or PKC agonists. Thus, AMPK-dependent CFTR phosphorylation renders the channel resistant to activation by PKA and PKC without preventing phosphorylation by these kinases. We found that Ser768, a CFTR R domain residue considered to be an inhibitory PKA site, is the dominant site of AMPK phosphorylation in vitro. Ser-to-Ala mutation at this site enhanced baseline CFTR activity and rendered CFTR resistant to inhibition by AMPK, suggesting that AMPK phosphorylation at Ser768 is required for its inhibition of CFTR. In summary, our findings indicate that AMPK-dependent phosphorylation of CFTR inhibits CFTR activation by PKA, thereby tuning the PKA-responsiveness of CFTR to metabolic and other stresses in the cell.

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A novel physical and functional association between nucleoside diphosphate kinase A and AMP-activated protein kinase α1 in liver and lung

Biochemical Journal ◽

10.1042/bj20050269 ◽

2005 ◽

Vol 392 (1) ◽

pp. 201-209 ◽

Cited By ~ 20

Author(s):

Russell M. Crawford ◽

Kate J. Treharne ◽

O. Giles Best ◽

Richmond Muimo ◽

Claudia E. Riemen ◽

...

Keyword(s):

Protein Kinase ◽

Mammalian Cells ◽

Nucleoside Diphosphate Kinase ◽

Cell Types ◽

Acid Synthesis ◽

Cellular Fatty Acid ◽

Energy Status ◽

Amp Activated Protein Kinase

Nucleoside diphosphate kinase (NDPK, NM23/awd) belongs to a multifunctional family of highly conserved proteins (∼16–20 kDa) containing two well-characterized isoforms (NM23-H1 and -H2; also known as NDPK A and B). NDPK catalyses the conversion of nucleoside diphosphates into nucleoside triphosphates, regulates a diverse array of cellular events and can act as a protein histidine kinase. AMPK (AMP-activated protein kinase) is a heterotrimeric protein complex that responds to cellular energy status by switching off ATP-consuming pathways and switching on ATP-generating pathways when ATP is limiting. AMPK was first discovered as an activity that inhibited preparations of ACC1 (acetyl-CoA carboxylase), a regulator of cellular fatty acid synthesis. We report that NM23-H1/NDPK A and AMPK α1 are associated in cytosol from two different tissue sources: rat liver and a human lung cell line (Calu-3). Co-immunoprecipitation and binding assay data from both cell types show that the H1/A (but not H2/B) isoform of NDPK is associated with AMPK complexes containing the α1 (but not α2) catalytic subunit. Manipulation of NM23-H1/NDPK A nucleotide transphosphorylation activity to generate ATP (but not GTP) enhances the activity of AMPK towards its specific peptide substrate in vitro and also regulates the phosphorylation of ACC1, an in vivo target for AMPK. Thus novel NM23-H1/NDPK A-dependent regulation of AMPK α1-mediated phosphorylation is present in mammalian cells.

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