Fateful triad of reactive oxygen species, mitochondrial dysfunction and lipid accumulation is associated with expression outline of the AMP-activated protein kinase pathway in bovine blastocysts

2017 ◽  
Vol 29 (5) ◽  
pp. 890 ◽  
Author(s):  
S. Prastowo ◽  
A. Amin ◽  
F. Rings ◽  
E. Held ◽  
D. Salilew Wondim ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Fatty Acids ◽  
Protein Kinase ◽  
Mitochondrial Dysfunction ◽  
Lipid Accumulation ◽  
Reactive Oxygen ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Bovine Embryos ◽  
Amp Activated Protein Kinase

Low cryotolerance is considered as the major drawback of in vitro-produced bovine embryos and is frequently associated with a triad encompassing increased cytoplasmic lipid accumulation, enhanced levels of reactive oxygen species (ROS) and mitochondrial dysfunction. The aim of the present study was to explore the role of the AMP-activated protein kinase (AMPK) pathway in the process resulting such phenotypes. Comparative analysis under different environmental conditions revealed downregulation of AMP-activated protein kinase cytalytic subunit 1alpha (AMPKA1), peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC1A) and carnitine palmitoyltransferase 1 (CPT1) genes and upregulation of acetyl-CoA carboxylase α (ACC). In contrast, the presence of fatty acids within the culture medium resulted in a distinct molecular profile in the embryo associated with enhanced levels of ROS, mitochondrial dysfunction and elevated lipid accumulation in bovine embryos. Because AMPKA1 regulates PGC1A, CPT1 and ACC, the results of the present study reveal that AMPK in active its form is the key enzyme promoting lipolysis. Because AMPK1 activity is, in turn, controlled by the AMP : ATP ratio, it is possible to speculate that excessive uptake of exogenous free fatty acids could increase cellular ATP levels as a result of the disturbed β-oxidation of these external fatty acids and could therefore bypass that molecular feedback mechanism. Subsequently, this condition would cause enhanced generation of ROS, which negatively affect mitochondrial activity. Both enhanced generation of ROS and low mitochondrial activity are suggested to enhance the accumulation of lipids in bovine embryos.

scholarly journals Arecoline inhibits the growth of 3T3-L1 preadipocytes via AMP-activated protein kinase and reactive oxygen species pathways

PLoS ONE ◽  
2018 ◽  
Vol 13 (7) ◽  
pp. e0200508
Author(s):  
Zi-Han Tian ◽  
Jueng-Tsueng Weng ◽  
Li-Jane Shih ◽  
An-Ci Siao ◽  
Tsai-Yun Chan ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Protein Kinase ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Amp Activated Protein Kinase

129 EFFECT OF SERUM SUPPLEMENTATION ON AMP-ACTIVATED PROTEIN KINASE (AMPK) ACTIVITY AND LIPID METABOLISM OF IN VITRO-CULTURED BOVINE EMBRYOS

2015 ◽  
Vol 27 (1) ◽  
pp. 156
Author(s):  
S. Prastowo ◽  
F. Rings ◽  
D. S. Wondim ◽  
E. Tholen ◽  
C. Looft ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Mitochondrial Biogenesis ◽  
Reactive Oxygen ◽  
Rna Isolation ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Bovine Embryos ◽  
Fluorescent Intensity ◽  
Ampk Activity

A major problem of embryos cultured in vitro with serum is cytoplasmic lipid accumulation resulting in lower cryotolerance compared with those derived from in vivo or in the absence of serum. AMPK is known as a master regulator of lipid, glucose, and protein metabolism in mammalian cells. Moreover, it has been reported as controller of acetyl-CoA carboxylase α (ACC), the gene responsible for lipid synthesis, and associated with mitochondrial biogenesis and activities in response to oxidative stress. In the present study we aimed to investigate the regulation of AMPK during serum supplementation in vitro. For this, bovine embryos were produced in vitro in SOF media supplemented with oestrous cow serum or fatty acid–free BSA as a system without serum. Triplicate pools (each 10 blastocysts) from each group were used for RNA isolation using Arcturus®PicoPure®RNA Isolation Kit (Life Technologies, USA). Reverse transcription was performed using a combination of Oligo(dT)23 and random primers. Quantification of AMPK catalytic α1 (AMPKA1), ACC, peroxisome proliferator-activated receptor gamma coactivator 1 α (PGC1A), and sterol regulatory element binding transcription factor 2 (SREBP2) transcripts were performed using ABI PRISM® 7000 SDS system (Applied Biosystems, Foster City, CA, USA) using GAPDH as internal control. Normalized log-transformed transcript amount data were statistically analysed using t-test. In addition, AMPK protein was detected by immunofluorescence, mitochondrial activity by MitoTracker® Red (Invitrogen, Carlsbad, CA, USA), and reactive oxygen species by H2DCFDA molecular probe (Life Technologies, USA), and fluorescent intensity signals were visualised under confocal laser scanning microscopy LSM 710 (Carl Zeiss, Germany). Results showed that the expression of AMPKA1, PGC1A, a mitochondrial biogenesis protein, and SREBP2, a regulator of lipid oxidation, were found to be lower (0.4-, 0.2-, and 0.7-fold, respectively; P < 0.05) in blastocysts derived from cultured with serum compared to without serum. By contrast, ACC was up-regulated in blastocysts cultured with serum by 1.8-fold (P < 0.05) compared to without serum. In comparison to blastocyst cultured without serum, a reduced fluorescent intensity was observed in AMPKA1 protein and mitochondrial activity in blastocyst cultured with serum. The presence of serum was also found to be involved in increasing reactive oxygen species accumulation in embryos cultured with serum. The reduced level of AMPK leads to increased ACC and subsequently enhanced conversion of fatty acids into lipid, which is associated with reduced mitochondrial biogenesis protein, elevated reactive oxygen species level, and reduced lipid oxidation by suppression of SREBP2. In conclusion, the presence of serum in in vitro culture environment affected the AMPK activity and thereby genes associated with lipid metabolism in early bovine embryos.

scholarly journals Reactive Oxygen Species-Induced Stimulation of 5'AMP-Activated Protein Kinase Mediates Sevoflurane-Induced Cardioprotection

Circulation ◽  
2009 ◽  
Vol 120 (11_suppl_1) ◽  
pp. S10-S15 ◽  
Author(s):  
R. R. Lamberts ◽  
G. Onderwater ◽  
N. Hamdani ◽  
M. J. A. Vreden ◽  
J. Steenhuisen ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Protein Kinase ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Amp Activated Protein Kinase ◽  
Stimulation Of

Exercise-induced immunosuppression: roles of reactive oxygen species and 5′-AMP-activated protein kinase dephosphorylation within immune cells

2010 ◽  
Vol 108 (5) ◽  
pp. 1284-1292 ◽  
Author(s):  
Hannah Moir ◽  
Michael G. Hughes ◽  
Stephen Potter ◽  
Craig Sims ◽  
Lee R. Butcher ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Protein Kinase ◽  
Immune Cells ◽  
Mononuclear Cells ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Ampk Phosphorylation ◽  
Intracellular Ros ◽  
Exercise Induced ◽  
Amp Activated Protein Kinase

We previously proposed 5′-AMP-activated protein kinase (AMPK) dephosphorylation within immune cells as an intracellular mechanism linking exercise and immunosuppression. In this study, AMPK phosphorylation underwent transient (<1 h) decreases (53.8 ± 7.2% basal) immediately after exercise (45 min of cycling at 70% V̇o2max) in a cohort of 16 adult male participants. Similar effects were seen with running. However, because exercise-induced inactivation of AMPK was previously shown to occur in an AMP-independent manner, the means by which AMPK is inactivated in this context is not yet clear. To investigate the hypothesis that exercise-induced inactivation of AMPK is mediated via signaling mechanisms distinct from changes in cellular AMP-to-ATP ratios, reactive oxygen species (ROS) and intracellular Ca2+ signaling were investigated in mononuclear cells before and after exercise and in cultured monocytic MM6 cells. In in vitro studies, treatment with an antioxidant (ascorbic acid, 4 h, 50 μM) decreased MM6 cell intracellular ROS levels (88.0 ± 5.2% basal) and induced dephosphorylation of AMPK (44.7 ± 17.6% basal). By analogy, the fact that exercise decreased mononuclear cell ROS content (32.8 ± 16.6% basal), possibly due to downregulation (43.4 ± 8.0% basal) of mRNA for NOX2, the catalytic subunit of the cytoplasmic ROS-generating enzyme NADPH oxidase, may provide an explanation for the AMPK-dephosphorylating effect of exercise. In contrast, exercise-induced Ca2+ signaling events did not seem to be coupled to changes in AMPK activity. Thus we propose that the exercise-induced decreases in both intracellular ROS and AMPK phosphorylation seen in this study constitute evidence supporting a role for ROS in controlling AMPK, and hence immune function, in the context of exercise-induced immunosuppression.

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