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The 5'-AMP-Activated Protein Kinase (AMPK) Is Involved in the Augmentation of Antioxidant Defenses in Cryopreserved Chicken Sperm.

Nguyen TM, Seigneurin F, Froment P, Combarnous Y, Blesbois E - PLoS ONE (2015)

Bottom Line: MET had similar effects as AICAR except that catalase activity was restored and that ATP and mitochondrial potential were further decreased.CC showed effects opposite to AICAR on SOD, ROS, LPO and AR and motility parameters.Taken together, our results strongly suggest that, upon freeze-thaw process, AMPK stimulated intracellular anti-oxidative defense enzymes through ATP regulation, thus reducing ROS and lipid peroxidation, and consequently partially restoring several essential sperm functions and leading to a better quality of cryopreserved sperm.

View Article: PubMed Central - PubMed

Affiliation: INRA-CNRS, Unité Mixte de Recherche de Physiologie de la Reproduction et des Comportements, F-37380 Nouzilly, France; Université François Rabelais, Tours, France.

ABSTRACT
Semen cryopreservation is a unique tool for the management of animal genetic diversity. However, the freeze-thaw process causes biochemical and physical alterations which make difficult the restoration of sperm energy-dependent functions needed for fertilization. 5'-AMP activated protein kinase (AMPK) is a key sensor and regulator of intracellular energy metabolism. Mitochondria functions are known to be severely affected during sperm cryopreservation with deleterious oxidative and peroxidative effects leading to cell integrity and functions damages. The aim of this study was thus to examine the role of AMPK on the peroxidation/antioxidant enzymes defense system in frozen-thawed sperm and its consequences on sperm functions. Chicken semen was diluted in media supplemented with or without AMPK activators (AICAR or Metformin [MET]) or inhibitor (Compound C [CC]) and then cryopreserved. AMPKα phosphorylation, antioxidant enzymes activities, mitochondrial potential, ATP, citrate, viability, acrosome reaction ability (AR) and various motility parameters were negatively affected by the freeze-thaw process while reactive oxygen species (ROS) production, lipid peroxidation (LPO) and lactate concentration were dramatically increased. AICAR partially restored superoxide dismutase (SOD), Glutathione Peroxidase (GPx) and Glutathione Reductase (GR), increased ATP, citrate, and lactate concentration and subsequently decreased the ROS and LPO (malondialdehyde) in frozen-thawed semen. Motility parameters were increased (i.e., + 23% for motility, + 34% for rapid sperm) as well as AR (+ 100%). MET had similar effects as AICAR except that catalase activity was restored and that ATP and mitochondrial potential were further decreased. CC showed effects opposite to AICAR on SOD, ROS, LPO and AR and motility parameters. Taken together, our results strongly suggest that, upon freeze-thaw process, AMPK stimulated intracellular anti-oxidative defense enzymes through ATP regulation, thus reducing ROS and lipid peroxidation, and consequently partially restoring several essential sperm functions and leading to a better quality of cryopreserved sperm.

No MeSH data available.


Related in: MedlinePlus

Effect of AMPK inhibitor CC and activators AICAR and MET in mitochondrial membrane potential of frozen-thawed sperm.Sperm were treated in the presence of an AMPK activator (2mM AICAR (in light gray) or 1mM MET (in diagonals)) or an AMPK inhibitor (5µM CC (in dark gray)) or Control (Ctrl, in white); Fresh semen (checkered pattern).Values represent means ± SEM from 6 different experiments (4 samples/experiment). Different superscripts indicate significant differences between Ctrl and treatments (AICAR, MET or CC) in frozen-thawed semen (a,b, P<0.05). Asterisks indicate statistically significant differences between fresh and frozen-thawed semen (*, P<0.001).
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pone.0134420.g003: Effect of AMPK inhibitor CC and activators AICAR and MET in mitochondrial membrane potential of frozen-thawed sperm.Sperm were treated in the presence of an AMPK activator (2mM AICAR (in light gray) or 1mM MET (in diagonals)) or an AMPK inhibitor (5µM CC (in dark gray)) or Control (Ctrl, in white); Fresh semen (checkered pattern).Values represent means ± SEM from 6 different experiments (4 samples/experiment). Different superscripts indicate significant differences between Ctrl and treatments (AICAR, MET or CC) in frozen-thawed semen (a,b, P<0.05). Asterisks indicate statistically significant differences between fresh and frozen-thawed semen (*, P<0.001).

Mentions: Cryopreservation induced a 68% loss of mitochondrial potential as measured with the JC1 dye and 70% decrease of ATP content (Fig 3). The presence of MET in the freezing medium led to a 36% decrease of mitochondrial potential compared to control whereas AICAR or CC had no effect. AICAR significantly increased the ATP concentration of the cryopreserved sperm but MET and CC significantly decreased it (Fig 4).


The 5'-AMP-Activated Protein Kinase (AMPK) Is Involved in the Augmentation of Antioxidant Defenses in Cryopreserved Chicken Sperm.

Nguyen TM, Seigneurin F, Froment P, Combarnous Y, Blesbois E - PLoS ONE (2015)

Effect of AMPK inhibitor CC and activators AICAR and MET in mitochondrial membrane potential of frozen-thawed sperm.Sperm were treated in the presence of an AMPK activator (2mM AICAR (in light gray) or 1mM MET (in diagonals)) or an AMPK inhibitor (5µM CC (in dark gray)) or Control (Ctrl, in white); Fresh semen (checkered pattern).Values represent means ± SEM from 6 different experiments (4 samples/experiment). Different superscripts indicate significant differences between Ctrl and treatments (AICAR, MET or CC) in frozen-thawed semen (a,b, P<0.05). Asterisks indicate statistically significant differences between fresh and frozen-thawed semen (*, P<0.001).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4519194&req=5

pone.0134420.g003: Effect of AMPK inhibitor CC and activators AICAR and MET in mitochondrial membrane potential of frozen-thawed sperm.Sperm were treated in the presence of an AMPK activator (2mM AICAR (in light gray) or 1mM MET (in diagonals)) or an AMPK inhibitor (5µM CC (in dark gray)) or Control (Ctrl, in white); Fresh semen (checkered pattern).Values represent means ± SEM from 6 different experiments (4 samples/experiment). Different superscripts indicate significant differences between Ctrl and treatments (AICAR, MET or CC) in frozen-thawed semen (a,b, P<0.05). Asterisks indicate statistically significant differences between fresh and frozen-thawed semen (*, P<0.001).
Mentions: Cryopreservation induced a 68% loss of mitochondrial potential as measured with the JC1 dye and 70% decrease of ATP content (Fig 3). The presence of MET in the freezing medium led to a 36% decrease of mitochondrial potential compared to control whereas AICAR or CC had no effect. AICAR significantly increased the ATP concentration of the cryopreserved sperm but MET and CC significantly decreased it (Fig 4).

Bottom Line: MET had similar effects as AICAR except that catalase activity was restored and that ATP and mitochondrial potential were further decreased.CC showed effects opposite to AICAR on SOD, ROS, LPO and AR and motility parameters.Taken together, our results strongly suggest that, upon freeze-thaw process, AMPK stimulated intracellular anti-oxidative defense enzymes through ATP regulation, thus reducing ROS and lipid peroxidation, and consequently partially restoring several essential sperm functions and leading to a better quality of cryopreserved sperm.

View Article: PubMed Central - PubMed

Affiliation: INRA-CNRS, Unité Mixte de Recherche de Physiologie de la Reproduction et des Comportements, F-37380 Nouzilly, France; Université François Rabelais, Tours, France.

ABSTRACT
Semen cryopreservation is a unique tool for the management of animal genetic diversity. However, the freeze-thaw process causes biochemical and physical alterations which make difficult the restoration of sperm energy-dependent functions needed for fertilization. 5'-AMP activated protein kinase (AMPK) is a key sensor and regulator of intracellular energy metabolism. Mitochondria functions are known to be severely affected during sperm cryopreservation with deleterious oxidative and peroxidative effects leading to cell integrity and functions damages. The aim of this study was thus to examine the role of AMPK on the peroxidation/antioxidant enzymes defense system in frozen-thawed sperm and its consequences on sperm functions. Chicken semen was diluted in media supplemented with or without AMPK activators (AICAR or Metformin [MET]) or inhibitor (Compound C [CC]) and then cryopreserved. AMPKα phosphorylation, antioxidant enzymes activities, mitochondrial potential, ATP, citrate, viability, acrosome reaction ability (AR) and various motility parameters were negatively affected by the freeze-thaw process while reactive oxygen species (ROS) production, lipid peroxidation (LPO) and lactate concentration were dramatically increased. AICAR partially restored superoxide dismutase (SOD), Glutathione Peroxidase (GPx) and Glutathione Reductase (GR), increased ATP, citrate, and lactate concentration and subsequently decreased the ROS and LPO (malondialdehyde) in frozen-thawed semen. Motility parameters were increased (i.e., + 23% for motility, + 34% for rapid sperm) as well as AR (+ 100%). MET had similar effects as AICAR except that catalase activity was restored and that ATP and mitochondrial potential were further decreased. CC showed effects opposite to AICAR on SOD, ROS, LPO and AR and motility parameters. Taken together, our results strongly suggest that, upon freeze-thaw process, AMPK stimulated intracellular anti-oxidative defense enzymes through ATP regulation, thus reducing ROS and lipid peroxidation, and consequently partially restoring several essential sperm functions and leading to a better quality of cryopreserved sperm.

No MeSH data available.


Related in: MedlinePlus