Calcium-independent phospholipase A2 modulates cytosolic oxidant activity and contractile function in murine skeletal muscle cells.
Phospholipase A2 (PLA2) activity supports production of reactive oxygen species (ROS) by mammalian cells. In skeletal muscle, endogenous ROS modulate the force of muscle contraction. We tested the hypothesis that skeletal muscle cells constitutively express the calcium-independent PLA2 (iPLA2) isoform and that iPLA2 modulates both cytosolic oxidant activity and contractile function. Experiments utilized differentiated C2C12 myotubes and a panel of striated muscles isolated from adult mice. Muscle preparations were processed for measurement of mRNA by real-time PCR, protein by immunoblot, cytosolic oxidant activity by the dichlorofluorescein oxidation assay, and contractile function by in vitro testing. We found that iPLA2 was constitutively expressed by all muscles tested (myotubes, diaphragm, soleus, extensor digitorum longus, gastrocnemius, heart) and that mRNA and protein levels were generally similar among muscles. Selective iPLA2 blockade by use of bromoenol lactone (10 microM) decreased cytosolic oxidant activity in myotubes and intact soleus muscle fibers. iPLA2 blockade also inhibited contractile function of unfatigued soleus muscles, shifting the force-frequency relationship rightward and depressing force production during acute fatigue. Each of these changes could be reproduced by selective depletion of superoxide anions using superoxide dismutase (1 kU/ml). These findings suggest that constitutively expressed iPLA2 modulates oxidant activity in skeletal muscle fibers by supporting ROS production, thereby influencing contractile properties and fatigue characteristics.
Gong
Ming C
MC
Dept. of Physiology, Univ. of Kentucky Medical Center, 800 Rose St., Lexington, KY 40536, USA.
Arbogast
Sandrine
S
Guo
Zhenheng
Z
Mathenia
Jeremy
J
Su
Wen
W
Reid
Michael B
MB
D000818:Animals; D002460:Cell Line; D003600:Cytosol / Q000201:enzymology; D016166:Free Radical Scavengers / Q000494:pharmacology; D054522:Group VI Phospholipases A2; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D009119:Muscle Contraction; D018763:Muscle Fatigue; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000201:enzymology; D018482:Muscle, Skeletal / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D009206:Myocardium / Q000201:enzymology; D009281:Naphthalenes / Q000494:pharmacology; D010084:Oxidation-Reduction; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D054467:Phospholipases A2; D011753:Pyrones / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D012132:Respiratory Muscles / Q000201:enzymology; D013482:Superoxide Dismutase / Q000494:pharmacology
D016166:Free Radical Scavengers; D009281:Naphthalenes; D010726:Phosphodiesterase Inhibitors; D011753:Pyrones; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; C068314:6-(bromomethylene)tetrahydro-3-(1-naphthaleneyl)-2H-pyran-2-one; D013482:Superoxide Dismutase; D010741:Phospholipases A; D054522:Group VI Phospholipases A2; D054467:Phospholipases A2; C516476:Pla2g6 protein, mouse