H₂O₂-Activated Mitochondrial Phospholipase iPLA₂γ Prevents Lipotoxic Oxidative Stress in Synergy with UCP2, Amplifies Signaling via G-Protein-Coupled Receptor GPR40, and Regulates Insulin Secretion in Pancreatic β-Cells.
OBJECTIVE
Pancreatic β-cell chronic lipotoxicity evolves from acute free fatty acid (FA)-mediated oxidative stress, unprotected by antioxidant mechanisms. Since mitochondrial uncoupling protein-2 (UCP2) plays antioxidant and insulin-regulating roles in pancreatic β-cells, we tested our hypothesis, that UCP2-mediated uncoupling attenuating mitochondrial superoxide production is initiated by FA release due to a direct H2O2-induced activation of mitochondrial phospholipase iPLA2γ.


RESULTS
Pro-oxidant tert-butylhydroperoxide increased respiration, decreased membrane potential and mitochondrial matrix superoxide release rates of control but not UCP2- or iPLA2γ-silenced INS-1E cells. iPLA2γ/UCP2-mediated uncoupling was alternatively activated by an H2O2 burst, resulting from palmitic acid (PA) β-oxidation, and it was prevented by antioxidants or catalase overexpression. Exclusively, nascent FAs that cleaved off phospholipids by iPLA2γ were capable of activating UCP2, indicating that the previously reported direct redox UCP2 activation is actually indirect. Glucose-stimulated insulin release was not affected by UCP2 or iPLA2γ silencing, unless pro-oxidant activation had taken place. PA augmented insulin secretion via G-protein-coupled receptor 40 (GPR40), stimulated by iPLA2γ-cleaved FAs (absent after GPR40 silencing).


CONCLUSIONS
The iPLA2γ/UCP2 synergy provides a feedback antioxidant mechanism preventing oxidative stress by physiological FA intake in pancreatic β-cells, regulating glucose-, FA-, and redox-stimulated insulin secretion. iPLA2γ is regulated by exogenous FA via β-oxidation causing H2O2 signaling, while FAs are cleaved off phospholipids, subsequently acting as amplifying messengers for GPR40. Hence, iPLA2γ acts in eminent physiological redox signaling, the impairment of which results in the lack of antilipotoxic defense and contributes to chronic lipotoxicity.
Department of Membrane Transport Biophysics, Institute of Physiology , Academy of Sciences of the Czech Republic, Prague, Czech Republic .
Jan
J
Ježek
Department of Membrane Transport Biophysics, Institute of Physiology , Academy of Sciences of the Czech Republic, Prague, Czech Republic .
Andrea
A
Dlasková
Department of Membrane Transport Biophysics, Institute of Physiology , Academy of Sciences of the Czech Republic, Prague, Czech Republic .
Jaroslav
J
Zelenka
Department of Membrane Transport Biophysics, Institute of Physiology , Academy of Sciences of the Czech Republic, Prague, Czech Republic .
Martin
M
Jabůrek
Department of Membrane Transport Biophysics, Institute of Physiology , Academy of Sciences of the Czech Republic, Prague, Czech Republic .
Petr
P
Ježek
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D045744:Cell Line, Tumor; D054501:Group II Phospholipases A2 / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000187:drug effects; D007473:Ion Channels / Q000378:metabolism*; D008055:Lipids / Q000633:toxicity; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects*; D051381:Rats; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013481:Superoxides / Q000378:metabolism; D000071246:Uncoupling Protein 2; D020122:tert-Butylhydroperoxide / Q000494:pharmacology
D000975:Antioxidants; C473035:G-protein-coupled receptor 40, rat; D007328:Insulin; D007473:Ion Channels; D008055:Lipids; D024101:Mitochondrial Proteins; D043562:Receptors, G-Protein-Coupled; C000604391:Ucp2 protein, rat; D000071246:Uncoupling Protein 2; D013481:Superoxides; D020122:tert-Butylhydroperoxide; D006861:Hydrogen Peroxide; D054501:Group II Phospholipases A2