Role of calcium-independent phospholipase A2 in oxidant-induced neural cell death
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Phospholipase A2 (PLA2) are esterases that hydrolyze the sn-2 bond of phospholipids resulting in the release of free fatty acids and lysophospholipids. This study determined the roles of calcium-independent PLA2 (iPLA2) in the regulation of phospholipid expression and oxidant-induced cell death in two types of neural cells, astrocytes and neurons. Cells expressed both cytosolic Group VIA (iPLA2²) and microsomal Group VIB (iPLA2³) PLA2 as determined by activity assays and immunoblot analysis. Inhibition of total iPLA2 activity using racemic bromoenol lactone (BEL, 2.5 µM) altered cell death induced by the oxidants hydrogen peroxide (H2O2) or tert-butylhydroperoxide (TBHP). In astrocytes, inhibition of iPLA2 accelerated the loss of ATP levels, but not the formation of reactive oxygen species and induced a significant increase in cells undergoing necrosis as determined by increases in propidium iodide staining. In contrast, inhibition of iPLA2 in neuronal cells protected against H2O2-induced cell death, but not against TBHP-induced cell death. The differential effects of BEL on H2O2 and TBHP-induced cell death correlated to the formation of reactive oxygen species, genesis of lipid peroxidation, and loss of ATP induced by these compounds. In addition, a lipidomic approach was used in combination with electrospray ionization mass spectrometry to determine the effect of oxidant exposure on the phospholipid profiles of astrocytes and neurons. In astrocytes, iPLA2³ was determined to play a greater role than iPLA2² in the maintenance of phosphatidylcholine phospholipids (PtdCho) under physiological conditions. However, oxidant-exposure only slightly altered these phospholipids. In contrast, exposure of neurons to oxidants resulted in significant changes in PtdCho species containing arachidonic acid at the sn-2 position. These changes were altered by the presence of inhibitors of both iPLA2 and cytosolic PLA2 and correlated to the formation of arachidonic acid. These data support the hypothesis that iPLA2 mediates oxidant-induced neural cell death and demonstrates differential roles of iPLA2 isoforms in physiological and pathological events. These data also identify the specific phospholipids and fatty acids targeted by oxidants in neural cell cultures and suggest that iPLA2 may be a therapeutic target for alteration of oxidant-induced neural cell death.