Abstract
Oxidant-induced lipid peroxidation and cell death are major components of ischemia/reperfusion and toxicant injury. Our previous studies showed that renal proximal tubular cells (RPTCs) express Ca(2+)-independent phospholipase A(2)γ (iPLA(2)γ) in endoplasmic reticulum (ER) and mitochondria and that iPLA(2)γ is cytoprotective. Our present studies reveal the role of ER-iPLA(2)γ in oxidant-induced ER lipid peroxidation, Ca(2+) release, and cell death. Oxidant tert-butyl hydroperoxide (TBHP) caused ER lipid peroxidation and Ca(2+) release in isolated rabbit kidney cortex microsomes. ER-iPLA(2)γ inhibition, using bromoenol lactone (BEL), potentiated both oxidant-induced ER lipid peroxidation and Ca(2+) release. Assessment of fatty acids using electrospray ionization-mass spectrometry revealed that ER-iPLA(2)γ mediates the TBHP-induced release of arachidonic acid (20:4), linoleic acid (18:2), and their oxidized forms (18:2-OH, 18:2-OOH, 20:4-OH, 20:4-OOH, 20:4-(OH)(3). iPLA(2)γ inhibition also accelerated oxidant-induced ER Ca(2+) release in RPTC. Depletion of ER Ca(2+) stores in RPTC with thapsigargin, an ER Ca(2+) pump inhibitor, prior to TBHP exposure reduced necrotic cell death and blocked the potentiation of TBHP-induced necrotic cell death by BEL. Together, these data provide strong evidence that ER-iPLA(2)γ protects renal cells from oxidant-induced necrotic cell death by releasing unsaturated and/or oxidized fatty acids from ER membranes, thereby preserving ER membrane integrity and preventing ER Ca(2+) release.