Arachidonic Acid Causes Cell Death through the Mitochondrial Permeability Transition

Abstract

We have investigated the effects of arachidonic and palmitic acids in isolated rat liver mitochondria and in rat hepatoma MH1C1 cells. We show that both compounds induce the mitochondrial permeability transition (PT). At variance from palmitic acid, however, arachidonic acid causes a PT at concentrations that do not cause PT-independent depolarization or respiratory inhibition, suggesting a specific effect on the PT pore. When added to intact MH1C1 cells, arachidonic acid but not palmitic acid caused a mitochondrial PT in situ that was accompanied by cytochrome c release and rapidly followed by cell death. All these effects of arachidonic acid could be prevented by cyclosporin A but not by the phospholipase A 2 inhibitor aristolochic acid. In contrast, tumor necrosis factor ␣ caused phospholipid hydrolysis, induction of the PT, cytochrome c release, and cell death that could be inhibited by both cyclosporin A and aristolochic acid. These findings suggest that arachidonic acid produced by cytosolic phospholipase A 2 may be a mediator of tumor necrosis factor ␣ cytotoxicity in situ through induction of the mitochondrial PT.The mitochondrial PT 1 is a regulated permeability increase to solutes with molecular masses up to 1,500 Da mediated by opening of a high conductance channel, the PTP, whose molecular nature remains debated (1). The PT is modulated by a variety of effectors of cell death, including reactive oxygen species that are produced early by mitochondria after stimulation of the TNF␣ receptor (2). Early evidence indicated that PTP opening can mediate cell death caused by oxidative stress and anoxia through ATP depletion and dysregulation of Ca 2ϩ homeostasis (3-5). The PT could also be instrumental in the release of intermembrane proteins such as cytochrome c (6, 7), which plays a key role in caspase 9 activation (8), and apoptosis-inducing factor (9), which causes nuclear degradation independent of caspase activation (10).Mitochondria play a role in apoptotic signaling in the TNF␣ pathway, which is activated by ligand binding to the TNF␣ receptor. Several cytosolic proteins are recruited to the receptor death domain, which in turn leads to the production of a variety of second messengers. As a result, multiple protein-mediated signaling cascades are activated, including caspase-dependent and caspase-independent pathways, phospholipases, protein kinases, and protein phosphatases (see Ref. 11 for a critical and comprehensive review). Lipid mediators may be involved both in signaling and in the late cellular responses by receptors of the TNF␣ superfamily, and solid evidence indicates that mitochondria can be the target of lipid mediators of pathophysiological relevance. Ceramide produced by acidic sphingomyelinase is essential for production of GD3 ganglioside, a cell death inducer (12) that is able to cause PT-dependent apoptosis (13) with cytochrome c release and caspase activation (14). Ceramide itself has prominent effects on mitochondria (14 -19) and may turn physiological Ca 2ϩ signals acting through ...

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