Role of Group VIA Calcium-independent Phospholipase A2 in Arachidonic Acid Release, Phospholipid Fatty Acid Incorporation, and Apoptosis in U937 Cells Responding to Hydrogen Peroxide

Abstract

Group VIA calcium-independent phospholipase A 2 (iPLA 2 ) has been shown to play a major role in regulating basal phospholipid deacylation reactions in certain cell types. More recently, roles for this enzyme have also been suggested in the destruction of membrane phospholipid during apoptosis and after oxidant injury. Proposed iPLA 2 roles have rested heavily on the use of bromoenol lactone as an iPLA 2 -specific inhibitor, but this compound actually inhibits other enzymes and lipid pathways unrelated to PLA 2 , which makes it difficult to define the contribution of iPLA 2 to specific functions. In previous work, we pioneered the use of antisense technology to decrease cellular iPLA 2 activity as an alternative approach to study iPLA 2 functions. In the present study, we followed the opposite strategy and prepared U937 cells that exhibited enhanced iPLA 2 activity by stably expressing a plasmid containing iPLA 2 cDNA. Compared with control cells, the iPLA 2 -overexpressing U937 cells showed elevated responses to hydrogen peroxide with regard to both arachidonic acid mobilization and incorporation of the fatty acid into phospholipids, thus providing additional evidence for the key role that iPLA 2 plays in these events. Long-term exposure of the cells to hydrogen peroxide resulted in cell death by apoptosis, and this process was accelerated in the iPLA 2 -overexpressing cells. Increased phospholipid hydrolysis and fatty acid release also occurred in these cells. Unexpectedly, however, abrogation of U937 cell iPLA 2 activity by either methyl arachidonyl fluorophosphonate or an antisense oligonucleotide did not delay or decrease the extent of apoptosis induced by hydrogen peroxide. These results indicate that, although iPLA 2

Group VIA Phospholipase A2 Mitigates Palmitate-induced β-Cell Mitochondrial Injury and Apoptosis

“…This proposed role for iPLA 2 ␤ in repair of oxidized phospholipids represents a special case of the originally proposed function of the enzyme in phospholipid remodeling (99 -102) and is consistent with the observations that oxidation of membranes accelerates iPLA 2 ␤-catalyzed fatty acid release from membranes and that iPLA 2 ␤ mediates oxidant-induced arachidonic acid release from cells (103)(104)(105). Moreover, iPLA 2 ␤ is active against phospholipids with short chain sn-2 substituents (106), such as those produced from polyunsaturated fatty acids by oxidation reactions (107).…”

Signaling Role for Lysophosphatidylcholine Acyltransferase 3 in Receptor-Regulated Arachidonic Acid Reacylation Reactions in Human Monocytes

“…Hence, inhibition of iPLA 2 results in reduced levels of lysophospholipid acceptors, which in turn leads to a decreased incorporation of AA into PLs under unstimulated conditions (14,(23)(24)(25). When the effect of zymosan on AA incorporation was assayed in monocytes treated with BEL, and hence exhibiting diminished lysophospholipid levels (44), the response was slightly but significantly reduced (Fig.…”

“…Under these conditions, Ca 2+ -independent PLA 2 (iPLA 2 ) is thought to account for most of the basal PLA 2 activity of the cells (6)(7)(8)(9)(10). iPLA 2 may, therefore, be a major contributor to the low levels of fatty acid liberated during the continuous recycling of membrane PLs that occurs under resting conditions (11)(12)(13)(14). Because the rate of fatty acid release by constitutive iPLA 2 is lesser than the rate of its reacylation back into PLs, no net accumulation of free fatty acid occurs.…”

“…By means of the former, low concentrations of free AA incorporate into PLs via direct acylation of preexisting PLA 2 -derived lysophospholipids. This pathway is believed to constitute the major pathway for AA incorporation into PLs in a variety of cells under physiological conditions; hence, the availability of lysophospholipid acceptors, particularly lysophosphatidylcholine (lysoPC), is a limiting factor (13,14,(23)(24)(25). The high-capacity/low-affinity pathway incorporates free AA via the de novo route, which ultimately results in the accumulation of AA into triacylglycerol and diarachidonoyl PLs (4,26).…”

Requirement of JNK-Mediated Phosphorylation for Translocation of Group IVA Phospholipase A2 to Phagosomes in Human Macrophages

“…Cells were then stimulated for 1 h in RPMI 1640 medium containing 0.5 mg/ml fatty acid-free BSA to blunt fatty acid reacylation (34), and supernatants were removed. When inhibitors were used, the cells were pretreated with them for 30 min before stimulation.…”