Induction of cytosolic phospholipase A(2) activity by phosphatidic acid and diglycerides in permeabilized human neutrophils: Interrelationship between phospholipases D and A(2)

Relationships between phospholipases are poorly understood, but phosphatidic acid (PA) and diglycerides (DCs), produced by phospholipase D (PLD) and phosphatidate phosphohydrolase actions, might function as second messengers coupling cell stimulation to cellular responses. This study investigates the role of PLD-mediated PA and DG formation in inducing phospholipase A(2) (PLA(2)) activity in intact human neutrophils (PMNs) and in PMNs permeabilized with Staphylococcus aureus alpha-toxin. PMNs were labelled with [H-3]arachidonic acid (AA) to assess AA release and metabolism and diacylglycerol formation, or with [H-3]1-O-hexadecyl-2-lyso-glycerophosphatidylcholine for the determination of platelet-activating factor (PAF), PA and alkylacylglycerol production, In intact PMNs primed with tumour necrosis factor alpha before stimulation with N-formyl-Met-Leu-Phe, AA release and metabolism and PAF formation increased in parallel with enhanced PA and DG formation, and inhibition of PA and DG production led to a decrease in both AA release and PAF accumulation. In a-toxin-permeabilized PMNs, AA release and PAF production result from the specific activation of cytosolic PLA(2) (cPLA(2)). In this system, PA and DG formation were always present when cPLA(2) activation occurred; blocking PA and DC production inhibited AA release and PAF accumulation. Adding either PA or DG back to permeabilized cells (with endogenous PA and DG formation blocked) led to a partial restoration of AA release and PAF formation; a combination of PA and DGs reconstituted full cPLA(2) activity. These results strongly suggest that products of PLD participate inactivating cPLA(2) in PMNs.