INDUCTION OF CYTOSOLIC PHOSPHOLIPASE A(2) ACTIVITY BY PHOSPHATIDIC-ACID AND DIGLYCERIDES IN PERMEABILIZED HUMAN NEUTROPHILS - INTERRELATIONSHIP BETWEEN PHOSPHOLIPASE-D AND PHOSPHOLIPASE-A(2)

Relationships between phospholipases are poorly understood, but phosph atidic acid (PA) and diglycerides (DCs), produced by phospholipase D ( PLD) and phosphatidate phosphohydrolase actions, might function as sec ond messengers coupling cell stimulation to cellular responses. This s tudy investigates the role of PLD-mediated PA and DG formation in indu cing phospholipase A(2) (PLA(2)) activity in intact human neutrophils (PMNs) and in PMNs permeabilized with Staphylococcus aureus alpha-toxi n. PMNs were labelled with [H-3]arachidonic acid (AA) to assess AA rel ease and metabolism and diacylglycerol formation, or with -3]1-O-hexad ecyl-2-lyso-glycerophosphatidylcholine for the determination of platel et-activating factor (PAF), PA and alkylacylglycerol production, In in tact 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 inhibiti on of PA and DG production led to a decrease in both AA release and PA F accumulation. In a-toxin-permeabilized PMNs, AA release and PAF prod uction 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 rel ease and PAF accumulation. Adding either PA or DG back to permeabilize d 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 sugges t that products of PLD participate inactivating cPLA(2) in PMNs.