Role of mitogen-activated protein kinase-mediated cytosolic phospholipase A(2) activation in arachidonic acid metabolism in human eosinophils

The objective of this investigation was to determine the role of secretory and cytosolic isoforms of phospholipase A(2) (PLA(2)) in the induction of a rachidonic acid (AA) and leukotriene synthesis in human eosinophils and the mechanism of PLA(2) activation by mitogen-activated protein kinase (MAPK) isoforms in this process. Pharmacological activation of eosinophils with fM LP caused increased AA release in a concentration (EC50 = 8.5 nM)- and time -dependent (t(1/2) = 3.5 min) manner. Both fMLP-induced AA release and leuk otriene C-4 (LTC4) secretion were inhibited concentration dependently by ar achidonic trifluoromethyl ketone, a cytosolic PLA(2) (cPLA(2)) inhibitor; h owever, inhibition of neither the 14-kDa secretory phospholipase A(2) by 3- (3-acetamide-1-benzyl-2-ethylindolyl-5-oxy)propanephosphonic acid nor cytos olic Ca2+-independent phospholipase A(2) inhibition by bromoenol lactone bl ocked hydrolysis of AA or subsequent leukotriene synthesis. Pretreatment of eosinophils with a mitogen-activated protein/extracellular signal-regulate d protein kinase (ERK) kinase inhibitor, U0126, or a p38 MAPK inhibitor, SB 203580, suppressed both AA production and LTC4 release. fMLP induced phosph orylation of MAPK isoforms, ERK1/2 and p38, which were evident after 30 s, maximal at 1-5 min, and declined thereafter. fMLP stimulation also increase d cPLA(2) activity in eosinophils, which was inhibited completely by 30 muM arachidonic trifluoromethyl ketone. Preincubation of eosinophils with U012 6 or SB203580 blocked fNILP-enhanced ePLA(2) activity. Furthermore, inhibit ion of Ras, an upstream GTP-binding protein of ERK, also suppressed fMLP-st imulated AA release. These findings demonstrate that cPLA(2) activation cau ses AA hydrolysis and LTC4 secretion. We also find that cPLA(2) activation caused by fMLP occurs subsequent to and is dependent upon ERK1/2 and p38 MA PK activation. Other PLA2 isoforms native to human eosinophils possess no s ignificant activity in the stimulated production of AA or LTC4. The Journal of Immunology, 2001.