DIFFERENTIAL ACTIVATION OF HUMAN NEUTROPHIL CYTOSOLIC PHOSPHOLIPASE A(2) AND SECRETORY PHOSPHOLIPASE A(2) DURING PRIMING BY 1,2-DIACYL-GLYCEROL AND 1-O-ALKYL-2-ACYLGLYCEROL

We have shown previously that both 1,2-diacylglycerol (AAG) and 1-O-al kyl-2-acylglycerol (EAG) prime neutrophil release of arachidonic acid via uncharacterized phospholipases A(2). Therefore, we investigated th e actions of EAG and AAG specifically on neutrophil cytosolic (cPLA(2) ) and secretory (sPLA(2)) phospholipase A(2)s We hypothesized that AAG as a protein kinase activator would activate cPLA(2) via phosphorylat ion events. EAG is antagonistic to the AAG activation of PKC, thus it was not expected to act via phosphorylation of cPLA2. Neutrophils were primed with either AAG or EAG and then stimulated with fMLP. When neu trophils were primed with 5-20 mu M 1,2-diacylglycerol, a shift was ob served in cPLA(2) migration on SDS-PAGE gels, consistent with phosphor ylation of the protein. This gel shift was not seen after exposure to EAG. AAG also caused a parallel increase in enzymatic activity of cPLA (2) that was not seen with EAG. We also investigated whether either di glyceride would cause similar priming or direct secretion of sPLA(2). Both AAG and EAG directly caused significant secretion of neutrophil s PLA(2). EAG also increased the release of sPLA(2) in cells subsequentl y stimulated with fMLP. Thus, AAG activated cPLA(2) and stimulated sec retion of sPLA2. In contrast, EAG did not activate cPLA(2), but direct ly activated secretion of sPLA(2). We also demonstrated that human syn ovial fluid sPLA(2) increased AA release from resting and fMLP-stimula ted neutrophils. Given that diglycerides prime for release of AA, PAF, and LTB4, these current data support the hypothesis that such priming may be mediated by phosphorylation dependent (cPLA(2)) or phosphoryla tion independent (e.g. secretion of sPLA(2)) events. (C) 1998 Publishe d by Elsevier Science B.V. All rights reserved.