MECHANISMS OF THE PRIMING EFFECT OF LIPOPOLYSACCHARIDES ON THE BIOSYNTHESIS OF LEUKOTRIENE B-4 IN CHEMOTACTIC PEPTIDE-STIMULATED HUMAN NEUTROPHILS

The goal of this study was to explain the priming effect of lipopolysa ccharides (LPS) in human polymorphonuclear leukocytes on leukotriene B -4 (LTB4) biosynthesis after stimulation with the receptor-mediated ag onist formyl-methionyl-leucyl-phenylalanine (fMLP), This priming effec t for LTB4 biosynthesis was maximal after a 30 min preincubation with LPS but was lost when incubations were extended to 90 min or longer, P riming with LPS resulted in an enhanced maximal activation of 5-lipoxy genase (5- to 15-fold above unprimed cells) as well as a prolonged act ivation of the enzyme after stimulation with fMLP compared to that mea sured in unprimed cells. The activation of 5-lipoxygenase was associat ed with its translocation to the nuclear fraction of the cell, after s timulation of LPS-primed cells but not of unprimed cells, Priming of c ells with LPS also resulted in an enhanced capacity (fivefold increase ) for arachidonic acid (AA) release after stimulation with fMLP compar ed to unprimed cells as measured by mass spectrometry, This release of AA was very efficiently blocked in a dose-dependent manner by the 85 kDa cytosolic phospholipase A(2) (PLA(2)) inhibitor MAFP (IC50=10nM) b ut not by the 14 kDa secretory PLA(2) inhibitor SE 203347 (up to 5 mu M), indicating that the 85 kDa cPLA(2) is the PLA(2) responsible for A A release in response to receptor-mediated agonists, In accord with in hibitor studies, the LPS-mediated phosphorylation of cPLA(2) followed the same kinetics as the priming for AA release, and a measurable fMLP -induced translocation of cPLA(2) was observed only in primed cells, A s with AA release and LTB4 biosynthesis, both the phosphorylation and capacity to translocate CPLA(2) were reversed when the preincubation p eriod with LPS was extended to 120 min. These results explain some of the cellular events responsible for the potentiation and subsequent de cline of functional responses of human polymorphonuclear leukocytes re cruited to inflammatory foci.