A heparin-sensitive phospholipase A₂ and prostaglandin endoperoxide synthase-2 are functionally linked in the delayed phase of prostaglandin D₂ generation in mouse bone marrow-derived mast cells

BALB/cJ mouse bone marrow-derived mast cells (BMMC) developed with interleukin (IL)-3 can be stimulated by c-kit ligand (KL) in the presence of IL-10 and IL-1β for sequential immediate and delayed generation of prostaglandin (PG) D₂ through utilization of constitutive prostaglandin endoperoxide synthase (PGHS) -1 and induced PGHS-2, respectively (Murakami, M., Matsumoto, R., Austen, K. F., and Arm, J.P. (1994) J. Biol. (Chem. 269, 22269-22275). We now report that BALB/cJ BMMC stimulated with KL + IL-10 + IL-1β also exhibit the biphasic release of [³H]arachidonic acid with an immediate phase over the first 10 min followed by a delayed phase from 2 to 7 h. The delayed phase of arachidonic acid release and of PGD₂ generation was inhibited by heparin, which concomitantly released a phospholipase (PL) A₂ from the cells into the supernatant. Both dexamethasone and a type II PLA₂ inhibitor, 12-epi-scalaradial, suppressed delayed-phase PGD₂ generation at concentrations that did not affect immediate eicosanoid generation. Transcripts for type IIA PLA₂, as assessed by reverse transcription- polymerase chain reaction, were progressively induced in BALB/cJ BMMC treated for 2 to 7 h with KL + IL-10 + IL-1β; the induction of these transcripts was down-regulated by l0^-6 M dexamethasone. The expression of steady-state transcripts and protein for cytosolic PLA₂ (cPLA₂) did not change. PGHS-2- dependent delayed-phase PGD₂ generation elicited by IgE-dependent activation of BALB/cJ BMMC primed with KL + IL-10 was also accompanied by the induction of type IIA PLA₂ transcripts and was suppressed by heparin, with concomitant release of PLA₂ into the supernatant. However, both the direct, cytokine- stimulated and the cytokine-primed, IgE-dependent, delayed-phase PGD₂ generation occurred in BMMC from C57BL/6J mice, which have a natural disruption of the type IIA PLA₂ gene. Thus, kinetic, pharmacologic, and genetic analyses suggest that an inducible, heparin-sensitive PLA₂, rather than cPLA₂, provides arachidonic acid to concomitantly induced PGHS-2 for delayed-phase PGD₂ biosynthesis in activated BMMC. Furthermore, this heparin-sensitive PLA₂ likely represents a novel PLA₂ or a new function for a known low molecular weight PLA₂.