C-KIT LIGAND MEDIATES INCREASED EXPRESSION OF CYTOSOLIC PHOSPHOLIPASE A(2), PROSTAGLANDIN ENDOPEROXIDE SYNTHASE-1, AND HEMATOPOIETIC PROSTAGLANDIN D-2 SYNTHASE AND INCREASED IGE-DEPENDENT PROSTAGLANDIN D-2 GENERATION IN IMMATURE MOUSE MAST-CELLS

We have examined the cytokine regulation of IgE-dependent prostaglandin (PG) D-2 generation in mouse mast cells by assessing the changes in the levels of the transcript, translated protein, and activity of the enzymes involved in the synthesis of PGD(2) from endogenous arachidonic acid. When mouse mast cells, derived by culture of bone marrow cells with WEHI-3 cell-conditioned medium as a source of interleukin (IL)-3 (BMMC) were cultured in recombinant c-kit ligand (KL), sensitized with IgE, and stimulated with antigen, PGD(2) generation increased 3-fold; when KL was combined with IL-3, IL-9, or IL-10, PGD(2) generation increased 6-8-fold above that produced by the cells cultured in IL -3 alone. The increased IgE-dependent PGD(2) generation by BMMC was apparent after 1 day of culture, reached a maximum after 2-4 days of culture, and was dose-dependent for KL and for each of the accessory cytokines. IgE-dependent generation of leukotriene C-4 increased 2-fold after the cells were cultured with KL and was not increased by the addition of IL-3, IL-9, or IL-10. Assays for steady-state transcripts by RNA blotting, for protein by SDS-PAGE/immunoblotting, and for function by enzymatic activities revealed that KL alone stimulated the increased expression of cytosolic phospholipase A(2) (cPLA(2)), prostaglandin endoperoxide synthase (PGHS)-1, and the terminal enzyme, hematopoietic PGD(2) synthase, without a change in expression of 5-lipoxygenase. IL-3, IL-9, and IL-10 each enhanced the KL-induced expression of PGHS-1. In contrast, transcripts for PGHS-2, which were detected transiently after the cells had been cultured for 5 h in KL + IL-3, were not expressed during the period of subsequent increase in IgE-dependent PGD(2) generation. These findings demonstrate that KL up-regulates expression of cPLA(2), PGHS-1, and hematopoietic PGD, synthase, leading to a relatively selective increase in IgE-dependent production of PGD(2) from endogenously released arachidonic acid in BMMC, and they provide the first example of cytokine regulation of hematopoietic PGD(2) synthase.