Generation of thromboxane A2 from highly purified human sinus mast cells after immunological stimulation

To better understand metabolites of arachidonic acid generated from human mast cells, the present study assessed the capacity of human mast cells to synthesize thromboxane B2 (TXB2). Anti-IgE challenge of human sinus mast cells resulted in the generation of TXB2 in a dose-dependent manner with a maximal generation of 8.2+/-4.4 ng/10(6) cells (n = 12), which is about 10-fold lower than the maximal generation of prostaglandin D2 (PGD2). Pretreatment of the cells with OKY-046, an inhibitor of TXA synthase, prevented formation of TXB2 in a dose-dependent manner without affecting the generation of PGD2 or leukotriene C4. Experiments using indomethacin or MK-591, a potent FLAP inhibitor, showed that shunting of arachidonic acid did not occur in a single-cell suspension of mast cells. Analysis by RT-PCR revealed that two species of TXA synthase, the full-length TXA synthase mRNA (TXAS-1, 570 BP) and a small quantity of the alternate-spliced form (400 BP), were present in mast cells. When cellular levels of TXAS-1 mRNA were normalized to those of G3PDH mRNA, the relative concentration of TXAS-1 was 2.06+/-0.60 (n = 7) in highly purified sinus mast cells (92.3+/-3.0% pure) and 3.66+/-0.98 (n = 5) in eosinophils.