Evidence for thromboxane biosynthesis in established cell lines derived from human lung adenocarcinomas

Thromboxane B2 (TxB2) is the stable nonenzymatic hydrolysis product of thromboxane A2, a substance implicated in the initiation of the facilitative role of thrombocytes in the metastatic process. TxB2 was isolated from protein-free culture medium of cell lines Calu-3, Calu-6, A549, and A549/Asc-1, derived from human lung adenocarcinomas. TxB2 and other 20-carbon fatty acid cyclooxygenase products synthesized from exogenous and endogenous arachidonic acid were identified by their characteristic retention indices and fragmentation of electron-capture derivatives of unlabeled and deuterium-labeled products during combined capillary gas chromatography-mass spectrometry. TxB2 comprised 2 to 6% of 20-carbon fatty acid cyclooxygenase products biosynthesized from endogenous arachidonic acid in calcium ionophore A23187-stimulated Calu-6 and A549/Asc-1 cells and 16 to 25% of these products in Calu-3 and A549 cells. The addition of 10(-5) M exogenous arachidonic acid to the cultured cells resulted in a 2- to 3-fold increase in TxB2 and bisenoic prostanoid production with no significant alterations in the proportion of TxB2 production. Prostaglandin E2 and prostaglandin F2 alpha, two prostanoids that can be formed either enzymatically or nonenzymatically from prostaglandin H2, accounted for greater than 75% of isolatable 20-carbon fatty acid cyclooxygenase products synthesized from endogenous and exogenous arachidonic acid.