Stimulation of arachidonic acid release and eicosanoid biosynthesis in an interleukin 2-dependent T cell line.

Abstract

Previous studies have provided pharmacologic evidence that T lymphocyte function may be regulated in part by the intracellular production of various arachidonic acid (AA) metabolites in response to cellular stimulation. However, the specific AA metabolic capabilities of homogeneous T cell populations have not been clearly defined. In the present studies, we have employed an accessory cell-free T cell line, HT-2, as a model system for the examination of stimulus-induced eicosanoid biosynthesis in T lymphocytes. HT-2 cells were biosynthetically labeled with [3H]-AA and challenged briefly with various agents that stimulate the hydrolytic release of AA from cellular phospholipids. The bee venom peptide melittin stimulated a profound AA release response in the cells and the concomitant synthesis of both cyclooxygenase (PGF2 alpha, PGE2 and PGD2) and lipoxygenase (5-,12-,15-HETE and possibly 5-,12-diHETE) metabolites of AA. The formation of PGs was blocked by 5 microM indomethacin, demonstrating that this cell line contains cyclooxygenase activity functionally similar to that described in macrophages and other cell types. The high activity of melittin in this system was shown to result largely from a synergy between the peptide itself and a persistent bee venom phospholipase A2 contaminant. However, experiments with melittin freed of detectable phospholipase A2 activity by heating, and with synthetic homopolymers of (L)-lysine and (L)-arginine demonstrated that HT-2 cells contain sufficient endogenous, stimulus-responsive phospholipase A2 to provide both the cyclooxygenase and lipoxygenase pathways of AA metabolism ith substrate. In contrast, Ca++ ionophores, which are known to stimulate AA release and metabolism in certain cell types, stimulated only AA release but no detectable eicosanoid biosynthesis in HT-2 cells. Experiments with exogenous bacterial phospholipase C suggested that this cell line can also generate free AA for eicosanoid biosynthesis from membrane-derived 1,2-diacylglycerol. These results indicate that multiple intracellular pathways of AA metabolism are present HT-2 cells, and that the stimulus-induced release of AA and the production of eicosanoid second messengers may result from activation of either phospholipase A2 or phospholipase C.