Abstract
Context: Bromoenol lactone (BEL) is an inhibitor of group VI phospholipases (iPLA2s), but has been shown to have severe side effects. Objective: iPLA2 characterization in osteoblasts and effect of BEL on prostaglandin (PG) E2 formation. Methods: iPLA2 expression: RT-PCR, Western Blotting. PGE2 formation: GC–MS after stimulation, treatment with inhibitors or gene silencing. Arachidonate (AA) reacylation into phospholipids, inhibitor reaction products, PGHS-1 modification proteomic analysis: HR-LC–MS/MS. AA accumulation: 14C-AA. Results: iPLA2ß and iPLA2γ were expressed and functionally active. BEL inhibition up to 20 μM caused AA accumulation and enhanced PGE2 formation, followed by a decrease at higher concentrations. BEL reacted with intracellular cysteine and GSH leading to GSH depletion and oxidative stress.Discussion: Initial PGE2 enhancement after BEL inhibition is due to iPLA2-independent accumulation of AA. GSH depletion caused by high BEL concentrations is responsible for the decrease in PGE2 production. Conclusion: BEL must be used with caution in a cellular environment due to conditions of extreme oxidative stress.
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