Cellular metabolism of brevetoxin (PbTx-2) by a monocyte cell line (U-937)

Blooms of Karenia brevis produce brevetoxins which cause neurotoxic shellfish poisoning and respiratory symptoms in humans as well as harmful effects on sea life. To investigate potential effects of brevetoxins on immune system components, a monocyte cell line (U-937) was exposed in vitro to PbTx-2. U-937 cells metabolized PbTx-2 through cellular detoxification mechanisms, as evidenced by depletion of intracellular glutathione and formation of glutathione and cysteine conjugates. Total intracellular glutathione was significantly decreased in toxin-treated cells compared to control cells, as measured using an enzymatic recycling method. LC/MS was used to detect the following brevetoxin metabolites: a cysteine-PbTx-2 conjugate (m/z 1018) and two putative glutathione-PbTx-2 conjugates (m/z 1204 and 1222). During 3h incubation, glutathione conjugates were detectable as early as 1h and increased in concentration after 2 and 3h. A cysteine-PbTx-2 conjugate appeared after 2h and increased in concentration after 3h. Detectable levels of brevetoxin conjugates were present in response to toxin concentrations of 1muM. Depletion of intracellular glutathione and formation of brevetoxin metabolites, with changes in concentrations over time, suggest immune cells (U-937) have important cellular detoxification pathways for PbTx-2.