Inactivation of Lipid Glyceryl Ester Metabolism in Human THP1 Monocytes/Macrophages by Activated Organophosphorus Insecticides Role of Carboxylesterases 1 and 2

Carboxylesterases (CES) have important roles in pesticide and drug metabolism and contribute to the clearance of ester containing xenobiotics in mammals Tissues with the highest levels of CES expression are the liver and small intestine In addition to xenobiotics CES also harness their broad substrate specificity to hydrolyze endobiotics such as cholesteryl esters and triacylglycerols Here we determined if two human CES isoforms CES1 and CES2 hydrolyze the endocannabinoids 2 arachidonoylglycerol (2AG) and anandamide (AEA) and two prostaglandin glyceryl esters (PG Gs) which are formed by COX mediated oxygenation of 2AG We show that recombinant CES1 and CES2 efficiently hydrolyze 2AG to arachidonic acid (AA) but not amide containing AEA Steady state kinetic parameters for CES I and CES2 mediated 2AG hydrolysis were respectively k(cat) 59 and 43 min(-1) K-m 49 and 46 mu M and k(cat)/K-m 1 2 and 0 93 mu M-1 min(-1) k(cat)/K-m values are comparable to published values for rat monoacylglycerol lipase (MAGL) catalyzed 2AG hydrolysis Furthermore we show that CES1 and CES2 also efficiently hydrolyze PGE2 G and PGF(2 alpha) G In addition when cultured human THP1 macrophages were treated with exogenous 2AG or PG G (10 mu M 1 h) significant quantities of AA or PGs were detected in the culture medium however the ability of macrophages to metabolize these compounds was inhibited (60-80%) following treatment with paraoxon the toxic metabolite of the insecticide parathion Incubation of THP1 cell lysates with small molecule inhibitors targeting CES1 (thieno[3 2 e][1]benzothiophene 4 5 dione or JZL184) significantly reduced lipid glyceryl ester hydrolase activities (40-50% for 2AG and 80-95% for PG Gs) Immunodepletion of CES1 also markedly reduced 2AG and PG G hydrolase activities These results suggested that CES1 is in part responsible for the hydrolysis of 2AG and PG Gs in THP1 cells although it did not rule out a role for other hydrolases especially with regard to 2AG metabolism since a substantial portion of its hydrolysis was not inactivated by the inhibitors An enzyme (M-r 31-32 kDa) of unknown function was detected by serine hydrolase activity profiling of THP1 cells and may be a candidate Finally the amounts of in situ generated 2AG and PG Gs in macrophages were enhanced by treating the cells with bioactive metabolites of OP insecticides Collectively the results suggest that in addition to MAGL and fatty acid amide hydrolase (FAAH) which have both been documented to terminate endocannabinoid signaling CES may also have a role Furthermore since PG Gs have been shown to possess biological activities in their own right CES may represent an important enzyme class that regulates their in vivo levels