Open AccessBiosynthesis and inactivation of the endocannabinoid 2‐arachidonoylglycerol in circulating and tumoral macrophages
Author(s) -
Di Marzo Vincenzo,
Bisogno Tiziana,
De Petrocellis Luciano,
Melck Dominique,
Orlando Pierangelo,
Wagner Jens A.,
Kunos George
Publication year - 1999
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1046/j.1432-1327.1999.00631.x
Subject(s) - anandamide , fatty acid amide hydrolase , endocannabinoid system , chemistry , 2 arachidonoylglycerol , biochemistry , cannabinoid receptor , arachidonic acid , lipopolysaccharide , monoacylglycerol lipase , extracellular , hydrolase , enzyme , biology , receptor , endocrinology , antagonist
The stimulus‐induced biosynthesis of the endocannabinoid 2‐arachidonoylglycerol (2‐AG) in intact mouse J774 macrophages and the inactivation of 2‐AG by the same cells or by rat circulating macrophages was studied. By using gas chromatography‐mass spectrometry, we found that ionomycin (5 µ m ) and lipopolysaccharide (LPS, 200 µg·mL −1 ) cause a 24‐fold and 2.5‐fold stimulation of 2‐AG levels in J774 cells, respectively, thus providing unprecedented evidence that this cannabimimetic metabolite can be synthesized by macrophages. In J774 cells, LPS also induced a 7.8‐fold increase of the levels of the other endocannabinoid, anandamide, and, in rat circulating macrophages, an almost twofold increase of 2‐AG levels. Extracellular [ 3 H]2‐AG was cleared from the medium of intact J774 macrophages ( t 1/2 = 19–28 min) and esterified to phospholipids, diacylglycerols and triglycerides or hydrolyzed to [ 3 H]arachidonic acid and glycerol. These catabolic processes were attenuated differentially by various enzyme inhibitors. Rat circulating macrophages were shown to contain enzymatic activities for the hydrolysis of 2‐AG, including: (a) fatty acid amide hydrolase (FAAH), the enzyme responsible for anandamide breakdown and previously shown to catalyse also 2‐AG hydrolysis, and (b) a 2‐AG hydrolase activity different from FAAH and down‐regulated by LPS. High levels of FAAH mRNA were found in circulating macrophages but not platelets, which, however, contain a 2‐AG hydrolase. Both platelets and macrophages were shown to express the mRNA for the CB1 cannabinoid receptor. A macrophage 2‐AG hydrolase with apparent K m = 110 µ m and V max = 7.9 nmol·min −1 ·(mg protein) −1 was partially characterized in J774 cells and found to exhibit an optimal pH of 6–7 and little or no sensitivity to typical FAAH inhibitors. These findings demonstrate for the first time that macrophages participate in the homeostasis of the hypotensive and immunomodulatory endocannabinoid 2‐AG through metabolic mechanisms that are subject to regulation.