Cyclopentenone Prostaglandin, 15-Deoxy-Δ^12,14-PGJ₂, Is Metabolized by HepG2 Cells via Conjugation with Glutathione

Abstract

15-deoxy-Delta12,14-prostaglandin J2 (15-d-PGJ2) is a dehydration product of PGD2. This compound possesses a highly reactive polyunsaturated carbonyl moiety that is a substrate for Michael addition with thiol-containing biomolecules such as glutathione and cysteine residues on proteins. By reacting with glutathione and proteins, 15-d-PGJ2 is believed to exert potent biological activity. Despite the large number of publications that have ascribed bioactivity to this molecule, it is not known to what extent 15-d-PGJ2 is formed in vivo. Levels of free 15-d-PGJ2 measured in human biological fluids such as urine are low, and the biological importance of this compound has thus been questioned. Because of its reactivity, we hypothesized that 15-d-PGJ2 is present in vivo primarily as a Michael conjugate. Therefore, we undertook a detailed study of the metabolism of this compound in HepG2 cells that are known to metabolize other cyclopentenone eicosanoids. We report that HepG2 cells primarily convert 15-d-PGJ2 to a glutathione conjugate in which the carbonyl at C-11 is reduced to a hydroxyl. Subsequently, the glutathione portion of the molecule is hydrolyzed with loss of glutamic acid and glycine resulting in a cysteine conjugate. These findings confirm a general route for the metabolism of cyclopentenone eicosanoids in HepG2 cells and may pave the way for new insights regarding the formation of 15-d-PGJ2 in vivo.

Cox-dependent fatty acid metabolites cause pain through activation of the irritant receptor TRPA1

“…The unsaturated carbonyl groups of cyclopentenone PGs rapidly react with thiol-containing proteins and peptides, and measurements of 15-d-PGJ 2 (or other cyclopentenone PGs) in vivo underestimates true concentrations. Thus, 15-d-PGJ2 is detectable in vivo primarily as a Michael conjugate to gluthatione (42). The same consideration applies to the measurement of 8-iso-PGA 2 levels at sites of inflammation or tissue injury (22).…”

Endogenous Interleukin-4 Regulates Glutathione Synthesis Following Acetaminophen-Induced Liver Injury in Mice

“…We speculate that glutathione transferase (GST)- pi in livers of IL-4 −/− mice may catalyze the conjugation of GSH with NAPQI more rapidly than in livers of WT mice (42). GSH may also form more conjugates with other electrophiles in livers of IL-4 −/− mice including those with 4-hydroxy-2-nonenal (43), 15-deoxy- 12,14 -PGJ2 (44), and perhaps oxidized proteins forming S-glutathionylated protein adducts (45). …”

Nonenzymatic free radical-catalyzed generation of 15-deoxy-Δ12,14-prostaglandin J2-like compounds (deoxy-J2-isoprostanes) in vivo

“…CID experiments confi rmed pounds, we sought to determine whether deoxy-J 2 -IsoPs would adduct GSH, an abundant cellular thiol, in the presence of GST. We have previously shown that 15-d-PGJ 2 and other cyclopentenone eicosanoids readily adduct GSH in the presence of GST in vitro (26)(27)(28)(29)(30). In the present study, we incubated deoxy-J 2 -IsoPs generated from the nonenzymatic oxidation of AA in vitro with GSH in the presence of GST.…”

“…This may be due to the rapid reactivity of 15-d-PGJ 2 . Previous studies have indicated that cyclopentenone eicosanoids rapidly undergo conjugation with GSH in vitro and in vivo (26)(27)(28)(29)(30). Brunoldi et al ( 26 ) reported that HepG2 cells convert 15-d-PGJ 2 to a GSH conjugate in which the carbonyl on the prostane ring is reduced to a hydroxyl, and the GSH portion of the molecule is subsequently hydrolyzed to a cysteine conjugate.…”

“…Previous studies have indicated that cyclopentenone eicosanoids rapidly undergo conjugation with GSH in vitro and in vivo (26)(27)(28)(29)(30). Brunoldi et al ( 26 ) reported that HepG2 cells convert 15-d-PGJ 2 to a GSH conjugate in which the carbonyl on the prostane ring is reduced to a hydroxyl, and the GSH portion of the molecule is subsequently hydrolyzed to a cysteine conjugate. Additionally, Milne et al ( 58 ) showed that the cyclopentone eicosanoid 15-A 2t -IsoP is metabolized primarily in vivo by conjugation with GSH and that the compound is excreted into the urine as the N -acetyl cysteine sulfoxide conjugate of 15-A 2t -IsoP in which the carbonyl on the prostane ring of the IsoP is reduced to a hydroxyl.…”