Peroxisome proliferator-activated receptors alpha and beta mediate the anti-inflammatory effects of the cyclopentenone prostaglandin 15-deoxy-?12,14-PGJ2 in fish granulocytes

Abstract

Prostaglandins (PGs) are highly reactive small lipophilic molecules derived from polyunsaturated fatty acids of the cell membrane and play a key role in the resolution of inflammation processes. 15-deoxy-?(12,14)-PGJ(2) (15dPGJ(2)) is a cyclopentenone PG (CyPG) of the J series with anti-inflammatory, anti-proliferative and pro-apoptotic effects. This CyPG can signal through: (i) the PGD(2) receptor (DP2) and peroxisome proliferator-activated receptor ? (PPAR?) or (ii) by covalent binding to protein nucleophiles, such as, thiols groups of cysteine, lysine or histidine via a Michael addition reaction, modifying its structure and function. In this work we show that acidophilic granulocytes (AGs) of gilthead seabream (Sparus aurata L.), the functional equivalent to mammalian neutrophils, constitutively expressed ppara, pparb and pparg genes, the latter showing the highest expression and up-regulation when stimulated by bacterial DNA. In addition, we tested the ability of 15dPGJ(2,) and its biotinylated analog, as well as several PPAR? ligands, to modulate reactive oxygen species (ROS) and/or cytokines production during a Toll like receptor (TLR)-mediated granulocyte response. Thus, 15dPGJ(2) was able to significantly decrease bacterial DNA-induced ROS production and transcript levels of pparg, interleukin-1? (il1b) and prostaglandin-endoperoxide synthase 2 (ptgs2). In contrast, its biotinylated analog was less potent and a higher dose was required to elicit the same effects on ROS production and cytokine expression. In addition, different PPAR? agonists were able to mimic the effects of 15dPGJ(2). Conversely, the PPAR? antagonist T007097 abolished the effect of 15dPGJ(2) on DNA bacterial-induced ROS production. Surprisingly, transactivation assays revealed that both 15dPGJ(2) and its biotinylated analog signaled via Ppar? and Ppar?, but not by Ppar?. These results were further confirmed by HPLC/MS analysis, where Ppar? was identified as an interactor of biotin-15dPGJ(2) in naïve and DNA-stimulated leukocytes. Taken together, our data show that 15dPGJ(2) acts both through Ppar activation and covalent binding to proteins in fish granulocytes and identify for the first time in vertebrates a role for Ppar? and Ppar? in the resolution of inflammation mediated by 15dPGJ(2).