Gallic acid-L-leucine conjugate corrects lipid mediator profiles in LPS-stimulated RAW264.7 via suppressing arachidonic acid metabolism

Abstract

Gallic acid-L-leucine conjugate (GL) is a synthesized compound by gallic acid and methyl-L-leucine. This study was designed to investigate the anti-inflammatory effect of GL on the generation of arachidonic acid (AA) pathway-derived inflammatory lipid mediators in RAW 264.7 cells. Based on the detection by LC/MS/MS, we found that GL profoundly suppressed LPS-induced release of AA metabolites such as prostaglandin E2 (PGE2), prostaglandin D2 (PGD2), prostaglandin F2 (PGF2) and thromboxane B2 (TXB2). To characterize the molecular mechanisms underlying the actions of GL on pro-inflammatory lipid mediators, we first investigated LPS-induced expression of AA-metabolizing enzymes such as cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX) as well as inducible NO synthase (iNOS). According to the results of Western blotting, GL decreased the expression of these three enzymes in a concentration-dependent manner. We further demonstrated that GL attenuated the LPS-induced activation of transcription factors NF-κB and AP-1. Along this line, we found GL prevented LPS-induced phosphorylation of p38, JNK and ERK1/2, and inhibited LPS-induced oxidative stress. Collectively, GL may inhibit LPS-induced inflammation via sequentially activating kinases JNK, p38 and ERK1/2, and transcription factors NF-κB and AP-1 in macrophages. Importantly, the results of this study indicate that the GL may exhibits its potential anti-inflammatory activity through correcting the profiles of AA-derived lipid mediators.