PROSTAGLANDINS AS NEGATIVE REGULATORS AGAINST LIPOPOLYSACCHARIDE, LIPOTEICHOIC ACID, AND PEPTIDOGLYCAN-INDUCED INDUCIBLE NITRIC OXIDE SYNTHASE/NITRIC OXIDE PRODUCTION THROUGH REACTIVE OXYGEN SPECIES-DEPENDENT HEME OXYGENASE 1 EXPRESSION IN MACROPHAGES

Although prostaglandins (PGs) were reported to exert proinflammatory and anti-inflammatory effects in macrophages, their action mechanisms remain unclear. The effects of PGs including PGJ(2) (J(2)), Delta(12)-PGJ(2) (Delta(12)), 15-deoxy-Delta(12,14) PGJ(2) (15d), PGE(2) (E-2), and PGF(2 alpha) (F-2 alpha) on lipopolysaccharide (LPS)-, lipoteichoic acid (LTA)-, and peptidoglycan (PGN)-induced inducible nitric oxide (NO) synthase (iNOS)/NO production by RAW264.7 macrophages were investigated. First, we found that induction of cyclooxygenase 2 (COX-2) protein occurred at a time earlier than that of heme oxygenase 1 (HO-1) protein, and the addition of the COX-2 inhibitor NS398 reduced HO-1 protein expression in LPS-, LTA-, and PGN-treated RAW264.7 macrophages. Incubation of RAW264.7 macrophages with the indicated PGs showed that J(2), Delta(12), and 15d significantly induced HO-1 protein expression; however, E-2 and F-2 alpha did not. Heme oxygenase 1 protein induced by J(2), Delta(12), and 15d was inhibited by the transcriptional inhibitor, actinomycin (Act) D; the translational inhibitor, cycloheximide; and the antioxidant, N-acetyl cysteine (NAC). Increases in intracellular peroxide levels by J(2), Delta(12), and 15d were detected via a 2', 7'-dichlorofluorescein diacetate (DCFH-DA) analysis, and they were prevented by the addition of NAC. In addition, J(2), Delta(12), and 15d produced significant inhibition of LPS-, LTA-, and PGN-induced iNOS protein and NO production by RAW264.7 cells, in accordance with increased HO-1 protein expression. Reductions of LPS-, LTA-, and PGN-induced phosphorylated c-Jun N-terminal kinase, c-Jun protein, and activator protein 1 luciferase activity by J(2), Delta(12), and 15d were identified, and the addition of the HO-1 inhibitor, tin protoporphyrin, reversed the inhibitory effects of Delta(12) and 15d on LPS- and LTA- induced iNOS/NO, phosphorylated c-Jun N-terminal kinase, and c-Jun protein expressions by macrophages. Knockdown of HO-1 protein expression by HO-1 small interfering RNA blocked Delta(12) and 15d inhibition of LPS- and LTA- induced events. Moreover, the compound, cyclopentenone (CP), which mimics the CP moiety of 15d, and its analog cyclohexenone were used, and cyclohexenone showed more potent induction of the HO-1 protein with effective inhibition of LPS-, LTA-, and PGN-induced iNOS/NO production than CP in macrophages. Reactive oxygen species-dependent HO-1 protein expression by PGs, which inhibited LPS-, LTA-, and PGN-induced iNOS/NO production, was identified in macrophages.