Bioactive oxylipins derived from DHA: neuroprostanes contribute to the anti-inflammatory action of DHA through the inhibition of NFkB and independently to PPARy

Oxylipins is a generic term representing the whole oxygenated metabolites produced through the enzymatic and non-enzymatic oxygenation of PUFAs. Oxylipins generated by free radical mediated peroxidation such as iso- and neuroprostanes have been restricted for a long time to oxidative stress biomarkers but several lines of evidence now suggest interesting signalling actions. In the present study, we aimed at investigating the anti-inflammatory properties of Neuroprostanes, a specific class of non-enzymatic oxylipins derived from DHA. Human peripheral blood mononuclear cells were isolated from healthy donors by Ficoll density gradient centrifugation. Monocytes were differentiated into resting macrophages (RM) for 6 days (37°C, 5% CO2) which were then exposed to 2 different types of NeuroPs (i.e. 14-A4-NeuroP and 4-F4t-NeuroP, 10 μM) or vehicle (ethanol 0.15%) during 30 min. Inflammatory response was induced by LPS (100 ng/mL) for 6 hrs. When RM macrophage were pre-exposed with 14-A4-NeuroP and 4-F4t-NeuroP, significant decrease of mRNA abundance for IL-6 (-49% and -26% respectively) and MCP-1 (-55% and -24 % respectively) were measured after LPS activation. Secretion of TNFα and MCP-1 was also reduced when RM were pre-exposed to 14-A4-NeuroP (-10%, ns and -34%, p<0.05) and 4-F4t-NeuroP (-12%, p<0.01 and 25%, ns). Preliminary results regarding the expression and phosphorylation of IkBa show a reduced level of phosphorylated IkBa with 4-F4t-NeuroP suggesting the inhibition of NFkB pathway. Finally, co-transfection of luciferase reporter vector with hPPARg expression vector performed on Cos-7 cells showed a weak interaction of NeuroPs with PPARg indicating that NeuroPs probably act independently of PPARg.In conclusion, these results suggest that the anti-inflammatory properties of DHA could be mediated, at least in part, by NeuroPs which corroborates the role of free-radical mediated reactions in cell signaling.