Peroxidized unsaturated fatty acids stimulate Toll-like receptor 4 signaling in endothelial cells

Although unsaturated fatty acids are assumed to be protective against inflammatory disorders that include a pathway involving Toll-like receptor 4 (TLR4) activation, they might actually be toxic because of their high susceptibility to lipid peroxidation. Here we studied the effects of peroxidized unsaturated fatty acids on the TLR4-nuclear factor (NF)-κB pathway in endothelial cells.

Confluent cultured endothelial cells from bovine aorta were incubated for 1h with fatty acids integrated into phosphatidylcholine vesicles. Lipopolysaccharide (LPS) or phosphatidylcholine vesicles without fatty acids were also applied as a positive control or a control for fatty acid groups, respectively. Activation of TLR4 and downstream signaling was assessed by membrane fractionation and Western blotting or immunofluorescent staining.

In the same way as LPS, application of sufficiently peroxidized unsaturated fatty acids like oleic acid or docosahexaenoic acid, acutely caused TLR4 translocation to caveolae/raft membranes, leading to activation of NF-κB signaling in endothelial cells. In contrast, saturated fatty acids did not show such effects. Applying well-peroxidized unsaturated fatty acids, but not saturated fatty acids, acutely activates the TLR4/NF-κB pathway.

Peroxidation of unsaturated fatty acid is essential for the acute activation of TLR4 by the fatty acids that follow the same pathway as the activation by LPS. Unsaturated fatty acids have been assumed to be protective against inflammatory disorders, and drugs containing unsaturated fatty acids are now developed and provided. Our result suggests that, for inflammatory disorders involving TLR4 signaling, using unsaturated fatty acids as anti-inflammatory drugs may cause contrary effects.