Dietary omega-3 polyunsaturated fatty acids (ω-3 PUFA) reduce sunburn, an acute inflammatory response, in humans. We assessed whether this may be mediated by reduced ultraviolet-B (UV-B) induction of proinflammatory mediators tumor necrosis factor–α (TNF-α), interleukin (IL)-1β, IL-6, IL-8 and prostaglandin (PG)E₂ in healthy skin. In a double-blind, randomized study, 28 humans received 4 g daily of 95% ethyl esters of eicosapentaenoic acid (EPA) or oleic acid (OA) orally for 3 months. Skin biopsies and suction blister fluid were taken from unexposed and UV-B–exposed skin and examined for mediator expression immunohistochemically and quantitatively by immunoassay; plasma levels were also assayed. The subjects taking EPA, but not OA, showed a significant rise in their minimal erythemal dose (MED) (data reported elsewhere). Before supplementation, irradiation with 3× MED UV-B increased blister fluid TNF-α, IL-6, IL-8 and PGE₂ at 16 h (all P < 0.001). No significant change occurred in baseline or UV-B–induced skin levels of cytokines after either supplement, whereas UV-B induction of PGE₂ was abolished after EPA but not OA. Immunohistochemical expression of the cytokines at baseline and after UV-B was unaltered by EPA and OA; circulating cytokine and PGE₂ levels were also unchanged. Hence, in healthy skin in vivo, there was no evidence that reduction of the sunburn response by EPA is mediated by the proinflammatory cytokines examined; abrogation of UV-B–generated PGE₂ may play a role.