Synergistic anti-inflammatory activity of ω-3 lipid and rofecoxib pretreatment on macrophage proinflammatory cytokine production occurs via divergent NF-κB activation

Tricia A. Babcock, W. Scott Helton, Khandaker N. Anwar, You Yang Zhao, N. Joseph Espat*, Paul Flakoll

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

15 Scopus citations

Abstract

ω-3 Lipid pretreatment significantly decreases TNF-α production in LPS-stimulated Mφs; however, this response is only a partial inhibition, suggesting that other nonsubstrate- (lipid) dependent mechanisms are involved. The cyclooxygenase (COX)-2 enzyme is principally responsible for lipid metabolism; thus, a selective COX-2 inhibitor (Rofecoxib) would clarify if it is an ω-3 lipid direct effect or a COX-2 enzyme-associated modulated reduction in TNF-α. Moreover, potential synergy between ω-3 lipids and selective COX-2 inhibition is postulated. Hypothesis: Through divergent regulatory mechanisms, ω-3 lipids in combination with Rofecoxib will synergistically decrease the LPS-stimulated Mφ inflammatory response. Methods: RAW 264.7 cells were pretreated with ω-3 lipids, Rofecoxib, or combination treatment and then washed and exposed to LPS. Supernatants were collected for ELISA, total proteins were obtained to determine COX-2 protein expression by Western blot, and nuclear extracts were isolated to determine NF-κB activation by electromobility shift assay. Results: TNF-α and PGE2 production was significantly decreased with ω-3 and Rofecoxib pretreatment, and with combination treatment a further decrease in TNF-α production was observed. COX-2 protein expression was demonstrated to increase in ω-3, Rofecoxib, and combination groups stimulated with LPS. No alteration in NF-κB activation was observed with Rofecoxib or combination pretreatment compared with LPS-stimulated control cells. Repletion of prostaglandin (PGE2) in the Mφ model significantly decreased TNF-α in all groups. Conclusions: ω-3 Lipids and Rofecoxib independently decrease TNF-α and PGE2 production in LPS-stimulated Mφ, yet in combination a synergistic reduction in TNF-α production is observed. Although the anti-inflammatory effects observed from ω-3 lipids are known to occur partially through decreasing NF-κB activation, we demonstrated that Rofecoxib or even a combination of ω-3 and Rofecoxib does not alter NF-κB activation, as seen with ω-3 lipids alone. These data support that combination treatment may result in decreased Mφ inflammation, yet this occurs via divergent mechanisms.

Original languageEnglish (US)
Pages (from-to)232-240
Number of pages9
JournalJournal of Parenteral and Enteral Nutrition
Volume28
Issue number4
StatePublished - Jul 1 2004

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Nutrition and Dietetics

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