1,8-CINEOLE ATTENUATES LIPOPOLYSACCHARIDE-INDUCED INFLAMMATORY RESPONSES IN RAW 264.7 MACROPHAGES THROUGH RESTORATION OF ANTIOXIDANT DEFENSE AND SUPPRESSION OF NF-ΚB-ASSOCIATED CYTOKINE SIGNALING

Abstract

Objectives: This study aimed to evaluate the anti-inflammatory effects of 1,8-cineole (eucalyptol) in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages by assessing inflammatory mediators, antioxidant status, gene expression, and nuclear factor kappa B (NF-κB)-associated cytokine signaling. Methods: RAW 264.7 macrophages were pre-treated with 1,8-cineole (10–100 μM) for 2 h before LPS (1 μg/mL) stimulation. Cell viability was determined using the MTT assay. Nitric oxide (NO), antioxidant markers (superoxide dismutase [SOD], catalase [CAT], and reduced glutathione [GSH]) and inflammatory mediators (Prostaglandin E2 [PGE2], tumor necrosis factor alpha [TNF-α], interleukin [IL]-6, and IL-1β) were quantified using standard assays and enzyme-linked immunosorbent assay. Inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) messenger ribonucleic acid expression was analyzed by quantitative real-time polymerase chain reaction, whereas NF-κB p65 phosphorylation and cytokine protein expression were evaluated by Western blotting. Results: 1,8-cineole was non-cytotoxic up to 100 μM. It significantly reduced LPS-induced NO (41.26±3.58–11.36±1.42 μM) and PGE2 (1894.28±142.74–624.18±66.92 pg/mL), restored SOD, CAT, and GSH levels, downregulated iNOS and COX-2 gene expression, and suppressed NF-κB activation with reduced TNF-α, IL-6, and IL-1β protein expression. Conclusion: 1,8-cineole attenuated LPS-induced inflammatory responses by suppressing inflammatory mediators, restoring antioxidant defense, inhibiting iNOS/COX-2 expression, and reducing NF-κB-associated cytokine signaling, supporting its potential as a promising anti-inflammatory agent.