Ecklonia stolonifera inhibits lipopolysaccharide-induced production of nitric oxide, prostaglandin E2, and proinflammatory cytokines in RAW264.7 macrophages

Abstract

As part of our ongoing alternative medicine program, we have directed our attention toward the identification of edible seaweeds in Korea. Here we report on the anti-inflammatory activities of Ecklonia stolonifera. The present study was undertaken to elucidate the pharmacological and biological effects of E. stolonifera extracts on the production of inflammatory mediators in macrophages. The results indicate that the hexane fraction of E. stolonifera extract (ESH) is an effective inhibitor of lipopolysccharide (LPS)-induced NO, prostaglandin E2, and proinflammatory cytokine production in RAW 264.7 cells. These inhibitory effects of ESH were accompanied by decreases in the expression of inducible nitric oxide synthase and cyclooxygenase-2 proteins. Furthermore, ESH inhibited the LPS-induced phosphorylation and degradation of IκB-α, which is required for the nuclear translocations of the p50 and p65 nuclear transcription factor kappa-B (NF-κB) subunits in RAW 264.7 cells. Our results suggest that ESH might exert an anti-inflammatory effect by inhibiting the expression of pro-inflammatory cytokines. Such an effect is mediated by a blocking of NF-κB activation, which consequently inhibits the generation of inflammatory mediators in RAW264.7 cells. Through HPLC fingerprinting of the E. stolonifera extract, the phloroglucinol was also identified and quantified as standard substance. Moreover, we tested the potential application of E. stolonifera extract as a cosmetic material by performing human skin primary irritation tests. In these assays, E. stolonifera extracts did not induce any adverse reactions. Based on these results, we suggest that E. stolonifera extracts be considered possible anti-inflammatory candidates for topical application.

Anti-inflammatory effects of apo-9′-fucoxanthinone from the brown alga, Sargassum muticum

“…RAW 264.7 cells were obtained from the Korean Cell Line Bank (KCLB; Seoul, Korea) and maintained at sub-confluence in a 95% air, 5% CO 2 humidified atmosphere at 37°C as described previously [11-14]. Cells at passages 10–20 were used for the experiments and subcultured every 2–3 days.…”

“…Cell viability was measured as described previously [11-14] with slight modification using MTT assay. RAW 264.7 cells were cultured in 96-well plates for 18 h, followed by treatment with LPS (1 μg/mL) in the presence of various concentrations of the sample.…”

“…RAW 264.7 cells were plated at 1.5 × 10 5 cells/well in 24-well plates and then incubated with or without LPS (1 μg/mL) in the absence or presence of various concentrations (12.5, 25, 50, and 100 μg/mL) of apo-9-fucoxanthinone for 24 h. Nitrite levels in culture media were determined as described previously [11-14] with slight modification using the Griess reaction and presumed to reflect NO levels. Briefly, the culture supernatant (100 μL) was mixed with the same volume of Griess reagent (1% sulphanilamide and 0.1% N-[1-naphthyl]-ethylenediamine dihydrochloride in 5% phosphoric acid %) for 10 min.…”

“…Western blotting was performed with a SDS-PAGE Electrophoresis System as described previously [11-14]. Briefly, the RAW 264.7 cells (5.0 × 10 5 cells/mL) were pre-incubated for 18 h and then treated with LPS (1 μg/mL) plus aliquots sample for 24 h. After incubation, the cells were washed twice with cold PBS.…”

“…It has various biological activities, including antioxidant, anti-inflammatory, and antibacterial activities [10,11]. Previously, our research group documented the anti-inflammatory properties of various seaweads [11-14]. During our on-going screening program designed to identify the anti-inflammatory potential of natural compounds, we have isolated apo-9′-fucoxanthinone from S. muticum , using activity-directed fractionation, and characterized apo-9′-fucoxanthinone’s structural identity using spectroscopy ( 1 H and 13 NMR) in this study.…”