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Enhanced anti-inflammatory effect of resveratrol and EPA in treated endotoxin-activated RAW 264.7 macrophages
Victor Pallarès1, Damien Calay, Lídia Cedó
1Departament de Bioquímica i Biotecnologia, Universitat Rovira i Virgili, Marcel·lí Domingo, s/n 43007, Tarragona, Spain.
Abstract:
Macrophages play an important role in immunogenic challenges by producing reactive oxygen species, NO and proinflammatory cytokines that can aggravate and propagate local inflammation. Multiple mechanisms regulate these inflammatory processes. NF-κB and activator protein 1 pathways are crucial in the expression of proinflammatory genes, such as TNF-α, IL-1 (α or β) and -6. Some polyphenols, which are present in beverages, vegetables and fruits, and PUFA, which are present in marine oils and fish food, possess anti-inflammatory effects in vivo and in vitro. Our aim in the present study was to assess whether polyphenols and PUFA have synergistic anti-inflammatory effects in murine macrophages in vitro. Inflammation in RAW 264.7 macrophages was induced by lipopolysaccharide at 100 ng/ml. The treatments with molecules were performed by co-incubation for 19 h. A NO production assay by Griess reaction, a phosphoprotein assay by Pathscan ELISA kit and gene expression analysis using the TaqMan® Low-density Array for ninety-one genes related to inflammation, oxidative stress and metabolism were performed to assess the synergistic anti-inflammatory effects of polyphenols, epigallocatechin gallate and resveratrol (Res; 2·5 μg/ml), and the PUFA, DHA and EPA (30 μm). Adding Res+EPA had an enhanced anti-inflammatory effect, in comparison with EPA and Res alone, leading to decreased NO levels; modulating the phospho-stress activated protein kinase/Jun N-terminal kinase (P-SAPK/JNK) level; down-regulating proinflammatory genes, such as IL, chemokines, transcription factors; and up-regulating several antioxidant genes. Therefore, this combination has a stronger anti-inflammatory effect than either of these molecules separately in RAW macrophages.
Insights
Resveratrol and omega-3 fatty acids (EPA/DHA) show synergistic anti-inflammatory effects in macrophages. This combination effectively reduces nitric oxide and inflammatory gene expression, offering enhanced benefits over individual compounds.
Area of Science:
- Immunology
- Molecular Biology
- Nutrition Science
Background:
- Macrophages are key players in immune responses, producing inflammatory mediators like nitric oxide (NO) and cytokines.
- Nuclear factor-kappa B (NF-κB) and activator protein 1 (AP-1) pathways are critical regulators of pro-inflammatory gene expression.
- Polyphenols and polyunsaturated fatty acids (PUFAs) are known for their individual anti-inflammatory properties.
Purpose of the Study:
- To investigate the synergistic anti-inflammatory effects of polyphenols (resveratrol) and PUFAs (EPA/DHA) in murine macrophages.
- To evaluate the impact of combined resveratrol and PUFA treatment on inflammatory pathways and gene expression.
Main Methods:
- RAW 264.7 murine macrophages were stimulated with lipopolysaccharide (LPS) to induce inflammation.
- Co-incubation with resveratrol (Res) and PUFAs (DHA/EPA) for 19 hours.
- Assays included Griess reaction for NO, ELISA for phosphoproteins, and TaqMan Low-density Array for gene expression analysis of 91 inflammation-related genes.
Main Results:
- Combined resveratrol and EPA treatment demonstrated enhanced anti-inflammatory effects compared to individual compounds.
- This combination significantly decreased NO production and modulated phospho-stress activated protein kinase/Jun N-terminal kinase (P-SAPK/JNK) levels.
- Pro-inflammatory gene expression (e.g., ILs, chemokines) was downregulated, while antioxidant gene expression was upregulated.
Conclusions:
- Resveratrol and EPA exhibit synergistic anti-inflammatory activity in macrophages.
- The combination therapy offers a more potent anti-inflammatory effect than either molecule alone.
- This finding suggests a potential therapeutic strategy for inflammatory conditions by combining specific polyphenols and PUFAs.
