Inflammatory responses of intestinal epithelial cells and their modulation by omega-3 polyunsaturated fatty acids

Abstract

Long-chain ω-3 polyunsaturated fatty acids (PUFAs) are bioactive nutrients found in foods and supplements that have been attributed to a range of health benefits in a range of conditions, including arthritis, cardiovascular disease, and inflammatory bowel disease (IBD). Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are two ω-3 PUFAs found in the flesh of fish, including salmon, mackerel, and herring, and in fish oil supplements. The essential ω-3 PUFA alpha-linolenic acid (ALA) is found mainly in plant-based dietary sources including walnuts, rapeseed oil, and flaxseeds. Due to poor cellular conversion, humans can only synthesise small amounts of EPA and DHA from ALA. Recently, interest has grown in alternative ω-3 PUFAs due to concerns over the sustainability of foods containing EPA and DHA, as well as diminishing concentrations of ω-3 PUFAs in these food sources. Docosapentaenoic acid (DPA) is an intermediate between EPA and DHA and is thought to possess similar bioactivity to EPA and DHA. Stearidonic acid (SDA) is an intermediate in the conversion of ALA to EPA. Documented benefits of ω-3 PUFA consumption are attributed to EPA and DHA, and to some extent ALA, whilst little is known about the effects of DPA and SDA. Intestinal epithelial cells form part of the gut barrier and are responsible for maintaining regulated intestinal permeability and immune responses. Epithelial cell dysfunction during inflammation is part of the pathogenesis of chronic inflammatory conditions such as IBD. Pro-inflammatory cytokine release by intestinal immune cells can drive inflammatory processes leading to altered epithelial cell function. In vitro models of intestinal epithelial cells, including the human adenocarcinoma cell line (Caco-2), have been utilised to assess changes to epithelial cells during inflammation. Experiments in this thesis aimed to describe the modulation of cytokine-induced inflammation in Caco-2 cells by different ω-3 PUFAs. Caco-2 cells were stimulated with a cocktail of inflammatory cytokines (TNF-α, IFN-γ, and IL-1β), with or without pre-treatment with EPA, DHA, DPA, ALA, or SDA. Cytokine stimulation induced hyper-permeability and increased inflammatory mediator production including intracellular adhesion molecule (ICAM)-1, vascular endothelial growth factor (VEGF), interleukin (IL)-6, CXCL8, and monokine-induced by γ-interferon (CXCL9). Cytokine stimulation also induced the nuclear factor kappa B (NF-κB) inflammatory pathway. Exogenous ω-3 PUFAs were accumulated by Caco-2 cells after 48 hours treatment. EPA was converted to DPA, DPA to EPA retro-conversion was seen, and SDA was converted to EPA, unlike ALA. ω-3 PUFA pre-treatment had variable effects on cytokine-induced inflammation, although no ω-3 PUFA treatments affected cytokine-induced permeability. DHA had the most potent effects on inflammation, significantly reducing CXCL8 concentration in the culture medium and inhibiting the activation of the NF-κB. Other ω-3 PUFAs were ineffective at inhibiting NF-κB expression and activation, and did not affect inflammatory mediator production. Future work could further explore alternative inflammatory pathways regulated by ω-3 PUFAs to elucidate the anti-inflammatory potential of lesser studied ω-3 PUFAs, DPA and SDA. Furthermore, the production and action of pro-resolving lipid mediators from different ω-3 PUFAs in this model could be compared and contrasted. Overall, ω-3 PUFAs had different efficacies on cytokine-induced inflammation in a Caco-2 cell model. However, increased EPA content after SDA treatment suggests SDA could be an important source of EPA within the human epithelium.

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ORCID for Philip Calder: orcid.org/0000-0002-6038-710X

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