Microbiota-independent effects of galactooligosaccharide fibres on human peripheral blood mononuclear cells

Abstract

Galactooligosaccharides (GOS) are prebiotic fibres known for their role in supporting intestinal health through modulation of the gut microbiota. GOS reach the colon intact and are fermented by commensal bacteria, resulting in the production of short-chain fatty acids with immunomodulatory properties. Supplementation with GOS is associated with reduced symptoms of gastrointestinal disorders and lower systemic and mucosal inflammation.
Prebiotics may bypass the gut barrier in individuals with pathologies that disrupt gut permeability (e.g. inflammatory bowel diseases, IBD) as well as in healthy subjects exposed to life-style associated stressors, and directly interact with intestinal and systemic immune cells. Among these cells, mucosal-associated invariant T (MAIT) cells play a key role in immune surveillance. Alterations in their blood frequencies and functions have been linked with inflammatory diseases, including IBD. More evidence is needed to understand whether GOS affect immunity solely via their prebiotic action on the microbiota or whether they also directly interact with immune cells.
The research described in this thesis investigates the effects of GOS upon phenotypes, soluble mediators, activation markers and toll-like receptors of ex-vivo peripheral blood mononuclear cells (PBMCs) and MAIT cells from healthy donors and those with IBD. Commercially available GOS (B-GOS®), which contain whole GOS as well as free sugars deriving from GOS production, and pure isolated GOS fractions were tested either alone or in combination with other inflammatory stimuli.
GOS consistently exerted anti-inflammatory effects on healthy PBMCs when combined with other immune challenges (riboflavin metabolite, lipopolysaccharide or polyinosinic:polycytidylic acid), as measured by a reduction in the levels of both pro-inflammatory mediators and activation markers. IBD PBMCs failed to respond to stimulation with a selected inflammatory stimulus (riboflavin metabolite) co-administered with GOS which is likely to relate to their significantly
higher baseline inflammation compared to healthy PBMCs. These data highlight the importance of performing donor immunophenotyping before assessing the effects of GOS on diseased donors.
When tested alone, GOS exerted direct immunomodulatory effects on healthy PBMCs and IBD PBMCs by enhancing the activation of T helper cells and by inducing the secretion of a wide range of anti-inflammatory as well as pro-inflammatory mediators. The effects of GOS were chain-length dependent, with fractions at lower degree of polymerisation inducing the strongest responses. The residual free sugars contained in B-GOS® displayed pro-inflammatory effects such as an increase in CD69 expression and IL-8 secretion by both healthy monocytes and IBD monocytes. These results suggest that the amount of free sugars present in commercially available prebiotics should be minimised to avoid pro-inflammatory responses by ex-vivo immune cells. Although further research is required to identify the pathways of GOS signalling, no involvement of toll-like receptor 4 was identified. Based on secreted cytokine data and activated cells, a mechanism via toll-like receptor 2 is proposed. Overall, this research confirms for the first time that GOS exert immunological effects on ex-vivo adult PBMCs with mechanisms that are independent from the action of the gut microbiota, and provide an insight into the cell types and potential pathways involved in their action.

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Identifiers

ORCID for Caroline Childs: orcid.org/0000-0001-6832-224X

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