Understanding the molecular mechanisms underpinning the immunomodulatory function of carbohydrates: focus on levan exopolysaccharide

Young, Ian (2018) Understanding the molecular mechanisms underpinning the immunomodulatory function of carbohydrates: focus on levan exopolysaccharide. Doctoral thesis, University of East Anglia.

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Polysaccharides (PS) found in plants, mushrooms, algae and gut microbes have been shown to modulate immune function in animals. However, the mechanisms underpinning their immunomodulatory properties remain unclear. C-type lectin receptors (CLRs) are carbohydrate-binding proteins, expressed by immune cells including dendritic cells (DCs). CLRs SIGNR1, Dectin-1 and Dectin-2 recognise mannose/fucose structures, -glucans and -mannans, respectively, which initiates cellular activation and an immune response.
We first hypothesised that the mechanisms underpinning immunomodulatory PS involve their interaction with CLRs. To test this, we established SIGNR1 reporter cells, and used Dectin-1 and Dectin-2 reporter cells to screen a range of food and microbial PS for their binding to CLRs.
We report several novel CLR-PS interactions including the interaction between microbial levan, a high molecular weight -(2,6) fructofuranose exopolysaccharide from gram-negative bacterium Erwinia herbicola, and Dectin-2. Crude E. herbicola levan also activated Toll-like receptor 4 (TLR4) reporter cells, suggesting an interaction with lipopolysaccharide (LPS).
Following the development of a robust method to remove LPS in levan, we confirmed that LPS was an important contributor to crude levan’s ability to bind to TLR4 and Dectin-2. Further, crude E. herbicola levan but not enzymatically synthesised (ES) levan or purified E. herbicola levan induced cytokine production in bone marrow-derived DCs. We also tested interaction of purified ES levan and intestinal epithelial cells in vitro using Caco-2 cells. We found that purified ES levan did not improve barrier integrity following LPS challenge and did not induce or modulate cytokine production following inflammatory challenge by TNF- treatment.
Overall, we demonstrated that the immunomodulatory properties of levan in vitro were largely dependent on the presence of LPS and we present a novel purification method to remove LPS in levan for future studies.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Science > School of Biological Sciences
Depositing User: Megan Ruddock
Date Deposited: 21 Mar 2019 13:49
Last Modified: 18 Mar 2022 01:38
URI: https://ueaeprints.uea.ac.uk/id/eprint/70294


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