Determining the Role of Eosinophils in Barrier Function

Abstract

Eosinophils are granular leukocytes associated with effector functions, propagation of immune responses and tissue damage in inflammation and infection. However, eosinophils also have regulatory roles including promoting remodelling processes, anti-inflammatory responses and tissue homeostasis. Some studies have also suggested eosinophils may play a role in gut barrier integrity. The gut barrier plays an important role in protecting the host from microbial incursion whilst maintaining permeability essential for nutrient absorption. Gut barrier dysfunction has been observed in intestinal inflammation with high levels of eosinophilia therefore we hypothesised that eosinophils are contributing to barrier function during inflammation. The work in this thesis uses three different gastrointestinal parasite infection models known to drive an intestinal eosinophilia, namely the large intestinal dwelling nematode parasite Trichuris muris, the small intestinal dwelling nematode parasite Trichinella spiralis and the protozoan parasite Toxoplasma gondii, and embraces the use of the eosinophil-deficient transgenic mouse the ∆dblGATA1−/− . My data revealed that eosinophils were trafficked to the site of inflammation in both T. muris and T. spiralis infection of wild type mice; however the lack of eosinophils in the deficient ∆dblGATA1−/− mice did not impact intestinal pathology in any of the infection models tested or worm expulsion kinetics. Although I was unable to assess eosinophil kinetics in T. gondii infection, both T. muris and T. spiralis showed increased recruitment of eosinophil to the gut with a significant recruitment of eosinophils into the epithelial layer. Subsequent research focused on T. spiralis infection in the small intestine. Confocal microscopy confirmed the intraepithelial positioning of these eosinophils, as well as cell to cell contact between eosinophils and epithelial cells. Although infection caused increased levels of markers indicative of gut leakiness, a lack of eosinophils did not affect this suggesting eosinophils were unlikely to be repairing the epithelial layer. However, expression of CD69 and EPX by eosinophils was indicative of activated eosinophils alongside deposition of Major Basic Protein in small intestinal crypts suggested a potential role in epithelial modulation by eosinophils perhaps by preventing microbiota incursion into the lamina propria. Mass cytometry also revealed that small intestinal eosinophil population were not apoptotic and but primed for degranulation. An in vitro model system was developed to explore the ability of eosinophils to respond to parasites antigens and to interact with epithelial cells. Through the use of the eosinophil cell line EOL-1, naïve and butyric acid differentiated eosinophils were shown to be able to respond to microbial associated molecular patterns including via the TLR ligands ssRNA and Poly(I:C) and the parasite derived product T. muris Excretory Secretory antigen. This supports an idea of eosinophils having the potential to have an active sentinel role. Collectively, the data presented in this thesis provides evidence supporting the hypothesis that eosinophils may directly modulate epithelial cell function. Furthermore, it highlights an eosinophil-epithelial cell interaction through which eosinophils could be modulating epithelial responses and/or performing an immunosurveillance role in the epithelial layer during active inflammation. The infection models used cause damage to the gut with potential for microbial incursion from the microbiota components and it may be postulated that the eosinophil perform an immunosurveillance role to prevent this from happening.

Date of Award	1 Aug 2021
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Kathryn Else (Supervisor), Sheena Cruickshank (Supervisor) & Ruth Forman (Supervisor)