The role of IL-33 in adenosine triphosphate-mediated human mast cells activation

Abstract

Mast cells (MCs) are potent immune tissue resident cells located at the interface between the host and environment. They express a plethora of receptors, allowing them to respond to a great variety of stimuli, such as extracellular ATP, a ligand of P2X receptors (P2XRs). IL-33, a cytokine released upon damage from epithelial cells, modulates activities such as mediator release and degranulation upon IgE sensitization. Due to the possible crosstalk between IL-33 and ATP, the aim of this thesis is to investigate the potentiating role of IL-33 on ATP mediated MC activities using an in vitro model of human primary MCs. In comparison to ADP, high concentrations of ATP initiated MC degranulation, which was further potentiated by IL-33 but not by TSLP, another epithelial cytokine. IL-33 also increased the release of IL-8, induced by both ATP and ADP, while showing no effect on the release of MCP-1 or IL-13. In ATP, but not BzATP mediated intracellular signalling, IL-33 led to an increase in ERK1/2 signalling, but to a decrease of STAT3 and NF-kB signalling for both stimuli. Although IL-33 potentiated ATP mediated MC degranulation, this effect didn’t correlate with a calcium flux in 1000µM BzATP. Use of orthosteric and allosteric P2X7 inhibitors helped to demonstrate that the activation of P2X7 leads to MC degranulation and intracellular signalling, which is then potentiated by IL-33 cross talking with P2X7. Use of NF449 and 5BDBD inhibitors revealed no involvement of P2X1 and P2X4 in MC mediated degranulation or its activation being potentiated by IL-33. While IL-33 upregulated gene expression of a great number of pro-inflammatory molecules, it did not affect the expression of P2XR transcript but led to increase in membrane expression of P2X1 and P2X7. When mimicking the inflammatory model, by stimulating IL-33 treated MCs by supernatants from damaged epithelial cells, MC degranulation wasn’t observed. This was either due to no presence or a presence of low concentrations of ATP in the supernatants. The novel data obtained in this thesis shows a unique potentiating effect of IL-33 on ATP mediated degranulation together with a release of specific mediators and components of intracellular signalling, suggesting an importance of ATP and IL-33 crosstalk in inflammatory conditions. 

Date of Award	1 Aug 2022
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Silvia Bulfone-Paus (Supervisor) & Tracy Hussell (Supervisor)