Defining the links between ROS and inflammaging in a human model of neutrophil-driven inflammation

Description

Neutrophils are cells of the immune system that provide protection against infection from micro-organisms (bacteria, fungi). Neutrophils produce large amounts of toxic reactive oxygen species (ROS) which are an important part of the killing repertoire that induce mutations to DNA, lipids and proteins. The production of ROS by the NADPH oxidase (NOX2) enzyme also activates granule enzymes (myeloperoxidase, elastase, gelatinase) which are microbiocidal and help to kill microorganisms. Intracellular signalling also induced by ROS is essential for the release of neutrophil extracellular traps (NETs). NETs are beneficial during infection but in the case of sterile inflammation (e.g. inflammation associated with ageing, “inflammageing”) NETs and ROS promote inflammation and damage host tissues. Mitochondrial ROS production (mtROS) also promotes NET release in the absence of NOX2-derived ROS.

Plasma membrane receptors that activate ROS production are well characterised on neutrophils. However the down-stream effects of intracellular ROS production are less well understood and there is evidence that ROS may be either pro-inflammatory (activating production of inflammatory molecules and NETs) or anti-inflammatory (inducing a stress response that induces cell death). Neutrophil ROS production clearly represents an important physiological mechanism in regulating the resolution of inflammation after infection and the dysregulation of inflammation during inflammaging.

Plan of work

Models: (1) Liverpool. Human neutrophils activated with cytokines or toll-like receptor agonists to induce inflammageing. (2) Newcastle. CRISPR mutant and wild-type C. elegans.

Objective 1: Identify the changes to the neutrophil proteome and phospho-proteome that are induced in response to ROS (neutrophil redox proteome) over 20 min using data-independent acquisition mass spectrometry (DIA MS). (Liverpool)

Objective 2: Use CRISPR mutant C. elegans to determine the effect of key phosphorylation events (identified Obj1) on the innate immune response to C. albicans and S. aureus. (Newcastle)

Objective 3: Inhibit ROS and mtROS chemically to determine the effect of ROS production on pro-inflammatory neutrophil functions (NET release, cell death, production of inflammatory molecules, gene expression) and bacterial killing. (Liverpool)

The project will increase our understanding of the signalling mechanisms regulating ROS production will provide important insight into neutrophil biology in healthy ageing and ways of restoring homeostasis during inflammaging. It will also provide training in human and non-human models of the immune system and transferrable skills in phospho-proteomics, statistical analysis, bioinformatics and CRISPR.

To apply to this project, email your CV and cover letter to the primary supervisor:

hlwright@liverpool.ac.uk