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Title: The role of neutrophils in autoimmune disease
Author: Glaser, Anna Elisa Andrea
ISNI:       0000 0004 7970 494X
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2019
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Background: Juvenile-onset systemic lupus erythematosus (JSLE) is a multisystem autoimmune disease with manifestations being very diverse between individuals. Neutrophils (PMN), the main innate phagocytic cells have been implied to contribute to disease development and show an Interferoninduced gene signatures (IGS) in patients. Contribution of PMN to disease development and patient stratification have mainly been investigated with genomic, transcriptomic, proteomic approaches. Aims: (1) To explore the ability to stratify patients with autoimmune diseases based on their IGS, including a metabolomics approach using urine and serum. (2) To investigate the phagocytosis-related gene (PRG) signature (PGS) and IGS in PMN of JSLE and healthy paediatric controls. (3) To determine if the PGS is translated onto protein and functional level in PMN of JSLE patients. (4) To determine the influence of factors present in the neutrophil environment including Tumour Necrosis Factor (TNF) α, Interferon (IFN) α, nucleosomes and signal released from apoptosing cells, on the PRG signature. Methods: Metabolite profiles of serum and urine were obtained from JSLE patients, Juvenile Idiopathic Arthritis (JIA) and paediatric controls using 1H NMR spectroscopy, bucket tables for serum samples were created with TopSpin® and Chenomx Profiler® whereas urine sample metabolites were annotated and quantified in Chenomx Profiler® for each sample individually. JSLE and paediatric controls were tested for differences in IGS and PGS using real-time PCR, flow cytometry and ELISA. Effects of stimulation with apoptotic supernatant, nucleosomes, IFNα and TNFα on IGS and PGS were also investigated. Phagocytosis was measured using pHrodo coated bioparticles (S.aureus, E.coli, zymosan) with flow cytometry and confocal microscopy. Results: JSLE, JIA and paediatric control patients and the JSLE IFN subgroups were distinguishable from each other in a model with partial-least squares discriminant analysis. Urine metabolites built better models than metabolites of serum. Pathway analysis suggested increased inflammation in patients despite low disease activity and differences for metabolites involved in phagocytosis. The PGS genes TLR2 and S100A9 had increased mRNA and protein expression in neutrophils of JSLE patients compared to paediatric control patients. The main difference between JSLE IFN high and IFN low patients was in FcγRIIIb, which was only high in IFN high patients. Increased PRG expression was reflected in increased phagocytosis of E.coli and zymosan particles. All stimulants tested contributed to the PGS and IGS, in particular the nucleosomes which increased IFNa, IGS expression and triggered S100A8/A9 release. Conclusions: Factors in the neutrophil cell environment of JSLE patients can contribute to increased expression of the PGS and IGS. Differences between JSLE, JSLE IFN high, JSLE IFN low and paediatric control patients can be measured at the mRNA, protein and metabolite level. Strong differences suggest that IFN high and IFN low patient subgroups may have different aetiopathogenesis and different cytokine profiles.
Supervisor: Beresford, Michael ; Midgley, Angela ; Peak, Matthew ; Wright, Helen Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral