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Title: Studies on the effect of interleukin-33 on gene expression and lipid composition of macrophages
Author: Buckley, Melanie
ISNI:       0000 0004 5351 1971
Awarding Body: Cardiff University
Current Institution: Cardiff University
Date of Award: 2014
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The atherosclerotic plaque is characterised by the presence of macrophage foam cells that arise from dysfunctional cholesterol metabolism and trafficking. Cytokines are highly expressed within the plaque and play a critical function in initiating and augmenting the disease state. Previous studies have shown that the novel cytokine, interleukin-33 (IL-33), exerts anti-atherogenic actions in animal and in vitro models of the disease. The effect of IL-33 on pro-atherosclerotic markers was assessed in human THP-1 and murine RAW264.7 macrophages and primary human monocyte-derived macrophages (HMDMs) by real time-quantitative polymerase chain reaction (RT-qPCR). The studies then focused on characterising the signalling pathways involved in the regulation of intercellular adhesion molecule-1 (ICAM-1) and monocyte chemotactic protein-1 (MCP-1) expression by IL-33. The expression of key signalling components implicated in atherosclerosis were knocked down by RNA interference (RNAi). These experiments demonstrated that the extracellular signal-regulated kinase (ERK)-1 and -2, p38α, c-Jun N-terminal kinase (JNK)-1 and -2, phosphoinositide-3-kinase (PI3K)-γ, p50 and p65 NF-κB were integral to the IL-33-mediated down-regulation of ICAM-1 and MCP-1 gene expression in THP-1 macrophages. Another key aim was to analyse the effects of IL-33 stimulation on the lipid profiles of macrophages. A combination of thin layer chromatography (TLC) and gas chromatography (GC) was used to assess the fatty acid composition of THP-1 and RAW264.7 macrophages following IL-33 treatment. The cytokine promoted the redistribution of fatty acids and caused a general increase in saturated fatty acids (SFAs), accompanied with a reduction in monounsaturated fatty acids (MUFAs). Additionally, IL-33 increased the content of n-3 polyunsaturated fatty acids (PUFAs) whereas the impact on n-6 PUFAs was more specific to particular fatty acids and varied between the two species. Overall, the cytokine enhanced the levels of PUFAs involved in eicosanoid production. Also, IL-33 influenced the precursors and products of desaturases and appropriately increased the activities of Δ-5 and Δ-6 desaturases but reduced stearoyl-CoA desaturase activity (SCD). The decrease in SCD activity was accompanied by a reduction in the mRNA expression of SCD-1 in RAW264.7 macrophages. The studies presented within this thesis provide new insights into the signalling pathways underlying the IL-33-mediated inhibition of gene expression in macrophages. Additionally, these experiments describe the novel effects of IL-33 stimulation on the lipid profile of macrophages.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Q Science (General) ; R Medicine (General)