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Title: The control of inflammation in airway epithelial cells
Author: Correia De Paiva, Claudia
ISNI:       0000 0004 6060 7816
Awarding Body: University of Sheffield
Current Institution: University of Sheffield
Date of Award: 2017
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COPD is a severe chronic and complex airway disease that represents a major financial burden on the healthcare and economic system. Environmental risk factors such as cigarette smoke have been associated with the predisposition to COPD. Other factors such as exposure to viral pathogens can exacerbate airway inflammation and tissue destruction generated by recruited neutrophils, culminating in altered epithelial cell responses in COPD. This thesis investigated the physiological role of Pellino-1, an E3-ubiquitin ligase, and its regulation in human primary bronchial epithelial cells (HBEpCs) in response to viral infection. The viral mimic poly(I:C) increased Pellino-1 protein and gene expression in HBEpCs. In addition, Pellino-1 gene expression was significantly increased by RV-16 and RV-1B infection in primary bronchial epithelial cells from COPD patients. Pellino-1 knockdown in HBEpCs led to a reduction in NF-κB regulated cytokines CXCL8, IL-1α and β gene expression and release of CXCL8 in response to poly(I:C) while having no measurable effect on IFNβ mRNA expression. Furthermore, the transient knockdown of Pellino-1 resulted in the decrease in IKKα/β phosphorylation. The role for Pellino-1 in the non-canonical NF-κB pathway was also investigated and while Pellino-1 knockdown did not alter the expression of the non-canonical NF-κB precursor protein NFKB1 following poly(I:C) stimulation, NFKB2 protein expression was suppressed. In contrast to Pellino-1 knockdown, the transient knockdown of NFKB2 resulted in significant increase in CXCL8 mRNA and protein expression and in turn did not regulate Pellino-1 mRNA expression to poly(I:C). These data suggest that following viral infection in airway epithelial cells, TLR3 activation culminates in the up-regulation of Pellino-1 leading to an increase in NFKB2 expression, resulting in the suppression of NF-κB specific gene transcription. In addition to activating non-canonical NF-κB, NFKB1 regulates the activation of ERK signalling via MEK1. Treatment of HBEpCs with MEK1 inhibitors, PD98059 and U0126, resulted in a significant reduction in Pellino-1 protein and gene expression which led to the suggestion of ERK as a potential Pellino-1 regulator. Proteomics analysis of primary epithelial cells obtained from COPD patient airways further identified a potential novel mechanism of action for Pellino-1 in the NF-κB signalling pathway, wherein it Pellino-1 may inhibit A20’s negative regulatory role or its adaptor proteins TNIP1 or TAX1BP. Taken in combination these data support Pellino-1 as a potential target to down-regulate neutrophilic inflammation whilst retaining antiviral immunity by selectively mediating the TLR3 TRIF-dependent NF-κB/MAPK pathway and not TLR3-mediated IRF3 and IFNβ activation.
Supervisor: Parker, Lisa ; Sabroe, Ian Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available