Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.568048
Title: The role of GSK3β in regulating corticosteroid function under conditions of oxidative stress in chronic obstructive pulmonary disease
Author: Ngkelo, Anta
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2013
Availability of Full Text:
Access through EThOS:
Full text unavailable from EThOS. Please try the link below.
Access through Institution:
Abstract:
Chronic obstructive pulmonary disease (COPD) is a progressive chronic inflammatory lung disease characterised by insensitivity to the anti-inflammatory actions of corticosteroids. Reactive oxygen species deriving from cigarette smoke or other environmental noxious particles or gases are a critical etiologic factor in the pathogenesis of COPD and contribute to the reduced corticosteroid action. In the lungs of COPD patients the increased levels of reactive oxygen species induce activation of the Phosphoinositide 3-kinase (PI3K)/Akt and mitogen activated protein kinase (MAPK) pathways which are known to be involved in regulating inflammatory responses as well as corticosteroid insensitivity. Members of these pathways have been implicated in phosphorylating glycogen synthase kinase (GSK3β) on serine 9 leading to inhibition of its activity. GSK3β is a constitutively active serine/threonine kinase that regulates a number of cellular processes, including inflammatory responses, cell cycle and metabolism. In this thesis I investigated whether GSK3β is modulated in primary peripheral macrophages and peripheral blood monocytes from COPD patients and/or under exposure to oxidative stress and how this modulation affects corticosteroid function. The results obtained showed that the levels of p-GSK3β-Ser9 are increased in peripheral lung macrophages and peripheral blood monocytes from COPD patients compared to smokers with normal lung function and non-smokers. This is mediated by increased levels of reactive oxygen species as in response to H2O2-derived oxidative stress, Akt, p38 MAPK and extracellular signal regulated kinase (ERK) 1/2 MAPK phosphorylate and inactivate GSK3β in a time- and concentration-dependent manner. Inhibition of GSK3β by treatment with the specific inhibitor CT99021, gene silencing using siRNA or overexpression of a kinase dead mutant reduced the anti-inflammatory effects of corticosteroids in monocytes and led to increased pro-inflammatory cytokine release upon LPS (Lipopolysaccharide) stimulation. In addition, overexpression of a constitutively active mutant GSK3β with a serine 9 to alanine mutation reversed H2O2-induced reduction of corticosteroid function. GSK3β-regulated corticosteroid function under exposure to oxidative stress does not involve impaired glucocorticoid receptor (GR) nuclear translocation, GR binding to (nuclear factor kappa B) NF-kB/p65 or p65 DNA binding capacity. By contrast, my data showed that GSK3β regulates the key co-repressor recruited by GR, histone deacetylase 2 (HDAC2). Inhibition of GSK3β led to reduction of HDAC2 activity which correlated with an increased phosphorylation on its serine 394 residue. This finding suggests that GSK3β regulates corticosteroid function by inhibiting histone deacetylation leading to chromatin condensation at the site of NF-kB-mediated transactivation of inflammatory gene expression. Analysis of the effects of CT99021 on global gene expression using microarrays confirmed that GSK3β is a key regulator of corticosteroid-sensitive inflammatory gene expression. Among the genes that were differentially expressed in response to CT99021, protein kinase A (PKA) was upregulated suggesting that it may play a significant role in mediating GSK3β-dependent corticosteroid function in monocytes. My findings suggest that there are multiple parallel mechanisms that mediate the oxidative stress-induced corticosteroid unresponsiveness and GSK3β is a key member of the redox-sensitive signalling pathways in COPD. Identification of the downstream targets of GSK3β that mediate phosphorylation and inhibition of HDAC2 activity will assist identification of novel key mediators with potential for therapeutic targeting.
Supervisor: Adcock, Ian Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.568048  DOI: Not available
Share: