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Title: The role of glucocorticoid receptor nuclear trafficking in oxidative stress induced corticosteroid insensitivity
Author: Hakim, Amir
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2013
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Corticosteroids are widely used in the treatment of chronic inflammatory diseases. However, in patients with chronic obstructive pulmonary disease (COPD), cystic fibrosis and a subset of patients with severe asthma, corticosteroids fail to improve the underlying inflammation. Recent advances in the cellular and molecular basis of the pathology in asthma and COPD have revealed several biological targets that may contribute to the relative corticosteroid insensitivity observed in these diseases. These targets include impaired glucocorticoid receptor (GR) nuclear translocation, reduced histone deactylase (HDAC)-2 activity and heightened pro-inflammatory transcription factor activity. Although in vitro and in vivo data support a role for impaired GR nuclear translocation as a potential mechanism of corticosteroid insensitivity in asthma, as of yet, no published data has confirmed this in COPD patients. Furthermore, the mechanisms that govern GR nuclear translocation are mostly unknown. There is growing evidence that excess reactive oxygen species (ROS) generated by inflammatory and immune cells of the lung leads to oxidative stress, which plays a crucial role in the amplification of redox-sensitive pro-inflammatory transcription factors that leads to corticosteroid insensitivity. In this PhD, I investigated the role of GR nuclear trafficking in oxidative stress-induced corticosteroid insensitivity. In a monocytic cell line, U937 cells, we showed that hydrogen peroxide (H2O2), an oxidant, induced intracellular and mitochondrial oxidative stress. This increase in oxidative stress was associated with enhanced IL-1β-induced NFϰB activity and IL-1β-induced CXCL8 release, pro-inflammatory markers shown to be elevated in COPD. Pre-treatment with H2O2 significantly reduced corticosteroid induced GR nuclear import for budesonide (BUD) and fluticasone propionate (FP). This reduction in GR nuclear import was associated with reduced suppression of CXCL8. In our ex vivo study in patients with severe asthma and COPD, we observed similar findings, where BUD was unable to induce GR binding to glucocorticoid response elements (GRE) in patients with severe asthma and also, BUD at a low concentration was unable to suppress CXCL8 release in patients with COPD. The degree of corticosteroid insensitivity observed in our patients with severe asthma and COPD was limited to the above mentioned observations. We then investigated the nuclear transport receptors that govern nucleocytoplasmic shuttling of macromolecules in U937 cells and revealed that importin-7 is essential for corticosteroid-induced GR nuclear import. Further experiments identified that RanGTP, an essential co-factor for importin-7 mediated GR nuclear import, was reduced in our oxidative stress-induced corticosteroid insensitivity model. This novel finding suggests that the importin-7-RanGTP complex may be a potential target to overcome corticosteroid insensitivity in COPD patients.
Supervisor: Adcock, Ian ; Usmani, Omar Sponsor: GlaxoSmithKline
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available