Airway inflammation plays a critical role in the pathogenesis of chronic asthma. Glucocorticoid (GC)s are the cornerstone of anti-inflammatory of anti-inflammatory therapy in this disease. However, not all asthmatics improve their pulmonary function following GC therapy. These patients are subjected to the unwanted side effects of prolonged systemic GC therapy, often in situations where there is no evidence that it is exerting any appreciable benefit. The mechanisms for corticosteroid resistance in these patients are poorly understood but may provide new insights into the basis of persistent airway inflammation in asthma. The present proposal will use cellular and molecular approaches to study the pathogenesis of corticosteroid resistant (CR) vs. corticosteroid sensitive (CS) asthma. The central hypothesis which we will pursue is that increased expression of glucocorticoid receptor (GCR) beta, an endogenous inhibitor of GC action, plays a key role in the pathogenesis of CR asthma. We postulate that in addition to inhibition of GCRalpha activity, increased GCRbeta results in enhanced activity of AP-1, a transcription factor involved in the activation of inflammation. Furthermore corticosteroid resistance may result from immune-directed activation of serine/arginine-rich (SR) proteins directing alternative splicing of the GCR pre-MRNA to GCRbeta.
Our specific aims are to determine whether GCRbeta, expression in CS mouse neutrophils or T cells, which naturally lack GCRbeta, will induce CR and augment airway inflammation in a mouse model of asthma. Conversely, we will determine whether inhibition of GCRbeta expression in human CR cells with antisense oligonucleotides leads to steroid sensitivity (specific aim 1); determine whether GCRbeta, but not GCRalpha, enhances AP-1 transcriptional activity and the mechanism by which this occurs in cell lines and cells from peripheral blood and airways of CR asthmatics (specific aim 2); identify splicing factors, SR protein(s) and cis-acting elements regulating selection of splice sites for alpha and beta forms of GCR mRNA in CR asthma. The elucidating of mechanisms underlying steroid resistance will have important consequences for more objective criteria to diagnose steroid resistance, and the development of novel therapeutic modalities in the treatment of CR asthma as well as other chronic inflammatory conditions where altered GC responsiveness contributes to persistent tissue inflammation.
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