Molecular mechanisms of anti-inflammatory glucocorticoid action: Evidence for a role of glucocorticoid-inducible genes and implications for glucocorticoid insensitivity

The anti-inflammatory activity of glucocorticoids, in diseases such as asthma, is attributed to their ability to reduce the expression of multiple inflammatory mediators. While glucocorticoid receptor (NR3C1)-mediated transcriptional activation, or transactivation, was believed to primarily mediate side-effects, accumulating evidence indicates that transactivation is also important for the inhibition of inflammatory gene expression. This illustrates the need to investigate possible functional roles for specific glucocorticoid-inducible genes. In this thesis, a repressive role for DUSP1, a phosphatase that inhibits MAPKs, was investigated. DUSP1 was induced by IL1B and dexamethasone in both human pulmonary A549 and primary HBE cells. IL1B-induced DUSP1 negatively regulated MAPK activity and in the further presence of dexamethasone, DUSP1 played a transient, typically partial, role in repressing expression of inflammatory genes, including CXCL1, CXCL2 and PTGS2. Thus, additional glucocorticoid-induced gene products are necessary for repression. Regulation of the mRNA destabilizing protein, ZFP36, by DUSP1 was examined. Following the loss of DUSP1, ZFP36 expression was enhanced and this attenuated IL1B-induced TNF expression. Despite a modest ability of dexamethasone to induce ZFP36, thereby off-setting loss of ZFP36 due to reduced MAPK activity, neither silencing of dexamethasone-induced ZFP36, DUSP1, nor both together, prevented repression of TNF by dexamethasone. Finally, while DUSP1 over-expression attenuated IL1B-induced expression of many inflammatory genes (e.g. IL8, CSF2 etc.), others, including, the inflammatory transcription factor, IRF1, and downstream target genes, such as CXCL10, were profoundly enhanced. While MAPK inhibition prolonged IRF1 expression, silencing of IL1B plus dexamethasone-induced DUSP1 reduced IRF1 and CXCL10 expression. Since, CXCL10 expression was largely unaffected by dexamethasone, these data suggest a mechanism whereby dexamethasone-induced DUSP1 expression maintains CXCL10 expression. In conclusion, this study demonstrates interlinked counterregulatory networks, in which IL1B-induced DUSP1 and ZFP36 regulate MAPK activation and inflammatory gene expression respectively. The transient and partial repressive effect of dexamethasone-induced DUSP1 on only few IL1B-induced inflammatory genes provides a rational for functional screening of glucocorticoid-inducible genes that may show repressive functions. Furthermore, by switching off MAPKs, DUSP1 maintains IRF1 expression and may contribute to glucocorticoid insensitivity.