Corticosteroid-Induced MKP-1 Represses Pro-Inflammatory Cytokine Secretion by Enhancing Activity of Tristetraprolin (TTP) in ASM Cells.

Exaggerated cytokine secretion drives pathogenesis of a number of chronic inflammatory diseases, including asthma. Anti-inflammatory pharmacotherapies, including corticosteroids, are front-line therapies and although they have proven clinical utility, the molecular mechanisms responsible for their actions are not fully understood. The corticosteroid-inducible gene, mitogen-activated protein kinase (MAPK) phosphatase 1 (MKP-1, DUSP1) has emerged as a key molecule responsible for the repressive effects of steroids. MKP-1 is known to deactivate p38 MAPK phosphorylation and can control the expression and activity of the mRNA destabilizing protein-tristetraprolin (TTP). But whether corticosteroid-induced MKP-1 acts via p38 MAPK-mediated modulation of TTP function in a pivotal airway cell type, airway smooth muscle (ASM), was unknown. While pretreatment of ASM cells with the corticosteroid dexamethasone (preventative protocol) is known to reduce ASM synthetic function in vitro, the impact of adding dexamethasone after stimulation (therapeutic protocol) had not been explored. Whether dexamethasone modulates TTP in a p38 MAPK-dependent manner in this cell type was also unknown. We address this herein and utilize an in vitro model of asthmatic inflammation where ASM cells were stimulated with the pro-asthmatic cytokine tumor necrosis factor (TNF) and the impact of adding dexamethasone 1 h after stimulation assessed. IL-6 mRNA expression and protein secretion was significantly repressed by dexamethasone acting in a temporally distinct manner to increase MKP-1, deactivate p38 MAPK, and modulate TTP phosphorylation status. In this way, dexamethasone-induced MKP-1 acts via p38 MAPK to switch on the mRNA destabilizing function of TTP to repress pro-inflammatory cytokine secretion from ASM cells. J. Cell. Physiol. 231: 2153-2158, 2016. © 2016 Wiley Periodicals, Inc.

Temporal regulation of cytokine mRNA expression by tristetraprolin: dynamic control by p38 MAPK and MKP-1.

Cytokines drive many inflammatory diseases, including asthma. Understanding the molecular mechanisms responsible for cytokine secretion will allow us to develop novel strategies to repress inflammation in the future. Harnessing the power of endogenous anti-inflammatory proteins is one such strategy. In this study, we investigate the p38 MAPK-mediated regulatory interaction of two anti-inflammatory proteins, mitogen-activated protein kinase phosphatase 1 (MKP-1) and tristetraprolin (TTP), in the context of asthmatic inflammation. Using primary cultures of airway smooth muscle cells in vitro, we explored the temporal regulation of IL-6 cytokine mRNA expression upon stimulation with TNF-α. Intriguingly, the temporal profile of mRNA expression was biphasic. This was not due to COX-2-derived prostanoid upregulation, increased expression of NLRP3 inflammasome components, or upregulation of the cognate receptor for TNF-α-TNFR1. Rather, the biphasic nature of TNF-α-induced IL-6 mRNA expression was regulated temporally by the RNA-destabilizing molecule, TTP. Importantly, TTP function is controlled by p38 MAPK, and our study reveals that its expression in airway smooth muscle cells is p38 MAPK-dependent and its anti-inflammatory activity is also controlled by p38 MAPK-mediated phosphorylation. MKP-1 is a MAPK deactivator; thus, by controlling p38 MAPK phosphorylation status in a temporally distinct manner, MKP-1 ensures that TTP is expressed and made functional at precisely the correct time to repress cytokine expression. Together, p38 MAPK, MKP-1, and TTP may form a regulatory network that exerts significant control on cytokine secretion in proasthmatic inflammation through precise temporal signaling.

The nucleotide-binding domain and leucine-rich repeat protein-3 inflammasome is not activated in airway smooth muscle upon toll-like receptor-2 ligation.

Inflammasomes have emerged as playing key roles in inflammation and innate immunity. A growing body of evidence has suggested that the nucleotide-binding domain and leucine-rich repeat protein-3 (NLRP3) inflammasome is important in chronic airway diseases such as asthma and chronic obstructive pulmonary disease. Inflammasome activation results, in part, in pro-IL-1ß processing and the secretion of the proinflammatory cytokine IL-1ß. Because asthma exacerbations are associated with elevated concentrations of secreted IL-1ß, we addressed whether the NLRP3 inflammasome is activated under in vitro conditions that mimic infectious exacerbations in asthma. Primary cultures of airway smooth muscle (ASM) cells were treated with infectious stimuli (mimicked using the Toll-like receptor-2 agonist Pam3CSK4, a synthetic bacterial lipopeptide). Whereas Pam3CSK4 robustly up-regulated ASM cytokine expression in response to TNF-α and significantly enhanced IL-1ß mRNA expression, we were unable to detect IL-1ß in the cell supernatants. Thus, IL-1ß was not secreted and therefore was unable to act in an autocrine manner to promote the amplification of ASM inflammatory responses. Moreover, Toll-like receptor-2 ligation did not enhance NLRP3 or caspase-1 expression in ASM cells, and NLRP3 and caspase-1 protein were not present in the ASM layer of tracheal sections from human donors. In conclusion, these data demonstrate that the enhanced synthetic function of ASM cells, induced by infectious exacerbations of airway inflammation, is NLRP3 inflammasome-independent and IL-1ß-independent. Activation of the NLRP3 inflammasome by invading pathogens may prove cell type-specific in exacerbations of airway inflammation in asthma.