Proteasomal inhibition upregulates the endogenous MAPK deactivator MKP-1 in human airway smooth muscle: mechanism of action and effect on cytokine secretion

Abstract

Asthma is a chronic inflammatory condition. Inhibition of the ubiquitin-proteasome system offers promise as a anti-inflammatory strategy, being responsible for the degradation of key proteins involved in crucial cellular functions, including gene expression in inflammation (e.g. inhibitory IkappaB-alpha and the endogenous MAPK deactivator - MKP-1). As MKP-1 inhibits MAPK-mediated pro-remodeling functions in human airway smooth muscle (ASM; a pivotal immunomodulatory cell in asthma) in this study we investigate the effect of the proteasome inhibitor MG-132 on MKP-1 and evaluate the anti-inflammatory effect of MG-132 on cytokine secretion from ASM cells. Examining the time-course of induction of MKP-1 mRNA and protein by MG-132 (10microM) we show that MKP-1 mRNA was first detected at 30min, increased to significant levels by 4h, resulting in a 12.6+/-1.5-fold increase in MKP-1 mRNA expression by 24h (P<0.05). MKP-1 protein levels corroborate the mRNA results. Investigating the effect of MG-132 on secretion of the cytokine IL-6 we show that while short-term pretreatment with MG-132 (30min) partially reduced TNFalpha-induced IL-6 via inhibition of IkappaB-alpha degradation and the NF-kappaB pathway, longer-term proteasome inhibition (up to 24h) robustly upregulated MKP-1 and was temporally correlated with repression of p38-mediated IL-6 secretion from ASM cells. Moreover, utilizing a cytokine array we show that MG-132 represses the secretion of multiple cytokines implicated in asthma. Taken together, our results demonstrate that MG-132 upregulates MKP-1 and represses cytokine secretion from ASM and highlight the potential of the proteasome as a therapeutic target in asthma.