Regulator of G protein signaling 2 is a key modulator of airway hyperresponsiveness
ABSTRACT Drugs targeting individual G protein-coupled receptors are used as asthma therapies, but this strategy is limited because of G protein-coupled receptor signal redundancy. Regulator of G protein signaling 2 (RGS2), an intracellular selective inhibitor of multiple bronchoconstrictor receptors, may play a central role in the pathophysiology and treatment of asthma.
We defined functions and mechanisms of RGS2 in regulating airway hyperresponsiveness (AHR), the pathophysiologic hallmark of asthma.
Real-time PCR and Western blot were used to determine changes in RGS2 expression in ovalbumin-sensitized/-challenged mice. We also used immunohistochemistry and real-time PCR to compare RGS2 expression between human asthmatic and control subjects. The AHR of RGS2 knockout mice was assessed by using invasive tracheostomy and unrestrained plethysmography. Effects of loss of RGS2 on mouse airway smooth muscle (ASM) remodeling, contraction, intracellular Ca(2+), and mitogenic signaling were determined in vivo and in vitro.
RGS2 was highly expressed in human and murine bronchial epithelium and ASM and was markedly downregulated in lungs of ovalbumin-sensitized/-challenged mice. Lung tissues and blood monocytes from asthma patients expressed significantly lower RGS2 protein (lung) and mRNA (monocytes) than from nonasthma subjects. The extent of reduction of RGS2 on human monocytes correlated with increased AHR. RGS2 knockout caused spontaneous AHR in mice. Loss of RGS2 augmented Ca(2+) mobilization and contraction of ASM cells. Loss of RGS2 also increased ASM mass and stimulated ASM cell growth via extracellular signal-regulated kinase and phosphatidylinositol 3-kinase pathways.
We identified RGS2 as a potent modulator of AHR and a potential novel therapeutic target for asthma.
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ABSTRACT: Glucocorticoids, also known as corticosteroids, induce effector gene transcription as a part of their anti-inflammatory mechanisms of action. Such genomic effects can be significantly enhanced by long-acting β2-adrenoceptor agonists (LABAs) and may contribute to the clinical superiority of inhaled corticosteroid (ICS)/LABA combinations in asthma and chronic obstructive pulmonary disease (COPD) over ICS alone. Using models of cAMP- and glucocorticoid-induced transcription in human bronchial epithelial BEAS-2B cells, we show that combining inhibitors of phosphodiesterase (PDE) 3 and PDE4 provides greater benefits compared to inhibiting either PDE alone. In respect of cAMP-dependent transcription, inhibitors of PDE3 (siguazodan, cilostazol) and PDE4 (rolipram, GSK256066, roflumilast N-oxide) each sensitized to the LABA, formoterol. This effect was magnified by dual PDE3 and PDE4 inhibition. Siguazodan plus rolipram was also more effective at inducing cAMP-dependent transcription than either inhibitor alone. Conversely, the concentration-response curve describing the enhancement of dexamethasone-induced, glucocorticoid response element-dependent transcription by formoterol was displaced to the left by PDE4, but not PDE3 inhibition. Overall similar effects were described for bona fides genes, including RGS2, CD200 and CRISPLD2. Importantly, the combination of siguazodan plus rolipram prolonged the duration of gene expression induced by formoterol, dexamethasone or dexamethasone plus formoterol. This was most apparent for RGS2, a bronchoprotective gene that may also reduce the pro-inflammatory effects of constrictor mediators. Collectively, these data provide a rationale for the use of PDE3 and PDE4 inhibitors in the treatment of COPD and asthma where they may enhance, sensitize and prolong the effects of LABA/ICS combination therapies.Molecular pharmacology 10/2014; 87(1). DOI:10.1124/mol.114.093393 · 4.12 Impact Factor
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ABSTRACT: Acting on the glucocorticoid receptor (NR3C1), glucocorticoids are widely used to treat inflammatory diseases. However, glucocorticoid resistance often leads to suboptimal asthma control. Since glucocorticoid-induced gene expression contributes to glucocorticoid activity, the aim of this study was to use a 2×glucocorticoid response element (GRE) reporter and glucocorticoid-induced gene expression to investigate approaches to combat cytokine-induced glucocorticoid resistance. Pre-treatment with tumor necrosis factor-α (TNF) or interleukin-1β inhibited dexamethasone-induced mRNA expression of the putative anti-inflammatory genes RGS2 and TSC22D3, or just TSC22D3, in primary human airway epithelial and smooth muscle cells, respectively. Dexamethasone-induced DUSP1 mRNA was unaffected. In human bronchial epithelial BEAS-2B cells, dexamethasone-induced TSC22D3 and CDKN1C expression (at 6 h) was reduced by TNF pre-treatment, whereas DUSP1 and RGS2 mRNAs were unaffected. TNF pre-treatment also reduced dexamethasone-dependent 2×GRE reporter activation. This was partially reversed by PS-1145 and c-jun N-terminal kinase (JNK) inhibitor VIII, inhibitors of IKK2 and JNK, respectively. However, neither inhibitor affected TNF-dependent loss of dexamethasone-induced CDKN1C or TSC22D3 mRNA. Similarly, inhibitors of the extracellular signal-regulated kinase, p38, phosphoinositide 3-kinase or protein kinase C pathways failed to attenuate TNF-dependent repression of the 2×GRE reporter. Fluticasone furoate, fluticasone propionate and budesonide were full agonists relative to dexamethasone, while GSK9027, RU24858, des-ciclesonide and GW870086X were partial agonists on the 2×GRE reporter. TNF reduced reporter activity in proportion with agonist efficacy. Full and partial agonists showed various degrees of agonism on RGS2 and TSC22D3 expression, but were equally effective at inducing CDKN1C and DUSP1, and did not affect the repression of CDKN1C or TSC22D3 expression by TNF. Finally, formoterol-enhanced 2×GRE reporter activity was also proportional to agonist efficacy and functionally reversed repression by TNF. As similar effects were apparent on glucocorticoid-induced gene expression, the most effective strategy to overcome glucocorticoid resistance in this model was addition of formoterol to high efficacy NR3C1 agonists.PLoS ONE 01/2015; 10(1):e0116773. DOI:10.1371/journal.pone.0116773 · 3.53 Impact Factor
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ABSTRACT: Inhaled glucocorticoids acting via the glucocorticoid receptor are a mainstay treatment option for individuals with asthma. There is a consensus that the remedial actions of inhaled glucocorticoids are due to their ability to suppress inflammation by modulating gene expression. While inhaled glucocorticoids are generally effective in asthma, there are subjects with moderate-to-severe disease in whom inhaled glucocorticoids fail to provide adequate control. For these individuals, asthma guidelines recommend that a long-acting β2-adrenoceptor agonist (LABA) be administered concurrently with an inhaled glucocorticoid. This so-called "combination therapy" is often effective and clinically superior to the inhaled glucocorticoid alone, irrespective of dose. LABAs, and another class of drug known as phosphodiesterase 4 (PDE4) inhibitors, may also enhance the efficacy of inhaled glucocorticoids in chronic obstructive pulmonary disease (COPD). In both conditions, these drugs are believed to work by elevating the concentration of cyclic adenosine-3',5'-monophosphate (cAMP) in target cells and tissues. Despite the success of inhaled glucocorticoid/LABA combination therapy, it remains unclear how an increase in cAMP enhances the clinical efficacy of an inhaled glucocorticoid. In this report, we provide a state-of-the-art appraisal, including unresolved and controversial issues, of how cAMP-elevating drugs and inhaled glucocorticoids interact at a molecular level to deliver enhanced anti-inflammatory benefit over inhaled glucocorticoid monotherapy. We also speculate on ways to further exploit this desirable interaction. Critical discussion of how these two drug classes regulate gene transcription, often in a synergistic manner, is a particular focus. Indeed, because interplay between glucocorticoid receptor and cAMP signaling pathways may contribute to the superiority of inhaled glucocorticoid/LABA combination therapy, understanding this interaction may provide a logical framework to rationally design these multicomponent therapeutics that was not previously possible.02/2015; 7. DOI:10.12703/P7-16