Haoxiao Zuo

The novel compound Sul-121 inhibits airway inflammation and hyperresponsiveness in experimental models of chronic obstructive pulmonary disease.

COPD is characterized by persistent airflow limitation, neutrophilia and oxidative stress from endogenous and exogenous insults. Current COPD therapy involving anticholinergics, β2-adrenoceptor agonists and/or corticosteroids, do not specifically target oxidative stress, nor do they reduce chronic pulmonary inflammation and disease progression in all patients. Here, we explore the effects of Sul-121, a novel compound with anti-oxidative capacity, on hyperresponsiveness (AHR) and inflammation in experimental models of COPD. Using a guinea pig model of lipopolysaccharide (LPS)-induced neutrophilia, we demonstrated that Sul-121 inhalation dose-dependently prevented LPS-induced airway neutrophilia (up to ~60%) and AHR (up to ~90%). Non-cartilaginous airways neutrophilia was inversely correlated with blood H2S, and LPS-induced attenuation of blood H2S (~60%) was prevented by Sul-121. Concomitantly, Sul-121 prevented LPS-induced production of the oxidative stress marker, malondialdehyde by ~80%. In immortalized human airway smooth muscle (ASM) cells, Sul-121 dose-dependently prevented cigarette smoke extract-induced IL-8 release parallel with inhibition of nuclear translocation of the NF-κB subunit, p65 (each ~90%). Sul-121 also diminished cellular reactive oxygen species production in ASM cells, and inhibited nuclear translocation of the anti-oxidative response regulator, Nrf2. Our data show that Sul-121 effectively inhibits airway inflammation and AHR in experimental COPD models, prospectively through inhibition of oxidative stress.

Epac Function and cAMP Scaffolds in the Heart and Lung

Evidence collected over the last ten years indicates that Epac and cAMP scaffold proteins play a critical role in integrating and transducing multiple signaling pathways at the basis of cardiac and lung physiopathology. Some of the deleterious effects of Epac, such as cardiomyocyte hypertrophy and arrhythmia, initially described in vitro, have been confirmed in genetically modified mice for Epac1 and Epac2. Similar recent findings have been collected in the lung. The following sections will describe how Epac and cAMP signalosomes in different subcellular compartments may contribute to cardiac and lung diseases.

Cigarette Smoke Up-regulates PDE3 and PDE4 to Decrease cAMP in Airway Cells

cAMP is a central second messenger that broadly regulates cell function and can underpin pathophysiology. In chronic obstructive pulmonary disease, a lung disease primarily provoked by cigarette smoke (CS), the activation of cAMP‐dependent pathways, via inhibition of hydrolyzing PDEs, is a major therapeutic strategy. Mechanisms that disrupt cAMP signalling in airway cells, in particular regulation of endogenous PDEs, are poorly understood.

Monitoring local pulmonary cAMP levels: combining precision cut lung slice (PCLS) and fluorescence resonance energy transfer (FRET) technologies in mice

RATIONALE Cyclic AMP (cAMP) is one of the most important second messengers and is involved as a target for the therapy of chronic obstructive pulmonary disease (COPD), an airway disease primarily provoked by cigarette smoke . Cyclic nucleotide hydrolyzing phosphodiesterases (PDEs) are able to degrade cAMP or cGMP within subcellular compartments, thereby potentially altering pulmonary responses including airway contractility and inflammation. In the present study, we combine the precision cut lung slice (PCLS) technique in mice with fluorescence resonance energy transfer (FRET) to monitor cAMP in real time. METHODS To monitor the cAMP levels in lung tissues, transgenic mice (CAG-Epac1-camps) that express the FRET-based cAMP sensor Epac1-camps were used for the preparation of PCLS. The β2-adrenergic receptor agonist fenoterol was applied to elevate intracellular cAMP. To achieve PDE subtype specific inhibition, the PDE4 inhibitor rolipram, the PDE3 inhibitor cilostamide and the PDE2 inhibitor BAY60-7550 were used. The nonselective PDE inhibitor 3-isobutyl-1-methylxanthine (IBMX) served as a control. Moreover, lung slices were exposed to 2.5 % cigarette smoke extract (CSE) for 24 hours used as a COPD model in vitro. RESULTS and CONCLUSIONS We provide evidence that the FRET and PCLS technologies can be combined in CAG-Epac1-camps mice to measure global cAMP. Moreover, we found in fenoterol-stimulated PCLS that PDE4 accounts for more than 80% of the total cAMP-PDE activity. Besides PDE4, PDE3 known as a cGMP-inhibited PDE also contributes to cAMP hydrolysis, indicating that cGMP may modulate the maintenance of local cAMP in PCLS. In contrast, the cGMP-activated PDE2 plays a limited role in cAMP hydrolysis. Exposure to CSE did not alter the FRET signal in the presence of the PDE4 inhibitor under fenoterol stimulated conditions. In contrast, we found that a significant increase could be observed in PDE3-dependent FRET responses (p<0.05). Under basal conditions, CSE treatment altered local cAMP levels by significantly increasing both PDE4 and PDE3 inhibitor effects. Therefore, as the major lung cyclic nucleotide hydrolyzing enzymes PDE3 and PDE4 are both involved in the local regulation of the cAMP levels in the β2-AR microdomain, our findings suggest that exposure to CSE induced alterations in the PDEs activity profile both under basal conditions and in the presence of the β2-agonist fenoterol.