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Oxidative stress-induced glucocorticoid resistance is prevented by dual PDE3/PDE4 inhibition in human alveolar macrophages


Javier Milara, Ph.D., Unidad de Investigación, Consorcio Hospital General Universitario, Avenida tres cruces s/n, E-46014 Valencia, Spain. E-mail:


Background Oxidative stress is present in airway diseases such as severe asthma or Chronic Obstructive Pulmonary Disease and contributes to the low response to glucocorticoids through the down-regulation of histone deacetylase (HDAC) activity.

Objective To study the effects of the phosphodiesterase (PDE)-3 and 4 inhibitors and their combination vs. glucocorticoids in a model of lipopolysaccharide (LPS)-induced cytokine release in alveolar macrophages under oxidative stress conditions.

Methods Differentiated U937 or human alveolar macrophages were stimulated with H2O2 (10–1000 μm) or cigarette smoke extract (CSE, 0–15%) for 4 h before LPS (0.5 μg/mL, 24 h) addition. In other experiments, cells were pre-treated with dexamethasone or budesonide (10−9–10−6 m), with the PDE4 inhibitor rolipram (10−9–10−5 m), PDE3 inhibitor motapizone (10 μm), 3′,5′-cyclic monophosphate enhancer PGE2 (10 nm), or with the combination of rolipram (10−6 m)+PGE2 (10 nm)+motapizone (10 μm) 15 min before oxidants. IL-8 and TNF-α were measured by ELISA and HDAC activity by a colorimetric assay.

Results Budesonide and dexamethasone produced a concentration-dependent inhibition of the LPS-induced IL-8 and TNF-α secretion with an Emax about 90% of inhibition, which was reduced by approximately 30% in the presence of H2O2 or CSE. Pre-treatment with rolipram, motapizone or PGE2 only reached about 20% of inhibition but was not affected by oxidative stress. In contrast, PDE4/PDE3 combination in presence of PGE2 effectively inhibited the LPS-induced cytokine secretion by about 90% and was not affected by oxidative stress. Combined PDE4 and PDE3 inhibition reversed glucocorticoid resistance under oxidative stress conditions. HDAC activity was reduced in the presence of oxidative stress, and in contrast to glucocorticoids, pre-treatment with PDE4/PDE3 combination was able to prevent HDAC inactivity.

Conclusions & Clinical Relevance This study shows that the combination of the PDE3/PDE4 inhibitors prevents alveolar macrophage activation in those situations of glucocorticoid resistance, which may be of potential interest to develop new effective anti-inflammatory drugs in airway diseases.

Cite this as: J. Milara, A. Navarro, P. Almudéver, J. Lluch, E. J. Morcillo and J. Cortijo, Clinical & Experimental Allergy, 2011 (41) 535–546.