Summary.
Phosphodiesterase (PDE) enzymes catalyze the breakdown of cAMP and thus may have potent immunomodulatory activity. We were interested to investigate whether PDE4 inhibitor rolipram and a non-specific PDE inhibitor theophylline modulate ERK1/2 signaling pathway in freshly isolated human PBMCs. To determine the effect of rolipram and theophylline on the phosphorylation and activation of ERK1/2 signaling pathway, human PBMCs (5 x 106) ml were serum-starved and treated with 1 μg/ml Der f. After 15 min incubation cells were harvested and lysed in cell lytic buffer. Equal amounts of lysates (40 μg) were subjected to SDS-PAGE and Western blot analysis using an antibody phospho-p44/42 MAP kinase (Thr 202/Tyr 204), specific for the activated form of ERK1/2. This antibody recognizes ERK when phosphorylated on both threonine and tyrosine residues. Cells treated with Der f exhibited detectable phosphorylation of ERK1/2 at 15 min. Our present study demonstrates that PDE4 inhibitor rolipram (10-5 M) and nonspecific phosphodiesterase inhibitor theophylline (10-5 M) decrease ERK phosphorylation induced by Der f. MEK/ERK inhibitor U0126 (10 μM) alone or in combination with rolipram (10-5 M) downregulated the gene expression of IL-5 mRNA of human PBMCs stimulated with Der f.