A phosphodiesterase 4 inhibitor, roflumilast N-oxide, inhibits human lung fibroblast functions in vitro

The PDE4 inhibitor roflumilast mitigates bleomycin-induced lung fibrotic remodeling in rodents. In the current study it was explored whether roflumilast N-oxide, the active metabolite of roflumilast influences functions of cultured lung fibroblasts. Cells of the human foetal lung fibroblast strain GM06114 were stimulated with TNF-α (5 ng ml−1) and cell surface ICAM-1 and eotaxin release were assessed. [methyl-3H] thymidine incorporation was measured following stimulation with bFGF (10 ng ml−1). α-Smooth muscle actin (protein), CTGF (mRNA) and fibronectin (mRNA) were determined secondary to TGFß1 (1 ng ml−1). In the presence of PGE2 (1 nM), roflumilast N-oxide reduced TNF-α-induced ICAM-1 and eotaxin by about 70% and >90% with half-maximum inhibition at 0.9 nM and 0.5 nM, respectively. Roflumilast N-oxide also attenuated [methyl-3H] thymidine incorporation secondary to bFGF by about 75% with half-maximum inhibition at 0.7 nM when cells were co-incubated with IL-1ß (10 pg ml−1). In the presence of this cytokine roflumilast N-oxide (1 μM) diminished TGFß1-induced expression of α-smooth muscle actin and transcripts of CTGF and fibronectin. In addition, IL-1ß up-regulated PDE4 activity in the lung fibroblasts. Taken together, these findings indicate that roflumilast N-oxide directly targets human lung fibroblasts, which may at least partially explain the efficacy of roflumilast to mitigate a pulmonary fibrotic response in vivo.