Agonist-specific desensitization of PGE(2)-stimulated cAMP signaling due to upregulated phosphodiesterase expression in human lung fibroblasts

Pulmonary fibrosis is characterized by fibroblasts persisting in an activated form, producing excessive fibrous material that destroys alveolar structure. The second messenger molecule cyclic 3′,5′-adenosine monophosphate (cAMP) has antifibrotic properties, and prostaglandin E(2) (PGE(2)) can stimulate cAMP production through prostaglandin E (EP)(2) and EP(4) receptors. Although EP receptors are attractive therapeutic targets, the effects of long-term exposure to PGE(2) have not been characterized. To determine the effects of long-term exposure of lung fibroblasts to PGE(2), human fetal lung (HFL)-1 cells were treated for 24 h with 100 nM PGE(2) or other cAMP-elevating agents. cAMP levels stimulated by acute exposure to PGE(2) were measured using a fluorescent biosensor. Pretreatment for 24 h with PGE(2) shifted the concentration-response curve to PGE(2) rightward by approximately 22-fold but did not affect responses to the beta-adrenoceptor agonist isoproterenol. Neither isoproterenol nor forskolin pretreatment altered PGE(2) responses, implying that other cAMP-elevating agents do not induce desensitization. Use of EP(2)- and EP(4)-selective agonists and antagonists suggested that PGE(2)-stimulated cAMP responses in HFL-1 cells are mediated by EP(2) receptors. EP(2) receptors are resistant to classical mechanisms of agonist-specific receptor desensitization, so we hypothesized that increased PDE activity mediates the loss of signaling after PGE(2) pretreatment. PGE(2) treatment upregulated messenger RNA for PDE3A, PDE3B, PDE4B, and PDE4D and increased overall PDE activity. The PDE4 inhibitor rolipram partially reversed PGE(2)- mediated desensitization and PDE4 activity was increased, but rolipram did not alter responses to isoproterenol. The PDE3 inhibitor cilostazol had minimal effect. These results show that long-term exposure to PGE(2) causes agonist-specific desensitization of EP(2) receptor-stimulated cAMP signaling through the increased expression of PDE isozymes, most likely of the PDE4 family.