Prostaglandin E2-induced intercellular adhesion molecule-1 expression is mediated by cAMP/Epac signalling modules in bEnd.3 brain endothelial cells

Background and Purpose Prostaglandin E2 (PGE2) has been implicated in the regulation of adhesion molecules, leukocyte adhesion and infiltration into inflamed site. However, the underlying mechanism therein involved remains ill-defined. In this study, we explored its cellular mechanism of action in the regulation of the intercellular adhesion molecule-1 (ICAM-1) expression in the brain endothelial cells. Experimental Approach bEnd.3 cells, the murine cerebrovascular endothelial cell line and primary mouse brain endothelial cells were treated with PGE2 with or without agonists/antagonists of PGE2 receptors and associated signalling molecules. ICAM-1 expression, Akt phosphorylation and activity of NF-B were determined by reverse transcription polymerase chain reaction (RT-PCR), immunoblot analysis, luciferase assay and immunocytochemistry. Key Results PGE2 significantly up-regulated the expression of ICAM-1, which was blocked by EP4 antagonist (ONO-AE2-227) and knock-down of EP4. PGE2 effects were mimicked by forskolin, dibutyryl cAMP (dbcAMP) and an exchange protein directly activated by cAMP (Epac) activator (8-Cpt-cAMP) but not a protein kinase A activator (N6-Bnz-cAMP). PGE2-induced ICAM-1 expression was reduced by knock-down of Epac1. A PI3K specific inhibitor (LY294002), Akt inhibitor VIII (Akti) and NF-B inhibitors (Bay-117082 and MG-132) attenuated the induction of ICAM-1 by PGE2. PGE2, dbcAMP and 8-Cpt-cAMP induced the phosphorylation of Akt, IB kinase and IB and the translocation of p65 to the nucleus and increased NF-B dependent reporter gene activity, which was diminished by Akti. Conclusion and Implications Our findings suggest that PGE2 induces ICAM-1 expression via EP4 receptor and Epac/Akt/NF-B signalling pathway in bEnd.3 brain endothelial cells, supporting its pathophysiological role in brain inflammation.