Endothelial ICAM-1 protein induction is regulated by cytosolic phospholipase A₂α via both NF-κB and CREB transcription factors

The regulated expression of ICAM-1 plays an important role in inflammatory processes and immune responses. The present study aimed to determine the in vivo involvement of cytosolic phospholipase A₂α (cPLA ₂α) in ICAM-1 overexpression during inflammation and to elucidate the cPLA₂α-specific role in signal events leading to ICAM-1 upregulation in endothelial cells. cPLA₂α and ICAM-1 upregulation were detected in inflamed paws of mice with collagen-induced arthritis and in periepididymal adipose tissue of mice fed a high-fat diet. Intravenous injection of 2 mg/kg oligonucleotide antisense against cPLA ₂α (AS) that reduced cPLA₂α upregulation also decreased ICAM-1 overexpression, suggesting a key role of cPLA ₂α in ICAM-1 upregulation during inflammation. Preincubation of endothelial ECV-304 cells that express ICAM-1 and of HUVEC that express ICAM-1 and VCAM-1 with 1 μM AS prevented cPLA₂α and the adhesion molecule upregulation induced by TNF-α and inhibited their adherence to phagocyte like-PLB cells. Whereas AS did not inhibit NADPH oxidase 4-NADPH oxidase activity, inhibition of oxidase activity attenuated cPLA ₂α activation, suggesting that NADPH oxidase acts upstream to cPLA₂α. Attenuating cPLA₂α activation by AS or diphenylene iodonium prevented the induction of cyclooxygenase-2 and the production of PGE₂ that were essential for ICAM-1 upregulation. Inhibition of cPLA₂α activity by AS inhibited the phosphorylation of both p65 NF-κB on Ser⁵³⁶ and protein kinase A-dependent CREB. To our knowledge, our results are the first to show that CREB activation is involved in ICAM-1 upregulation and suggest that cPLA ₂α activated by NADPH oxidase is required for sequential phosphorylation of NF-κB by an undefined kinase and CREB activation by PGE₂-mediated protein kinase A.