A ceramide analog inhibits cPLA2 activity and consequent PGE2 formation in LPS-stimulated macrophages

Prostaglandin E2 (PGE2) is an important mediator of the inflammatory response. Phospho-ceramide analogue-1 (PCERA-1), a synthetic phospholipid-like molecule, was previously reported to modulate pro- and anti-inflammatory cytokine production. We show here that PCERA-1 inhibited LPS-stimulated PGE2 production in RAW264.7 macrophages, without affecting COX-2 expression. Furthermore, PCERA-1 efficiently suppressed arachidonic acid (AA) release in response to LPS. The dephosphorylated derivative of PCERA-1, ceramide analogue-1 (CERA-1), mimicked the inhibitory effect of PCERA-1 on AA release and PGE2 production in macrophages. Inhibition of PGE2 production by CERA-1 was completely rescued by addition of exogenous AA. Importantly, PCERA-1 and ceramide-1-phosphate (C1P) stimulated the enzymatic activity of cPLA2α in an in vitro assay, whereas CERA-1 and ceramide inhibited both basal and C1P-stimulated cPLA2α activity. Collectively, these results indicate that CERA-1 suppresses AA release and subsequent PGE2 production in LPS-stimulated macrophages by direct interaction with cPLA2, and suggest that ceramide may similarly counteract C1P effect on cPLA2 activity in cells. The suppression of PGE2 production is suggested to contribute to the anti-inflammatory action of PCERA-1.