Suzuki_2026_J.Biol.Chem__113615

Soluble epoxide hydrolase exhibits two distinct activities: C-terminal epoxide hydrolase (C-EH) and N-terminal phosphatase (N-phos). Epoxy fatty acids are the primary substrates of C-EH and exhibit anti-inflammatory properties. N-phos hydrolyses lipid phosphate monoesters, such as isoprenoid pyrophosphates, lysophosphatidic acid (LPA), and sphingosine-1-phosphate (S1P). We found that inhibiting both sEH activities, rather than just C-EH, suppressed vascular endothelial cell (VEC) activation. A negatively charged polar lipid (NCPL) fraction from mouse liver extracts also suppressed VEC activation. This effect was canceled by sEH treatment. The inhibition of isoprenoid and nitric oxide synthesis and the treatment with epoxyeicosatrienoic acid, LPAs, and typical S1P did not inhibit VEC activation, suggesting that a metabolite other than these is responsible for suppressing VEC activation. To identify the sEH substrate responsible for this activity, we performed a non-targeted LC-MS/MS analysis using NCPL. As a result, phospho-N-acylethanolamine emerged as a candidate. Enzyme kinetic analysis confirmed that species in this class, including arachidonoyl ethanolamide (anandamide) phosphate, were favorable N-phos substrates. The exogenous addition of anandamide phosphate inhibited VEC activation-associated expression of cell adhesion molecules at concentrations of 3 microM or more.