Abstract 18400: Opposite Roles of the Soluble Epoxide Hydrolase Domains in the Development of Vascular Calcification

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Abstract

Background: Soluble epoxide hydrolase (sEH) metabolizes endothelium-derived epoxyeicosatrienoic acids (EETs) through its hydrolase domain and also possesses a phosphatase domain, whose biological role remains uncertain. We aimed to study the role of sEH enzymatic activities in the vascular calcification process.

Methods and results: The inhibition of sEH hydrolase using trans-4-[4-(3-adamantan-1-yl-ureido)-cyclohexyloxy]-benzoic acid (t-AUCB) increased the calcium content of rat aortic rings cultured in procalcifying conditions during 7 days (addition of inorganic phosphate, 3.8 mM Pi). This increased calcification was confirmed by Alizarin Red and Von Kossa stainings. This was associated with an increase in tissue-nonspecific alkaline phosphatase (TNAP) activity and subsequent decrease in the calcification inhibitor pyrophosphate (PPi) in culture supernatants but not prevention of the increase in mRNA expression levels of the osteochondrogenic markers Msx2 and Sox9 induced by 3.8 mM Pi. All these effects were prevented by the inhibition of the EETs-synthesizing enzyme cytochrome P450 using fluconazole. In vivo,t-AUCB also increased the aortic calcification of mice with subtotal nephrectomy. At the opposite, inhibition of sEH phosphatase using N-acetyl-S-farnesyl-L-cysteine (AFC) or ebselen reduced the calcification of aortic rings cultured with 3.8mM Pi. This was associated with a decreased Msx2 and Sox9 expression levels. Nor t-AUCB or AFC altered PPi-induced calcification of deendothelialized aortic rings or isolated human VSMC.

Conclusion: Taken together, our results demonstrate that the phosphatase and hydrolase domains of sEH regulate the vascular calcification process through endothelium-dependent mechanisms. The inhibition of the phosphatase domain of sEH represents a new pharmacological target in the prevention of vascular calcification.

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