EDHF function in the ductus arteriosus: evidence against involvement of epoxyeicosatrienoic acids and 12S-hydroxyeicosatetraenoic acid

Abstract

We have previously shown (Ref. 2) that endothelium-derived hyperpolarizing factor (EDHF) becomes functional in the fetal ductus arteriosus on removal of nitric oxide and carbon monoxide. From this, it was proposed that EDHF originates from a cytochrome P-450 (CYP450)-catalyzed reaction being inhibited by the two agents. Here, we have examined in the mouse ductus whether EDHF can be identified as an arachidonic acid product of a CYP450 epoxygenase and allied pathways. We did not detect transcripts of the mouse CYP2C subfamily in vessel, while CYP2J subfamily transcripts were expressed with CYP2J6 and CYP2J9. These CYP2J hemoproteins were also detected in the ductus by immunofluorescence microscopy, being colocalized with the endoplasmic reticulum in both endothelial and muscle cells. Distinct CYP450 transcripts were also detected and were responsible for omega-hydroxylation (CYP4A31) and 12R-hydroxylation (CYP4B1). Mass spectrometric analysis showed formation of epoxyeicosatrienoic acids (EETs) in the intact ductus, with 11,12- and 14,15-EETs being more prominent than 5,6- and 8,9-EETs. However, their yield did not increase with nitric oxide/carbon monoxide suppression, nor did it abate with endothelium removal. No evidence was obtained for formation of 12R-hydroxyeicosatrienoic acid and omega-hydroxylation products. 2S-hydroxyeicosatetraenoic acid was instead detected, and, contrary to data implicating this compound as an alternative EDHF, its suppression with baicalein did not modify the EDHF-mediated relaxation to bradykinin. We conclude that none of the more common CYP450-linked arachidonic acid metabolites appears to qualify as EDHF in mouse ductus. We speculate that some novel eicosanoid or a totally unrelated compound requiring CYP450 for its synthesis accounts for EDHF in this vessel.

Cimetidine-associated patent ductus arteriosus is mediated via a cytochrome P450 mechanism independent of H2 receptor antagonism

“…2B). In contrast to prior reports on limited CYP expression in the DA [18, 19], these results demonstrate the presence of multiple CYP enzymes that may be inhibited by cimetidine.…”

“…However, only limited information is available on the expression of CYP enzymes in the DA [18, 19]. Conventional PCR demonstrated the presence of Cyp2d22 (similar to human CYP2D6 ), and the CYP3A mouse homologues Cyp3a11, Cyp3a13, Cyp3a16, Cyp3a25 and Cyp3a44 in the fetal and newborn DA.…”

Ocular cytochrome P450s and transporters: roles in disease and endobiotic and xenobiotic disposition

“…The precise signaling pathway by which 12( R )-HETrE influences corneal neovascularization is uncertain, however, some insights have been derived from the rabbit eye hypoxia model developed by Laniado-Schwartzman and colleagues (Baragatti et al, 2009; Mastyugin et al, 2004; Mezentsev et al, 2005; Seta et al, 2007). These researchers have suggested that vascular endothelial growth factor (VEGF) expression depends on 12( R )-HETrE levels and that CYP4B1 expression parallels VEGF expression in the cornea.…”

Hydrogen sulfide in the mouse ductus arteriosus: a naturally occurring relaxant with potential EDHF function

“…As expected, this newly emerging EDHF is suppressed by excess potassium or the combined treatment with drugs interfering with small-and intermediate-conductance Ca 2ϩ -activated K ϩ channels, i.e., apamin plus 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole (2). Based on these findings and data from other vascular districts, we then examined whether ductal EDHF may be identified with an epoxyeicosatrienoic acid or 12(S)-hydroxyeicosatrienoic acid, but results were negative (4). Nevertheless, we speculated at the time on H 2 S playing such role, and this possibility has now become more plausible with the demonstration of EDHF-like properties in this agent (17,19).…”