Nature of Endothelium-Derived Hyperpolarizing Factor, EDHF

In arteries, the endothelium modulates precapillary resistance, maintains patency and regulates the structure and growth of blood vessels. Among the mediators involved in these endothelial functions, a endothelium-derived hyperpolarizing factor (EDHF) induces relaxation of vascular smooth muscle (VSM). So far, pharmacological evidence suggest epoxyeicosatrienoic acids (Its) account for EDHF However, inhibitors of epoxygenases (i.e. enzyme generating EETs) have numerous undesirable side effects, which have been s source of doubt. A cascade bioassay method will be used to discriminate the site of action of EDHF-inhibitors. In addition, spectrofluorometric assays for intracellular Ca2+, free fatty acids and epoxygenase activity will help study the mechanisms that control the output o , EDHF/EETs (measured by gas chromatography/mass spectrometry) by endothelial cells. thus, in a first series of experiments, in bioassay experiments using cultured bovine aortic endothelial cells (BAEC) a I a source of EDHF, and a ring of bovine coronary artery without endothelium, we will define suitable EDHF-inhibitors. Secondly, experiments will asses the role of receptor-dependent activation of phospholipases in the release EDHF/EETs independently of the de novo synthesis by endothelia epoxygenase(s). In a second series of experiments, we will assess the presence of receptors for EDHF/EETs. We will characterize the basis for organ and/or species-related differences that have bee documented in the pharmacology of EDHF. Finally, we will address the role of EDHF/EET as determinant of endothelial reactivity to agonists. Risk factors for cardiovascular diseases (hypentension diabetes, atherogenic stimuli) are associated with impaired endothelial functions. Endothelial cells n longer respond to agonists to release vasodilators and anti-thrombotic mediators. To delineate the cellular and molecular basis of the types of endothelial dysfunction, it is instrumental to identify the different mediators that are involved and their role in healthy and diseased blood vessels. Hence, it is important to determine whether or not EETs are EDHF.