Objective: A major driver of cardiovascular morbidity and mortality in primary aldosteronism (PA) is endothelial dysfunction. Endothelial dysfunction in PA is well established, but the precise mechanism underlying this phenomenon is incompletely investigated. Specifically, the cytochrome P450 monoxygenase (CYP)-dependent epoxyeicosatrienoic (EET) acid pathway has not been studied comprehensively. Promiscuity of mineralocorticoid receptors due to activation by glucocorticoids further complicates understanding of mechanisms. Design and method: Primary human coronary artery smooth muscle and endothelial cells were challenged with aldosterone concentrations typically found in primary aldosteronism (1 nM) in a background of physiological cortisol levels (140 nM). In both cell types, we analysed expression levels of transcripts which are vital for the generation and inactivation of EETs as well as their proposed mode of action. Smooth muscle cell responses to EETs were assessed by calcium imaging with Fura2. Release of EETs by aldosterone and cortisol-treated endothelial cells were measured by high-pressure liquid chromatography followed by tandem mass spectrometry. Results: Glucocorticoids and mineralocorticoids synergistically reduced BKCa potassium channel transcription in smooth muscle cells. Antagonist studies demonstrated that this effect was predominantly related to glucocorticoid receptor stimulation. The aldosterone-mediated decrease in BKCa mRNA levels was not reflected at the protein level. In line with preserved BKCa protein expression, aldosterone treatment did not affect the calcium response of smooth muscle cells to 14,15 EET stimulation. Endothelial cells challenged with aldosterone showed unchanged mRNA levels of the CYP monoxygenases and epoxide hydrolases, which are relevant for the synthesis and degradation of EETs, respectively. Mass spectrometry of supernatants from acetylcholine-stimulated endothelial cells did not reveal significant differences in concentrations of any of the 4 EET regioisomeres or their respective degradation products (dihydroxyeicosatrienoic acids). Conclusions: In endothelial dysfunction related to mineralocorticoid excess, targeting the EET pathway may be a promising therapeutic approach because all constituents of EET signalling appear to be expressed and functionally intact. Soluble epoxide hydrolase inhibition may thus represent a feasible specific therapy of endothelial dysfunction in PA. The effects of glucocorticoids on BKCa channel expression warrant further investigation under conditions such as glucocorticoid co-secretion in PA and Cushing syndrome.
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