Impact of aldosterone synthase inhibitor FAD286 on steroid hormone profile in human adrenocortical cells.

Inhibition of aldosterone synthase (CYP11B2) is an alternative treatment option to mineralocorticoid receptor antagonism to prevent harmful aldosterone effects. FAD286 is the best characterized aldosterone synthase inhibitor. However, to date, no study has used sensitive liquid chromatography-tandem mass spectrometry to characterize in detail the effect of FAD286 on the secreted steroid hormone profile of adrenocortical cells. Basal aldosterone production in NCI-H295R cells was detectable and 9-fold elevated after stimulation with angiotensin II. FAD286 inhibited this increase, showing a maximal effect at 10 nmol/l. Higher concentrations of FAD286 did not further reduce aldosterone concentrations, but showed a parallel reduction in corticosterone, cortisol and cortisone levels, reflecting additional inhibition of steroid-11 beta-hydroxylase (CYP11B1). Pregnenolone, progesterone and 17-OH-progesterone levels remained unaffected. In conclusion, the aldosterone synthase inhibitor FAD286 lowers angiotensin II-induced aldosterone concentrations in adrenocortical cells but the relative lack of selectivity over CYP11B1 is evident at higher FAD286 concentrations

An LC-MS/MS method for steroid profiling during adrenal venous sampling for investigation of primary aldosteronism

Item does not contain fulltextBACKGROUND: Steroid profiling for diagnosis of endocrine disorders featuring disordered production of steroid hormones is now possible from advances in liquid chromatography with tandem mass spectrometry (LC-MS/MS). Adrenal venous (AV) measurements of aldosterone and cortisol are a standard practice in the clinical work-up of primary aldosteronism, but do not yet take advantage of steroid profiling. METHODS: A novel LC-MS/MS based method was developed for simultaneous measurement of 15 adrenal steroids: aldosterone, corticosterone, 11-deoxycorticosterone, progesterone, pregnenolone, cortisone, cortisol, 11-deoxycortisol, 17-hydroxyprogesterone, androstenedione, dehydroepiandrosterone, dehydroepiandrosterone-sulfate, 21-deoxycortisol, 18-oxocortisol and 18-hydroxycortisol. These were compared in peripheral venous (pV) and AV plasma from 70 patients undergoing AV sampling with and without cosyntropin stimulation. Aldosterone and cortisol levels measured by LC-MS/MS were compared with those measured by immunoassay. RESULTS: Reproducibility of measurements with coefficients of variation </=10% as well as analytical sensitivity sufficient to measure low pV levels particularly of aldosterone demonstrate the utility of the assay for profiling adrenal steroids in primary aldosteronism. Method comparisons indicated assay and concentration dependent differences of cortisol and aldosterone concentrations measured by immunoassay and LC-MS/MS. Median AV/pV ratios of 11-deoxycortisol (53.0), 17-hydroxyprogesterone (33.4), pregnenolone (62.4), androstenedione (40.6) and dehydroepiandrosterone (33.3) were 2.9- to, 5.4-fold larger than those for cortisol (11.6), with additionally generally larger increases than for cortisol with than without cosyntropin stimulation. CONCLUSION: Our LC-MS/MS assay, in addition to improvements over existing immunoassay measurements of aldosterone and cortisol, offers profiling of 13 other adrenal steroids, providing a potentially useful method for the clinical work-up of patients with primary aldosteronism. In particular, the larger AV/pV ratios of several steroids compared to cortisol suggest more sensitive alternatives to the latter for assessing positioning of AV sampling catheters

Mass Spectrometry-Based Adrenal and Peripheral Venous Steroid Profiling for Subtyping Primary Aldosteronism

BACKGROUND: Differentiating patients with primary aldosteronism caused by aldosterone-producing adenomas (APAs) from those with bilateral adrenal hyperplasia (BAH), which is essential for choice of therapeutic intervention, relies on adrenal venous sampling (AVS)-based measurements of aldosterone and cortisol. We assessed the utility of LC-MS/MS-based steroid profiling to stratify patients with primary aldosteronism. METHODS: Fifteen adrenal steroids were measured by LC-MS/MS in peripheral and adrenal venous plasma from AVS studies for 216 patients with primary aldosteronism at 3 tertiary referral centers. Ninety patients were diagnosed with BAH and 126 with APAs on the basis of immunoassay-derived adrenal venous aldosterone lateralization ratios. RESULTS: Among 119 patients confirmed to have APAs at follow-up, LC-MS/MS-derived lateralization ratios of aldosterone normalized to cortisol, dehydroepiandrosterone, and androstenedione were all higher (P < 0.0001) than immunoassay-derived ratios. The hybrid steroids, 18-oxocortisol and 18-hydroxycortisol, also showed lateralized secretion in 76% and 35% of patients with APAs. Adrenal venous concentrations of glucocorticoids and androgens were bilaterally higher in patients with BAH than in those with APAs. Consequently, peripheral plasma concentrations of 18-oxocortisol were 8.5-fold higher, whereas concentrations of cortisol, corticosterone, and dehydroepiandrosterone were lower in patients with APAs than in those with BAH. Correct classification of 80% of cases of APAs vs BAH was thereby possible by use of a combination of steroids in peripheral plasma. CONCLUSIONS: LC-MS/MS-based steroid profiling during AVS achieves higher aldosterone lateralization ratios in patients with APAs than immunoassay. LC-MS/MS also enables multiple measures for discriminating unilateral from bilateral aldosterone excess, with potential use of peripheral plasma for subtype classification

The role of Toll-like receptors in the immune-adrenal crosstalk

Sepsis and septic shock remain major health concerns worldwide, and rapid activation of adrenal steroid release is a key event in the organism's first line of defense during this form of severe illness. Toll-like receptors (TLRs) are critical in the early immune response upon bacterial infection, and recent data from our lab demonstrate a novel link between the innate immune system and the adrenal stress response mediated by TLRs. Glucocorticoids and TLRs regulate each other in a bidirectional way. Bacterial toxins acting through TLRs directly activate adrenocortical steroid release. TLR-2 and TLR-4 are expressed in human and mice adrenals and TLR-2 deficiency is associated with an impaired glucocorticoid response. Furthermore, TLR-2 deficiency in mice is associated with marked cellular alterations in adrenocortical tissue. TLR-2-deficient mice have an impaired adrenal corticosterone release following inflammatory stress induced by bacterial cell wall compounds. This defect appears to be associated with a decrease in systemic and intraadrenal cytokine expression. In conclusion, TLRs play a crucial role in the immune–adrenal crosstalk. This close functional relationship needs to be considered in the treatment of inflammatory diseases requiring an intact adrenal stress response.Fil: Bornstein, S. R.. Heinrich Heine University; AlemaniaFil: Ziegler, C. G.. Heinrich Heine University; AlemaniaFil: Krug, A. W.. Heinrich Heine University; AlemaniaFil: Kanczkowski, W.. Heinrich Heine University; AlemaniaFil: Besuhli, Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: McCann, S. M.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Wirth, M.. University of Dresden; AlemaniaFil: Zacharowski, K.. Heinrich Heine University; Alemani