Endothelial epoxyeicosatrienoic acid release is intact in aldosterone excess.

BACKGROUND AND AIMS: Endothelial dysfunction (ED) is considered to be a major driver of the increased incidence of cardiovascular disease in primary aldosteronism (PA). The functionality of the epoxyeicosatrienoic acid (EET) pathway, involving the release of beneficial endothelium-derived lipid mediators, in PA is unknown. Evidence suggests this pathway to be disturbed in various models of experimental hypertension. We therefore assessed EET production in primary human coronary artery endothelial cells exposed to aldosterone excess and measured circulating EET in patients with PA. METHODS: We used qPCR to investigate changes in the expression levels of essential genes for the synthesis and degradation of EET, calcium imaging to address the functional impact on overall endothelial function, as well as mass spectrometry to determine endothelial synthetic capacity to release EET upon stimulation. RNA-seq was performed to gain further mechanistic insights. Eicosanoid concentrations in patient's plasma were also determined by mass spectrometry. RESULTS: Aldosterone, while eliciting proinflammatory VCAM1 expression and disturbed calcium response to acetylcholine, did not negatively affect stimulated release of endothelial EET. Likewise, no differences were observed in eicosanoid concentrations in plasma from patients with PA when compared to essential hypertensive controls. However, an inhibitor of soluble epoxide hydrolase abrogated aldosterone-mediated VCAM1 induction and led to a normalized endothelial calcium response probably by restoring expression of CHRNE. CONCLUSION: EET release appears intact despite aldosterone excess. Epoxide hydrolase inhibition may revert aldosterone-induced functional changes in endothelial cells. These findings indicate a potential new therapeutic principle to address ED, which should be explored in future preclinical and clinical trials.

Fast and reliable quantification of aldosterone, cortisol and cortisone via LC-MS/MS to study 11ß-hydroxysteroid dehydrogenase activities in primary cell cultures.

Cell culture experiments can support characterization of enzymatic activities in healthy and tumorous human tissues. Liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) enables simultaneous measurement of several steroids from a single sample, facilitating analysis of molecular pathways involved in steroid biosynthesis. We developed a reliable but fast method for quantification of cortisol, cortisone and aldosterone in cell culture supernatant. Validation, including investigation of matrix-matched calibration, was performed for two different cell types. Utility of the method was demonstrated in the study of 11ß-hydroxysteroid dehydrogenase type 2 (HSD11B2) activity under conditions of glucocorticoid and mineralocorticoid excess in different cell types. Aldosterone, cortisol and cortisone were extracted by liquid-liquid extraction (LLE) with methyl tert-butyl ether from 1 mL of cell culture supernatant. Steroids were separated on a Kinetex biphenyl column (50 ×2.1 mm, 2.6 µm) with gradient elution of water and methanol containing 2 mM ammonium format and analysed in multiple reaction monitoring mode after positive electrospray ionization. Application of the method included cell culture experiments with two different primary cell types, human coronary artery smooth muscle cells (HCSMC) and human coronary artery endothelial cells (EC). Cells were treated with different concentrations of cortisol, aldosterone and mifepristone, a glucocorticoid receptor antagonist and quantitative PCR was performed. The method exhibits high precision (CV ≤ 6 %) and accuracy (deviation from nominal concentration ≤ 6 %) for concentrations above the limit of quantification (LoQ) which is 0.11, 0.56 and 0.69 nmol/L for aldosterone, cortisone and cortisol, respectively. Calibration curves did not differ when prepared in media or solvent. The method enabled us to confirm activity of HSD11B2 and concentration dependent conversion of cortisol to cortisone in HCSMC (median conversion ratio at 140 nM cortisol = 1.46 %). In contrast we did not observe any HSD11B2 activity in EC. Neither addition of high aldosterone, nor addition of 1 µM mifepristone had impact on glucocorticoid concentrations. Quantitative PCR revealed expression of HSD11B1 and HSD11B2 in HCSMC but not in EC. We present a fast and reliable method for quantification of cortisol, cortisone and aldosterone in cell culture supernatants. The method enabled us to study HSD11B2 activity in two different cell types and will support future experiments investigating mechanisms of target organ damage in conditions of glucocorticoid and mineralocorticoid excess.

Moderate salt restriction in primary aldosteronism improves bone metabolism through attenuation of urinary calcium and phosphate losses.

Accumulating evidence links osteoporosis and dietary salt consumption. Primary aldosteronism (PA) is a model disease with increased dietary salt intake and constitutes an independent risk factor for osteoporosis. We, thus, assessed whether a short-term moderate reduction in salt intake in PA results in detectable osteoanabolic effects. Forty-one patients with PA on stable mineralocorticoid receptor antagonist therapy were subjected to a 12-week salt restriction. Serum and urinary electrolytes, markers of bone turnover, and a 15 steroids plasma profile were registered. After 12 weeks, urinary calcium and phosphate decreased, while plasma testosterone, serum phosphate, and bone alkaline phosphatase (BAP) all increased significantly. Longitudinal changes in BAP were independently correlated with changes in serum phosphate, parathyroid hormone, and urinary calcium in multivariate analysis. Salt restriction in PA limits urinary calcium and phosphate losses and may confer favorable osteoanabolic effects. Our findings suggest that salt restriction should be considered in patients with PA to improve bone health.

Sleep disturbances in primary aldosteronism are associated to depressive symptoms - Could specific mineralocorticoidreceptors be a common pathway?

Symptoms of depression and anxiety are frequent in patients with primary aldosteronism (PA) and are supposed to be independent risk factors for cardiovascular diseases (CVD). As patients with PA have an increased cardiovascular risk compared to patients with essential hypertension, sleep disturbances, which often accompany depressive and anxiety symptoms, may be an additional contributor to the cardiometabolic consequences of PA. To clarify this possible link we investigated 132 patients with PA at baseline and after one year after initiation of treatment either by adrenalectomy (ADX) or mineralocorticoid-receptor-antagonist (MRA). Sleep disturbances and daytime sleepiness were assessed with Pittsburg sleep Inventory (PSQI) and Epworth sleepiness scale (ESS). Patients with PA showed pathological scores for sleep disturbances at baseline according to PSQI, with females being more affected (8.1 vs. 5.7 p < 0.001), which was significantly improved after initiation of specific treatment (p = 0.002). For ESS we found scores within the normal range, but higher than the general population, which significantly improved at follow-up (p < 0.001). The intensity of sleep disturbances was highly correlated with scores of anxiety and depression at baseline and follow-up. However, clinical and biochemical markers of PA (e.g. aldosterone, blood pressure) and metabolic markers did not show a consistent association with sleep changes. The degree of improvement in PSQI was significantly associated with the improvement of brief patients health questionnaire (PHQD) (p = 0.0151). Sleep disturbances seem not to be an independent risk factor for cardiovascular and metabolic problems in PA. They are strongly associated to depressive symptoms and maybe mediated by the same mineralocorticoid receptor circuits.

Benchmarking of Coatings for Cathode Active Materials in Solid-State Batteries Using Surface Analysis and Reference Electrodes.

Fast and reliable evaluation of degradation and performance of cathode active materials (CAMs) for solid-state batteries (SSBs) is crucial to help better understand these systems and enable the synthesis of well-performing CAMs. However, there is a lack of well-thought-out procedures to reliably evaluate CAMs in SSBs. Current approaches often rely on X-ray photoelectron spectroscopy (XPS) for the evaluation of degradation. Unfortunately, XPS sensitivity is not very high, and minor but relevant degradation products may not be detected and distinguished. Furthermore, degradation caused by the current collector (CC) itself is usually not distinguished from CAM-induced degradation. This study uses a modified CC, which allows us to separate electrochemical degradation caused by the CC from degradation at the CAM itself. Using this CC, we present an approach using time-of-flight secondary ions mass spectrometry (ToF-SIMS) that offers high sensitivity and reliability. Principal component analysis (PCA) is applied to differentiate secondary ions as well as identify those mass fragments that correlate with degradation products. This approach also enables distinguishing between different pathways of degradation. To evaluate the kinetic performance of the samples, three-electrode rate tests are performed. Electrochemical characterization evaluates the kinetic performance of the samples under investigation. The samples are finally rated with a score that allows a reliable comparison between the different materials and offers a complete picture of the materials' characteristics in terms of electrochemical performance and degradation.

Salt and Aldosterone - Reciprocal and Combined Effects in Preclinical Models and Humans.

Primary aldosteronism is an endocrine disorder caused by excessive production of aldosterone by the adrenal glands, and is recognized as the most important cause of endocrine hypertension. With specific therapy, this type of hypertension is potentially curable. In the general population, high salt intake increases the risk for cardiovascular diseases like stroke. In populations with aldosterone excess, observational and experimental data suggest that aldosterone-induced organ damage requires a combination of high dietary salt intake and high plasma aldosterone, i.e., plasma aldosterone levels inappropriately high for salt status. Therefore, understanding the relationship between plasma aldosterone levels and dietary salt intake and the nature of their combined effects is crucial for developing effective prevention and treatment strategies. In this review, we present an update on findings about primary aldosteronism and salt intake and the underlying mechanisms governing their interaction.

Major immunophenotypic abnormalities in patients with primary adrenal insufficiency of different etiology.

Introduction: Patients with primary adrenal insufficiency (PAI) suffer from increased risk of infection, adrenal crises and have a higher mortality rate. Such dismal outcomes have been inferred to immune cell dysregulation because of unphysiological cortisol replacement. As the immune landscape of patients with different types of PAI has not been systematically explored, we set out to immunophenotype PAI patients with different causes of glucocorticoid (GC) deficiency. Methods: This cross-sectional single center study includes 28 patients with congenital adrenal hyperplasia (CAH), 27 after bilateral adrenalectomy due to Cushing's syndrome (BADx), 21 with Addison's disease (AD) and 52 healthy controls. All patients with PAI were on a stable GC replacement regimen with a median dose of 25 mg hydrocortisone per day. Peripheral blood mononuclear cells were isolated from heparinized blood samples. Immune cell subsets were analyzed using multicolor flow cytometry after four-hour stimulation with phorbol myristate acetate and ionomycin. Natural killer (NK-) cell cytotoxicity and clock gene expression were investigated. Results: The percentage of T helper cell subsets was downregulated in AD patients (Th1 p = 0.0024, Th2 p = 0.0157, Th17 p < 0.0001) compared to controls. Cytotoxic T cell subsets were reduced in AD (Tc1 p = 0.0075, Tc2 p = 0.0154) and CAH patients (Tc1 p = 0.0055, Tc2 p = 0.0012) compared to controls. NKCC was reduced in all subsets of PAI patients, with smallest changes in CAH. Degranulation marker CD107a expression was upregulated in BADx and AD, not in CAH patients compared to controls (BADx p < 0.0001; AD p = 0.0002). In contrast to NK cell activating receptors, NK cell inhibiting receptor CD94 was upregulated in BADx and AD, but not in CAH patients (p < 0.0001). Although modulation in clock gene expression could be confirmed in our patient subgroups, major interindividual-intergroup dissimilarities were not detected. Discussion: In patients with different etiologies of PAI, distinct differences in T and NK cell-phenotypes became apparent despite the use of same GC preparation and dose. Our results highlight unsuspected differences in immune cell composition and function in PAI patients of different causes and suggest disease-specific alterations that might necessitate disease-specific treatment.

Postoperative ACTH-stimulated aldosterone predicts biochemical outcome in primary aldosteronism.

OBJECTIVE: Primary aldosteronism (PA) is the most common surgically curable cause of hypertension. Unilateral aldosterone-producing adenoma can be treated with adrenalectomy. Clinical and biochemical outcomes are assessed 6-12 months after adrenalectomy according to primary aldosteronism surgical outcome (PASO) consensus criteria. Earlier prediction of biochemical remission would be desirable as it could reduce cumbersome follow-up visits. We hypothesized that postoperative adrenocorticotropic hormone (ACTH) stimulated plasma aldosterone concentrations (PAC) measured shortly after adrenalectomy can predict PASO outcomes. DESIGN: Retrospective cohort study. METHODS: We analyzed 100 patients of the German Conn's registry who underwent adrenalectomy and postoperative ACTH stimulation tests within the first week after adrenalectomy. Six to twelve months after adrenalectomy we assessed clinical and biochemical outcomes according to PASO criteria. Serum cortisol and PAC were measured by immunoassay at baseline and 30 min after the intravenous ACTH infusion. We used receiver operating characteristics (ROC) curve analysis and matched the parameters to PASO outcomes. RESULTS: Eighty-one percent of patients had complete, 13% partial, and 6% absent biochemical remission. Complete clinical remission was observed in 28%. For a cut-off of 58.5 pg/mL, stimulated PAC could predict partial/absent biochemical remission with a high sensitivity (95%) and reasonable specificity (74%). Stimulated PAC's area under the curve (AUC) (0.89; confidence interval (CI) 0.82-0.96) was significantly higher than other investigated parameters. CONCLUSIONS: Low postoperative ACTH stimulated PAC was predictive of biochemical remission. If confirmed, this approach could reduce follow-up visits to assess biochemical outcome.

p-Phenylene Diisothiocyanate-Based Covalent Immobilization of ß-d-Galactosidase and Determination of Enzyme Activity by Cleavage of X-Gal and ONPG on Solid Support.

Herein, we present the immobilization of a technical grade ß-d-galactosidase on amino-functionalized microtiter plates. Afterward, we transferred the results to a resin-based approach. For the covalent binding of the enzyme, an amino-functionalized microtiter plate was prefunctionalized with 1,4-phenylendiisothiocyanate. The cleavage of the substrate 5-bromo-4-chloro-3-indoxyl-ß-d-galactopyranoside (X-Gal) produces a deep blue dye, which was quantified in a microtiter plate reader at 595 nm. The maximum reaction rates and the Michaelis-Menten constant were calculated. In addition, the unwanted blue precipitate formed during the experiments could be minimized by optimizing the experiments. When transferring the immobilization method to Rink amide resin, o-nitrophenyl-ß-d-galactopyranoside was used as the substrate and the measurement was carried out in a photometer at 420 nm.

Moderate dietary salt restriction improves blood pressure and mental well-being in patients with primary aldosteronism: The salt CONNtrol trial.

BACKGROUND: Primary aldosteronism (PA) is a frequent cause of hypertension. Aldosterone excess together with high dietary salt intake aggravates cardiovascular damage, despite guideline-recommended mineralocorticoid receptor antagonist (MRA) treatment. OBJECTIVES: To investigate the antihypertensive impact of a moderate dietary salt restriction and associated physiological changes, including mental well-being. METHODS: A total of 41 patients with PA on a stable antihypertensive regimen-including MRA-followed a dietary salt restriction for 12 weeks with structured nutritional training and consolidation by a mobile health app. Salt intake and adherence were monitored every 4 weeks using 24-h urinary sodium excretion and nutrition protocols. Body composition was assessed by bioimpedance analysis and mental well-being by validated questionnaires. RESULTS: Dietary salt intake significantly decreased from 9.1 to 5.2 g/d at the end of the study. In parallel, systolic (130 vs. 121 mm Hg) and diastolic blood pressure (BP) (84 vs. 81 mm Hg) improved significantly. Patients' aptitude of estimating dietary salt content was refined significantly (underestimation by 2.4 vs. 1.4 g/d). Salt restriction entailed a significant weight loss of 1.4 kg, improvement in pulse pressure (46 vs. 40 mm Hg) and normalization of depressive symptoms (PHQD scale, p < 0.05). Salt restriction, cortisol after dexamethasone suppression test and dosage of renin-angiotensin-aldosterone-system (RAAS) blockers were independently associated with BP reduction. CONCLUSION: A moderate restriction of dietary salt intake in patients with PA substantially reduces BP and depressive symptoms. Moreover, the findings underline that a sufficient RAAS blockade seems to augment the effects of salt restriction on BP and cardiovascular risk.

Deoxyguanosine kinase mutation F180S is associated with a lean phenotype in mice.

BACKGROUND: Deoxyguanosine kinase (DGUOK) deficiency is one of the genetic causes of mitochondrial DNA depletion syndrome (MDDS) in humans, leading to the hepatocerebral or the isolated hepatic form of MDDS. Mouse models are helpful tools for the improvement of understanding of the pathophysiology of diseases and offer the opportunity to examine new therapeutic options. METHODS: Herein, we describe the generation and metabolic characterization of a mouse line carrying a homozygous DguokF180S/F180S mutation derived from an N-ethyl-N-nitrosourea-mutagenesis screen. Energy expenditure (EE), oxygen consumption (VO2) and carbon dioxide production (VCO2) were assessed in metabolic cages. LC-MS/MS was used to quantify plasma adrenal steroids. Plasma insulin and leptin levels were quantified with commercially available assay kits. RESULTS: Mutant animals displayed significantly lower body weights and reduced inguinal fat pad mass, in comparison to unaffected littermates. Biochemically, they were characterized by significantly lower blood glucose levels, accompanied by significantly lower insulin, total cholesterol, high density lipoprotein and triglyceride levels. They also displayed an almost 2-fold increase in transaminases. Moreover, absolute EE was comparable in mutant and control mice, but EE in mutants was uncoupled from their body weights. Histological examination of inguinal white adipose tissue (WAT) revealed adipocytes with multilocular fat droplets reminiscent of WAT browning. In addition, mRNA and protein expression of Ucp1 was increased. Mutant mice also presented differing mitochondrial DNA content in various tissues and altered metabolic activity in mitochondria, but no further phenotypical or behavioral abnormalities. Preliminary data imply normal survival of DguokF180S/F180S mutant animals. CONCLUSION: Taken together, DGUOK mutation F180S leads to a lean phenotype, with lower glucose, insulin, and lipid levels rendering this mouse model not only useful for the study of MDDS forms but also for deciphering mechanisms resulting in a lean phenotype.

Differential central regulatory mineralocorticoidreceptor systems for anxiety and depression - Could KCNJ5 be an interesting target for further investigations in major depression?

The mineralocorticoid receptor (MR) is suggested to play a role in the pathophysiology of depression and anxiety. Main support comes from studies in patients with primary aldosteronism (PA) which suggested different central pathways for depression and anxiety mediated via the MR and gender differences. We investigated 118 patients with PA over 3 years using self-rating questionnaires for anxiety (GAD-7) and depression (PHQD) at baseline and once a year under specific treatment with adrenalectomy (ADX; n = 48) or a MR antagonist (MRA; n = 70). Genotyping for KCNJ5 mutation was performed in resected tumors. At baseline, patients treated by ADX or MRA had comparable scores for anxiety and depression. Females showed a better metabolic profile but higher scores of depression and anxiety, compared to males. Initiation of specific treatment for PA resulted in a better response in depressive symptoms after ADX and of anxiety under MRA treatment. However, GAD-7 and PHQD remained high in women over the three-year follow-up. KCNJ5 mutation, linked to co-secretion of hybrid steroids as 18-oxocortisol and 18-hydroxycortisol, was detected in 10 female and 2 male patients. They tended to have higher GAD and PHQD scores at baseline compared to patients without KNCJ5 mutation, but showed a significant better reduction in symptoms of anxiety during the 3-year follow up compared to patients without this mutation (all p < 0.05). These data support a differentiated regulation of depression and anxiety by the MR. Moreover, genetic mutations such as KCNJ5 could affect the pathophysiology of these disorders by impacting in adrenal steroidogenesis.

Influence of cortisol cosecretion on non-ACTH-stimulated adrenal venous sampling in primary aldosteronism: a retrospective cohort study.

Objective: Cortisol measurements are essential for the interpretation of adrenal venous samplings (AVS) in primary aldosteronism (PA). Cortisol cosecretion may influence AVS indices. We aimed to investigate whether cortisol cosecretion affects non-adrenocorticotrophic hormone (ACTH)-stimulated AVS results. Design: Retrospective cohort study at a tertiary referral center. Methods: We analyzed 278 PA patients who underwent non-ACTH-stimulated AVS and had undergone at least a 1-mg dexamethasone suppression test (DST). Subsets underwent additional late-night salivary cortisol (LSC) and/or 24-h urinary free cortisol (UFC) measurements. Patients were studied from 2013 to 2020 with follow-up data of 6 months following adrenalectomy or mineralocorticoid antagonist therapy initiation. We analyzed AVS parameters including adrenal vein aldosterone/cortisol ratios, selectivity, lateralization (LI) and contralateral suppression indices and post-operative ACTH-stimulation. We classified outcomes according to the primary aldosteronism surgical outcome (PASO) criteria. Results: Among the patients, 18.9% had a pathological DST result (1.9-5 µg/dL: n = 44 (15.8%); >5 µg/dL: n = 8 (2.9%)). Comparison of AVS results stratified according to the 1-mg DST (≤1.8 vs >1.8 µg/dL: P = 0.499; ≤1.8 vs 1.8 ≤ 5 vs >5 µg/dL: P = 0.811) showed no difference. Lateralized cases with post DST serum cortisol values > 5 µg/dL had lower LI (≤1.8 µg/dL: 11.11 (5.36; 26.76) vs 1.9-5 µg/dL: 11.76 (4.9; 31.88) vs >5 µg/dL: 2.58 (1.67; 3.3); P = 0.008). PASO outcome was not different according to cortisol cosecretion. Conclusions: Marked cortisol cosecretion has the potential to influence non-ACTH-stimulated AVS results. While this could result in falsely classified lateralized cases as bilateral, further analysis of substitutes for cortisol are required to unmask effects on clinical outcome.

Renin, a marker for left ventricular hypertrophy, in primary aldosteronism: a cohort study.

CONTEXT: Primary aldosteronism (PA) causes left ventricular hypertrophy (LVH) via hemodynamic factors and directly by aldosterone effects. Specific treatment by mineralocorticoid receptor antagonists (MRA) or adrenalectomy (ADX) has been reported to improve LVH. However, the cardiovascular benefit could depend on plasma renin concentration (PRC) in patients on MRA. PATIENTS AND OBJECTIVE: We analyzed data from 184 patients from the Munich center of the German Conn's Registry, who underwent echocardiography at the time of diagnosis and 1 year after treatment. To assess the effect of PRC on cardiac recovery, we stratified patients on MRA according to suppression (n = 46) or non-suppression of PRC (n = 59) at follow-up and compared them to PA patients after ADX (n = 79). RESULTS: At baseline, patients treated by ADX or MRA had comparable left ventricular mass index (LVMI, 61.7 vs 58.9 g/m2.7, P = 0.591). Likewise, patients on MRA had similar LVMI at baseline, when stratified into treatment groups with suppressed and unsuppressed PRC during follow-up (60.0 vs 58.1 g/m2.7, P = 0.576). In all three groups, we observed a significant reduction in LVMI following treatment (P < 0.001). However, patients with suppressed PRC had no decrease in pro-BNP levels, and the reduction of LVMI was less intense than in patients with unsuppressed PRC (4.1 vs 8.2 g/m2.7, P = 0.033) or after ADX (9.3 g/m2.7, P = 0.019). Similarly, in multivariate analysis, higher PRC was correlated with the regression of LVH. CONCLUSION: PA patients with suppressed PRC on MRA show impaired regression of LVH. Therefore, dosing of MRA according to PRC could improve their cardiovascular benefit.

Patients with low IGF-I after curative surgery for Cushing's syndrome have an adverse long-term outcome of hypercortisolism-induced myopathy.

BACKGROUND: Glucocorticoid excess leads to muscle atrophy and weakness in patients with endogenous Cushing's syndrome. Insulin-like growth factor I (IGF-I) is known to have protective effects on muscle loss. We hypothesized that individual serum IGF-I concentrations might be predictive for long-term myopathy outcome in Cushing's syndrome. PATIENTS AND METHODS: In a prospective longitudinal study of 31 patients with florid Cushing's syndrome, we analyzed IGF-I and IGF binding protein 3 (IGFBP 3) concentrations at the time of diagnosis and following surgical remission over a period of up to 3 years. We assessed muscle strength by grip strength measurements using a hand grip dynamometer and muscle mass by bio-impedance measurements. FINDINGS: Individual serum IGF-I concentrations in the postoperative phase were strongly predictive of long-term grip strength outcome (rs = 0.696, P ≤ 0.001). Also, lower IGF-I concentrations were associated with a lower muscle mass after 3 years (rs = 0.404, P = 0.033). While patients with high IGF-I s.d. scores (SDS; >1.4) showed an improvement in grip strength within the follow-up period (P = 0.009), patients with lower IGF-I SDS (≤-0.4) had a worse outcome with persisting muscle dysfunction. In contrast, preoperative IGF-I concentrations during the florid phase of Cushing's syndrome did not predict long-term muscle function outcome (rs = 0.285, P = 0.127). CONCLUSION: Lower individual IGF-I concentrations 6 months after curative surgery for Cushing's syndrome are associated with adverse long-term myopathy outcome and IGF-I might be essential for muscle regeneration in the early phase after correction of hypercortisolism.

Altered Taste Perception for Sodium Chloride in Patients With Primary Aldosteronism: A Prospective Cohort Study.

[Figure: see text].

The mitochondrial thioredoxin reductase system (TrxR2) in vascular endothelium controls peroxynitrite levels and tissue integrity.

The mitochondrial thioredoxin/peroxiredoxin system encompasses NADPH, thioredoxin reductase 2 (TrxR2), thioredoxin 2, and peroxiredoxins 3 and 5 (Prx3 and Prx5) and is crucial to regulate cell redox homeostasis via the efficient catabolism of peroxides (TrxR2 and Trxrd2 refer to the mitochondrial thioredoxin reductase protein and gene, respectively). Here, we report that endothelial TrxR2 controls both the steady-state concentration of peroxynitrite, the product of the reaction of superoxide radical and nitric oxide, and the integrity of the vascular system. Mice with endothelial deletion of the Trxrd2 gene develop increased vascular stiffness and hypertrophy of the vascular wall. Furthermore, they suffer from renal abnormalities, including thickening of the Bowman's capsule, glomerulosclerosis, and functional alterations. Mechanistically, we show that loss of Trxrd2 results in enhanced peroxynitrite steady-state levels in both vascular endothelial cells and vessels by using a highly sensitive redox probe, fluorescein-boronate. High steady-state peroxynitrite levels were further found to coincide with elevated protein tyrosine nitration in renal tissue and a substantial change of the redox state of Prx3 toward the oxidized protein, even though glutaredoxin 2 (Grx2) expression increased in parallel. Additional studies using a mitochondria-specific fluorescence probe (MitoPY1) in vessels revealed that enhanced peroxynitrite levels are indeed generated in mitochondria. Treatment with Mn(III)tetrakis(1-methyl-4-pyridyl)porphyrin [Mn(III)TMPyP], a peroxynitrite-decomposition catalyst, blunted intravascular formation of peroxynitrite. Our data provide compelling evidence for a yet-unrecognized role of TrxR2 in balancing the nitric oxide/peroxynitrite ratio in endothelial cells in vivo and thus establish a link between enhanced mitochondrial peroxynitrite and disruption of vascular integrity.

Atherosclerotic Burden and Arterial Stiffness are Not Increased in Patients with Milder Forms of Primary Aldosteronism Compared to Patients with Essential Hypertension.

Patients with primary aldosteronism (PA) are at increased cardiovascular risk, compared to patients with essential hypertension (EH). Cardiovascular damage could depend on PA phenotype, potentially being lower in milder forms of PA. Our aim was to assess atherosclerotic burden and arterial stiffness in 88 prospectively recruited patients, including 44 patients with mild PA and EH respectively. All patients underwent a structured study program, including measurements of ankle-brachial index, oscillometric measurement of central pulse wave velocity (cPWV) and vascular ultrasound examination of the supraaortic arteries, the abdominal aorta, and the femoropopliteal arteries. A plaque score was calculated to estimate atherosclerotic burden for each patient. This is a prospective case-control study set at a tertiary care hospital. Patients with PA and EH matched well for age, gender, blood pressure, BMI, and cardiovascular risk factors such as diabetes mellitus and smoking status. Common carotid intima-media thickness (0.77 vs. 0.75 mm; p=0.997) and cPWV (7.2 vs. 7.1 m/s; p=0.372) were comparable between patients with PA and EH. The atherosclerotic burden, as expressed by the plaque score, did not differ between the two groups (p=0.159). However, after initiation of treatment cPWV was significantly decreased in patients with PA (p=0.017). This study shows that subclinical atherosclerotic burden and arterial stiffness in patients with milder forms of PA is comparable to patients with EH. Nevertheless, specific treatment for PA significantly improved cPWV, which argues for a more liberal use of mineralocorticoid receptor antagonists in patients with arterial hypertension.