Aldosterone Research Articles

Radiofrequency catheter ablation combined with spironolactone in the treatment of atrial fibrillation: A single-center randomized controlled study.

At present, the question of whether radiofrequency ablation (RFA) combined with spironolactone can reduce the levels of plasma angiotensin II (AngII) and aldosterone (ALD) in patients with atrial fibrillation (AF) and reduce the recurrence of AF has not been reported.HypothesisThe present study evaluates the effect of spironolactone as an ALD antagonist on the short‐term and long‐term recurrence of AF after RFA. A total of 203 patients were enrolled in the present study, with 102 patients in the spironolactone therapy group (Group PVI/SP) and 101 patients in the control group (Group PVI alone). The AngII and ALD levels and the size of the left atrium in patients with AF were observed in order to evaluate the relationship between the combination therapy of spironolactone with RFA and the success rate in AF treatment. After therapy, the levels of AngII (52.8 vs. 64.3 pg/ml, p < .001), ALD (45.7 vs. 60.6 pg/ml, p = .016), and N‐terminal of B‐type natriuretic peptide (NT‐proBNP) (73.5 vs. 110 pg/ml, p = .016), along with the size of the left atrium (35.8 vs. 37.2 mm, p = .007), were all significantly lower in Group PVI/SP compared with Group PVI alone. The cumulative AF‐free survival rate was higher in Group PVI/SP than in Group PVI alone after treatment (85.3% vs.73.3%, p = .033). In RFA combined with spironolactone treatment, spironolactone can directly antagonize the effects of ALD and AngII and the recurrence of AF and improve left ventricular function.

MO033WHOLE-EXOME SEQUENCING AS A FIST-LINE DIAGNOSTIC TOOL IN BARTTER AND GITELMAN SYNDROME

Abstract Background and Aims Bartter (BS) and Gitelman syndrome (GS) are autosomal recessive rare inherited disorders characterized by hypokalemic metabolic alkalosis and secondary hyperaldosteronism. The primary defect is a genetically determined impairment of sodium chloride reabsorption in the renal tubule, thus resulting in salt loss, dehydration and acid-base homeostasis perturbations. Although the diagnosis can be suspected based on presenting features, the clinical diagnosis of BS and GS can be challenging, as they are rare and phenotypically overlapping. As a consequence, the current clinical classification lacks of specificity and genetic testing represents the gold standard for the diagnosis. Driven by the rapidly decreasing costs and turn-around time, next-generation sequencing technologies are increasingly utilized in diagnostics and research of inherited tubulopathies, including BS and GS. Recently, sequencing of selected gene panels provided the advantage of achieving high coverage of genes of interest at lower costs, providing high diagnostic yield and new insights into the phenotypic spectrum of these rare disorders. However, whole-exome (WES) is not routinely performed for the molecular diagnosis of BS and GS. The aim of our study was to assess the diagnostic performance of WES in BS and GS and to establish genotype-phenotype correlations. Method We performed WES in all consecutive patients referred for genetic testing with a clinical suspect of BS or GS. Variant prioritization was carried out according to the American College of Medical Genetics and Genomics guidelines (ACMG). Parents and first-degree relatives were included, whenever available. Demographic, clinical and laboratory data were collected retrospectively, in order to establish genotype-phenotype correlations. Results We enrolled 50 patients (22 males, 46 Caucasians) with a clinical diagnosis of BS (19), GS (24) or BS/GS (7). All the patients showed hypokalemic metabolic alkalosis at onset (serum bicarbonate=29.5 mEq/l ± 4.4, potassium= 2.7 mEq/l ± 0.6). The median age at clinical diagnosis was 7 years (range 0-67 years). Three patients had familial history of tubulopathies. WES showed pathogenic variants in 42/50 patients (84%), thus establishing a conclusive diagnosis. Interestingly, a dedicated analytic pipeline allowed us to identify copy number variations (CNVs) in 7/42 patients with a confirmed genetic diagnosis. In detail, WES allowed us to confirm the clinical diagnosis in 33/50 patients, with an improvement in classification in at least 14 cases (i.e. subtype I-V of BS). In 9 additional patients, genetic testing changed the clinical diagnosis: 6 patients with a clinical of BS turned out to have pathogenic variants in SLC12A3, resulting in GS; in 3 patients, genetic testing revised the clinical diagnosis indicating inherited disorders outside the BS/GS spectrum (HELIX syndrome, Primary familial hypoparatiroidism, Type 2 renal hypomagnesemia). Only 38% of patients with a genetic diagnosis of BS showed nephrocalcinosis. Strikingly, this was present in 8% of patients with GS. On the other hand, hypomagnesemia, a distinctive feature of GS, was similarly distributed among BS and GS patients (45% vs. 68%, respectively). Finally, although patients with GS showed a median age at onset higher than patients with BS, some overlap did exist, making differential diagnosis challenging at single-patient level. Conclusion The results of our study demonstrate that WES ensures a high diagnostic yield (84%) in patients with a clinical diagnosis of BS or GS, especially if coupled with analysis of CNVs. This approach showed to be useful in dealing with the phenotypic heterogeneity typical of these rare disorders, improving differential diagnosis by detecting phenocopies also outside the BS/GS spectrum, enabling additional specific work-up, genetic counseling, and screening of at-risk relatives.

Unravelling the Genetic Basis of ACTH-Mediated Aldosterone Hypersecretion in Hypertensive Patients Without Primary Aldosteronism

Introduction: Primary aldosteronism (PA), a condition characterized by autonomous aldosterone hypersecretion, constitutes the most common cause of secondary hypertension. PA includes both sporadic and familial forms, inherited in an autosomal dominant manner. Recent evidence suggests a higher prevalence of aldosterone excess among hypertensive patients than previously thought, while chronic stress-related ACTH-mediated aldosterone hypersecretion has also been implicated in the pathogenesis of essential hypertension. Objective: To determine whether genetic variations of aldosterone regulating genes could be implicated in the ACTH-mediated aldosterone hypersecretion in hypertensive patients without PA. Methods: Twenty-one hypertensive patients without PA, who exhibited ACTH-mediated aldosterone hypersecretion, underwent Whole Exome Sequencing (WES) on Novaseq 6000 platform (Illumina). As hyper-responders were defined patients whose aldosterone (ALD) and aldosterone-to-renin ratio (ARR) response to ultra-low ACTH stimulation test was above the 97.5th percentile values of controls. The cutoff levels for ALD and ARR were 1300 pmol/L and 77 pmol/mIU, respectively. Variant calling was performed according to GATK best practices and VCF files were filtered for variants in 25 genes associated with PA. To identify new susceptibility genes for PA, VCF files were also intersected for variants in ion channels encoding genes involved in pathways responsible for PA. The analysis was restricted to rare variants with gnomAD frequency < 1%. Qualifying variants and pathogenicity were established by employing in silico tools. Copy Number Variant analysis was performed using ExomeDepth algorithm. Results: Eight out of twenty-one patients were heterozygous for rare variants in genes associated with PA, while two patients carried potentially damaging variants in genes encoding ion channels. Specifically, one patient was heterozygous for p.V259M in KCNK5 and one patient was heterozygous for the novel variant p.V221M in KCNK9. Two additional patients carried a predicted pathogenic variant p.R492W in SLC26A2, a gene that has been associated with PA through GWAS. Germline variants in calcium channel genes were also detected in three patients: p.V249I in CACNA1H, p.R462Q in CACNA1D and p.L1801M in CACNA1I, while one patient carried an ultra-rare variant (p.R26L) in ATP13A3. Finally, in two patients we identified rare, likely pathogenic variants in two new susceptibility genes for PA: KCNK16 (p.P255H) and CACNA2D3 (p.V55I). Conclusion: These findings support the notion that mutations in aldosterone synthesis/secretion regulating genes may sensitize zone glomerulosa cells to ACTH stimulation, leading to aldosterone hypersecretion under conditions of stress. We also report two novel candidate susceptibility genes for PA, KCNK16 and CACNA2D3, and one novel variant in KCNK9.

Adrenoleukodystrophy and Secondary Hyperaldosteronism; A Clinical Demonstration of the Zona Glomerulosa Persistence

Adrenoleukodystrophy (ALD) is a peroxisomal disorder which leads to the accumulation of very long-chain fatty acids in all tissues. Age at onset of symptoms vary depending on the phenotype severity. It can present with progressive symptoms of neurological defects and/or primary adrenal insufficiency.We report a case of 31 yo man diagnosed in childhood with ALD and treated with hydrocortisone and fludrocortisone who eventually developed resistant hypertension due to secondary hyperaldosteronism.He was diagnosed with ALD at 9-year-old and have received an allogenic hematopoietic cell graft 4 years later. After transplantation, he developed a bronchiolitis obliterans which was treated with high dose of glucocorticoids for 3 years.In 2014, at first evaluation with our team, he was on hydrocortisone 10mg/m2 and fludrocortisone 0.05 mg daily. Fludrocortisone was started at 14 yo for a clinical suspicion of mineralocorticoids deficiency. At that time, the patient was normokaliemic (3.8 mmol\\L), his aldosterone was, 2 days apart, 245pmol/L lying down and 68 pmol/L (N 138 - 413) sitting, his renin activity was 0.11 ng/L/s (N 0.14 - 0.31) and no orthostatic hypotension was documented.In July 2015, high blood pressure (BP) was noticed, and fludrocortisone was decreased at 0.05 mg every other day. However, his BP continued to increase progressively, thereby fludrocortisone was discontinued in June 2018. Thereafter, amlodipine 10 mg daily, hydrochlorothiazide 12.5 mg daily and terazosin 2 mg daily were progressively introduced. Even with those three anti-hypertensive drugs his BP wasn’t controlled (190/100 mmHg) and he also developed persistent hypokalemia (3.0 - 3.3 mmol/L). Furthermore, significant aortic atheromatosis was described on abdominal computed tomography. In that context, his serum aldosterone/renin ratio was measured. The aldosterone was 632 pmol/L (N < 350) and the renin mass was 93.4 ng/L (N 3 - 16). An assessment of renal arteries was done by doppler ultrasonography, which was compatible with a hemodynamically significant right renal artery stenotic lesion. The patient had never smoke, had no diabetes (HbA1c 5.1%) or dyslipidemia (LDL 2.02 mmol/L). Based on the diagnosis of secondary aldosteronism, spironolactone was introduced in June 2020. Spironolactone was titrated to 37.5 mg daily and BP significantly improved with values around 136/75 and potassium return in normal range (3.8 mmol/L).This interesting case illustrates 1) the persistence of a functioning zona glomerulosa in some patient with ALD and the possible development of secondary hyperaldosteronism in response to renovascular disorder and 2) the particularly high burden of atherosclerosis in this young patient, without classic risk factor, raises questions on the effect of the metabolic defect of ALD itself on the development of atheromatosis.

Cushing’s Syndrome Associated With an Adrenal Cavernous Hemangioma

Introduction: Adrenal cavernous hemangiomas are rare benign tumors that arise from vascular endothelium and are often discovered incidentally on abdominal imaging. The majority are nonfunctioning; however, we present a case of adrenal Cushing’s syndrome in a patient with a cavernous adrenal hemangioma.Case: A 72-year-old woman was referred for an incidental right adrenal mass. On questioning, she endorsed abdominal pain, sixty-pound unintentional weight gain over five years, truncal obesity, and easy bruising. Past medical history was relevant for hypertension. Her surgical history was extremely complicated, having had a perforated peptic ulcer, open cholecystectomy complicated by injury to the right ureter, incisional hernia repair, appendectomy, and hysterectomy with bilateral salpingo-oophorectomy.She was first noted to have a right adrenal mass on a CT done eight years ago, measuring 3.8 x 3.2 cm. A repeat CT abdomen and pelvis now showed this mass to be 6.5 x 6.3 x 8.1 cm with unenhanced Hounsfield units of 29.6. Radiographically, this was a heterogeneous, solid, and cystic appearing mass with peripheral brisk arterial enhancement areas, which appeared to fill in on delayed imaging. The enhancement pattern of the lesion was consistent with an adrenal cavernous hemangioma. An MRI of the abdomen also demonstrated similar peripheral nodular enhancement favoring an adrenal cavernous hemangioma.Functional testing for the adrenal mass was undertaken. Morning cortisol was 23.4 mcg/dl with ACTH low at 5.3 pg/ml, and DHEA-S 3 mcg/dl. She failed to suppress with overnight 1 mg dexamethasone with AM cortisol of 3.6 mcg/dl. Midnight salivary cortisol levels were high at 0.237 mcg/dl and 0.419 mcg/dl while a 24-hour urine free cortisol was normal at 15.2 mcg/d. She tested negative for pheochromocytoma and primary aldosteronism.The patient was deemed a poor surgical candidate due to her history of multiple prior abdominal surgeries and a BMI of 46. Therefore, she underwent an IR angioembolization of the right adrenal mass instead. On follow-up CT, there was no significant change in the size of the lesion; however the degree of rim enhancement was slightly decreased. Post procedurally, her a.m. cortisol remained high-normal at 18.3 mcg/dL.. She is currently enrolled in a study for medical treatment of Cushing’s syndrome.Discussion: Adrenal cavernous hemangiomas usually present incidentally in the 6th-7th decade of life with a female predominance. These lesions are often asymptomatic; however, abdominal pain is the most common presenting symptom. The majority of adrenal cavernous hemangiomas are hormonally quiescent and mineralocorticoid excess and/or subclinical Cushing’s syndrome is exceedingly rare. Our patient is unique in her presentation of adrenal Cushing’s with this lesion and the novel use of angioembolization to decrease the size of this vascular tumor.

Concurrent Pseudohyperaldosteronism and Primary Glucocorticoid Deficiency From Posaconazole

Abstract Background: Posaconazole can cause pseudohyperaldosteronism via inhibition of 11-beta hydroxylase and 11-beta-hydroxysteroid dehydrogenase type 2 (1). The accumulation of 11-deoxycorticosterone and increased cortisol-to-cortisone ratio in the kidney causes apparent mineralocorticoid excess. The effect of posaconazole on the glucocorticoid axis is less established. Clinical Case: A 56-year-old Hispanic man with a history of chronic septic arthritis of the left ankle from Coccidioides presented with 3 months of malaise, nausea, weight loss of 30 pounds, and recurrent hypokalemia. He was recently switched from long-term fluconazole therapy to posaconazole around the time his symptoms began. His initial labs at our hospital were notable for low potassium (2.9 mmol/L, nl 3.5–5.5 mmol/L) and a random cortisol of 5.8 mcg/dL (nl ≥2.0 mcg/dL). A Cosyntropin stimulation test revealed elevated ACTH (168 pg/mL, nl 7.2–63.3 pg/mL) with minimal rise of cortisol from 4.6 to 7.2 mcg/dL at 1 hour after Cosyntropin administration (nl ≥18 mcg/dL at 1-hour post-Cosyntropin). His plasma renin activity was below detection (&amp;lt;0.6 ng/ml/h, nl 0.6–3.0 ng/mL/h), consistent with renin suppression from apparent mineralocorticoid excess. Hydrocortisone for glucocorticoid deficiency was started. A posaconazole determination indicated elevation (5240 ng/mL, usual therapeutic range ≥1000 ng/mL). His posaconazole was stopped, and he was switched back to fluconazole. Three months later, his symptoms were improved with regain of lost weight. Repeat Cosyntropin stimulation test showed ongoing primary glucocorticoid deficiency (ACTH 123 pg/mL, cortisol 4.1 mcg/dL at 1-hour post-Cosyntropin) but normal levels of plasma renin activity (1.2 ng/mL/h), aldosterone (8.6 ng/dL, nl ≤21 ng/dL), and potassium. Quantiferon TB and 21-hydroxylase antibody tests were negative. Hydrocortisone has been continued with plans to repeat Cosyntropin testing in 3 months to reassess. Conclusion: Pseudohyperaldosteronism with glucocorticoid deficiency requiring hydrocortisone treatment has thus far not been reported with posaconazole. Our case shows that posaconazole may lead to true primary glucocorticoid deficiency that can persist after discontinuation of posaconazole and reversal of pseudohyperaldosteronism. Reference: (1) Sanchez-Niño MD, Ortiz A. Unravelling drug-induced hypertension: Molecular mechanisms of aldosterone-independent mineralocorticoid receptor activation by posaconazole. Clin Kidney J 2018;11(5):688–90.

Late Onset of Abiraterone Severe Hypokalemia Due to Unsuspected ACTH-Secreting NEC

Clinical Case: 68-year-old male, Active smoker (30-40 packs/year), and moderate alcohol drinker. An increase in urinary frequency, nocturia, and a PSA of 116 µg/L led to the diagnosis of disseminated castration-sensitive prostate cancer (CSPC), advanced Gleason score (3 + 4 pT2), with bone and retroperitoneal metastases (CT). By 02/2018, he started androgen deprivation therapy (ADT) with Abiraterone (AB) 1000 mg, prednisone, and LHRH agonist. 15 months later, PSA levels decreased to 0.75 ng/ml without side effects and normal K+(3.7 mEq/L), when a CT scan revealed both liver and bone metastasis. During the progression study, two months later, the patient was admitted to the hospital for severe hypoK+ (1.7 mEq / L), normal renal function and metabolic alkalosis. Although abiraterone was discontinued, up to 460 mEq iv per day of K+ and spironolactone were required to maintain serum K+ above 2.5. 3 days later, the hormonal study revealed TSH 1.13 mU /L, ACTH 162 pmol/L, cortisol 258 nmol/L, aldosterone 105 pmol/L, renin (protein) 0.7 µU/mL and deoxycorticosterone 507 pmol/L. 11 days later, plasma cortisol was 967 nmol/L, ACTH 132pmol/L, and cortisoluria 1456 nmol/d. The suppression test with 1 mg of DXM for cortisol was 1162,5 nmol/L. DHEAS was < 407 nmol/L. Liver biopsy showed a small cell neuroendocrine carcinoma (NEC), chromogranin (+), synaptophysin (+), CD56 (+), TTF-1 (+), PSA (-), Ki-67 90% and the granular cytoplasmic ACTH (+). The octreotide scan (+) revealed pathological uptake in L4, multiple uptakes in the liver, and in the axial skeleton. Treatment with Carboplatin plus Paclitaxel for NEC was started, completing 3 weekly doses with good tolerance and clinical benefit, but with the persistence of severe hypoK+ (2.5 mEq/L). Treatment with lanreotide 120 mg every 4 weeks was added, following a feeling of clinical improvement, the disappearance of edema, asthenia, and normalization of plasma K+ (3,4mEq/L). The patient died two months later from respiratory sepsis. Discussion: Several clinical trials have demonstrated that the combination of AB plus ADT prolongs overall survival in DCSPC. The CYP17 inhibition by AB increases ACTH leading to early secondary mineralocorticoid excess with hypokalemia and hypertension while the cortisol levels remain normal or low. Cortisol serum levels increased after AB was discontinued, whereas aldosterone serum levels remained low due to K+ regulatory feedback. The hormonal profile and the pathological and radiological studies revealed an ACTH-producing small cell NEC. In this patient, the previous treatment with BA has masked the clinical and hormonal profile of an ectopic Cushing syndrome. Therefore, Cyp17 inhibitors can mask adrenal or extra-adrenal processes characterized by alterations in steroid metabolism. This case has suggested a more thorough assessment of the adrenal hormonal profile including ACTH during BA treatment.

Limited Efficacy of Mifepristone Due to Poor Tolerance: A Clinical Challenge in Managing Cortisol Excess

A 53-year male with incidental bilateral adrenal masses along with symptoms of proximal muscle weakness, anxiety, and depression. Past medical history of uric acid nephrolithiasis, hypertension, hyperlipidemia, type 2 diabetes mellitus, anxiety, and depression. A right adrenal mass of 2.8cm and a left adrenal mass of 1.5cm both of <10 HU on non-enhancing CT. Hormonal activity workup was ordered which showed cortisol levels of 4.7mcg/dL on 1mg ONDST suggestive of autonomous cortisol excess and workup for primary hyperaldosteronism was negative. On follow up CT adrenal masses remained stable, but the patient underwent the annual hormonal workup for incidentaloma with 1mg ONDST and found with cortisol 4.5mcg/dL, eventually repeated with 2-day LDDST which showed no suppression of cortisol with levels at 4.5 mcg/dL confirming the diagnosis of autonomous cortisol excess. ACTH found to be suppressed at 3.1pg/dL (7.2–63.3 pg/dL) which helped confirm the diagnosis of non-ACTH dependent autonomous cortisol excess. Referred to surgery evaluation due to complications associated with cortisol excess as osteoporosis and uncontrolled hyperglycemia however surgical intervention was deferred until the source of cortisol excess could be identified within the bilateral adrenal masses and it was recommended to continue with medical therapy.As the cortisol excess source cannot be identified by localizing procedures and despite adequate medical therapy and stable adrenal masses on imaging, patient persisted with uncontrolled DM/hypertension and worsening BMD in the spine and hip with surgery not being an option. The patient was started on glucocorticoid receptor antagonist mifepristone in which upon reevaluation weakness and fatigue were noticeable along with hypokalemia of 3.1mmol/L after 2 weeks of therapy that was eventually replaced but 2 weeks later the patient discontinued therapy as he could not tolerate side effects related to adrenal insufficiency associated with mifepristone, such as weakness, fatigue, and dependence of potassium supplementation due to hypokalemia.This effect is physiologically important, because cortisol binds as avidly as aldosterone to the mineralocorticoid receptor, and the plasma cortisol concentration is approximately 100-fold higher than the plasma aldosterone concentration and it can lead to hypokalemia by the mechanism of apparent mineralocorticoid excess. Also, as mifepristone blocks cortisol action, the levels of ACTH and cortisol increase so high that hormonal measurement cannot be used to judge either therapeutic efficacy or adrenal insufficiency, we must go based on signs and symptoms.

Investigating the Transitional Cell Type in the Kidney Collecting Duct

The kidney cortical collecting duct (CCD) has long been thought to contain two distinct cell types, principal cells (PC) and intercalated cells (IC), which facilitate 4-5% of sodium and 10% of water reabsorption in the kidney. Recent single cell RNA-Seq data has identified a distinct transitional cell expressing both PC and IC markers and a number of distinct markers. In this study, the presence of CCD cell types was investigated in mice and rats under the disease model of the syndrome of apparent mineralocorticoid excess (SAME, conferred by a mutation of the Hsd11b2gene, expressed by PCs) and also in two CCD derived cell lines. Inactivation of HSD11B2in humans results in a severe, and often fatal, phenotype in early neonatal life, whilst milder mutations, with residual enzyme activity cause hypertension, sodium retention and hypokalemia in adulthood. The Hsd11b2 gene knockout (KO) model of SAME in mice and rats was studied in the context of CCD cell plasticity and it was hypothesised to reduce the proportion of PCs within the CCD. Cells in the CCD were counted in WT and Hsd11b2 KO animals following immunofluorescent staining for PC- and IC-specific markers (Aqp2 and V-ATPase B1, respectively) in paraffin-embedded kidney sections. ImageJ and GraphPadPrism were used to analyse data, which are expressed as mean ± SEM. Significance was assessed using Tukey's multiple comparisons test, p<0.0001. PC proportion was significantly reduced in KO mice CCDs compared to controls at 4 weeks (52.1 ± 1.9 % vs. 73.0 ± 1.9 %) and 17 weeks (54.1 ± 1.8 % vs. 72.2 ± 1.3 %). A significant increase in the number of cells expressing both PC and IC markers was observed between WT and KO at 4 weeks (8.4 ± 1.2 % vs.25.9 ± 2.0 %) and at 17 weeks (6.5 ± 0.8 % vs. 22.1 ± 2.0 %). Similarly, in the rat KO a significant reduction in the proportion of PCs was noted compared to controls at 5 weeks (60.69 ± 2.51 % vs. 73.30 ± 1.84 %) and 24 weeks (38.40 ± 2.07 % vs. 66.80 ± 1.94 %). This study shows CCD cell plasticity in models of SAME, but also provides further evidence for a PC+/IC+cell in the kidney. The mCCDcl1­ and mpkCCDcl4­ cell lines are typically used as models of PC function. Immunofluorescent staining using antibodies against PC, IC and transitional cell markers (Aqp2, V-ATPase B1 and Parm1, respectively) was carried out on fixed, polarised cells and images analysed using ImageJ software. It was found that both cell line lines contained cells that co-expressed PC and IC markers, similar to the cell type seen in mouse and rat kidneys. Furthermore, a large proportion of cells in both cell lines expressed the transitional cell marker, Parm1. These results provide further support for the presence of a transitional cell type in the CCD, especially in pathophysiological states. The observation of similar cells in CCD cell lines also indicates their use as in vitro models for investigating the role and plasticity of CCD cell types.

Aldosterone up-regulates basolateral Na+ -K+ -2Cl- cotransporter-1 to support enhanced large-conductance K+ channel-mediated K+ secretion in rat distal colon.

Na+ -K+ -2Cl- cotransporter-1 (NKCC1) facilitates basolateral K+ and Cl- uptake, supporting their efflux across mucosal membranes of colonic epithelial cells. NKCC1 activity has also been shown to be critical for electrogenic K+ secretion induced by aldosterone, which is known to stimulate large-conductance K+ (BK) channel expression in mucosal membranes. This study was aimed to (1) identify whether aldosterone enhances NKCC1 expression specifically to support BK-mediated K+ secretion and (2) to determine whether increased NKCC1 supports electrogenic Cl- secretion in parallel to K+ secretion. Dietary Na+ depletion was used to induce secondary hyperaldosteronism in rats, or aldosterone was administered ex vivo to rat distal colonic mucosae. NKCC1-dependent electrogenic K+ or Cl- secretion was measured as a function of short circuit current (ISC ). qRT-PCR, western blot, and immunofluorescence analyses were performed using standard techniques. Aldosterone enhanced NKCC1 and BKα expression and electrogenic K+ secretion in the distal colon, which was inhibited by either serosal bumetanide (NKCC1 inhibitor) or mucosal iberiotoxin (IbTX; BK channel blocker), but not TRAM-34 (IK channel blocker). Expression of NKCC1 and BKα proteins was enhanced in crypt cells of hyper-aldosterone rats. However, neither NKCC1-dependent Cl- secretion nor CFTR (apical Cl- channel) expression was enhanced by aldosterone. We conclude that aldosterone enhances NKCC1 to support BK-mediated K+ secretion independently of Cl- secretion in the distal colon. The regulation of NKCC1 expression/K+ secretion by aldosterone may be a therapeutic target in treating gastrointestinal disorders associated with alterations in colonic K+ transport, such as colonic pseudo-obstruction, and hyperkalemia associated with renal disease.

The association between serum calcium and chronic heart failure: A cross-sectional observational study

Background: Chronic heart failure (CHF) is characterized by sustained activation of neurohormonal and cytokine systems leading to a series of end-organ changes within the myocardium referred to as left ventricular remodeling. Renin-Angiotensin system activation leading to secondary hyperaldosteronism is accompanied by ionized hypocalcemia with secondary hyperparathyroidism which causes dyshomeostasis of extra and intracellular calcium leading to cardiomyocyte necrosis. Objectives: To study the levels of serum calcium in patients of heart failure and its association with severity and duration of chronic heart failure. Methods: The study was conducted on 50 patients with CHF after taking informed consent. All patients met inclusion and exclusion criteria and underwent blood sampling, urine examination and other relevant investigations. Serum calcium levels were correlated with the severity and duration of CHF statistically. Results: Among the patients studied, 96% with ejection fraction (EF)≤ 35%, 87.5% with EF between 35-40% and 44.4% with EF ≥ 40% had low serum calcium values ≤9 mg/dl (p-value &lt;0.001). All patients with duration of heart failure ≥ 2 years and 70.4% patients with duration of heart failure 1-2 years had low serum calcium levels ≤ 9 mg/dl (p value =0.001). Conclusion: There is a significant positive correlation of serum calcium levels with severity of heart failure as measured by EF and significant negative correlation with duration of CHF and NYHA functional grades. The degree of hypocalcemia correlates with severity of cardiomyocyte injury and extent of the neurohormonal response, and accordingly the corresponding risk of adverse cardiovascular events.

Diuretic effect of Lagopsis supina fraction in saline-loaded rats is mediated through inhibition of aquaporin and renin-angiotensin-aldosterone systems and up-regulation of atriopeptin

Lagopsis supina (Steph. ex Willd.) lk. -Gal. ex Knorr. has been used as a diuretic agent in China for centuries with limited scientific evidence. This study investigated the diuretic efficacy and underlying mechanism of a macroporous adsorption resin with 30% ethanol elution fraction from L. supina (LSC) in saline-loaded rats and to identify its phytochemicals by ultra-high-performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UHPLC-qTOF-MS/MS). As a result, 18 phenylpropanoids, 14 flavonoids and 15 others were identified in LSC, among which stachysoside A and acteoside could be the main bio-active constituents responsible for the diuretic effect. In parallel, the daily administration of LSC (16, 32 and 64 mg/kg) markedly promoted urinary excretion after 2 h of treatment. Moreover, LSC had no effect on urinary Na+ and K+ concentrations, as well as on serum Na+-K+-ATPase activity. Meanwhile, LSC significantly decreased the serum levels of angiotensin II (Ang II), anti-diuretic hormone (ADH), aldosterone (ALD), aquaporin (AQP) 1, AQP2 and AQP3, suppressed renal AQP1, AQP2, and AQP3 mRNA expressions, down-regulated AQP1, AQP2 and AQP3 protein levels, and up-regulated serum atriopeptin (ANP) level in a dose-dependent manner. These findings suggest that LSC has acute and prolonged diuretic effects by inhibiting the AQPs, RAAS, and upregulation of atriopeptin in saline-loaded rats, and this finding support LSC as a novel diuretic agent.

EFFECT OF L- AND N-TYPE CALCIUM CHANNEL BLOCKER CILNIDIPINE ON SERUM SOLUBLE (PRO)RENIN RECEPTOR CONCENTRATION

Objective: Cilnidipine (Cil) is an L- and N-type calcium channel blocker that inhibits the release of norepinephrine (NE) from the sympathetic nerve endings. It has inhibitory effects on SNA and the renin-angiotensin-aldosterone system (RAAS), as well as its strong renal and vascular protective effects. (Pro)renin receptor [(P)RR] activates the tissue RAAS and contributes to the pathogenesis of organ damages. The soluble form of (P)RR [s(P)RR] is produced via cleavage of (P)RR, and its blood concentration is a biomarker of organ damage. The present study investigated the effects of Cil on serum s(P)RR concentration, as well as the relationships between the serum s(P)RR concentration and changes in NE and aldosterone (ALDO) levels. Design and method: Patients with essential hypertension were randomly assigned to 2 groups: the Cil treatment (C) group (n = 19) and the amlodipine treatment (A) group (18). Changes in various parameters (urinary, biochemistry, and endocrine data and cardio-ankle vascular index (CAVI), an index of arterial stiffness, during the 6 months-treatment period were compared. Results: In both groups, there was a significant decrease in blood pressure by the treatments, while there was no difference in changes in blood pressure between the two groups. In the C group, urinary albumin excretion and CAVI decreased significantly. There were no changes in s(P)RR levels in both groups. Serum NE levels (rate of decline: 13.6 ± 33.3%) and ALDO levels (18.4 ± 33.6%) significantly decreased in the C group. In the C group, while the decline rate of NE was significantly greater in the low s(P)RR group (24.6 ± 31.4%, n = 9) than in the high s(P)RR group (1.2 ± 33.0%, n = 10), changes in ALDO was not significantly different between the groups. Conclusions: The present study reconfirmed that Cil reduces NE and ALDO levels, and has renal and vascular protective effects. Although changes in serum s(P)RR concentration by Cil were not demonstrated, there was an association between the serum s(P)RR levels and the rate of decline of NE, suggesting involvement of (P)RR expression in the regulation of SNA. Future studies are needed to determine the underlying mechanism of this involvement.

CARDIORENAL CONSEQUENCE IN A CHILD WITH LATENT HEPATITIS B

Objective: Approximately 20% of patients with HBV infection have a wide range of extrahepatic damages that include renal and cardiovascular abnormalities. The purpose: is to highlight the cause of hypertension in a child with viral hepatitis B and multiorgan damage. Design and method: Describing a clinical case of a child with occult hepatitits. Results: A 1-year-old girl with toxic syndromes was hospitalized in Pediatric Department. Anamnestic data: the mother was diagnosed with HBsAg positive 4 years ago. The child was vaccinated with all 4 scheduled doses of anti-HBV vaccine. At first examination - increased BP values (160–220/100–120 mm/Hg), tachycardia, hepatomegaly. Laboratory tests: anemia gr. II, hypoproteinemia, hypoalbuminemia, hypocalcemia, slightly elevated transaminases. Ecocardiography - cardiac remodeling with dilated lef ventricle (LV); hypokinesia, and mild hypertrophy of interventricular septum; LV ejection fraction - 47%. The secondary causes of arterial hypertension have been investigated: concentration of fractionated urinary and plasma metanephrines - N, direct plasma renin - &gt; 5000 UI/mL, and serum aldosterone - 128 ng / dL. The aldosterone/renin ratio was 0.025 - suggestion of renal ischemia with secondary hyperaldosteronism. Cardiac and abdominal angioCT - severe hypoplasia of the common hepatic artery, moderate diffuse stenosis of the abdominal aorta at the renal and infrarenal levels, moderate diffuse stenosis of the bilateral renal arteries, more pronounced on the right, bilateral renal atrophy changes; cateterism cardiac - the aorta is permeable throughout, stenosis on the renal arteries, celiac trunk, and mesenteric artery was not determined, data that allowed the exclusion of the renovascular cause of hypertension. Abdominal MRI - focal areas 7x8 mm of diffusion restriction in the bilateral renal parenchymal. The patient was suspected of an HBV infection with a risk of perinatal transmission, after in dynamics, there was identified hepatomegaly (+3.0 cm) and elevated ALAT and ASAT. Quantitative plasma HBV DNA -1 418 771 300 copies/ml or 834 571 352 IU/ml, which confirmed the diagnosis - Occult viral hepatitis B, gr. III of activity, seropositive form, with extrahepatic cardiorenal impairment. Conclusions: Cardiorenal impairment should be suspected in patients with HBV infection, especially in presence of clinical signs.

GLUCO- AND MINERALOCORTICOIDS SYNERGISTICALLY DOWNREGULATE BKCA CHANNEL TRANSCRIPTION IN VASCULAR SMOOTH MUSCLE CELLS BUT DO NOT AFFECT THE EPOXYEICOSATRIENOIC ACID PATHWAY

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.

Genotypic Sex and Severity of the Disease Determine the Time of Clinical Presentation in Steroid 17α-Hydroxylase/17,20-Lyase Deficiency

Context: Steroid 17α-hydroxylase/17,20-lyase deficiency (17OHD) is characterized by decreased sex steroids and cortisol, and excessive mineralocorticoid action. The clinical symptoms of hypocortisolemia are subtle. Aim: The clinical, biochemical, and molecular characteristics of patients with 17OHD were evaluated to determine the factors influencing the time of diagnosis and the management. Patients and Methods: Clinical data, steroid profiles by liquid chromatography-tandem mass spectrometry, and Sanger sequencing of the CYP17A1 gene was evaluated in 12 patients with 17OHD diagnosed between 2004 and 2020. Results: Median age of diagnosis was 13.9 (range: 0.04–29.5) years. Ten of 12 patients had 46,XY karyotype. Except for one boy with partial 17OHD, all patients had female external genitalia hence raised as females. The clinical presentation of 17OHD was earlier (median age: 7 years) in patients, who presented with severe hypertension, atypical genitalia, or positive family history (n = 6, 50%) than those without (median age: 15.3 years; p = 0.0005). The latter group presented with amenorrhea (n = 6, 50%). Steroid profile of patients uniformly showed a typical pattern of 17OHD regardless of the age at diagnosis. Serum gonadotropin concentrations were elevated in patients >12 years (n = 7), normal in pre-adolescents (n = 4), and low in a patient, who had a digenic inheritance of homozygous CYP17A1 and KISS1R mutations. Conclusions: Early clinical presentation and diagnosis in 17OHD are associated with symptomatic hypertension in both 46,XX and 46,XY patients or inadequate virilization of external genitalia in 46,XY partial 17OHD. In the absence of these, the clinical presentation is at late pubertal ages at which time amenorrhea and elevated gonadotropins are the hints for diagnosis.

Relationship between plasma aldosterone and left ventricular structure and function in patients with heart failure with preserved ejection fraction

Aim. To study the relationship between plasma aldosterone level and left ventricular (LV) structure and function in heart failure with preserved ejection fraction (HFpEF).Material and methods. This prospective study included 158 patients (58 men and 100 women, mean age, 62,3±7,4 years) with compensated HFpEF. Patients had no history of primary aldosteronism and did not use the mineralocorticoid receptor antagonists during the last 6 weeks. The plasma aldosterone was determined by enzyme immunoassay in all patients and the severity of structural and functional cardiac changes was assessed. The concentration of 40160 pg/ml was considered the reference values. Assessment of cardiac structure and function was carried out using transthoracic echocardiography.Results. According to the laboratory results, all patients were divided into two groups: group 1 — 99 (62,7%) patients (95% confidence interval (CI), 55,0-70,0%) with normal aldosterone levels; group 2 — 59 (37,3%) patients (95% Cl, 30,0-45,0%) with hyperaldosteronism. End-diastolic volume, left atrial volume, LV mass index, severity of LV diastolic dysfunction and the prevalence of concentric hypertrophy were significantly higher in group 2 patients compared with group 1 (p&lt;0,05 for all). Blood aldosterone levels positively correlated with E/e’ (r=0,63, p&lt;0,001). Regression analysis, adjusted for age and comorbidity, demonstrated that plasma aldosterone levels were closely associated with E/e’ (odds ratio, 3,42; 95% CI, 1,65-9,64, p=0,001) and LV concentric hypertrophy (odds ratio, 1,12; 95% CI, 1,08-3,16, p=0,042).Conclusion. The development of secondary hyperaldosteronism in patients with HFpEF is an independent predictor of LV diastolic dysfunction and unfavorable prognostic types of LV remodeling.

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

[Figure: see text].

Hyperglycemia Potentiates Prothrombotic Effect of Aldosterone in a Rat Arterial Thrombosis Model

We investigated the role of aldosterone (ALDO) in the development of arterial thrombosis in streptozotocin-induced diabetic rats. To evaluate the effect of endogenous ALDO, the rats underwent adrenalectomy (ADX). ADX reduced the development of arterial thrombosis. A 1 h infusion of ALDO (30 μg/kg/h) enhanced thrombosis in adrenalectomized rats, while this effect was potentiated in diabetic rats. ALDO shortened bleeding time, increased plasma levels of tissue factor (TF) and plasminogen activator inhibitor, decreased plasma level of nitric oxide (NO) metabolites, and increased oxidative stress. Moreover, 2 h incubation of human umbilical vein endothelial cells (HUVECs) with ALDO (10−7 M) disrupted hemostatic balance in endothelial cells in normoglycemia (glucose 5.5 mM), and this effect was more pronounced in hyperglycemia (glucose 30 mM). We demonstrated that the acute ALDO infusion enhances arterial thrombosis in rats and hyperglycemia potentiates this prothrombotic effect. The mechanism of ALDO action was partially mediated by mineralocorticoid (MR) and glucocorticoid (GR) receptors and related to impact of the hormone on primary hemostasis, TF-dependent coagulation cascade, fibrinolysis, NO bioavailability, and oxidative stress balance. Our in vitro study confirmed that ALDO induces prothrombotic phenotype in the endothelium, particularly under hyperglycemic conditions.

The chemical profiling of aqueous soluble fraction from Lagopsis supina and its diuretic effects via suppression of AQP and RAAS pathways in saline-loaded rats

Ethnopharmacological relevanceLagopsis supina (Steph.) Ik. -Gal. ex Knorr. has been widely used as a remedy treatment for diuresis and edema in China over 2500 years. Our previous results showed that the aqueous soluble fraction from L. supina (LSB) possessed acute diuretic effect. Aim of the studyThe aim of this study was to appraise the acute (6 h) and prolonged (7 d) diuretic effects, underlying mechanisms, and chemical profiling of LSB. Materials and methodsThe chemical profiling of LSB was performed by ultra-high-performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UHPLC-qTOF-MS/MS). Then, oral administration of LSB (40, 80, 160 and 320 mg/kg) and furosemide (10 mg/kg) once daily for 7 consecutive days to evaluate the diuretic effects in saline-loaded rats. The body weight, food consumption, and water intake were recorded once daily. The urinary volume, pH and electrolyte concentrations (Na+, K+, Cl−, and Ca2+) were measured after administration drugs for acute and prolonged diuretic effects. In addition, the serum levels of Na+-K+-ATPase, angiotensin II (Ang II), anti-diuretic hormone (ADH), aldosterone (ALD), atriopeptin (ANP), aquaporins (AQPs)-1, 2 and 3 were determined by ELISA kits. The mRNA expressions and protein levels of AQPs-1, 2 and 3 were analyzed by real-time quantitative PCR and Western blot assays, respectively. Results30 compounds were identified in LSB based on accurate mass and MS/MS fragmentation compared to literature, among which phenylpropanoids and flavonoids could be partly responsible for the major diuretic effect. Daily administration of LSB (160 or 320 mg/kg) prominently increased urinary excretion volume after the 2 h at the first day of treatment, remaining until the 7th day. LSB did not cause Na+ and K+ electrolyte abnormalities, and has minor effect on Cl− and Ca2+ concentrations at 320 mg/kg. Furthermore, LSB observably suppressed renin-angiotensin-aldosterone system (RAAS) activation, including decreased serum levels of Ang II, ADH, and ALD, and prominently increased serum level of ANP in rats. LSB treatment significantly down-regulated the serum levels, mRNA expressions and protein levels of AQP1, AQP2, and AQP3. ConclusionLSB has a prominent acute and prolonged diuretic effects via suppression of AQP and RAAS pathways in saline-loaded rats, and support the traditional folk use of this plant. Taken together, LSB might be a potential diuretic agent.