Urinary Aldosterone Research Articles

Abstract 17781: Spironolactone Improves Left Atrial Function and Atrioventricular Coupling in Resistant Hypertension

Purpose: To determine the blood pressure independent effects of spironolactone on left atrial (LA) size and function in patients with resistant hypertension (RHTN). Methods: Patients with RHTN (N=36, 55±7 years) were prospectively recruited. Spironolactone was initiated at 25 mg/day and increased to 50 mg/day after four weeks. Other antihypertensives were withdrawn to maintain constant blood pressure. Cardiac magnetic resonance imaging was performed at baseline and after six months of spironolactone treatment and changes in LA functional metrics were assessed (Figure 1). The statistical analysis was conducted using two-sided tests of significance. Results: LA size and function parameters improved from baseline to month-six: LA volumes indexed to body surface area (LAVI) were reduced (LAVI max 41.4±12 vs 33.2±9.7 ml/m 2 ; LAVI pre-A 32.6±9.8 vs 25.6±8.1 ml/m 2 ; LAVI min 18.5 [13.9 - 24.8] vs 14.1 [10.9 - 19.2] ml/m 2 ; all p < 0.05); left atrioventricular coupling index was reduced (28.2±11.5 vs 22.7±9.2 %, p < 0.05); LA emptying fractions (LAEF) were increased (total LAEF 52.4 [48.7 - 60.3] vs 55.9 [50.3 - 61.1] %; active LAEF 40.2±8.6 vs 43.1±7.8 %, both p < 0.05). There was a substantial increase in reservoir strain (29.1 ± 8.5 % vs 30.9 ± 5.5 %, p = 0.068) whereas active strain was significantly increased from baseline (16.3 ± 4.1 % vs 17.8 ± 4.2 %, p < 0.05). Changes in passive strain and strain rates were not statistically significant (p > 0.1). The effect of spironolactone was similar in patients with high (N = 18) and normal (N = 18) aldosterone status (defined by plasma renin activity and 24-hour urine aldosterone). Conclusion: Treatment of RHTN with spironolactone is associated with improvements in LA size and function regardless of whether aldosterone levels were normal or high. This study suggests the need for larger prospective studies examining effects of mineralocorticoid receptor antagonists on atrial function and atrioventricular coupling.

Circadian Rhythm Misalignment Induced Differential Expression of Mineralocorticoid Receptor Type I and Aldosterone in Kidneys and Urine of Male Rats

Circadian Rhythm Misalignment Induced Differential Expression of Mineralocorticoid Receptor Type I and Aldosterone in Kidneys and Urine of Male Rats

Structural and clinical characterization of CYP11B2 inhibition by dexfadrostat phosphate

Aldosterone synthase (CYP11B2) represents a promising drug target because its genetic dysregulation is causally associated with cardiovascular disease, its autonomous activity leads to primary aldosteronism, and its deficiency leads to salt wasting syndromes. The serendipitous discovery that the dextro-rotatory stereoisomer of the racemic aromatase (CYP19A1) inhibitor CGS16949A mediates potent CYP11B2 inhibition led to the purification and clinical development of dexfadrostat phosphate. To characterize the pharmacophore of dexfadrostat phosphate, structure-based enzyme coordination with CYP11B2, CYP11B1 and CYP19A1 was combined with steroid turnover upon in vitro and clinical treatment. Dexfadrostat, but not its 5S-enantiomer (5S-fadrozole), precisely coordinates with the catalytic heme moiety in the space of the CYP11B2 substrate binding pocket forming a tight and stable complex. Conversely, neither rigid nor flexible docking led to a plausible coordination geometry for dexfadrostat in steroid 11β-hydroxylase (CYP11B1 – orthologue to CYP11B2) or in CYP19A1. The inhibitory preference of dexfadrostat was confirmed in vitro using an adrenal cortex-derived cell line. Dexfadrostat phosphate treatment of healthy subjects in the context of a clinical phase 1 study led to a dose-dependent decrease in urinary aldosterone secretion, accompanied by an increase in urinary corticosterone and deoxycorticosterone metabolites. Increased urinary corticosterone metabolites are indicative of CYP11B2 (18-oxidase) inhibition with clinical features reminiscent of patients with inborn corticosterone methyloxidase type II deficiency. An off-target effect on CYP19A1 was not observed as indicated by no clinical changes in testosterone and estradiol levels. Therefore, dexfadrostat exhibits the ideal structural features for binding and catalytic inhibition of CYP11B2 but not CYP11B1. Clinically, treatment with dexfadrostat phosphate leads to suppression of aldosterone levels by inhibiting predominantly one or both final CYP11B2-mediated reactions.

FRI165 BIUXX

Abstract Disclosure: A. Haider: None. M. Sadiq: None. J. Shakesprere: None. G.K. Sidhu: None. Background: Syndrome of inappropriate anti-diuretic hormone (ADH) secretion (SIADH) is characterized by inappropriate ADH secretion causing hyponatremia due to free water retention. Plasma volume expansion leads to renin inhibition by the renal juxtaglomerular complex, resulting in decreased aldosterone secretion with inappropriate natriuresis. However, when autonomous renin-independent aldosterone production is present, natriuresis can be suppressed, causing normal sodium levels despite inappropriately high ADH. Case: 58-year-old male with history of resistant hypertension & bipolar disorder on Oxcarbazepine presented with dizziness, lightheadedness, and severe hypokalemia (2.7 mmol/L, n 3.5-5.0 mmol/L). CT Adrenals revealed an incidental 1.2 cm left-sided adrenal adenoma [HU 6, 40% washout]. Serum aldosterone and plasma renin activity (PRA) were obtained in the supine position via LC/MS/MS assay; concurrent serum K+ was 3.4 mmol/L. Spironolactone was started and titrated to 50 mg BID for blood pressure control due to high suspicion of hyperaldosteronism. The patient was continued on Oxcarbazepine 900 mg BID. A 3 mg Dexamethasone suppression test showed suppressed morning cortisol (1.0 μg/dL, n <1.8 μg/dL). Urine catecholamines and metanephrines were obtained and normal. 3-4 days after starting Spironolactone, the patient developed severe subacute hyponatremia (119 mmol/L, n 135-145 mmol/L) from an admission level of 137 mmol/L. No active renal or GI losses, diuretic use, or untreated hypothyroidism were documented. Serum osmolality was low (262 mOsm/kg, n 275-295 mOsm/kg) with urine osmolality of 357 mOsm/kg (n 50-1200 mOsm/kg) and urine sodium of 92 mEq/L (n >20 mEq/L). Serum aldosterone and PRA levels returned with normal aldosterone (7 ng/dL, n 2-9 ng/dL) but suppressed PRA (0.07 ng/mL/hr, n 0.7-3.3 ng/mL/hr) and elevated aldosterone-renin ratio (100 ng/dL per ng/mL/hr, n ≤ 30 ng/dL per ng/mL/hr). As renin was very low despite mild hypokalemia and active Losartan use at the time of testing, these findings were suggestive of primary aldosteronism. In-hospital 24-hour urine aldosterone collection showed urine aldosterone of 29.3 mcg/24 h (n 5-19 mcg/24 h) with urine sodium of 93 mmol/ 24 h (n 40-220 mmol/24 h). Oxcarbazepine was subsequently tapered off; Spironolactone was intermittently held until complete discontinuation. Fluid restriction and oral salt repletion were started and the patient’s sodium ultimately normalized. Outpatient saline suppression testing and adrenal vein sampling were scheduled. Conclusion: Oxcarbazepine was likely causing high ADH release but co-occurring untreated hyperaldosteronism inhibited natriuresis. Use of Spironolactone unmasked natriuresis by blocking aldosterone activity on the intercalated cells of the renal nephron. Stopping Oxcarbazepine ultimately resulted in hyponatremia resolution. Presentation Date: Friday, June 16, 2023

FRI132 Aldosterone Secretory Capacity Immediately After Surgery Predicts Biochemical Outcome 1 Year After Surgery In Primary Aldosteronism

Abstract Disclosure: Y. Ishida: None. K. Nakai: None. Y. Tsurutani: None. K. Watanabe: None. J. Saito: None. T. Nishikawa: None. Background and Objective: In primary aldosteronism (PA), the biochemical outcome of the Primary Aldosteronism Surgical Outcome (PASO) study is used to assess aldosterone hypersecretion six months to one year after surgery. However, there are few reports on whether outcomes can be predicted in the early postoperative period. This retrospective study evaluated aldosterone secretory capacity immediately after surgery using the adrenocorticotropin (ACTH) stimulation test (AST) and oral salt loading test (OST). We hypothesized that these tests would predict biochemical outcomes one year after surgery. Methods: This study assessed 282 patients with PA who underwent adrenalectomy in our hospital between 2008 and 2020 and determined biochemical PASO outcomes 1 year after surgery. Patients with serum cortisol levels ≥ 5 μg/dL after a 1 mg overnight dexamethasone suppression test (n = 9) were excluded. Patients were divided into biochemical complete success (COM) and incomplete (partial + absent) success (INC) groups. AST and OST were performed within 15 days of surgery. The relationships between various AST and OST values and outcomes were analyzed. Results: Two hundred and twenty patients and 53 patients were classified into the COM and INC groups, respectively. The INC group had significantly higher plasma aldosterone concentration (PAC) and higher PAC/cortisol ratio (A/C) at baseline, 30 min, and 60 min after ACTH loading than the COM group. The INC group showed a significantly lower level of plasma renin activity. In the OST, the INC group had a significantly higher 24-hour urine aldosterone level. Of these values, A/C at 30 minutes after ACTH loading (area under the curve (AUC) = 0.77) and 24-hour urine aldosterone (AUC = 0.78) were relatively superior predictors of the outcome, with higher AUCs than plasma renin activity (AUC = 0.63), baseline PAC (AUC = 0.71), baseline ARR (AUC = 0.72), and baseline A/C (AUC = 0.70). A/C at 30 minutes after ACTH loading had 83% sensitivity and 62% specificity when 6.03 was set as the cutoff; 24-hour urine aldosterone had 84% sensitivity and 61% specificity when 2.6 was set as the cutoff. Conclusion: AST and OST immediately after surgery are useful for predicting biochemical outcomes 1 year after surgery in patients with PA. Presentation: Friday, June 16, 2023

Aldosterone-progesterone relationship in sexually intact Chihuahua bitches

BackgroundAldosterone represents an important target of heart failure therapy and may be a valuable indicator of the renin-angiotensin-aldosterone system activity. However, its assessment might be challenging because of the effect of individual factors. In a recent study, intact female dogs showed the highest value of urinary aldosterone-to-creatinine ratio (UAldo:C) compared to other sex categories. In humans and rodents, an influence of progesterone has been reported by several studies. To our knowledge, the relationship between aldosterone and progesterone has not yet been investigated in dogs. The aim of this prospective study was to investigate this relationship in sexually intact Chihuahua females, measuring both hormones twice in the same bitch, that is in anoestrus when progesterone concentrations are baseline and in dioestrus when they are high.ResultsThe study population consisted of 14 sexually intact Chihuahua bitches. Serum progesterone (34.06 (21.17–44.90) vs. 0.19 [0.13–0.38] ng/ml; P < 0.001) and urinary aldosterone (9886.98 ± 5735.22 vs. 5005.72 ± 2127.73 pg/ml; P = 0.01) were significantly higher in dioestrus compared to anoestrous. Urinary aldosterone-to-creatinine ratio was higher in dioestrus compared to anoestrus (4.16 [3.17–6.80] vs. 3.39 ± 1.64 µg/g), but it did not reach the statistical significance (P = 0.056). Serum progesterone showed a moderate positive correlation with urinary aldosterone (ρ = 0.638, P < 0.001) and UAldo:C (ρ = 0.516, P = 0.005).ConclusionsThe results of the present study suggest the existence of a progesterone-aldosterone relationship in canine species, indicating that sex and phase of reproductive cycle should be taken into account when interpreting aldosterone concentrations. Further studies are needed to confirm these results on a larger canine population and to identify the underlying mechanisms in this species.

Simultaneous quantitation of urine aldosterone and tetrahydroaldosterone in healthy Chinese subjects using a validated LC-MS/MS method.

Aldosterone (ALD) is excreted in urine mainly as glucuronide conjugates of ALD and tetrahydroaldosterone. Measuring these urinary metabolites might be an alternative screening test to plasma ALD for primary aldosteronism. We report a validated LC-MS/MS method to measure both analytes simultaneously. Urine samples underwent enzymatic hydrolysis to release the analytes from their glucuronide conjugates followed by organic solvent extraction and LC-MS/MS. The analytical performance of this method was evaluated. The within-batch and between-batch coefficients of variation for urine ALD and urine THA were all ≤5.2 and ≤3.7%. The lower limit of quantification was 0.5nmol/L, and the linearity was up to at least 2770 nmol/L for both analytes. No significant matrix interference and carryover were observed. Both analytes in urine were stable for at least 48 h at 10°C and at least 18 months at -80°C. Local reference intervals were established from 126 healthy normotensive Chinese subjects (53% women, age: 20-65 years). Reference intervals for urine ALD and tetrahydroaldosterone were 2-38 and 9-139 nmol/day, respectively. This validated method can be applied to screening and diagnosing primary aldosteronism.

#5527 ALDOSTERONE, CORTISOL AND URINE VOLUME CAN EXPLAIN DISCREPANCIES BETWEEN DIETARY AND URINARY NA/K RATIO

Abstract Background and Aims Increased urinary sodium-to-potassium (Na/K) ratio has been associated with chronic kidney disease. However, we demonstrated that urinary Na/K ratio does not accurately reflect dietary Na/K ratio. Rhythmic patterns of sodium storage and release from the skin interstitium are associated with aldosterone and cortisol levels and previous studies linked tissue sodium accumulation to water conservation. Therefore, we investigated whether the discrepancy between urinary and dietary Na/K ratio could be explained by these factors. Method For this post-hoc analysis, we used data from the long-term sodium balance studies Mars105 and Mars520. These studies collected 24-hour urine samples and controlled dietary salt intake for 105 (4 men) and 205 days (6 men) at levels of 6, 9 and 12 grams per day. We calculated the difference between urinary and dietary Na/K ratio. We tested whether this difference varied across salt intake levels in a linear mixed-effects model. Furthermore, we fitted two linear mixed-effect models to explain discrepancies between urinary and dietary Na/K ratio. In the first model, sodium intake, potassium intake, urinary aldosterone and urinary cortisol were fixed effects and for each factor random slopes per participant were allowed. In the second model, sodium intake, potassium intake and urine volume were fixed effects and for each factor random slopes per participant were allowed. Each salt intake level was assessed separately. Results The median difference between 24-hour urinary and dietary Na/K ratio was −0.21 (IQR −0.47 to 0.09). At higher salt intake levels the underestimation was significantly larger (mean difference −0.11, −0.22 and −0.36 for 6, 9 and 12 grams salt intake, respectively; P&amp;lt;0.001). At each salt intake level, higher urinary aldosterone was associated with underestimation of the dietary Na/K ratio (Figure 1A). In contrast, higher urinary cortisol and higher urine volume were associated with relative overestimation of the dietary Na/K ratio (Figure 1B and C, respectively). Conclusion The discrepancy between dietary and urinary Na/K ratio can be explained by sodium intake, aldosterone, cortisol and urine volume. This suggests that tissue sodium accumulation contributes to the inaccuracy of a single 24-hour urine collection for estimation of dietary sodium and potassium intake.

A Novel Enzymatic Hydrolysis Method for Urine Aldosterone Quantification: A Case for Reassessing Clinical Cut-Offs of Primary Aldosteronism.

Primary aldosteronism (PA) is a common endocrine cause of secondary hypertension. The aldosterone/renin ratio is an important tool for PA screening, and dynamic testing in serum or urine is used to confirm the diagnosis. While LC-MS/MS is considered the gold standard for testing, there is significant interlaboratory variability between the extraction procedures, which can impact diagnostic interpretation. To help overcome this, we present a simple and accurate LC-MS/MS method for the quantification of both serum and urine aldosterone using a novel enzymatic hydrolysis procedure. Serum and urine aldosterone was extracted and measured by LC-MS/MS. Urine-conjugated aldosterone glucuronide was hydrolyzed using a genetically modified glucuronidase enzyme. The assay precision, accuracy, limit of quantification, recovery, and carryover were evaluated and the new assay cut-offs were proposed. The liquid chromatography method allowed for adequate separation of the aldosterone peak from closely eluting peaks. Significant in vitro aldosterone loss was observed during acid-catalyzed hydrolysis of urine, which was corrected with the addition of the internal standard to the urine before the hydrolysis step. Glucuronidase catalyzed hydrolysis of urine aldosterone glucuronide displays good correlation with the corrected acid-catalyzed hydrolysis. Serum aldosterone showed good agreement with reference values and the consensus range reported for external quality assessment specimens. A simple, fast, and highly accurate method for the detection of serum and urine aldosterone has been developed. The proposed novel enzymatic procedure allows for short hydrolysis time and compensates for urine aldosterone loss during the hydrolysis step.

DISCREPANCIES BETWEEN DIETARY AND URINE NA/K RATIO CAN BE EXPLAINED BY ALDOSTERONE, CORTISOL AND URINE VOLUME

Objective: Increased urine sodium-to-potassium (Na/K) ratio is associated with hypertension and cardiovascular disease. However, we demonstrated that urine Na/K ratio does not accurately reflect dietary Na/K ratio. As rhythmic patterns of sodium storage and release are associated with aldosterone and cortisol levels and previous studies linked tissue sodium accumulation to water conservation, we investigated whether the discrepancy between urine and dietary Na/K ratio could be explained by these factors. Design and method: We performed a post-hoc analysis of the long-term sodium balance studies Mars105 and Mars520. Ten healthy participants consumed a diet with known sodium and potassium content. For 105 or 205 days 24-hour urine samples were collected. We calculated the difference between urine and dietary Na/K ratio. A linear mixed-effect model was used to determine whether this difference varied across salt intake levels. Furthermore, we fitted two linear mixed-effect models to explain discrepancies between urine and dietary Na/K ratio. In the first, sodium intake, potassium intake, urine aldosterone and urine cortisol were fixed effects and for each factor random slopes per participant were allowed. In the second, sodium intake, potassium intake and urine volume were fixed effects and for each factor random slopes per participant were allowed. We analysed the salt intake levels 6, 9 and 12 grams separately. Results: Urine Na/K ratio underestimated dietary Na/K ratio (median difference -0.21, IQR -0.47 to 0.09). The discrepancy was significantly larger at higher salt intake levels (mean difference -0.11, -0.22 and -0.36 for 6, 9 and 12 grams salt intake, respectively, P &lt; 0.001). At all salt intake levels, higher aldosterone levels were associated with underestimation of the dietary Na/K ratio (Figure 1A). In contrast, higher urine cortisol and higher urine volume were associated with overestimation of the dietary Na/K ratio (Figure 1B and C respectively). Conclusions: Sodium intake, aldosterone, cortisol and urine volume can explain the discrepancy between dietary and urine Na/K ratio. These data suggest that tissue sodium accumulation contributes to the inability to estimate dietary intake using the Na/K ratio in a single 24-hour urine collection.

Association Between Life's Simple 7 and Biomarkers of Cardiovascular Disease: Aldosterone, Interleukin-6, C-Reactive Protein.

Background To promote ideal cardiovascular health, the American Heart Association recommends adhering to Life's Simple 7 (LS7)-achieving healthy targets for body mass index, physical activity, dietary intake, blood pressure, fasting plasma glucose, and cholesterol, along with smoking abstinence. Poorer achievement of LS7 (lower score) has been associated with the development of hypertension and cardiovascular disease. However, less is known about the associations between LS7 and specific biomarkers linked to cardiovascular health: aldosterone, CRP (C-reactive protein), and IL-6 (interleukin-6). Methods and Results We analyzed 379 individuals (age 18-66 years) from the HyperPATH (International Hypertensive Pathotype), who were maintained on ≥200 mEq of sodium daily for 1 week. We calculated a 14-point summative LS7 score according to participants' baseline data. Based on the range of LS7 score in this population (3-14), we classified participants as "inadequate" (3-6), "average" (7-10), and "optimal" (11-14). Regression analyses found that a higher LS7 score group was associated with lower levels of serum and urinary aldosterone (Ptrend<0.001 and Ptrend=0.001, respectively), lower plasma renin activity (Ptrend<0.001), and a blunted increase in serum aldosterone with angiotensin II infusion (Ptrend=0.023). Being in the "optimal" LS7 score group was associated with lower serum CRP (Ptrend=0.001) and IL-6 (Ptrend=0.001). Conclusions A higher LS7 score was associated with a lower activity of the renin-angiotensin-aldosterone system and lower levels of the inflammatory markers CRP and IL-6. These findings offer a possible link between ideal cardiovascular health targets and biomarkers known to play a central role in the development of cardiovascular disease.

Bmal1 contributes to mean blood pressure rhythms in rats on a low salt diet

Bmal1 is a core component of the molecular clock which is responsible for generating circadian rhythms in many physiological parameters, including blood pressure (BP). Sodium intake can affect circadian rhythms, as high salt consumption can increase BP and alter the expression of the molecular clock. However, the effect of low salt consumption on the circadian rhythm of BP and hormones crucial to sodium homeostasis has not been extensively studied. Using a whole-body Bmal1-knockout (KO) rat, we wanted to determine the contribution of the molecular clock to circadian BP rhythms during low salt diet (LSD, 0.0049%).We hypothesized that Bmal1-KO rats on a LSD would have lower mean BP and higher aldosterone levels in comparison to littermate controls on LSD. Male and female Bmal1-KO rats and littermate controls (WT) had telemeters implanted in the abdominal aorta at 8-10 weeks old. After a 10-day surgical recovery period, BP was continuously recorded while rats were maintained on ad libitum normal salt diet (0.49% NaCl, NSD) for 1 week. Rats were placed in metabolic cages and 12-hour urine samples collected at ZT0 and ZT12 for two days. Next, rats were placed on LSD for one week while maintaining BP measurements. This was again followed by two day, 12-hour urine collections. BP was averaged hourly and analyzed for circadian rhythmicity (cosinor). The MESOR for mean arterial pressure was significantly higher in WT (n=10, 4/10 male) and KO groups (n=8, 4/8 male) on NSD (WT: 108±2 mmHg; KO: 104±2 mmHg) compared to LSD (WT: 104±2 mmHg; KO: 102±2 mmHg; p&lt;0.05 for diet, 2-way ANOVA). BP amplitude was significantly higher in WT rats (NSD: 7.6±0.6 mmHg; LSD: 6.9±0.7 mmHg) compared to KO rats (NSD: 5.2±0.4 mmHg; LSD: 5.3±0.9 mmHg) (p&lt;0.05 for genotype). BP acrophase was similar between groups on both diets. Urinary aldosterone excretion (ELISA) was similar between WT and KO rats on NSD (Daytime: WT 26±5 pg/12hrs, KO 16±1 pg/12hrs; Nighttime: WT 48±8 pg/12hrs, KO 46±4 pg/12hrs) and on LSD (Daytime: WT 95±38 pg/12hrs, KO 109±44 pg/12hrs; Nighttime: WT 174±63 pg/12hrs, KO 155±47 pg/12hrs), with statistically significant overall effects of diet and time of day (p&lt;0.05, 3-way ANOVA). Together, these data support a role for Bmal1 in control of BP amplitude in rats on NSD and LSD. This does not appear to involve changes in aldosterone since urinary excretion followed predicted patterns by time of day and salt diet, but were not impacted by loss of Bmal1 in ad libitum fed rats. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Sub-clinical injury in AKI prior to pregnancy induces renal inflammation, renal fibrosis and suppresses the renin-angiotensin aldosterone system (RAAS) in female rats

Acute kidney injury (AKI), a sudden loss of kidney function, is responsible for 1 in 5 US hospitalizations. Although women are considered to be protected from AKI, they make up 40% of AKI patients. Recent clinical data report a 3-5-fold increase in adverse pregnancy outcomes among women with a history of AKI, despite apparent clinical recovery of renal function before pregnancy. These women have increased odds for developing preeclampsia and fetal growth restriction, via mechanisms unknown. To address this clinical issue, we developed a rat model of pregnancy post-AKI. We show that female Sprague-Dawley (SD) rats recover renal function as measured by creatinine clearance 30 days post-AKI. These rats exhibit decreased creatinine clearance, increased blood pressure, reduced plasma volume expansion and fetal growth restriction during pregnancy compared to sham rats. This study tested the hypothesis that AKI prior to pregnancy induces renal inflammation, fibrosis and dysregulates the renin-angiotensin aldosterone system (RAAS). Female SD rats were randomized to 45-minute warm sham or bilateral renal ischemia reperfusion (N=6/grp) and mated 30 days post sham/AKI surgery. Gestational day 1 (GD1) was identified through vaginal smear. Rats were placed in metabolic cages on GD19 for urine collection and euthanized on GD20. Urinary volume/day (30±5 vs 19±2 ml/day, P=0.08) and water intake (55±5 ml vs 41±1 ml, *P&lt;0.05) increased in post-AKI pregnant rats compared to sham. In addition, urinary aldosterone levels measured by ELISA decreased in post-AKI pregnant rats compared to sham (188±34 pg/ml/day vs 312±35, *P&lt;0.05). These data suggest disruption of fluid homeostasis and RAAS by pre-pregnancy AKI. Kidney injury marker-1 (KIM-1) measured via ELISA trended to increase in pre-pregnant AKI rats compared to sham at 30 days post AKI (580±169 pg/ml vs 257±17, P=0.09) and in pregnancy (283±28 vs 216±12 pg/ml, P=0.06), suggesting a lingering kidney injury despite creatinine clearance returning to sham levels. Mason Trichrome renal staining at GD20 revealed increases in both glomerular fibrosis (0.93±0.08 vs 0.04±0.04 score, *P&lt;0.05) and tubular fibrosis (1.7±0.19 vs 0.0±0.0 score, *P&lt;0.05) in post-AKI pregnant rats compared to sham. T regulatory immune (Tregs) cell populations in blood and kidney were measured via flow cytometry on GD20. Blood Tregs (16195±2935 cells/ml vs 26740±2508, *P&lt;0.05) as well as renal Tregs (1811±364 cells/gram kidney vs 3241±405.6, *P&lt;0.05) decreased in post-AKI pregnant rats compared to sham, suggesting AKI prior to pregnancy increases renal inflammation. Taken together, these data suggest that history of AKI predisposes to pregnancy induced dysregulation of fluid hemodynamics, renal inflammation, renal fibrosis and sustained renal injury mediating adverse fetal and maternal outcomes. R00 HL146948-03 to DM; AHA CDA858380 to JLF; AHA DIVSUP0224732 to DM This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Effects of Mineralocorticoid Receptor Blockade and Statins on Kidney Injury Marker 1 (KIM-1) in Female Rats Receiving L-NAME and Angiotensin II.

Kidney injury molecule-1 (KIM-1) is a biomarker of renal injury and a predictor of cardiovascular disease. Aldosterone, via activation of the mineralocorticoid receptor, is linked to cardiac and renal injury. However, the impact of mineralocorticoid receptor activation and blockade on KIM-1 is uncertain. We investigated whether renal KIM-1 is increased in a cardiorenal injury model induced by L-NAME/ANG II, and whether mineralocorticoid receptor blockade prevents the increase in KIM-1. Since statin use is associated with lower aldosterone, we also investigated whether administering eiSther a lipophilic statin (simvastatin) or a hydrophilic statin (pravastatin) prevents the increase in renal KIM-1. Female Wistar rats (8-10 week old), consuming a high salt diet (1.6% Na+), were randomized to the following conditions for 14 days: control; L-NAME (0.2 mg/mL in drinking water)/ANG II (225 ug/kg/day on days 12-14); L-NAME/ANG II + eplerenone (100 mg/kg/day p.o.); L-NAME/ANG II + pravastatin (20 mg/kg/day p.o.); L-NAME/ANG II + simvastatin (20 mg/kg/day p.o.). Groups treated with L-NAME/ANG II had significantly higher blood pressure, plasma and urine aldosterone, cardiac injury/stroke composite score, and renal KIM-1 than the control group. Both eplerenone and simvastatin reduced 24-h urinary KIM-1 (p = 0.0046, p = 0.031, respectively) and renal KIM-1 immunostaining (p = 0.004, p = 0.037, respectively). Eplerenone also reduced renal KIM-1 mRNA expression (p = 0.012) and cardiac injury/stroke composite score (p = 0.04). Pravastatin did not affect these damage markers. The 24-h urinary KIM-1, renal KIM-1 immunostaining, and renal KIM-1 mRNA expression correlated with cardiac injury/stroke composite score (p < 0.0001, Spearman ranked correlation = 0.69, 0.66, 0.59, respectively). In conclusion, L-NAME/ANG II increases renal KIM-1 and both eplerenone and simvastatin blunt this increase in renal KIM-1.

Reference intervals for LC-MS /MS measurements of plasma renin activity, aldosterone, angiotensin II, and 24-hour urinary aldosterone in Northern Chinese Han population

BackgroundExamination of aldosterone to Renin Ratio (ARR) and plasma aldosterone concentration (PAC) or 24-h urinary aldosterone excretion (24-h UALD) was the necessary tests in confirmatory tests for primary aldosteronism (PA). We developed a combined liquid chromatography-tandem mass spectrometry method (LC-MS/MS) for plasma renin activity (PRA), PAC, and angiotensin II (Ang II) and investigated their reference intervals (RIs) in northern Chinese Han population. The RIs of 24-h UALD excretion were also studied using LC-MS/MS. MethodsA total of 309 healthy volunteers were recruited in 3 cities in China. PRA, PAC, Ang II, and 24-h UALD were measured using the laboratory-developed LC-MS/MS. Multiple linear regression and the variance component model were applied to determine if the RI needed to be split. The RIs of PRA, PAC, and Ang II were determined using the nonparametric percentile method. ResultsThe laboratory-developed LC-MS/MS method was verified and showed good performance. Standard deviation ratio (SDR) sex for PAC and SDR region for Ang II are 0.466 and 0.407, respectively, indicating that the RIs of PAC and Ang II must be divided by sex and region, respectively. In addition, the SDR 24hUK for 24-h UALD is 0.579, indicating that the RI of 24-h UALD must be partitioned by urine potassium. ConclusionRIs were established for tests related to the renin-angiotensin-aldosterone system in the apparently healthy northern Chinese Han population by the LC-MS/MS method.

Primary Aldosteronism and COVID-19-related Management, Disease Severity, and Outcomes: A Retrospective Cohort Study.

The SARS-CoV-2 virus is dependent on components of the renin-angiotensin-aldosterone system for infectivity. Primary aldosteronism (PA) is a form of secondary hypertension mediated by autonomous aldosterone production. The intersection of COVID-19 and PA, both which may involve components of the renin-angiotensin-aldosterone system, remains unknown. We assessed PA as a risk factor for COVID-19 infection and compared management, severity of disease, and outcomes during COVID-19 with a matched population of patients with essential hypertension (EH) by conducting a retrospective observational cohort study. Of the patients with PA, 81 had a negative PCR test for COVID-19, whereas 43 had a documented positive PCR test for COVID-19. Those patients with PA who tested positive for COVID-19 tended to be female (P = .08) and the majority of those with COVID-19 infection identified as non-White race (P = .02) and Hispanic ethnicity (P = .02). In a subanalysis, 24-hour urine aldosterone on initial PA diagnosis tended to be higher those in the PA group who developed COVID-19 compared with those in the PA group who did not develop COVID-19 [median (interquartile range): 36.5 (16.9, 54.3) vs 22.0 (15.8, 26.8) mcg, P = .049] and was an independent predictor of COVID-19 infection controlling for sex, race, and ethnicity. Angiotensin-converting enzyme inhibitor, angiotensin II receptor blocker, and mineralocorticoid receptor antagonist use did not differ between those patients with PA who did and did not have COVID-19 infection. Comparing those patients with PA and matched patients with EH (n = 286) who were COVID-19 PCR positive, there was a significantly higher incidence of cardiovascular complications (12 vs 2%, P = .004) in the PA vs EH group. These data begin to inform us as to whether PA should be a newly identified subpopulation at risk for COVID-19-related cardiovascular disease sequelae.

Biochemical Evaluation by Confirmatory Tests after Unilateral Adrenalectomy for Primary Aldosteronism.

Primary aldosteronism (PA) is the most common cause of endocrine hypertension. Unilateral PA can be cured using unilateral adrenalectomy (Adx). PA surgery outcome (PASO) criteria, which include clinical and biochemical outcomes, have been proposed to evaluate PA cure after Adx. However, clinical outcomes are often inconsistent with biochemical outcomes. In addition, although confirmatory tests are included as endpoints of biochemical outcomes in the PASO criteria, their clinical usefulness has not yet been established. We evaluated clinical parameters and confirmatory test results before and after Adx in 16 patients with PA and assessed the usefulness of the confirmatory tests. The following were the clinical outcomes after Adx: 37.5% complete success, 62.5% partial success, and 0% absent success. The ratio of biochemical complete success was as follows: 69% aldosterone/renin ratio and basal plasma aldosterone concentration, 19% as assessed by the captopril challenge test, 47% as assessed by the saline infusion test, 30% as assessed by the furosemide upright test, and 100% urine aldosterone. Of these, biochemical complete success was judged in four cases by aldosterone/renin ratio and basal plasma aldosterone concentration, one case by captopril challenge test, five cases by saline infusion test, and one case by furosemide upright test. Although clinical outcomes and urine aldosterone levels improved after Adx, confirmatory tests failed to improve in some cases. The current criteria are not considered useful for biochemical evaluation after Adx. To determine whether additional treatment with mineralocorticoid receptor antagonists is required, more accurate biochemical criteria should be established after Adx.

JS-HR-9: A NEW ERA OF PROGRESS IN PRIMARY ALDOSTERONISM TREATMENT, A NEW ALDOSTERONE ASSAY, AND A CLINICAL PRACTICE GUIDELINE

Primary aldosteronism (PA), one of the mineralocorticoid receptor (MR)-associated hypertension, is a highly prevalent secondary hypertension that accounts for 5–15% of all hypertension and is a severe hypertension that is prone to cerebrovascular and cardiovascular disease. Therefore, it is expected that the pathophysiology and treatment strategies for PA will be elucidated. In recent years, the major changes in PA have been in treatment and diagnosis in Japan. Bilateral or non-surgical unilateral PA is treated with mineralocorticoid receptor antagonist (MRA). Esaxerenone, a non-steroidal MRA, is now widely used in Japan for hypertension including PA. It has been reported that add-on treatment using esaxerenone with maximal tolerable doses of a renin-angiotensin system inhibitor reduced the urinary albumin creatinine ratio in patients with type 2 diabetes mellitus (ESAX-DN), suggesting a renoprotective effect against diabetic nephropathy. Several clinical studies of another non-steroidal MRA, finerenone(FIDELIO-DKD, FIGARO-DKD, and FIDELITY), have shown its renoprotective effect against diabetic nephropathy as well as cardiovascular events, particularly hospitalization for heart failure in patients with type 2 diabetes and chronic kidney disease. Although there are no reports of clinical studies on finerenone for PA, finerenone may be used in the future for type 2 diabetes patients with PA, as obesity, glucose intolerance, and sleep apnea are common complications in patients with PA. Secondly, radiofrequency ablation of macroscopic adrenal tumors has been reported as an alternative treatment for PA to lower blood pressure and plasma aldosterone levels. This treatment has been covered by health insurance providers in Japan since April 2022, but long-term outcomes need to be validated. In diagnosis, serum and urine aldosterone has been measured in Japan since early 2021 by the two-step sandwich chemiluminescent enzyme immunoassay (CLEIA) method. This new CLEIA method correlates well with LC-MS/MS values and is expected to be more accurate. As CLEIA serum aldosterone levels are lower than those measured with a previous radioimmunoassay, the Japan Endocrine Society issued the 2021 Primary Aldosteronism Clinical Guidelines, which included a provisional positive value for the diagnosis of PA. More than ever, the diagnosis of PA needs to be carefully considered. This CLEIA method is currently available only in Japan. This presentation will cover the latest treatments, diagnostic procedures, and other hot topics in PA as described above.

PS-R07-5: A CASE OF PRIMARY ALDOSTERONISM WITH DECLINED GLOMERULAR FILTRATION RATE AFTER ADRENALECTOMY

Background: Primary aldosteronism (PA) has been known to cause organ damage, such as renal and cardiovascular disorders more frequently than essential hypertension (EH), independently of blood pressure. Among renal disorders, albuminuria is more prevalent in patients with PA than EH, while GFR tends to be higher in PA due to glomerular hyperfiltration. Surgical and medical treatments for PA correct these effects of excess aldosterone, which sometimes uncover preexisting chronic kidney disease (CKD). Here we present a case of PA with CKD who manifested decline of glomerular filtration rate (GFR) and overt renal failure following resection of aldosterone-producing adenoma. Case report: A 53-year-old woman was diagnosed with hypertension at the age of 40. Her primary care physician started treatment with a calcium channel blocker (CCB). At the age of 49, she was diagnosed with Sjogren's syndrome (SS). Blood tests revealed severe hypokalemia and eGFR of 50 mL/min/1.73m2, while 24-hr urine collection showed urinary protein of 0.5 g per day. SS was suspected to underlie her renal disease. A renal biopsy was performed, and suggested nephrosclerosis and IgA nephropathy. On the other hand, a 11mm-sized tumor in the left adrenal gland was discovered during the investigation of secondary hypertension. PA was confirmed based on active renin concentration (ARC) of 4.3 pg/mL, plasma aldosterone concentration (PAC) of 342 pg/mL, an aldosterone-to-renin ratio of 79.5 (&gt; 40), and 24-hr urinary aldosterone levels of 15.9 μg per day under the adequate salt-loaded condition. Adrenal venous sampling yielded aldosterone hypersecretion from the left adrenal vein, which indicated aldosterone-producing adenoma. Laparoscopic partial left adrenalectomy was performed and fully normalized ARC, PAC, and urinary aldosterone levels, which indicated biochemical cure of PA. Consequently, blood pressure had been well controlled with a single CCB. However, while her preoperative serum creatine level ranged from 0.9 to 1.1 mg/dL and eGFR was around 40–50 mL/min/1.73m2, her postoperative serum creatine level increased to 1.4 to 1.9 mg/dL and eGFR decreased to 25 to 35 mL/min/1.73m2 on average. In contrast, urinary protein level improved to less than 0.1 g/day, suggesting that post-treatment increase in creatine level indicated unveiling CKD rather than deterioration of renal function. Conclusion: PA treatment may unmask CKD through improving glomerular hyperfiltration from aldosterone excess.

PS-C26-8: UNIQUE PROPERTIES OF DEPRESSOR, RENAL, AND ALDOSTERONE RESPONSES TO ALISKIREN THERAPY IN HYPERTENSIVE PATIENTS WITH NEPHROPATHY

Objective: Blood pressure (BP) control using renin-angiotensin system (RAS) inhibitor as a first choice is recommended in hypertension accompanied by chronic kidney disease (CKD), and RAS localized in the kidney is known to exist independent of circulating RAS. Aliskiren, direct renin inhibitor (DRI), is most excellent in kidney tissue migration among RAS inhibitors despite it has been reported to cause aldosterone breakthrough like angiotensin II receptor blocker (ARB) in essential hypertension. Therefore, DRI-induced antihypertensive, renoprotective and aldosterone-suppressing effects were investigated as compared to those of ARB in hypertensive CKD patients. Design and method: 168 hypertensive patients with CKD (stage G1–4) were employed in this study. Home BP in the morning, estimated glomerular filtration rate (eGFR), urinary protein/creatinine ratio (UPCR), urinary liver-type fatty-acid-binding protein (U-LFABP) as an indicator of tubular dysfunction, plasma and urinary aldosterone concentration (PAC and UAC) were measured before and 0.5 to 5 years after administration of ARB or DRI and assessed retrospectively. Aldosterone breakthrough was defined as any increase in PAC from the baseline level following treatment with a RAS inhibitor. Results: In ARB-treated group, BP and UPCR decreased significantly at 6 months, but sufficient declines were not observed 1st year onward. In DRI-treated group, a significant reduction in BP was sustained until 3 years and BP unchanged decreasing tendency even 4 years later, meanwhile UPCR decreased significantly for 2 years and was below the pretreatment value 3 years after. ARB raised BP and UPCR again in patients with CKD G3–4 prior to those with CKD G1–2, however, DRI expressed no difference between stages. Although PAC diminished up to 6 months in ARB group and 1 year in DRI group, it turned up 1 year and 2 years after, respectively. PAC-ascensive rate and maximum elevation in PAC were higher and greater in ARB group than in DRI group (48.7 vs 22.6%, 56.1 vs 29.3pg/ml). All significant differences in rising PAC cases disappeared by 5 years, whereas in non-rising PAC cases, only DRI reduced U-LFABP without deterioration in eGFR and UAC further lowered while PAC was maintained. Conclusions: In comparison with ARB, aliskiren prolongs hypotensive and proteinuria-lowering effects with retained eGFR regardless of CKD severity possibly via improvement of tubular disorders and less PAC reascent in hypertensive nephropathy. Aliskiren given to CKD patients might elicit mild and delayed aldosterone breakthrough rather than ARB due to a direct action against aldosterone derived from the kidney.