Chemistry, toxicology & urinalysis

• Aldosterone is a mineralocorticoid hormone derived from cholesterol in the zona glomerulosa (the outer layer) of the adrenal cortex

• Aldosterone helps control water and salt balance; production is controlled by the renin angiotensin aldosterone system (RAAS), adrenocorticotropic hormone (ACTH) and extracellular potassium concentration
• Most patients with hyperaldosteronism present with normokalemic hypertension; hypoaldosteronism should be considered in any patient with persistent hyperkalemia
• Adrenal adenoma and bilateral adrenal hyperplasia are the most common causes of hyperaldosteronism; causes of hypoaldosteronism include disorders that reduce aldosterone synthesis and cause aldosterone resistance
• Screening test is a concurrent measurement of plasma renin, aldosterone and calculation of aldosterone to renin ratio (ARR); confirmatory testing and subtype classification should be followed with positive screening results
• Many variables in preanalytical, analytical and postanalytical steps can affect plasma aldosterone and renin results; these factors should be considered when interpreting test results

Images hosted on other servers:

Causes of hyperaldosteronism (Jameson: Harrison's Endocrinology, 3rd Edition, 2013, Eur J Endocrinol 2019;180:R45)

Primary hyperaldosteronism (isolated excess production of aldosterone)
Bilateral (micronodular) adrenal hyperplasia	60%
Adrenal (Conn) adenoma	40%
Glucocorticoid remediable aldosteronism (ACTH driven)	1%
Secondary hyperaldosteronism (excessive activation of the RAAS)
Renin producing tumor
Renal artery stenosis
Edematous disorders (e.g., heart failure, cirrhosis with ascites, nephrotic syndrome)
Other rare causes (pseudoprimary aldosteronism due to exogenous aldosterone or enhanced mineralocorticoid activity)
Syndrome of apparent mineralocorticoid excess
Cushing syndrome
Glucocorticoid resistance
Adrenocortical carcinoma
Congenital adrenal hyperplasia
Progesterone induced hypertension
Liddle syndrome

• Clinical manifestations of hyperaldosteronism
  • Primary and secondary hyperaldosteronism present similarly but can be differentiated by laboratory tests and diagnostic studies
  • Most patients present with normokalemic hypertension; hypokalemia can be only seen in the more severe cases (J Clin Endocrinol Metab 2016;101:1889)

Causes of hypoaldosteronism (adapted from UpToDate: Etiology, Diagnosis, and Treatment of Hypoaldosteronism [Accessed 8 August 2023], StatPearls: Hypoaldosteronism [Accessed 8 August 2023])

Reduced aldosterone production
Hyporeninemic hypoaldosteronism	Kidney disease, most often diabetic nephropathy	Most common acquired causes
	Nonsteroidal anti-inflammatory drugs
	Calcineurin inhibitors
Angiotensin inhibitors (ACE inhibitors, angiotensin II receptor blockers and direct renin inhibitors)
Chronic heparin therapy
Primary adrenal insufficiency		Infrequent causes
Severe illness
Inherited disorders	Congenital hypoaldosteronism (21 hydroxylase deficiency and isolated hypoaldosteronism)
	Pseudohypoaldosteronism type 2 (Gordon syndrome)
Aldosterone resistance
Inhibition of the epithelial sodium channel	Potassium sparing diuretics
	Antibiotics (trimethoprim and pentamidine)
Voltage defects	Markedly reduced distal sodium delivery
	Acquired or congenital defects in sodium reabsorption by the distal tubule principal cells (obstructive uropathy), systemic lupus erythematosus (SLE) and sickle cell disease
Inherited disorders	Pseudohypoaldosteronism type 1

• Clinical manifestations of hypoaldosteronism
  • Persistent hyperkalemia with no obvious cause
  • Mild metabolic acidosis with a normal anion gap

• Hyperaldosteronism (J Clin Endocrinol Metab 2016;101:1889)
  • Screening test of hyperaldosteronism is concurrent measurement of plasma renin activity (PRA) or direct renin concentration (DRC) and aldosterone concentration (PAC) and calculation of aldosterone to renin ratio (ARR)
  • Confirmatory testing with a positive ARR screening result includes saline infusion, oral sodium loading, fludrocortisone suppression and captopril challenge
    • For oral sodium loading, aldosterone is measured in the 24 hour urine collection
  • Adrenal CT and adrenal vein sampling (AVS) are used for subtype classification of confirmed hyperaldosteronism
  • AVS should be performed to lateralize adrenal mass when surgical treatment is feasible, with or without cosyntropin stimulation
    • Cortisol corrected aldosterone ratio of high side to low side > 4:1 indicates unilateral aldosterone production
    • Ratio of < 3:1 suggests bilateral aldosterone production (Surgery 2004;136:1227)
• Hypoaldosteronism
  • PRA or PAC, PAC and serum cortisol can be used to differentiate different causes of hypoaldosteronism

Differential diagnosis of hyper or hypoaldosteronism based on laboratory tests (Endotext: Hyperaldosteronism [Accessed 8 August 2023], UpToDate: Etiology, Diagnosis, and Treatment of Hypoaldosteronism [Accessed 8 August 2023])

Disease	Laboratory test
	PAC	PRA or PRC	ARR	Serum cortisol
Primary hyperaldosteronism	↑	↓	↑	x
Pseudoprimary hyperaldosteronism	↓	↓	/	x
Secondary hyperaldosteronism	↑	↑	↑	x
Hyporeninemic hypoaldosteronism	↓	↓	/	→
Primary adrenal insufficiency	↓	↑	/	↓
Congenital adrenal hyperplasia	↓	↑	/	→

* Symbols: ↑ = increase; ↓ = decrease; → = normal; x = not used; / = variable

• Preanalytical requirements (J Clin Endocrinol Metab 2016;101:1889)
  • Normalize serum potassium
  • Unrestricted dietary salt intake
  • Withdraw interfering medications that markedly or moderately affect the ARR if possible or commence medications that minimally affect the ARR
  • Collect blood samples midmorning after the patient has been up (sitting, standing or walking) for a minimum of 2 hours and then seated for 5 - 15 min prior to collection
  • For either renin mass or activity, it is recommended that samples are collected and processed at room temperature (not chilled), followed by rapid freezing of separated plasma at -20 °C to prevent cryoactivation
    • Cryoactivation may result in a false elevation of renin mass or activity

Medications that markedly affect the ARR (withdraw for at least 4 weeks)	Spironolactone, eplerenone, amiloride and triamterene
	Potassium wasting diuretics
	Products are derived from licorice root
Medications that moderately affect the ARR (withdraw for at least 2 weeks)	Beta adrenergic blockers, central alpha 2 agonists (e.g., clonidine, alpha methyldopa) and nonsteroidal anti-inflammatory drugs
	Angiotensin converting enzyme inhibitors, angiotensin receptor blockers, renin inhibitors and dihydropyridine calcium channel antagonists
Medications that minimally affect ARR	Verapamil slow release, hydralazine, prazosin, doxazosin, terazosin

• Analytical limitations (Clin Biochem 2015;48:377)
  • DRC assay crossreactivity with prorenin is most significant in the low renin state
  • Lack of assay standardization is a common problem with both CLIA and LC MS / MS assay for aldosterone and renin measurement
• Postanalytical considerations
  • PRA: reporting units of ng/mL/h is most commonly used
  • Plasma aldosterone concentration (PAC): reporting patient posture and providing posture specific (supine and upright) reference intervals
  • ARR cutoff values depend on assay and reporting units; numbers with asterisks indicate the most commonly adopted cutoff values (J Clin Endocrinol Metab 2016;101:1889)

	PRA, ng/mL/h	PRA, pmol/L/min	DRC, mU/L	DRC, ng/L
PAC (as ng/dL)	20	1.6	2.4	3.8
	30*	2.5	3.7	5.7
	40	3.1	4.9	7.7
PAC (as pmol/L)	750*	60	91	144
	1000	80	122	192

• Comparison studies of platforms or methodologies
  • Methodology to measure aldosterone and renin assay
    • Assays have evolved from radioimmunoassay (RIA) and chemiluminescent immunoassay (CLIA) to liquid chromatography tandem mass spectrometry (LC MS / MS) assay
    • CLIA is commonly used on automated instruments; the results from RIA and CLIA are shown to be comparable (J Hypertens 2016;34:920, Prague Med Rep. 2021;122:80)
    • LC MS / MS method is available in large reference laboratories
      • Aldosterone was substantially lower by LC MS / MS than immunoassay, presumably due to antibody crossreactivity with structurally similar metabolites in immunoassay but other contributing factors may also play a role (J Endocr Soc 2022;6:bvac049)
    • Clinicians should be aware of assay characteristics
      • As the current diagnostic cutoffs are from previous studies based on RIA and CLIA, there is a need to update clinical guidelines to reflect the differences between CLIA and modern LC MS / MS assays
• Different renin assays (PRA versus DRC)
  • PRA assay measures the concentration of angiotensin I produced
  • DRC assay directly measures renin mass (concentration)
  • For most patients, both assays give comparable information; however, renin activity and mass assays may give conflicting results in patients taking direct renin inhibitors and in patients with significantly high renin activity, high estrogen states (e.g., pregnancy) or congestive heart failure (Pract Lab Med 2021;25:e00229)

Which of the following is commonly associated with hyperaldosteronism?

1. Adrenal nodules
2. Easily managed hypertension
3. High plasma potassium
4. Hypertension after 50 years of age
5. Increased plasma sodium

A. Adrenal nodules. Bilateral (micronodular) adrenal hyperplasia is the most common cause of hyperaldosteronism. Answer B is incorrect because resistant hypertension is presented in hyperaldosteronism. Answer D is incorrect because early onset hypertension before 40 years old is a clinical indication of hyperaldosteronism. Answer C is incorrect because most patients with hyperaldosteronism present with normokalemic hypertension. Answer E is incorrect because plasma sodium tends to be normal due to the concurrent fluid retention.

Comment Here

Reference: Aldosterone

Which of the following factors can cause misinterpretation of aldosterone, renin or the aldosterone to renin ratio (ARR)?

1. Interferences with the testing methods
2. Patients without restriction on dietary salt intake
3. Sample collected and processed at room temperature, followed by rapid freezing
4. Upright position during blood collection
5. Use of selective serotonin reuptake inhibitors (SSRIs) prior to testing

A. Interferences with the testing methods. Assay specific interferences can cause erroneous results. Answer E is incorrect because SSRIs can affect metanephrines but are not known to affect aldosterone or renin. Answer B is incorrect because the endocrinology practice guideline does not recommend dietary salt restriction. Sodium restriction can significantly raise plasma renin activity (PRA), normalize the ARR and could cause falsely negative screening results (J Clin Endocrinol Metab 2016;101:3989). Answer D is incorrect because samples should be collected after the patient has been up (sitting, standing or walking) for a minimum of 2 hours. Answer C is incorrect because the sample should be collected and processed at room temperature and separated plasma should be rapidly frozen to prevent cryoactivation and false elevation of renin.

Comment Here

Reference: Aldosterone