Adrenal Cortical Hormones ENDO 412

Objectives of the Lecture 1- Identifying the general structure of the adrenal cortex. 2- Characterizing the chemical nature of the adrenal cortical hormones 3- Identifying the hypothalamic-pituitary-adrenal (HPA) axis. 4- Describing in brief the actions of glucocorticoids & mineralocorticoids. 5- Identifying and describing the regulation of actions of glucocorticoids & mineralocorticoids. 6- Listing causes of adrenocortical hyperfunction (Cushing`s syndrome). 8- Describing the biochemical & clinical concepts of adrenocortical hyperfunction. 9- Identifying and describing laboratory investigations for detection of suspected adrenocortical hyperfunction 10- Identifying the causes of adrenocortical hypofunction 11- Describing the biochemical & clinical concepts of Addison`s disease. 12- Identifying and describing investigations of suspected cases of Addison’s disease.

Structure of the adrenal cortex Outer: Zona Glomerulosa (produces mineralocorticoids: aldesterone) Middle: Zona Fasciculata (produces glucocorticoids : cortisol) Inner: Zona Reticularis (produces sex steroid hormones)

Synthesis of adrenal cortical hormones Cholesterol is the precursor of all classes of steroid hormones

Mechanism of action of adrenal cortical (steroid) hormones Adrenal Cortical (Steroid) hormones belong to group I hormones Cytosolic Receptors Hormone Receptor Complex Transcription of genes is increased HRE of genes

Glucocorticoids (as Cortisol) Action: on Metabolism Carbohydrates: increase of gluconeogenesis (in liver). Proteins: increase amino acids uptake by the liver for gluconeogenesis. So, increase proteolysis in skeletal muscles to give amino acids. Lipids: increase ketogenesis in liver increase lipolysis in adipose tissue

Glucocorticoids (as Cortisol) cont. Regulation 1- ACTH (adrenocorticotrophic hormone) of anterior pitutary: by feedback control by corticitrophin releasing hormone (CRH): increased cortisol secretion (or synthetic glucocorticoids) suppresses secretion of CRH. (HPA axis is the main regulation of cortisol secretion by adrenal cortex). 2- Stress induces sudden large increase in CRH that increases ACTH & cortisol. 3- Diurnal rhythm of plasma cortisol: Levels of cortisol in blood is highest at the start of day & lowest at sleep.

Mineralocorticoids (as Aldesterone) Action: Electrolyte balance (Na+ & K+) BY: Renin-Angiotensin System Angiotensinogen (in liver, inactive) Renin (synth. by kidney) Angiotensin I Angiotensin Converting Enzyme (ACE) Angiotensin II Aldesterone (from adrenal cortex) In tubules of kidney Decrease Na+ excretion Increase K+ excretions Hypernatremia hypokalemia Increase BP

Mineralocorticoids (as Aldesterone) cont. Regulation 1-Renin (of the kidney) increased in response to low blood volume or sodium loss. 2- Potassium (hyperkalemia) hyperkalemia (increase blood K+) stimulates release of aldesterone from adrenal cortex. 3- ACTH (ONLY IN STRESS)

Adrenal hyperfunction Hpercortisolism Endogenous i.e. over secretion of CRH, ACTH or glucocorticoids (cortisol) Or: Exogenous intake of cortisol (or ACTH) (in all these cases, blood cortisol is elevated) Cushing`s Syndrome describes a group of signs & symptoms resulting from excess glucocorticoids (cortisol) production or prolonged exogenous steroid use.

Causes of adrenal hyperfunction (Cushing`s syndrome) 1- ACTH dependent 1- ACTH secreting pituitary adenoma , 68% 2- Ectopic ACTH or ectopic CRH, 15% (usually malignant) 3- ACTH therapy (Iatrogenic Cushing`s Syndrome) 2- ACTH independent 1- Adrenal adenoma, 17% (ACTH is suppressed) 2- Glucocorticoids therapy

Biochemical & clinical concepts of Cushing`s syndrome Clinical Effects Biochemical Changes Hyperglycemia (may be DM) Increased gluconeogenesis Truncal obesity (Buffalo hump) Disturbed fat metabolism with redistribution Thinning of skin Wasting of muscles Osteoporosis of bones Severe catabolic effects on proteins Poor wound healing Reduced resistance to infection (low immunity) Suppressed immune response Hpernatremia (increased Na+ in blood) Hypokalemia (decreased K+ in blood) Alkalosis Hypertension Mineralocorticoid effects of cortisol

Laboratory investigations of adrenal hyperfunction (Cushing`s syndrome) Stage I: Diagnosis of Cushing`s syndrome Suspected cases by clinical examination are checked in Clinical Chemistry Laboratory for Cushing`s syndrome. 1- Screening tests 1- Cortisol excess 2- Loss of diurnal rhythm determination 3- Suppression resistance determination 2- If screening tests are positive, diagnosis is confirmed by confirmatory tests Insulin hyperglycemic test Stage II: If Cushing`s syndrome is confirmed, tests for determining the cause of Cushing`s syndrome (ACTH dependent or ACTH independent): 1- Plasma ACTH 2- CRH stimulation test (CRH-stimulated BIPSS & peripheral veins sampling)

Laboratory investigations of adrenal hyperfunction (Cushing`s syndrome) Clinical manifestations (symptoms & signs) Screening test Positive Result Confirmatory tests Positive result Plasma ACTH Positive Negative ACTH dependent ACTH independent

By: Urine free cortisol (and/or metaboites) Measurement Laboratory investigations of adrenal hyperfunction (Cushing`s syndrome) Screening test-1 Cortisol excess By: Urine free cortisol (and/or metaboites) Measurement Free cortisol (& metabolites) is excreted in urine if blood cortisol exceeds capacity of its carrier protein. Urine free cortisol (or metabolites) is a sensitive indicator of endogenous cortisolism. Advantage of urine free cortisol: It reflects free cortisol level during the period of urine collection. 17-hydroxycorticosteroid (metabolite of cortisol), is preferred as it is not affected by urine volume. (Other metabolites are secreted in higher amounts with increase urine volume). Urine collection period: 24 hours (or only 10 hours from 10 PM till 8 AM) NB: Random plasma cortisol measurement is of little value in diagnosis of Cushing`s syndrome as levels of normal people vary widely during the day & may overlap with levels found in patients of Cushing`s syndrome.

Laboratory investigations of adrenal hyperfunction (Cushing`s syndrome) Screening tests-2 By: Loss of diurnal rhythm determination Principle of the test: Normally, blood cortisol is at its highest levels between 6 - 8 AM & at its lowest levels between 10 PM – 12 AM (midnight). This variation is lost in Cushing`s syndrome (i.e. increased all over the day) Loss can be determined by measuring plasma cortisol 11 PM – 12 AM (midnight). This test is more sensitive than urine cortisol in diagnosing Cushing` syndrome. Or by: Saliva cortisol (instead of plasma cortisol) - Cortisol is stable at room temperature in saliva (easy storing of samples) - Non-invasive (no sampling by puncture etc…) - Patient can collect the samples by himself - Many samples can be collected over a defined period. BUT: less sensitive than urine cortisol

Laboratory investigations of adrenal hyperfunction (Cushing`s syndrome) Screening tests-3 Loss of normal cortisol suppression by dexamethazone By: Overnight dexamethazone suppression test Principle of the test Dexamethasone act as an exogenous cortisol substitute that suppresses endogenous cortisol secretion if adrenal cortex is normal (through suppressing ACTH if ant. pit. is normal) Procedure Dexamethazone 1 mg is given at 11 PM (should suppress early morning cortisol high secretion). Then, 8-9 AM: serum free cortisol is measured. Results: In normal individuals : cortisol is less than 3.6 mg/dl (cortisol is normally suppressed by dexameth.) In Cushing`s syndrome: cortisol level in blood is higher than 3.6 mg/dl. (cortisol secretion is not suppressed by dexamethazone in these cases). NB: Dexamethazone levels in blood is measured (for checking compliance of the patient).

Laboratory investigations of adrenal hyperfunction (Cushing`s syndrome) Interpretation of Screening Tests Positive results for screening test 1, 2 & 3 Cushing`s syndrome or Pseudo-Cushing`s syndrome Depression Extremely anxious patients Severe illness Alcoholism So, confirmatory tests should be performed to rule out pseudo-Cushing`s syndrome

Insulin hypoglycemic test Laboratory investigations of adrenal hyperfunction (Cushing`s syndrome) Confirmatory Test Insulin hypoglycemic test Principle: Hypoglycemia induces CRH that induces ACTH that induces cortisol secretion. i.e. normal HPA axis) In Cushing`s syndrome (for any cause), no response to hypoglycemia & accordingly no effect on CRH, ACTH or cortisol. Procedure (IN HOSPITAL UNDER RECAUSIONS) Insulin IV (0.15 U/kg) will reduce blood glucose to 2.2 mmol/l or less Normally, serum cortisol reaches its maximum 60-90 minutes after injection Blood samples for cortisol is withdrawn before injection & then 60 and 90 minutes after injection (together with blood glucose measurement) Results: Increase in blood cortisol in after-injection samples: Negative for Cushing`s syndrome. No difference between before & after samples: Positive for Cushing`s (defect in HPA axis ----verify??)

Determining the Cause of Cushing`s Syndrome Laboratory investigations of adrenal hyperfunction (Cushing`s syndrome) Determining the Cause of Cushing`s Syndrome Once Cushing`s syndrome is confirmed, cause is to be decided (i.e. ACTH dependent or ACTH independent) By: Plasma ACTH (At 8 AM & 10 PM) Undetectable Normal or Increased ACTH independent ACTH dependent Adrenal Cause Pituitary cause Ectopic ACTH (e.g. Adrenal tumor) to differentiate : CRH stimulated BIPSS & peripheral vein sampling BIPSS ACTH / peripheral ACTH > 3 in pituitary causes BIPSS ACTH / peripheral ACTH < 2.5 in ectopic causes (BIPSS = bilateral inferior petrosal sampling)

Adrenal insufficiency (Addison`s disease) Low cortisol result from: 1- Primary adrenal problem (destruction of 90% of adrenal cortex) mainly caused by autoimmune destruction of adrenal cortex (more than 70% of cases of adrenal insufficiency) 2- Secondary to ACTH deficiency (abnormal of HPA axis) Main clinical manifestation: 1- Weakness, fatigue, anorexia, weight loss (failure to thrive) 2- Hyponatremia, hyperkalemia & mild metabolic acidosis

Adrenal insufficiency (Addison`s disease) Diagnosis Low Base-line Cortisol (at 8 AM) Cosyntropin Test (Cortisol Stimulation Test) Normal response to stimulation No response to stimulation Secondary adrenal insufficiency ?? Primary adrenal insufficiency ?? LOW ACTH HIGH ACTH SECONDARY ADRENAL INSUFFICIENCY PRIMARY ADRENAL INSUFFICIENCY

Adrenal insufficiency (Addison`s disease) Cosyntropin test: Principle: Cosyntropin is a synthetic stimulator of cortisol secretion by adrenal cortex. Cosyntrpoin test checks the capacity of adrenal gland to increase of hormone production in response to stimulation by cosyntropin Procedure: 1- Base-line cortisol is measured 2- Then, cosyntropin is IV or IM administered 3- Cortisol is measured 30 & 60 minutes after cosyntropin administration. Results: Normally, cortisol secretion is increased after stimulation of the adrenal gland by cosyntropin. In primary adrenal insufficiency, cosyntropin fails to increase cortisol secretion by the adrenal cortex.