3.4 Endocrinology: Hormone Physiology and Endocrine Function Testing

Endocrinology: Hormone Physiology and Endocrine Function Testing

Endocrine testing is a small but reliable slice of the Clinical Chemistry board subject, which RA 5527 weights at 20% of the MTLE. Questions rarely ask for obscure hormones. They test whether you can read a feedback pattern (which gland failed first), pick the right screening or confirmatory test, and recognize the immunoassay artifacts that send a clinician the wrong way. This section covers hormone classes and assay pitfalls, the thyroid and adrenal cortex axes, the adrenal medulla, mineralocorticoids, parathyroid and posterior pituitary hormones, growth hormone and prolactin, and reproductive hormones including hCG.

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1. Hormone Classes, Feedback Logic, and Immunoassay Pitfalls

Three Chemical Classes

Reading a Feedback Loop

• Primary disease starts in the target gland. The target hormone and the pituitary trophic hormone move in opposite directions (low cortisol with high ACTH in Addison disease).
• Secondary disease starts in the pituitary, and tertiary disease in the hypothalamus. Both hormones move in the same direction (low cortisol with low ACTH).
• Rule for dynamic tests: suspect deficiency, stimulate (cosyntropin for adrenal insufficiency, insulin tolerance or arginine for GH deficiency); suspect excess, suppress (dexamethasone for Cushing syndrome, glucose load for acromegaly).

Immunoassay Pitfalls the Board Likes

• High-dose hook (prozone) effect: In one-step sandwich immunoassays, an extremely high antigen concentration saturates both capture and detection antibodies, so the signal is falsely low. Classic examples are very high hCG (molar pregnancy, choriocarcinoma) and giant prolactinomas. Re-assay after serial dilution.
• Heterophile antibodies and HAMA: Human antibodies against animal immunoglobulins bridge the capture and detection antibodies and create a false-positive signal. Suspect it when a result does not fit the clinical picture; blocking reagents or a different platform resolve it.
• Biotin interference: High-dose biotin supplements interfere with streptavidin-biotin assays. Sandwich assays (TSH, troponin, hCG) read falsely low, and competitive assays (free T4, free T3) read falsely high, which can mimic hyperthyroidism.
• Macroprolactin: Prolactin bound to IgG is biologically inactive but reacts in most assays. Polyethylene glycol (PEG) precipitation separates it from monomeric prolactin.

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2. Thyroid and Adrenal Cortex Axes

The Hypothalamic-Pituitary-Thyroid (HPT) Axis

                                  HYPOTHALAMUS
                                        |
                                   Thyrotropin-Releasing Hormone (TRH)
                                        |
                                        v
                              ANTERIOR PITUITARY
                                        |
                                   Thyroid-Stimulating Hormone (TSH)
                                        |
                                        v
                                  THYROID GLAND
                                        |
                             +----------+----------+
                             |                     |
                         Thyroxine (T4)     Triiodothyronine (T3)
                             |                     |
                             +----------+----------+
                                        |
                                        v
                         Negative Feedback on Pituitary & Hypothalamus

Thyroid hormones are synthesized from tyrosine residues on thyroglobulin within thyroid follicles, iodinated to form monoiodotyrosine (MIT) and diiodotyrosine (DIT), and coupled into thyroxine ($T_4$, 90%) and triiodothyronine ($T_3$, 10%). In circulation, >99.9% of $T_4$ and >99.7% of $T_3$ are bound to plasma carrier proteins (Thyroid-Binding Globulin [TBG], transthyretin, and albumin). Only the unbound free fractions (Free $T_4$, Free $T_3$) enter cells to exert metabolic actions and provide negative feedback inhibition on pituitary TSH secretion.

Diagnostic Interpretation of Thyroid Profiles

The Hypothalamic-Pituitary-Adrenal (HPA) Axis and Cortisol Dynamics

Cortisol is a glucocorticoid synthesized in the zona fasciculata of the adrenal cortex under the control of hypothalamic Corticotropin-Releasing Hormone (CRH) and anterior pituitary Adrenocorticotropic Hormone (ACTH).

• Circadian / Diurnal Rhythm: Cortisol exhibits a pronounced diurnal secretion pattern in healthy individuals. Peak circulating concentrations occur immediately after waking in the early morning (6:00 AM – 8:00 AM, reference $10-20\text{ }\mu\text{g/dL}$); concentrations fall steadily throughout the day, reaching an evening nadir at midnight (midnight trough, reference $<5\text{ }\mu\text{g/dL}$, <50% of morning peak). Loss of this normal diurnal variation is an early biochemical hallmark of Cushing syndrome.
• Cushing Syndrome (Hypercortisolism):
  • First-Line Screening Tests: (1) 24-hour urinary free cortisol (UFC), (2) late-night salivary cortisol (demonstrating loss of midnight nadir), or (3) low-dose ($1\text{ mg}$) overnight dexamethasone suppression test (DST). Dexamethasone is a potent synthetic glucocorticoid that normally suppresses ACTH and early morning cortisol to $<1.8\text{ }\mu\text{g/dL}$. Failure to suppress confirms autonomous hypercortisolism.
  • Etiological Differentiation: Plasma ACTH is measured:
    • ACTH-Dependent Cushing: High or normal ACTH. Caused by a pituitary ACTH-secreting microadenoma (Cushing Disease, $\approx 70\%$) or ectopic ACTH secretion (small cell lung carcinoma).
    • ACTH-Independent Cushing: Suppressed/undetectable ACTH. Caused by autonomous adrenal adenoma, adrenal carcinoma, or exogenous corticosteroid intake.
• Addison Disease (Primary Adrenal Insufficiency): Autoimmune destruction of all three zones of the adrenal cortex, causing deficiency of cortisol, aldosterone, and adrenal androgens.
  • Laboratory Profile: Inappropriately low baseline morning cortisol with markedly elevated ACTH (elevated ACTH processing generates melanocyte-stimulating hormone, producing cutaneous and mucosal hyperpigmentation). Electrolytes show hyponatremia, hyperkalemia, and mild non-anion gap metabolic acidosis due to lack of aldosterone.
  • Confirmatory Test: Cosyntropin (synthetic ACTH) stimulation test. Baseline cortisol is measured, $250\text{ }\mu\text{g}$ of cosyntropin is administered intramuscularly or intravenously, and cortisol is measured at 30 and 60 minutes. Healthy adrenals respond with a brisk rise to $>18-20\text{ }\mu\text{g/dL}$; in primary Addison disease, the atrophic adrenals show no response.

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3. Adrenal Medulla: Catecholamines and Their Metabolites

Chromaffin cells of the adrenal medulla convert tyrosine to dopamine, norepinephrine, and epinephrine. Catecholamines are metabolized by catechol-O-methyltransferase (COMT) to metanephrine and normetanephrine, and by monoamine oxidase (MAO) and COMT to the end product vanillylmandelic acid (VMA). Dopamine is metabolized to homovanillic acid (HVA).

• Pheochromocytoma and paraganglioma: Tumors cause paroxysmal hypertension, headache, sweating, and palpitations. Plasma free metanephrines or 24-hour urine fractionated metanephrines are the first-line screening tests because tumors metabolize catecholamines to metanephrines continuously, even between episodes. Urine VMA is less sensitive.
• Neuroblastoma: A childhood tumor of neural crest origin. Urine VMA and HVA, often measured on a random specimen and reported per gram of creatinine, are elevated in most cases.
• Specimen handling: Collect 24-hour urine with an acid preservative (commonly hydrochloric acid) to keep catecholamines stable. Strenuous exercise, stress, caffeine, and several drugs (such as tricyclic antidepressants and MAO inhibitors) raise results. Older colorimetric VMA methods also reacted with dietary vanilla, banana, and coffee compounds, which is why older texts list diet restrictions.

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4. Mineralocorticoids, Parathyroid Hormone, and Antidiuretic Hormone

Aldosterone and the Renin-Angiotensin System

• Aldosterone from the zona glomerulosa promotes sodium reabsorption and potassium and hydrogen ion secretion in the distal nephron.
• Primary aldosteronism (Conn syndrome): Hypertension, hypokalemia, and metabolic alkalosis with high aldosterone and suppressed renin. The screening test is the aldosterone-to-renin ratio (ARR).
• Secondary aldosteronism (renal artery stenosis, heart failure, cirrhosis): Renin and aldosterone are both high.

Parathyroid Hormone and Calcium

Antidiuretic Hormone (ADH, Vasopressin)

• SIADH: Hyponatremia with low serum osmolality, inappropriately concentrated urine, and urine sodium usually above 30 mmol/L in a euvolemic patient. Causes include small cell lung carcinoma, CNS disease, and drugs.
• Diabetes insipidus: Polyuria with dilute urine and high-normal or high serum osmolality. A water deprivation test followed by desmopressin separates central DI (urine concentrates after desmopressin) from nephrogenic DI (no response).

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5. Anterior Pituitary: Growth Hormone and Prolactin

• Growth hormone (GH) is secreted in pulses, so a random GH level is not useful. Insulin-like growth factor 1 (IGF-1) reflects average GH secretion and is the screening test for acromegaly (adults) and gigantism (before epiphyseal closure). Failure of GH to fall below about 1 µg/L after a 75-g oral glucose load confirms excess; laboratories using ultrasensitive assays apply lower cutoffs. GH deficiency is confirmed with stimulation tests such as the insulin tolerance, arginine, or glucagon test.
• Prolactin is the only anterior pituitary hormone under predominantly inhibitory (dopamine) control. Pregnancy, breastfeeding, stress, dopamine antagonists (antipsychotics, metoclopramide), and primary hypothyroidism (TRH stimulates prolactin) raise it. A prolactinoma causes galactorrhea, amenorrhea, and infertility in women and hypogonadism in men.

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6. Reproductive Hormones and Human Chorionic Gonadotropin (hCG)

hCG

• A glycoprotein made by the syncytiotrophoblast. Its alpha subunit is identical to that of LH, FSH, and TSH, so assays target the beta subunit for specificity.
• Serum hCG becomes detectable within about 8 to 11 days after conception. In an early viable intrauterine pregnancy, it classically doubles about every 48 to 72 hours, peaks at about 8 to 11 weeks of gestation, and then falls to a lower plateau.
• A slow rise or plateau suggests an ectopic or failing pregnancy. Very high levels suggest hydatidiform mole, choriocarcinoma, or multiple gestation. hCG is also a tumor marker for testicular germ cell tumors.
• For qualitative urine tests, a first-morning specimen is preferred because it is the most concentrated.

Ovarian, Testicular, and Gonadotropin Tests

• The LH surge triggers ovulation about 24 to 36 hours later. A mid-luteal (about day 21 of a 28-day cycle) progesterone level confirms that ovulation occurred.
• Menopause and primary ovarian failure show high FSH and LH with low estradiol. Hypogonadotropic hypogonadism shows low gonadotropins with low sex steroids.
• Testosterone has a diurnal rhythm, so draw it in the morning. Most circulating testosterone is bound to sex hormone-binding globulin (SHBG) and albumin.

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7. Endocrine Test Selection Summary