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🧬 COMPLETE BIOCHEMISTRY GUIDE: Hormones, Metabolism & Diabetes

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PART 1: STEROID HORMONE SYNTHESIS

What is a steroid hormone? Start from scratch.

Where are they made?

The Big Picture Pathway

CHOLESTEROL (C27)
      ↓  [RATE-LIMITING STEP: StAR protein moves cholesterol into mitochondria]
      ↓  [Enzyme: CYP11A1 = P450scc, in inner mitochondrial membrane]
PREGNENOLONE  ← This is the "mother of all steroids"
      ↓  [3β-HSD enzyme, in smooth ER]
PROGESTERONE
      ↓ splits into 3 pathways based on which zone of adrenal gland
  ┌───┴────────────────────────────┐
  ↓                                ↓                  ↓
GLUCOCORTICOIDS            MINERALOCORTICOIDS      ANDROGENS/ESTROGENS
(Zona Fasciculata)          (Zona Glomerulosa)      (Zona Reticularis)
CORTISOL                    ALDOSTERONE             DHEA → Testosterone → Estradiol

Step-by-Step: Cholesterol to Each Hormone

Step 1 - The Rate-Limiting Step

• StAR protein (Steroidogenic Acute Regulatory protein) physically shuttles cholesterol from the outer to the inner mitochondrial membrane
• This is the first thing ACTH triggers - no StAR activity = no steroid synthesis
• Think of StAR as the "gate opener" - without it, the raw material (cholesterol) can't enter the factory (mitochondria)

Step 2 - Cholesterol → Pregnenolone

• Enzyme: P450scc (CYP11A1) - "scc" = side chain cleavage
• It snips off a 6-carbon piece from cholesterol (C27) to make pregnenolone (C21)
• Needs: NADPH + Oxygen
• Location: Inner mitochondrial membrane

Step 3 - Pregnenolone → Progesterone

• Enzyme: 3β-HSD (3-beta hydroxysteroid dehydrogenase) + isomerase
• Converts the Δ5 compound (pregnenolone) into a Δ4 compound (progesterone)
• Location: Smooth endoplasmic reticulum

Now the 3 Pathways Diverge:

Pathway A - CORTISOL (Zona Fasciculata - stress hormone)

Progesterone
  ↓ CYP17A1 (17α-hydroxylase) - adds OH at C17
17α-Hydroxyprogesterone (17-OHP)
  ↓ CYP21A2 (21-hydroxylase) - adds OH at C21
11-Deoxycortisol
  ↓ CYP11B1 (11β-hydroxylase) - adds OH at C11, in mitochondria
CORTISOL ✓

• Important exam fact: CYP21A2 deficiency = most common CAH (Congenital Adrenal Hyperplasia). If you can't make cortisol/aldosterone, all precursors flood into the androgen pathway → virilization + salt-wasting crisis.

Pathway B - ALDOSTERONE (Zona Glomerulosa - salt hormone)

Progesterone
  ↓ CYP21A2 (21-hydroxylase)
11-Deoxycorticosterone (DOC)
  ↓ CYP11B1 (11β-hydroxylase)
Corticosterone
  ↓ CYP11B2 (Aldosterone synthase) ← ONLY exists in zona glomerulosa!
ALDOSTERONE ✓

• Key: Zona glomerulosa lacks CYP17A1 (17α-hydroxylase) so it can't make cortisol or androgens - it's wired only for aldosterone.

Pathway C - ANDROGENS/ESTROGENS (Zona Reticularis + Gonads)

17-Hydroxypregnenolone
  ↓ CYP17A1 (now acting as 17,20-lyase - second function)
DHEA (C19 - weak androgen)
  ↓ 3β-HSD
Androstenedione
  ↓ 17β-HSD (in gonads)
TESTOSTERONE
  ↓ CYP19A1 (Aromatase) in ovarian granulosa cells
ESTRADIOL (C18 - most potent estrogen)
  
OR
  ↓ 5α-Reductase in prostate/skin
DHT (Dihydrotestosterone - most potent androgen)

Carbon Number Rule (Easy Memory)

• C21 = Glucocorticoids (cortisol) + Mineralocorticoids (aldosterone) + Progestogens
• C19 = Androgens (testosterone, DHEA)
• C18 = Estrogens (estradiol) - have aromatic ring A

Regulation - Who Controls All This?

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PART 2: THYROID HORMONE SYNTHESIS

What are thyroid hormones?

• T4 (Thyroxine) = 4 iodine atoms, made exclusively in thyroid, is a prohormone (not very active)
• T3 (Triiodothyronine) = 3 iodine atoms, the active hormone, 80% comes from peripheral conversion of T4

The 6 Steps of Thyroid Hormone Synthesis

Where it happens

BLOOD (Iodide I⁻) → [STEP 1: TRAPPING] → Inside thyrocyte → [STEP 3: ORGANIFICATION]
→ COLLOID (Thyroglobulin + Iodine) → [STEP 4: COUPLING] → T4/T3 stored on Tg
→ [STEP 5: SECRETION] → Free T4/T3 released into blood
→ [STEP 6: PERIPHERAL CONVERSION] → T4 → T3 in liver/kidney

Step 1 - Iodide Trapping (Active Transport)

• Transporter: NIS (Sodium-Iodide Symporter) on the basolateral membrane (blood side)
• Brings 2 Na⁺ + 1 I⁻ into the cell together (symport)
• Concentrates iodide 20-40 times higher inside the cell than in blood
• Driven by the Na⁺ gradient (created by Na/K-ATPase)
• Pendrin transports I⁻ from cell into the colloid (apical membrane)
• Regulated by: TSH increases NIS activity; excess iodine inhibits it (Wolff-Chaikoff effect)

Step 2 - Thyroglobulin Synthesis

• Thyroglobulin (660 kDa glycoprotein) made in rough ER of thyrocyte
• Processed in Golgi, then secreted by exocytosis into the colloid
• Contains 123 tyrosine residues, but only 4-8 will actually become thyroid hormones
• Think of Tg as a "tyrosine delivery board" waiting to be iodinated

Step 3 - Organification of Iodide

• Enzyme: TPO (Thyroid Peroxidase) at the apical membrane (colloid side)
• TPO uses H₂O₂ (made by DUOX2 enzyme) to oxidize I⁻ → reactive iodine (I⁰)
• Reactive iodine attaches to tyrosine residues on Tg:
  • One iodine added → MIT (Monoiodotyrosine)
  • Two iodines added → DIT (Diiodotyrosine)
• Blocked by: Propylthiouracil (PTU), Methimazole (anti-thyroid drugs)

Step 4 - Coupling (Making T3 and T4)

• Same enzyme: TPO catalyzes coupling
• Two iodinated tyrosines join together on the Tg scaffold:
  • DIT + DIT → T4 (thyroxine, 4 iodines) - most abundant
  • MIT + DIT → T3 (3,5,3′-triiodothyronine, 3 iodines) - active form
  • DIT + MIT → rT3 (reverse T3, biologically inactive)
• This all happens while tyrosines are still attached to thyroglobulin

Step 5 - Secretion

1. TSH stimulates thyrocytes to endocytose (eat) the colloid
2. Colloid vesicles fuse with lysosomes → proteases digest thyroglobulin
3. Free T4 and T3 are released and diffuse into the bloodstream
4. MIT and DIT are NOT secreted - iodotyrosine deiodinase strips their iodine off and recycles it back into new hormone synthesis

Step 6 - Peripheral Conversion (T4 → T3)

• T4 is a prohormone; it must be converted to T3 to be active
• 80% of circulating T3 comes from this conversion in liver, kidney, muscle
• Deiodinases contain selenocysteine (selenium):

How Thyroid Hormones Work (Receptor Mechanism)

• T3 enters the cell → binds nuclear thyroid hormone receptor (TR)
• TR-T3 complex binds to Thyroid Hormone Response Elements (TREs) in DNA
• Results: Upregulates genes for metabolic enzymes, Na/K-ATPase, cardiac proteins, growth hormones

Transport in Blood

• TBG (Thyroxine-binding globulin) - binds 67% of T4, 46% of T3
• Transthyretin - binds 20% of T4
• Albumin - binds 13% of T4, 53% of T3
• Only the free fraction (0.02% of T4, 0.2% of T3) is biologically active

HPT Axis (Regulation)

Hypothalamus: TRH (tripeptide)
      ↓
Anterior Pituitary (thyrotrophs): TSH (glycoprotein, α+β subunits)
      ↓
Thyroid Gland: T4 (mainly) + small T3
      ↑ Negative feedback (free T3/T4 suppress both TRH and TSH)

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PART 3: THYROID FUNCTION TESTS

Why do we test thyroid function?

The Tests - One by One

1. TSH (Thyroid-Stimulating Hormone)

• Normal range: 0.35-5.0 mIU/L
• The most sensitive test for thyroid dysfunction
• HIGH TSH = thyroid is failing = the pituitary is screaming louder = Primary Hypothyroidism
• LOW TSH = thyroid is overactive = the pituitary is quiet = Primary Hyperthyroidism
• Exception: If the pituitary itself is diseased (central hypothyroidism), TSH may be low despite low T4 - always check TSH + FT4 together if you suspect pituitary disease

2. Free T4 (FT4)

• Normal range: 12-28 pmol/L (0.8-2.4 ng/dL)
• Measures only the unbound, biologically active T4
• More reliable than Total T4 because it's not affected by binding protein changes (e.g., pregnancy raises TBG → raises Total T4 but Free T4 stays normal)

3. Free T3 (FT3)

• Normal range: 3-9 pmol/L
• Used when: T3 toxicosis suspected (patient is hyperthyroid with normal FT4 but elevated FT3)

4. Total T4 and Total T3

• Measures bound + free hormone
• Less useful because binding protein changes confuse the results
• Estrogen (pregnancy, OCP) → ↑ TBG → ↑ Total T4/T3 (but patient is euthyroid, FT4 normal!)
• Androgens, glucocorticoids → ↓ TBG → ↓ Total T4/T3 (but patient is still euthyroid)

5. Reverse T3 (rT3)

• Elevated rT3 + Low T3 + Normal T4 = Euthyroid Sick Syndrome (nonthyroidal illness)
• In critical illness, starvation, major surgery: the body shunts T4 away from T3 (active) toward rT3 (inactive) to conserve energy

6. TRH Stimulation Test

• Inject 500 μg TRH IV → measure TSH at 0, 30, 60 min
• Normal: TSH rises by ≥6 mIU/L at 30 min
• Blunted response: primary hyperthyroidism OR pituitary disease
• Exaggerated response: primary hypothyroidism
• Largely replaced by modern sensitive TSH assays

7. Thyroid Antibodies

• Anti-TPO (aTPO): Positive in Hashimoto's (>80%) and Graves' disease
• TSI (Thyroid-Stimulating Immunoglobulin): Diagnostic for Graves' disease - these antibodies mimic TSH and stimulate the gland continuously
• Anti-thyroglobulin (ATG): Hashimoto's; also interferes with Tg measurements used in thyroid cancer monitoring

Interpretation Table (Memorize This)

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PART 4: PTH AND CALCIUM METABOLISM

Why is calcium so important?

• Muscle contraction (including heart muscle)
• Nerve impulse transmission
• Blood clotting (coagulation cascade)
• Cell signaling (second messenger)
• Enzyme activation

Three Organs Control Calcium: Bone, Kidney, Intestine

1. PTH (Parathyroid Hormone) - raises calcium
2. Calcitriol (1,25-(OH)₂-Vitamin D) - raises calcium
3. Calcitonin - lowers calcium (minor role in adults)

PTH: The Emergency Calcium Raiser

Structure

• PTH is an 84 amino acid polypeptide made by chief cells of the 4 parathyroid glands (tiny glands behind the thyroid)
• The biologically active portion is the first 34 amino acids (N-terminal)

What triggers PTH release?

• Low blood Ca²⁺ → chief cells detect this via CaSR (Calcium-Sensing Receptor) on their surface
• Low Ca²⁺ → CaSR inactive → PTH released
• High Ca²⁺ → CaSR activated → PTH suppressed

PTH's 3 Target Actions

PTH Mechanism of Action

• PTH receptor (PTH1R) is a GPCR (G-protein coupled receptor)
• Binds PTH → activates Gs → cAMP ↑ → PKA activation
• Also activates Gq → PLC → IP₃ → Ca²⁺ release + PKC activation

Vitamin D - The Intestinal Calcium Hero

Activation Pathway

Skin: UV light + 7-dehydrocholesterol → Cholecalciferol (Vitamin D3)
  ↓ Liver: 25-hydroxylase
25-hydroxyvitamin D (Calcidiol) ← this is what you measure in blood tests!
  ↓ Kidney: 1α-hydroxylase (activated by PTH, low PO₄, low Ca)
1,25-dihydroxyvitamin D (Calcitriol) ← the ACTIVE form

Actions of Calcitriol

• Intestine: Increases synthesis of calbindin-D (calcium transport protein) and TRPV6 channels → increases Ca²⁺ absorption (up to 80% of dietary calcium!)
• Bone: At normal levels, promotes bone mineralization; at high levels, works with PTH to release Ca from bone
• Kidney: Mild increase in Ca²⁺ reabsorption
• Parathyroid gland: Suppresses PTH synthesis (negative feedback)

Calcitonin - The Bone Protector

• Made by C-cells (parafollicular cells) of the thyroid gland
• Released when Ca²⁺ is HIGH
• Actions: Inhibits osteoclasts → reduces bone resorption; increases renal Ca excretion
• Clinical note: Calcitonin's role in adult humans is minor; it's more important in growing children and pregnant/lactating women. Used therapeutically in Paget's disease and hypercalcemia.

Calcium and Phosphate - The Inverse Relationship

• In hyperparathyroidism: Ca ↑, PO₄ ↓
• In hypoparathyroidism: Ca ↓, PO₄ ↑
• In Vitamin D toxicity: Both Ca AND PO₄ rise (because Vit D raises both)
• In Chronic Kidney Disease: Kidneys can't make calcitriol + can't excrete PO₄ → PO₄ ↑, Ca ↓ → Secondary hyperparathyroidism

Phosphate Regulation Summary

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PART 5: INSULIN - STRUCTURE AND RECEPTOR

What is insulin? The basics.

• α cells (25%) - make Glucagon
• β cells (60%) - make Insulin
• δ cells (10%) - make Somatostatin
• PP cells (5%) - make Pancreatic polypeptide

Insulin Structure - Built Like a Two-Strand Rope

• Molecular weight: 5,808 Da
• Two chains: A chain (21 amino acids) + B chain (30 amino acids)
• Connected by: 2 disulfide bonds (between chains) + 1 within the A chain
• If you split the two chains apart, insulin completely loses its activity

How Insulin is Made - The Biosynthesis Pathway

Gene on chromosome 11
      ↓ Transcription + Translation on ribosomes (RER)
PREPROINSULIN (110 amino acids, MW ~11,500)
      ↓ Signal peptide cleaved in ER lumen
PROINSULIN (86 amino acids, MW ~9,000) = A chain + B chain + C peptide
      ↓ Packaged in Golgi vesicles
      ↓ Cleavage by prohormone convertases (PC1 and PC2) + CPE
INSULIN (A+B chains, MW 5,808) + C PEPTIDE (31 amino acids)
      ↓ Both secreted together in equimolar amounts from secretory granules

What is C-Peptide? (Role of C-Peptide)

1. Diagnostic use: Injected insulin has no C-peptide. So if a diabetic patient has high insulin but low C-peptide, they're injecting insulin (factitious hypoglycemia or insulin misuse). If insulin AND C-peptide are both high, the body is making too much insulin (insulinoma).
2. Beta cell function marker: C-peptide measurement tells you how much insulin the pancreas is making. Because C-peptide has a longer half-life (~35 min vs. ~5 min for insulin), it's easier to measure.
3. Biological activity: C-peptide binds a G-protein-coupled receptor and activates Na/K-ATPase and endothelial nitric oxide synthase (eNOS) → may protect against diabetic nephropathy and neuropathy. This biological role is still being studied.

Insulin Secretion - How Does Glucose Trigger It?

Glucose enters β-cell via GLUT2 (passive)
      ↓
Glucokinase (Hexokinase IV) phosphorylates glucose → Glucose-6-phosphate
      ↓
Glycolysis + TCA cycle → ATP production
      ↓
↑ ATP/ADP ratio
      ↓
ATP-sensitive K⁺ channel (KATP channel) CLOSES
      ↓
β-cell membrane DEPOLARIZES (K⁺ can't leave)
      ↓
Voltage-gated Ca²⁺ channels OPEN
      ↓
Ca²⁺ floods into β-cell
      ↓
Insulin granules FUSE with membrane → EXOCYTOSIS of insulin + C-peptide

• Phase 1 (First phase, 3-5 min): Rapid dump of pre-stored insulin granules (immediate response to glucose rise)
• Phase 2 (15 min to 2 hrs): Slower, sustained secretion from newly synthesized + stored insulin

The Insulin Receptor - How Insulin Acts

Structure

• Heterotetramer: 2α subunits (extracellular, bind insulin) + 2β subunits (transmembrane + intracellular kinase domain)
• Connected by disulfide bonds

Signal Transduction Cascade

Insulin binds to α subunit
      ↓
Conformational change activates β subunit tyrosine kinase
      ↓
β subunit AUTOPHOSPHORYLATES (phosphorylates itself on tyrosine residues)
      ↓
Phosphorylates IRS-1/IRS-2 (Insulin Receptor Substrate proteins)
      ↓
       ┌─────────────────────────────┐
       ↓                             ↓
PI3K → PIP3 → PDK1 → Akt          Grb2/SOS → Ras → Raf → MEK → ERK
(Metabolic effects:                 (Mitogenic effects:
 GLUT4 translocation,                cell growth,
 glycogen synthesis,                 gene expression,
 lipogenesis,                        differentiation)
 protein synthesis)

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PART 6: GLUCAGON

What is glucagon? The opposite of insulin.

• Insulin = fed state hormone (glucose ↑ → insulin ↑ → store energy)
• Glucagon = fasting state hormone (glucose ↓ → glucagon ↑ → release energy)

Glucagon Structure

• 29 amino acid polypeptide
• Made from proglucagon (158 amino acids) gene
• Same proglucagon gene, but processed differently in intestinal L-cells → makes GLP-1 (glucagon-like peptide 1) instead

What Stimulates Glucagon?

Glucagon Receptor and Mechanism

Glucagon binds glucagon receptor on hepatocyte
      ↓
Gαs protein activates Adenylyl Cyclase
      ↓
ATP → cAMP ↑↑
      ↓
Protein Kinase A (PKA) activated
      ↓ (phosphorylates multiple enzymes simultaneously)
     ┌────────────────────────────────────────────┐
     ↓                                            ↓
GLYCOGENOLYSIS                             GLUCONEOGENESIS
(glycogen phosphorylase activated;         (PEP carboxykinase,
 phosphofructokinase-2 inhibited)          fructose-1,6-bisphosphatase activated)
     ↓                                            ↓
Glucose released from glycogen          New glucose made from
into blood                              amino acids, lactate, glycerol

• Lipolysis: Activates hormone-sensitive lipase → releases fatty acids from adipose
• Ketogenesis: Promotes fatty acid oxidation → acetyl-CoA → ketone bodies in liver
• Protein catabolism: Promotes amino acid release from muscle for gluconeogenesis

Glucagon Metabolism (How is it broken down?)

• Half-life: 3-6 minutes
• Degraded primarily by: liver (first pass, ~60% removed) and kidneys
• Mechanism: Proteolytic degradation by liver proteases and renal clearance
• The short half-life means glucagon levels change very quickly in response to blood glucose

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PART 7: SOMATOSTATIN

What is somatostatin? The "off switch."

• δ (delta) cells of the pancreatic islets
• D cells of the stomach and intestine
• Hypothalamus (inhibits growth hormone from pituitary)

Structure

• Two forms: SS-14 (14 amino acids) and SS-28 (28 amino acids)
• Both are cyclic peptides (amino and carboxy termini linked by a disulfide bridge)
• SS-14 predominates in pancreas; SS-28 predominates in intestine

What Triggers Somatostatin Release?

• High glucose, amino acids, fatty acids after a meal
• Other GI hormones (CCK, secretin)
• High glucagon or insulin (local paracrine effect)

Somatostatin's Effects - "SOMATOSTATIN Stops Everything"

Mechanism of Action

• Somatostatin receptors (SSTR1-5) are GPCRs coupled to Gi (inhibitory G-protein)
• Gi → inhibits adenylyl cyclase → cAMP ↓ → PKA activity ↓
• Also: Opens K⁺ channels → hyperpolarization → inhibits Ca²⁺ entry → inhibits exocytosis
• In α and β cells: the reduced cAMP + hyperpolarization directly suppresses hormone granule release

Somatostatin Metabolism

• Half-life: extremely short, ~1-3 minutes in circulation
• Degraded by peptidases in the blood and tissues
• This is why octreotide (a synthetic analog with half-life 1.5 hours) and lanreotide (long-acting, monthly injection) were developed for clinical use
• Clinical uses: acromegaly (excess GH), carcinoid tumors, VIPomas, glucagonomas, bleeding esophageal varices

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PART 8: GLUT TRANSPORTERS

What are GLUT transporters?

The Key GLUT Transporters - A Complete Table

The Magic of GLUT4 - Why Insulin Matters

• GLUT4 vesicles sit in the cytoplasm, docked to intracellular membranes
• Very few GLUT4 molecules on the cell surface
• Little glucose enters muscle/fat cells

Insulin → IR → IRS-1 → PI3K → PIP3 → Akt → phosphorylates AS160
      ↓
TBC1D4 (AS160) is inactivated
      ↓
Rab-GTPase proteins activated
      ↓
GLUT4 vesicles traffic to plasma membrane
      ↓
GLUT4 fuses with membrane (exocytosis)
      ↓
GLUT4 on surface increases 10-20 fold
      ↓
Glucose floods into muscle/fat cell

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PART 9: LABORATORY DIAGNOSIS OF DIABETES MELLITUS

ADA Diagnostic Criteria (any ONE of these criteria on TWO separate days):

Oral Glucose Tolerance Test (OGTT) - In Detail

Procedure

1. Preparation: Patient fasts for 8-14 hours overnight; no heavy exercise 3 days prior; normal carbohydrate diet (≥150g/day) for 3 days prior
2. Fasting blood sample taken
3. Patient drinks 75g anhydrous glucose dissolved in 250-300 mL water (within 5 minutes)
4. Blood samples taken at 1 hour and 2 hours
5. Patient sits quietly, no smoking during test

Interpretation

Why is OGTT used?

• Better than FPG for detecting early Type 2 DM
• Essential for diagnosing Gestational Diabetes Mellitus (GDM)
• Reveals glucose regulation under physiological stress (a real meal)

• 2-step approach (USA): 50g glucose screen at 24-28 weeks → if ≥130-140 mg/dL at 1hr → do 100g 3-hr OGTT
• 1-step approach (international): 75g OGTT directly; GDM if fasting ≥92, 1-hr ≥180, 2-hr ≥153 mg/dL

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PART 10: HbA1c (GLYCATED HEMOGLOBIN)

What is HbA1c?

High blood glucose + Hemoglobin A
      ↓ (non-enzymatic, irreversible, SLOW process)
Schiff base (unstable)
      ↓ Amadori rearrangement
HbA1c (stable ketoamine)

Why does HbA1c reflect 3-month glucose control?

• RBCs (red blood cells) live for approximately 120 days (3 months)
• As the RBC is born, it has zero HbA1c
• Every day it lives, more glucose attaches to hemoglobin
• The amount of HbA1c at the end reflects the average glucose the RBC has been exposed to over its entire lifetime
• HbA1c is a "memory" of blood glucose for the past 2-3 months
• It reflects the last 30 days more heavily than earlier months (because younger RBCs are more abundant)

Reference Values

How HbA1c Correlates to Average Blood Glucose

Conditions That Falsely Alter HbA1c

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PART 11: METABOLISM OF INSULIN, GLUCAGON, AND SOMATOSTATIN

Insulin Metabolism

Secretion

• Normal fasting insulin: 5-15 μU/mL (35-105 pmol/L)
• After a meal: peaks at 50-100 μU/mL
• Daily secretion: ~40-50 units/day

Half-Life

• ~5-8 minutes in circulation (very short!)

Degradation

1. Liver - most important site (~50% of portal insulin removed in FIRST PASS through liver before it even reaches systemic circulation)
2. Kidneys - second most important (~30-40% of peripheral insulin)
3. Muscle, adipose - minor

Mechanism of Degradation

• Insulin-degrading enzyme (IDE) = insulysin - a zinc metalloprotease
• IDE degrades insulin in the cytosol after receptor-mediated endocytosis
• Also, insulin-receptor complex is internalized → insulin degraded in lysosomes → receptor recycled to surface

Why does this matter clinically?

• Kidney disease → insulin half-life prolonged → LESS insulin needed (risk of hypoglycemia)
• Liver disease → reduced first-pass extraction → HIGHER systemic insulin levels

Glucagon Metabolism

Secretion

• Normal fasting glucagon: 75-150 pg/mL
• After protein meal: rises significantly
• During hypoglycemia: rises markedly (key counter-regulatory response)

Half-Life

• 3-6 minutes (even shorter than insulin)

Degradation

• Liver is the primary site (60% removed in first pass)
• Kidneys are secondary
• Mechanism: Proteolytic degradation by liver enzymes + renal clearance
• This rapid degradation means glucagon works quickly but briefly - perfect for the acute hypoglycemia response

Somatostatin Metabolism

Secretion

• SS-14 predominates in pancreatic δ cells
• SS-28 predominates in intestinal D cells and hypothalamus

Half-Life

• Remarkably short: 1-3 minutes
• This is why it mainly acts locally (paracrine) rather than as a true circulating hormone

Degradation

• Peptidases in plasma (mainly endopeptidases)
• Liver and kidney clearance
• Clinical consequence: The natural hormone is useless therapeutically → synthetic analogs needed:
  • Octreotide (t½ ~1.5 hours, SubQ injection 3x/day)
  • Lanreotide (depot injection, monthly)
  • Pasireotide (broader SSTR subtype binding, used in Cushing's)

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QUICK REFERENCE SUMMARY TABLE

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Key Concepts to Always Remember

1. All steroid hormones come from cholesterol; the rate-limiting step is StAR protein moving cholesterol into mitochondria.
2. Thyroid hormone synthesis needs iodine + TPO enzyme; T4 is a prohormone, T3 is active.
3. TSH is the most sensitive thyroid test; low TSH = overactive gland, high TSH = underactive gland.
4. PTH raises calcium AND lowers phosphate; Vitamin D raises both.
5. Insulin is made as preproinsulin → proinsulin → insulin + C-peptide (released in equal amounts).
6. GLUT2 is the glucose sensor in β cells (low affinity, high capacity); GLUT4 is insulin-regulated in muscle/fat.
7. C-peptide helps distinguish endogenous insulin production from exogenous injection.
8. HbA1c reflects average blood glucose over 120 days (RBC lifespan); ≥6.5% = diabetes.
9. Glucagon → GPCR → cAMP → PKA → glycogenolysis + gluconeogenesis.
10. Somatostatin = master inhibitor of both insulin and glucagon via Gi → ↓cAMP pathway; half-life 1-3 minutes.

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What's inside (20 slides, wide 16:9 format)

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Presentation Overview (20 slides, wide 16:9)

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🧬 COMPLETE BIOCHEMISTRY GUIDE: Hormones, Metabolism & Diabetes

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PART 1: STEROID HORMONE SYNTHESIS

What is a steroid hormone?

Where are they made?

The Big Picture Pathway

CHOLESTEROL (C27)
      ↓  [RATE-LIMITING STEP: StAR protein moves cholesterol into mitochondria]
      ↓  [Enzyme: CYP11A1 = P450scc, inner mitochondrial membrane]
PREGNENOLONE  ← "Mother of all steroids"
      ↓  [3β-HSD enzyme, smooth ER]
PROGESTERONE
      ↓ splits into 3 pathways based on adrenal zone
  ┌──────────────────┬────────────────────┬──────────────────────┐
  ↓                  ↓                    ↓
GLUCOCORTICOIDS   MINERALOCORTICOIDS   ANDROGENS → ESTROGENS
(Zona Fasciculata) (Zona Glomerulosa)  (Zona Reticularis)
CORTISOL           ALDOSTERONE         DHEA → Testosterone → Estradiol

Step-by-Step Enzymatic Pathway

Step 1 - The Rate-Limiting Step

• StAR protein (Steroidogenic Acute Regulatory protein) shuttles cholesterol from the outer to the inner mitochondrial membrane
• ACTH triggers StAR immediately - no StAR = no steroid synthesis
• Think of StAR as the "gate opener"

Step 2 - Cholesterol → Pregnenolone

• Enzyme: P450scc (CYP11A1) - "scc" = side chain cleavage
• Snips off a 6-carbon piece from cholesterol (C27) → pregnenolone (C21)
• Needs NADPH + Oxygen
• Location: Inner mitochondrial membrane

Step 3 - Pregnenolone → Progesterone

• Enzyme: 3β-HSD + isomerase
• Converts Δ5 compound (pregnenolone) into Δ4 compound (progesterone)
• Location: Smooth ER

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Pathway A - CORTISOL (Zona Fasciculata)

Progesterone
  ↓ CYP17A1 (17α-hydroxylase) → adds OH at C17
17α-Hydroxyprogesterone (17-OHP)
  ↓ CYP21A2 (21-hydroxylase) → adds OH at C21
11-Deoxycortisol
  ↓ CYP11B1 (11β-hydroxylase) → adds OH at C11, in mitochondria
CORTISOL ✓

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Pathway B - ALDOSTERONE (Zona Glomerulosa)

Progesterone
  ↓ CYP21A2 (21-hydroxylase)
11-Deoxycorticosterone (DOC)
  ↓ CYP11B1 (11β-hydroxylase)
Corticosterone
  ↓ CYP11B2 (Aldosterone synthase) ← ONLY in zona glomerulosa!
ALDOSTERONE ✓

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Pathway C - ANDROGENS & ESTROGENS

17-Hydroxypregnenolone
  ↓ CYP17A1 (17,20-lyase activity)
DHEA (C19 - weak androgen)
  ↓ 3β-HSD
Androstenedione
  ↓ 17β-HSD
TESTOSTERONE (C19)
  ↓ CYP19A1 (Aromatase) in ovarian granulosa cells
ESTRADIOL (C18 - most potent estrogen)

OR
  ↓ 5α-Reductase in prostate/skin
DHT (most potent androgen, 5× more potent than testosterone)

Carbon Number Rule

• C21 = Glucocorticoids (cortisol) + Mineralocorticoids (aldosterone) + Progestogens
• C19 = Androgens (testosterone, DHEA)
• C18 = Estrogens (estradiol) - have aromatic ring A

Key Enzyme Summary Table

Regulation

CAH - Enzyme Deficiency Clinical Table

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PART 2: THYROID HORMONE SYNTHESIS

What are thyroid hormones?

• T4 (Thyroxine) = 4 iodine atoms, made exclusively in thyroid, is a prohormone (not very active)
• T3 (Triiodothyronine) = 3 iodine atoms, the active hormone, 80% comes from peripheral conversion of T4

Where it happens

The 6 Steps of Thyroid Hormone Synthesis

Step 1 - Iodide Trapping (Active Transport)

• Transporter: NIS (Sodium-Iodide Symporter) on the basolateral membrane (blood side)
• Brings 2 Na⁺ + 1 I⁻ into the cell together (symport)
• Concentrates iodide 20-40 times higher inside the cell than in blood
• Driven by the Na⁺ gradient (created by Na/K-ATPase)
• Pendrin transports I⁻ from cell into the colloid (apical membrane)
• Regulated by TSH (increases NIS activity); excess iodine inhibits it (Wolff-Chaikoff effect)

Step 2 - Thyroglobulin Synthesis

• Thyroglobulin (660 kDa glycoprotein) made in rough ER of thyrocyte
• Processed in Golgi, secreted by exocytosis into the colloid
• Contains 123 tyrosine residues, but only 4-8 actually become thyroid hormones
• Think of Tg as a "tyrosine delivery board" waiting to be iodinated

Step 3 - Organification of Iodide

• Enzyme: TPO (Thyroid Peroxidase) at the apical membrane (colloid side)
• TPO uses H₂O₂ (made by DUOX2 enzyme) to oxidize I⁻ → reactive iodine (I⁰)
• Reactive iodine attaches to tyrosine residues on Tg:
  • One iodine added → MIT (Monoiodotyrosine)
  • Two iodines added → DIT (Diiodotyrosine)
• Blocked by: PTU, Methimazole (antithyroid drugs)

Step 4 - Coupling (Making T3 and T4)

• Same enzyme: TPO catalyzes coupling
• Two iodinated tyrosines join together on the Tg scaffold:
  • DIT + DIT → T4 (thyroxine, 4 iodines) - most abundant
  • MIT + DIT → T3 (3,5,3′-triiodothyronine, 3 iodines) - active form
  • DIT + MIT → rT3 (reverse T3, biologically inactive)

Step 5 - Secretion

1. TSH stimulates thyrocytes to endocytose (eat) the colloid
2. Colloid vesicles fuse with lysosomes → proteases digest thyroglobulin
3. Free T4 and T3 released and diffuse into the bloodstream
4. MIT and DIT are NOT secreted - iodotyrosine deiodinase strips their iodine off and recycles it

Step 6 - Peripheral Conversion (T4 → T3)

• T4 is a prohormone; must be converted to T3 to be active
• 80% of circulating T3 comes from this conversion in liver, kidney, muscle
• Deiodinases contain selenocysteine (selenium):

Transport in Blood

Half-Lives

HPT Axis (Regulation)

Hypothalamus → TRH (tripeptide)
      ↓
Anterior Pituitary (thyrotrophs) → TSH (glycoprotein, α+β subunits)
      ↓
Thyroid Gland → T4 (mainly) + small T3
      ↑ Negative feedback (free T3/T4 suppress both TRH and TSH)

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PART 3: THYROID FUNCTION TESTS

The Tests - One by One

1. TSH (Thyroid-Stimulating Hormone)

• Normal range: 0.35-5.0 mIU/L
• The most sensitive test for thyroid dysfunction
• HIGH TSH = thyroid is failing = Primary Hypothyroidism
• LOW TSH = thyroid is overactive = Primary Hyperthyroidism
• Exception: If pituitary is diseased (central hypothyroidism), TSH may be low despite low T4

2. Free T4 (FT4)

• Normal range: 12-28 pmol/L (0.8-2.4 ng/dL)
• Measures only the unbound, biologically active T4
• More reliable than Total T4 because not affected by binding protein changes

3. Free T3 (FT3)

• Normal range: 3-9 pmol/L
• Used when: T3 toxicosis suspected (hyperthyroid with normal FT4 but elevated FT3)

4. Total T4 and Total T3

• Measures bound + free hormone
• Less useful because binding protein changes confuse results
• Estrogen (pregnancy, OCP) → ↑TBG → ↑Total T4/T3 (but patient is euthyroid, FT4 normal!)
• Androgens, glucocorticoids → ↓TBG → ↓Total T4/T3 (still euthyroid)

5. Reverse T3 (rT3)

• Elevated rT3 + Low T3 + Normal T4 = Euthyroid Sick Syndrome (nonthyroidal illness)
• In critical illness, starvation, major surgery: body shunts T4 → rT3 (inactive) to conserve energy

6. TRH Stimulation Test

• Inject 500 μg TRH IV → measure TSH at 0, 30, 60 min
• Normal: TSH rises by ≥6 mIU/L at 30 min
• Blunted response: Primary hyperthyroidism OR pituitary disease
• Exaggerated response: Primary hypothyroidism
• Largely replaced by modern sensitive TSH assays

7. Thyroid Antibodies

• Anti-TPO (aTPO): Positive in Hashimoto's (>80%) and Graves'
• TSI (Thyroid-Stimulating Immunoglobulin): Diagnostic for Graves' disease (~90% sensitivity)
• Anti-thyroglobulin (ATG): Hashimoto's

Complete Interpretation Table

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PART 4: PTH AND CALCIUM METABOLISM

Why is calcium important?

• Muscle contraction (including heart muscle)
• Nerve impulse transmission
• Blood clotting (coagulation cascade)
• Cell signaling (second messenger)
• Enzyme activation

Three Hormones Control Calcium

1. PTH (Parathyroid Hormone) - raises calcium
2. Calcitriol (1,25-(OH)₂-Vitamin D) - raises calcium
3. Calcitonin - lowers calcium (minor role in adults)

PTH Structure and Trigger

• 84 amino acid polypeptide made by chief cells of the 4 parathyroid glands
• Biologically active portion = first 34 amino acids (N-terminal)
• CaSR (Calcium-Sensing Receptor) on chief cells detects blood calcium:
  • Low Ca²⁺ → CaSR inactive → PTH released
  • High Ca²⁺ → CaSR activated → PTH suppressed

PTH's 3 Target Actions

PTH Mechanism of Action

• PTH receptor (PTH1R) is a GPCR
• PTH → Gαs → Adenylyl cyclase → cAMP ↑ → PKA activation
• Also activates Gq → PLC → IP₃ → Ca²⁺ release + PKC

Vitamin D Activation Pathway

Skin: UV light + 7-dehydrocholesterol → Cholecalciferol (Vitamin D3)
  ↓ Liver: 25-hydroxylase
25-hydroxyvitamin D (Calcidiol) ← what you measure in blood tests!
  ↓ Kidney: 1α-hydroxylase (activated by PTH, low PO₄, low Ca)
1,25-dihydroxyvitamin D (Calcitriol) ← the ACTIVE form

Actions of Calcitriol

• Intestine: Increases calbindin-D (calcium transport protein) + TRPV6 channels → ↑ Ca²⁺ absorption (up to 80% of dietary calcium)
• Bone: At normal levels, promotes mineralization; at high levels, releases Ca with PTH
• Kidney: Mild increase in Ca²⁺ reabsorption
• Parathyroid gland: Suppresses PTH synthesis (negative feedback)

Calcitonin

• Made by C-cells (parafollicular cells) of the thyroid
• Released when Ca²⁺ is HIGH
• Inhibits osteoclasts → reduces bone resorption; increases renal Ca excretion

Calcium and Phosphate Inverse Relationship

Phosphate Regulation

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PART 5: INSULIN - STRUCTURE AND RECEPTOR

What is insulin?

• α cells (25%) - Glucagon
• β cells (60%) - Insulin
• δ cells (10%) - Somatostatin
• PP cells (5%) - Pancreatic polypeptide

Insulin Structure

• Molecular weight: 5,808 Da
• Two chains: A chain (21 amino acids) + B chain (30 amino acids)
• Connected by: 2 disulfide bonds between chains + 1 within the A chain
• If you split the two chains apart, insulin completely loses its activity

Insulin Biosynthesis Pathway

Gene on chromosome 11
      ↓ Transcription + Translation on ribosomes (RER)
PREPROINSULIN (110 amino acids, MW ~11,500)
      ↓ Signal peptide cleaved in ER lumen
PROINSULIN (86 amino acids, MW ~9,000) = A chain + B chain + C peptide
      ↓ Packaged in Golgi vesicles
      ↓ Cleavage by prohormone convertases (PC1 and PC2) + CPE
INSULIN (A+B chains, MW 5,808) + C PEPTIDE (31 amino acids)
      ↓ Both secreted together in EQUIMOLAR amounts from secretory granules

How Glucose Triggers Insulin Secretion

Glucose enters β-cell via GLUT2 (passive)
      ↓
Glucokinase (Hexokinase IV) phosphorylates glucose → G-6-P
      ↓
Glycolysis + TCA cycle → ATP production
      ↓
↑ ATP/ADP ratio
      ↓
ATP-sensitive K⁺ channel (KATP channel) CLOSES
      ↓
β-cell membrane DEPOLARIZES
      ↓
Voltage-gated Ca²⁺ channels OPEN
      ↓
Ca²⁺ floods into β-cell
      ↓
Insulin granules FUSE with membrane → EXOCYTOSIS of insulin + C-peptide

• Phase 1 (3-5 min): Rapid dump of pre-stored insulin granules
• Phase 2 (15 min to 2 hrs): Slower, sustained secretion from newly synthesized insulin

Insulin Receptor Signal Transduction

Insulin binds α subunit
      ↓
β subunit AUTOPHOSPHORYLATES (on tyrosine residues)
      ↓
Phosphorylates IRS-1/IRS-2 (Insulin Receptor Substrate proteins)
      ↓
       ┌─────────────────────────────────┐
       ↓                                 ↓
PI3K → PIP3 → PDK1 → Akt            Grb2/SOS → Ras → ERK
(Metabolic effects:                   (Mitogenic effects:
 GLUT4 translocation,                  cell growth,
 glycogen synthesis,                   gene expression,
 lipogenesis,                          differentiation)
 protein synthesis)

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PART 6: ROLE OF C-PEPTIDE

Why Does C-Peptide Matter?

• Injected insulin has no C-peptide
• High insulin + Low C-peptide → patient is injecting insulin (factitious hypoglycemia / insulin misuse)
• High insulin + High C-peptide → body is making too much insulin (insulinoma)

• C-peptide measurement tells you how much insulin the pancreas is actually making
• C-peptide has longer half-life (~35 min) vs. insulin (~5 min) → easier to measure
• Used to distinguish Type 1 DM (C-peptide absent/very low) from Type 2 DM (C-peptide normal/high)

• Binds a G-protein-coupled receptor
• Activates Na/K-ATPase and eNOS (endothelial nitric oxide synthase)
• May protect against diabetic nephropathy and neuropathy (role still being studied)

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PART 7: GLUCAGON

What is glucagon?

• Insulin = fed state hormone (glucose ↑ → insulin ↑ → store energy)
• Glucagon = fasting state hormone (glucose ↓ → glucagon ↑ → release energy)

Structure

• 29 amino acid polypeptide
• Made from proglucagon (158 amino acids)
• Same proglucagon gene, but processed differently in intestinal L-cells → makes GLP-1 instead

Stimulation and Inhibition

Glucagon Receptor and Mechanism

Glucagon binds glucagon receptor on hepatocyte
      ↓
Gαs protein activates Adenylyl Cyclase
      ↓
ATP → cAMP ↑↑
      ↓
Protein Kinase A (PKA) activated
      ↓
     ┌──────────────────────────────────────────┐
     ↓                                          ↓
GLYCOGENOLYSIS                           GLUCONEOGENESIS
(glycogen phosphorylase activated;       (PEPCK, fructose-1,6-
 PFK-2 inhibited)                        bisphosphatase activated)
     ↓                                          ↓
Glucose released from glycogen         New glucose from
into blood                             amino acids, lactate, glycerol

• Lipolysis: Activates hormone-sensitive lipase → releases fatty acids from adipose
• Ketogenesis: Promotes fatty acid oxidation → ketone bodies in liver
• Protein catabolism: Promotes amino acid release from muscle for gluconeogenesis

Glucagon Metabolism

• Half-life: 3-6 minutes
• Degraded primarily by: liver (first pass, ~60% removed) and kidneys
• Mechanism: Proteolytic degradation by liver proteases + renal clearance

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PART 8: SOMATOSTATIN

What is somatostatin?

• δ (delta) cells of the pancreatic islets
• D cells of the stomach and intestine
• Hypothalamus (inhibits growth hormone from pituitary)

Structure

• Two forms: SS-14 (14 amino acids) and SS-28 (28 amino acids)
• Both are cyclic peptides (linked by a disulfide bridge)
• SS-14 predominates in pancreas; SS-28 predominates in intestine

What Triggers Somatostatin Release?

• High glucose, amino acids, fatty acids after a meal
• Other GI hormones (CCK, secretin)
• High glucagon or insulin (local paracrine effect)

Somatostatin's Effects

Mechanism of Action

• Somatostatin receptors (SSTR1-5) are GPCRs coupled to Gi (inhibitory)
• Gi → inhibits adenylyl cyclase → cAMP ↓ → PKA activity ↓
• Also: Opens K⁺ channels → hyperpolarization → inhibits Ca²⁺ entry → inhibits exocytosis

Somatostatin Metabolism

• Half-life: extremely short, 1-3 minutes in circulation
• Degraded by peptidases in blood and tissues
• This is why octreotide (t½ 1.5 hours) and lanreotide (monthly depot) were developed
• Clinical uses: acromegaly, carcinoid tumors, VIPomas, glucagonomas, bleeding esophageal varices

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PART 9: GLUT TRANSPORTERS

What are GLUT transporters?

The Key GLUT Transporters

The Magic of GLUT4

Insulin → IR → IRS-1 → PI3K → PIP3 → Akt → phosphorylates AS160
      ↓
TBC1D4 (AS160) inactivated
      ↓
Rab-GTPase proteins activated
      ↓
GLUT4 vesicles traffic to plasma membrane
      ↓
GLUT4 fuses with membrane (exocytosis) - increases 10-20 fold
      ↓
Glucose floods into muscle/fat cell

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PART 10: LABORATORY DIAGNOSIS OF DIABETES MELLITUS

ADA Diagnostic Criteria (any ONE on TWO separate days, except symptomatic random glucose):

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PART 11: ORAL GLUCOSE TOLERANCE TEST (OGTT)

Procedure

1. Patient fasts 8-14 hours overnight; normal carbohydrate diet (≥150g/day) for 3 days prior; no heavy exercise 3 days prior
2. Fasting blood sample taken
3. Patient drinks 75g anhydrous glucose dissolved in 250-300 mL water (within 5 minutes)
4. Blood samples taken at 1 hour and 2 hours
5. Patient sits quietly, no smoking during test

Interpretation

For Gestational Diabetes (GDM)

• 2-step approach (USA): 50g glucose screen at 24-28 weeks → if ≥130-140 mg/dL at 1 hr → 100g 3-hour OGTT
• 1-step approach (international): 75g OGTT directly; GDM if fasting ≥92, 1-hr ≥180, 2-hr ≥153 mg/dL

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PART 12: HbA1c (GLYCATED HEMOGLOBIN)

What is HbA1c?

High blood glucose + Hemoglobin A
      ↓ (non-enzymatic, irreversible, SLOW process)
Schiff base (unstable)
      ↓ Amadori rearrangement
HbA1c (stable ketoamine)

Why does it reflect 3-month glucose control?

• RBCs live for approximately 120 days (3 months)
• Every day the RBC lives, more glucose attaches to hemoglobin
• The amount of HbA1c reflects the average glucose the RBC has been exposed to over its entire lifetime
• HbA1c is a "memory" of blood glucose for the past 2-3 months
• It reflects the last 30 days more heavily (because younger RBCs are more abundant)

Reference Values

HbA1c to Average Blood Glucose Correlation

Conditions That Falsely Alter HbA1c

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PART 13: METABOLISM OF INSULIN, GLUCAGON & SOMATOSTATIN

Insulin Metabolism

• Normal fasting insulin: 5-15 μU/mL
• After a meal, peaks at 50-100 μU/mL
• Daily secretion: ~40-50 units/day
• Half-life: ~5-8 minutes

1. Liver - most important (~50% of portal insulin removed in FIRST PASS)
2. Kidneys - second (~30-40% of peripheral insulin)
3. Muscle, adipose - minor

• Kidney disease → insulin half-life prolonged → LESS insulin needed (risk of hypoglycemia)
• Liver disease → reduced first-pass extraction → HIGHER systemic insulin levels

Glucagon Metabolism

• Normal fasting glucagon: 75-150 pg/mL
• Half-life: 3-6 minutes
• Liver is the primary degradation site (60% removed in first pass)
• Kidneys are secondary
• Mechanism: Proteolytic degradation by liver enzymes + renal clearance

Somatostatin Metabolism

• Half-life: 1-3 minutes (shortest of all three)
• Mainly acts locally (paracrine) rather than as a true circulating hormone
• Degradation: Plasma peptidases + liver + kidney clearance
• Clinical consequence: Natural hormone is useless therapeutically → synthetic analogs needed:
  • Octreotide (t½ ~1.5 hours, SubQ injection 3x/day)
  • Lanreotide (depot injection, monthly)
  • Pasireotide (broader SSTR subtype binding, used in Cushing's disease)

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QUICK REFERENCE MASTER TABLE

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10 Key Concepts to Always Remember

1. All steroid hormones come from cholesterol; the rate-limiting step is StAR protein moving cholesterol into mitochondria.
2. Thyroid hormone synthesis needs iodine + TPO enzyme; T4 is a prohormone, T3 is the active hormone.
3. TSH is the most sensitive thyroid test; low TSH = overactive gland, high TSH = underactive gland.
4. PTH raises calcium AND lowers phosphate; Vitamin D raises both; FGF-23 lowers phosphate.
5. Insulin is made as preproinsulin → proinsulin → insulin + C-peptide (released in equimolar amounts).
6. GLUT2 is the glucose sensor in β cells (low affinity, high capacity); GLUT4 is insulin-regulated in muscle/fat.
7. C-peptide helps distinguish endogenous insulin production from exogenous injection; also a marker of beta cell reserve.
8. HbA1c reflects average blood glucose over 120 days (RBC lifespan); ≥6.5% = diabetes; target <7% in treatment.
9. Glucagon → GPCR → cAMP → PKA → glycogenolysis + gluconeogenesis in liver.
10. Somatostatin = master inhibitor of both insulin and glucagon via Gi → ↓cAMP; half-life only 1-3 minutes.

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