THYROID FUNCTION TESTS DR. SMILY PRUTHI PAHWA (MD-BIOCHEMISTRY) WATCH VIDEO ON YOUTUBE………. BY TULIP ACADEMY OF MEDICAL SCIENCES OR GET VIDEO FROM MOBILE APP – SEARCH ON PLAY STORE : TULIP ACADEMY WEBSITE : WWW.TAMSMED.COM
THYROID TESTS
TOTAL T-4 TOTAL T-3 FREE Thyroxine FREE T-3 Thyrotropin/ TSH Reverse T-3 FREE T-4 Fraction (% FT-4) FREE T-3 Fraction (% FT-3) Thyroid Hormone Binding Ratio (THBR)
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THYROID TESTS
FREE HORMONE CONCENTRATION FREE T-4 Index (T4 x THBR) FREE T-3 Index (T3 x THBR) T-4/TBG Ratio and T-3/TBG Ratio
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THYROID TESTS
SERUM BINDING PROTEINS THYROXINE BINDING GLOBULIN THYROXINE BINDING PREALBUMIN THYROXINE BINDING ALBUMIN
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THYROID TESTS
FOR AUTOIMMUNE THYROID DISEASE : ANTI-THYROGLOBIN Ab ANTI-MICROSOMAL Ab ANTI-THYROID PEROXIDASE Ab(TPO-Ab) TSH receptor Ab
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THYROID TESTS
Other hormone & thyroid related proteins : TRH Thyroglobulin Calcitonin
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FOR PROPER SELECTION & USE OF THESE TESTS NORMAL THYROID PHYSIOLOGY
ANATOMY OF THE THYROID GLAND
First described by Galen who thought its function was to lubricate the phyranx Derived from the Greek word Thyreos which means shield
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HYPOTHALAMIC-PITUITARY-THYROID AXIS NEGATIVE FEEDBACK MECHANISM
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HYPOTHALAMIC-PITUITARY-THYROID AXIS PHYSIOLOGY Hypothalamus
TRH
–
Pituitary –
TSH
T4
Target Tissues
T3
Heart Thyroid Gland T4
T3
TR
Liver
T4 T3 Liver Adapted from Merck Manual of Medical Information. ed. R Berkow. 704:1997. Website : www.tamsmed.com
Bone CNS
PITUITARY-THYROTROPE CELL
HM Goodman, BASIC MEDICAL ENDOCRINOLOGY 3rd Ed. Website : www.tamsmed.com
TSH REGULATION OF THYROID FUNCTION
TSH binds to specific cell surface receptors that stimulate adenylate cyclase to produce cAMP. TSH increases metabolic activity that is required to synthesize Thyroglobulin (Tg) and generate peroxide. TSH stimulates both I- uptake and iodination of tyrosine residues on Tg.
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FOLLICLES: THE FUNCTIONAL UNITS OF THE THYROID GLAND Follicles Are the Sites Where Key Thyroid Elements Function: • Thyroglobulin (Tg) • Tyrosine • Iodine • Thyroxine (T4) • Triiodotyrosine (T3) Website : www.tamsmed.com
Thyroglobulin (prohormone )
T4 (pre-hormone)
T3 (active hormone )
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THYROID HORMONES OH
OH
I
I
I
I
I
O
O NH2
I O
Thyroxine (T4)
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OH
NH2
I O
OH
3,5,3’-Triiodothyronine (T3)
T4
T3
Potency
1
10
Protein Bound
10-20
1
Half-Life
5-7d
< 24h
Secreted by thyroid
100 ug/d
6 ug/d
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FORMATION & SECRETION OF THYROID
HORMONES
ION TRANSPORT BY THE THYROID FOLLICULAR CELL
ClO4-, SCN-
BLOOD I-
I
-
NaI symporter (NIS) Thyroid peroxidase (TPO) Website : www.tamsmed.com
organification
COLLOID
Propylthiouracil (PTU) blocks iodination of thyroglobulin
THYROGLOBULIN SYNTHESIS IN THE THYROID FOLLICULAR CELL Iodination of Tyr residues of Tg
COLLOID
TSH TSH receptor TPO Website : www.tamsmed.com
THYROID HORMONE SECRETION BY THE THYROID FOLLICULAR CELL
T4 T3 COLLOID DIT MIT
TSH TSH receptor Website : www.tamsmed.com
I-
THYROID HORMONES IN THE BLOOD
Approximately 99.98% of T4 is bound to 3 serum proteins: Thyroid binding globulin (TBG) ~75%; Thyroid binding prealbumin (TBPA or transthyretin) 15-20%; albumin ~5-10% Only ~0.02% of the total T4 in blood is unbound or free. Only ~0.4% of total T3 in blood is free.
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BASICS OF THYROID HORMONE ACTION IN THE CELL
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OH
“Step up”
I
OH
I
NH
2
THYROID HORMONE METABOLISM
O I
O
I
“Step down”
I O
I
rT3 I
R
O
OH
I
I
T3
R
O
OH
R
I
OH
I
I
T4
OH
NH
2
3,3’-T2
O
R=
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THYROID HORMONE DEIODINASES
Three deiodinases (D1, D2 & D3) catalyze the generation and/disposal of bioactive thyroid hormone. D1 & D2 “bioactivate” thyroid hormone by removing a single “outerring” iodine atom. D3 “inactivates” thyroid hormone by removing a single “innerring”iodine atom. All family members contain the novel amino acid selenocysteine (SeC) in their catalytic center.
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BASIC ORGANIZATION OF THE SELENODEIODINASES NH2
extracellular domain
A
intracellular domain
B E
C
EXISTS AS A DIMER
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D
COOH
THYROID HORMONE DEIODINASES
D1- in thyroid , liver & kidneys. Low affinity for T4 D2 â&#x20AC;&#x201C; in pituitary gland, brain, brown fat & thyroid gland. Higher affinity for T4 Expression of D2 allows regulation of T3 concentration locally
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SPECIFIC ACTIONS OF THYROID HORMONE: METABOLIC Regulates of Basal Metabolic Rate (BMR). Increases oxygen consumption in most target tissues. Permissive actions: TH increases sensitivity of target tissues to catecholamines, thereby elevating lipolysis, glycogenolysis, and gluconeogenesis.
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SPECIFIC ACTIONS OF THYROID HORMONE: DEVELOPMENT TH is critical for normal development of the skeletal system and musculature. TH is also essential for normal brain development and regulates synaptogenesis, neuronal integration, myelination and cell migration.
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IN PREGNANCY: - TBG ↑ in first 2 wks to reach a plateau at 20 wks due to ↑ Oestrogen level . - ↑ TBG → ↑ serum T4 & T3 with no change in free T4 & T3. -Iodine deficiency (↑GFR + fetal thyroid uptake) → ↑ thyroid gland uptake & enlargement (goitre). -hCG occupies TSH receptors → ↓TSH but it stimulates the gland → ↑ T4. -Placental conversion of T4 to T3 to prevent excess T4 & even low T4 level in late pregnancy.
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PATHOPHYSIOLOGY
Pathophysiology OBJECTIVES Hyperthyroidism Hypothyroidism
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TYPICAL THYROID HORMONE LEVELS IN THYROID DISEASE
TSH
T4
T3
Hypothyroidism Hyperthyroidism High
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High
Low Low
Low High
Functional Disorders Website : www.tamsmed.com
Thyrotoxicosis
Hypothyroidism
-Grave’s Disease - Toxic Adenoma - Toxic Multinodular Goiter -Thyroiditis -Exogenous -TSH Mediated
-Hashimoto’s Disease - Post-op/Post-ablative -I deficiency
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Hoarseness/ Nervousness/Tremor
Deepening of Voice Persistent Dry or Sore Throat
Mental Disturbances/ Irritability
Difficulty Swallowing
Difficulty Sleeping Bulging Eyes/Unblinking Stare/ Vision
Palpitations/
Changes
Tachycardia
Enlarged Thyroid (Goiter)
Impaired Fertility Weight Loss or Gain
Menstrual Irregularities/
Heat Intolerance Increased Sweating
Light Period Frequent Bowel Movements
Sudden Paralysis
Warm, Moist Palms First-Trimester Miscarriage/
Family History of
Excessive Vomiting in Pregnancy
Thyroid Disease Website : www.tamsmed.com
or Diabetes
INCREASED SYNTHESIS: HYPERTHYROIDISM High T4 & Low TSH Increased T4/T3 release: • Grave’s • Toxic MNG • Toxic Adenoma High RAIU
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GRAVEâ&#x20AC;&#x2122;S DISEASE Most common cause in US AutoAb against TSH receptor Diffuse Goiter, Thyrotoxicosis, High RAIU Thyroid Scan: Increased activity Ophthalmopathy, Dermopathy, Acropathy
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GRAVE’S DISEASE - TREATMENT
Medication: 50% remission @ 1 year • Methimazole • PTU • Beta blockers Radioactive Iodine Ablation • Not for pts with severe ophthalmopathy Surgical Removal
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TOXIC MULTINODULAR GOITER
Sporadic Goiter Multinodular Euthyroid Subclinical Overt Thyrotoxicosis Increased RAIU (autonomous production) • Rest of Gland suppressed Treatment: Radioactive Iodine
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THYROID NODULES
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TOXIC ADENOMA HOT Nodule: Autonomous function Activating Mutation of TSH Receptor Size = Hormone production • >3 cm Treatment: • Hemithyroidectomy • Radioactive Iodine
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NON-HYPERTHYROIDISM (DAMAGED GLAND) Low RAIU Subacute Thyroiditis • Firm & painful gland • Post-viral Drug-Induced • Amiodarone, Lithium, α-IFN, IL-2 Postpartum Thyroiditis Ectopic thyroid tissue
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THYROTOXICOSIS TREATMENT Beta-blockers (hyperadrenergic symptoms) Hyperthyroidism: • Anti-thyroid Drugs • Propylthiouracil (PTU), Methimazole • Radioiodine Ablation • Surgical Thyroidectomy
Thyroiditis: • ASA, NSAIDS, +/- corticosteroids
Iodine (high doses Wolff Chaikoff effect)
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Functional Disorders
Thyrotoxicosis
Hypothyroidism
-Grave’s Disease - Toxic Adenoma - Toxic Multinodular Goiter -Thyroiditis -Exogenous -TSH Mediated
-Hashimoto’s Disease - Post-op/Post-ablative -I deficiency
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HYPOTHYROIDISM
Low T4 & High TSH More common than Thyrotoxicosis Treatment: Synthroid - goal TSH 1-2 mU/L
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IODINE DEFICIENCY REMAINS THE MOST COMMON CAUSE HASHIMOTO’s – in developed countries Website : www.tamsmed.com
Tiredness
Puffy Eyes Enlarged Thyroid (Goiter)
Forgetfulness/Slower Thinking Moodiness/ Irritability
Hoarseness/ Deepening of Voice Persistent Dry or Sore Throat
Depression Inability to Concentrate Thinning Hair/Hair Loss
Difficulty Swallowing
Loss of Body Hair
Slower Heartbeat
Dry, Patchy Skin
Menstrual Irregularities/ Heavy Period Infertility
Weight Gain Cold Intolerance Elevated Cholesterol
Muscle Weakness/ Family History of Thyroid Disease or Diabetes
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Cramps
NON THYROIDAL ILLNESS Euthyroid sick syndrome Reduced peripheral conversion of T4 to T3 Increased production of Rt3 Reduced production of thyroid horm,one binding proteins Circulating inhibitors of thyroid hormone binding Mild elevation of TSH during recovery Mild depression of TSH - acute phase
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DRUGS AND CONDITIONS THAT INCREASE SERUM T4 AND T3 LEVELS BY INCREASING TBG Drugs that increase TBG • Oral contraceptives and other sources of estrogen • Methadone • Clofibrate • 5-Fluorouracil • Heroin • Tamoxifen
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Conditions that increase TBG • Pregnancy • Infectious/chronic active hepatitis • HIV infection • Biliary cirrhosis • Acute intermittent porphyria • Genetic factors
DRUGS AND CONDITIONS THAT DECREASE SERUM T4 AND T3 BY DECREASING TBG LEVELS OR BINDING OF HORMONE TO TBG Drugs that decrease serum T4 and T3 Conditions that decrease serum T4 and T3 • Glucocorticoids • • • • •
Androgens L-Asparaginase Salicylates Mefenamic acid Antiseizure medications, eg, phenytoin, carbamazepine • Furosemide Website : www.tamsmed.com
• Genetic factors • Acute and chronic illness
ANALYTICAL METHODS
TSHTHYROID STIMULATING HORMONE
THYROTROPIN
SERUM TSH a glycoprotein MW- 30,000 daltons Formed in basophil cells of ant. pituitary 2 subunits: Alpha â&#x20AC;&#x201C; species specific & has identical a.a. sequence to the alpha chains of LH, FSH & hCG Beta- carries immunological & biological information Secretion occurs in circadian fashion ď&#x192; maximum b/w 0200 and 0400 hrs Lowest b/w 1700-1800 hrs
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SERUM TSH Single best or initial test of the thyroid function. Central to the negative-feedback system. Inverse, log-linear relationship with thyroid hormone. Small changes in FT4 result in large changes in TSH level Normal range 0.5 â&#x20AC;&#x201C; 5.0 mU/L Third generation TSH chemiluminometric assays have detection limits of about 0.01mU/L. A normal TSH is sufficed to halt further testing unless suspect of possible hypothalamic pituitary disease.
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SERUM TSH 1st generation – RIA (with functional detection limit of 1-2 mIU/L) 2nd generation – Immunometric assays (0.1-0.2 mIU/L ) 3rd generation – Chemiluminescence assays (< 0.02 mIU/L) 4th generation assays 0.001- 0.002 mIU/L
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SERUM TOTAL T4 & T3 measured by radioimmunoassay (RIA), chemiluminometric assay, or similar immunometric technique. Most is bound to TBG (thyroxine binding globulin), transthyretin or TBPA (thyroxine-binding prealbumin), or albumin. measure both bound and unbound (â&#x20AC;&#x153;freeâ&#x20AC;?) Levels are high in approximately 90% of hyperthyroid patients and low in approximately 85% of hypothyroid patients.
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SERUM FREE T4 & FREE T3
2 direct methods Free hormone competetion with radiolabelled hormone for binding to a solid phase Ab Physical separation of free hormone fraction by ultracentrifugation or equilibrium dialysis or estimated indirectly by calculation of free-thyroxine index (FTI) from total hormone conc & THBR
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FREE T3, AND RT3
FT3 - Useful to distinguish T3 toxicosis from subclinical thyrotoxicosis. Reverse T3 (rT3) - increased in NTI. - inactive. - helpful to exclude central hypothyroidism Website : www.tamsmed.com
THYROID HORMONE BINDING PROTEINS
THYROGLOBULIN LARGE GLYCOPROTEIN MW- 660 k Da Made by follicular cells , secreted into follicular lumen
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METHOD OF ANALYSIS
THYROGLOBULIN RIA Double Ab Immunoassays Two site immunometric assays ï&#x192;¼Interference due to anti-thyroglobulin Ab (+nt in 15-35 % of thyroid CA patients). To overcome use multisite immuno radiometric assay
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THYROID ANTIBODIES
Anti-Thyroid Peroxidase (Anti- microsomal) 100% in Hashimato thyroiditis & idiopathic myxedema 87% with graves disease Anti Thyroglobulin Antibody 76% of Graves Disease TSH receptor antibody Normally present in 12 â&#x20AC;&#x201C;18 % of female population Anti non-Tg colloid Ag Antibody to T4 & T3
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MOST commonly used Ab in evaluating thyroid autoimmune disease â&#x20AC;&#x201C; TPO-Ab Thyroglobulin Ab is used for detecting interference with Tg measurement
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THYROID ANTIBODIES Analytical methods
Antithyroglobulin Ab Passive haemagglutination Agar gel diffusion precipitin technique Immunofluorescence of tissue sections ELISA Radioassay Chemiluminescence based immuno metric assays
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THYROID ANTIBODIES Analytical methods Antithyroid peroxidase Ab Complement fixation Immunofluorescence ELISA Radioassay Passive haemagglutination
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RADIOIODINE UPTAKE
Useful to differentiate the type of thyrotoxicosis A set dose of I123 is given and 24 hr later a radiation detector is placed over the thyroid to determine the percentage of the dose that was taken up by the thyroid high: Graves disease, TSH-secreting pituitary tumor, hot nodules, hCG secreting tumor, iodine deficiency low : thyroiditis, thyroiditis factitia, iodine excess (contrast dye, amiodarone-induced thyrotoxicosis type 2)
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THYROID SCANS Useful in nodular disease to determine whether a nodule is hot or cold. A “hot” functioning nodule is nearly always benign. A “cold” nodule has 5% chance of being malignant and should be further evaluated
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THYROID SCAN Thyroid nodule: risk of malignancy 6.5% only 5-10% of nodules
Cold nodule
16-20% malignant
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â&#x20AC;&#x153;Warmâ&#x20AC;? Nodule
(indeterminant) 5% malignant
Hot Nodule
Tc-99m < 5% malignant I123 < 1% malignant
THYROID ULTRASOUND determine the size and the content of nodules (cystic or solid, or mixed.) Select site for FNA biopsy Provide assistance in therapeutic procedure (cyst aspiration, ETOH injection, laser therapy) and facilitate the monitoring of the effects of treatment. Worrisome characteristics: hypoechogenicity, microcalcification, irregular margins, increased nodular blood flow and evidence of margin or regional lymphadenopathy
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FINE NEEDLE ASPIRATION
First test of choice in euthyroid patient with a (palpable) nodule. Sampling errors can be minimize by using ultrasound-guided biopsy. Avoids unnecessary surgery
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SUMMARY
TSH
Low
High
FT4 & FT3
FT4
Low
High
1° Hypothyroid
Low Central Hypothyroid
2° thyrotoxicosis TRH Stim.
•FT3, rT3 •MRI Website : www.tamsmed.com
MRI, etc.
High 1° Thyrotoxicosis
RAIU
LOW FREE T4
NORMAL FREE T4
HIGH FREE T4
Secondary hypothyroidism
Subclinical hyperthyroidism
Hyperthyroi dism
Severe NTI
NTI
NORMAL TSH
Secondary hypothyroidism
NORMAL
Artifact, Pituitary hyperthyroidism, Lab draw within 6 hrs of thyroxine dose
HIGH TSH
Primary hypothyroidism
Subclinical hypothyroidism
Artifact Pituitary hyperthyroidism Thyroid hormone Resistance
LOW TSH
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Severe NTI
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