THYROID

HORMONES

M.Prasad Naidu
MSc Medical Biochemistry, Ph.D,.
 Thyroid gland produces two principal
hormones … thyroxine & tri iodo
thyronine which regulate the metabolic rate of
the body.

 Iodine is essential for the synthesis of thyroid

hormones

 More than half of the body’s total content is

found in the thyroid gland
Hypothalamo pituitary axis

The hypothalamo-pituitary
axis is a classical
negative feedback
regulatory mechanism in
which secretion of TSH
is modulated by thyroid
hormones. Release of
TSH from the pituitary
gland is stimulated by
thyrotropin releasing
hormone (TRH) from the
hypothalamus.
Hypothalamo pituitary axis
 A small increase in T3
and T4 produces a  dopamine
diminished TSH physiologically inhibits
response to TRH at the TSH secretion
pituitary level.
 T3 and T4 act at the  glucocorticoids have
hypothalamic level by been shown to dull the
inhibiting mRNA for TRH response of the
synthesis. pituitary to TRH
 Only unbound fractions  oestrogens increase
of hormone are the sensitivity of
metabolically active and thyrotrophs to TRH
only this free hormone
has an inhibitory effect
on the secretory activity
of the thyroid.
Mechanism of thyroid hormone
receptor action
 Actions of thyroid hormones
 Brain----growth&development of  Adipose tissue –increased
nervous system lipolysis

 Bone&tissue growth– linear growth &  Muscle –increased protein

maturation of bones catabolism in skeletal muscle

 CVS-- increased contractility,heart  Kidney -increased erythropoietin

rate &cardiac output synthesis

 GUT—increased absorption of  Respiration- increased central

nutrients, increased motility stimulation of respiration

 Liver -increased  Energy metabolism -increased

gluconeogenesis&glyco genolysis BMR,increased oxygen
consumption,increased heat
production stimulation of Na-K-
ATP ase
Wolff-chaikoff effect
 Iodine deficiency increases thyroid blood flow
& upregulates the NIS , stimulating more
efficient uptake.
 Excess iodide transiently inhibits thyroid iodide
organification ,a phenomenon known as the
wolff-chaikoff effect
The functional unit of thyroid is
thyroid follicle. Normal
follicle
Thyroid follicle with high TSH
stimulation Thyroid follicle with out TSH
High T3 or T4 gives

 decreased TSH
subunit synthesis
 inactive
thyrotrophs may
lose the capacity
to respond to
reduced T3 or T4
levels
somatostatin TRH

 inhibits TSH release

 potentiates the  derives from the
effect of thyroid median eminence of
hormones on the hypothalamus
thyrotrophs, ie  thyrotropin
thyroid hormone has releasing
inhibitory effects on hormone, ie
TSH release stimulates TSH
release
Primary hypo thyroidism
 Iodine deficiency
 Hasimoto’s thyroiditis
 Thyroidectomy
 Radiation therapy
 Drugs-lithium,antithyroid
drugs and PAS
 Absent or ectopic thyroid
gland
 Dyshormonogenesis
 TSH receptor mutation
Secondary hypotyroidism
Hypopituitarism
 Tumors,pituitary
surgery, irradiation/infi
ltration, sheehan’s
syndrome & isolated
TSH deficiency
Hypothalamic disease
 Trauma & infiltration
cretinism
- congential absence of T3 and T4 or
chronic iodine deficiency during childhood

- retarded growth

- sluggish movements

- mental deficiencies
myxedema
- low rate of metabolism and lethargy

- decreased body temp

- decreased heart rate

- outer skin becomes scaley

- myxodema – swelling of sub-cu connective

tissues
Primary hyperthyroidism Secondary hyper thyroidism

 Grave’ disease  TSH secreting

 Toxic multinodular pituitary adenoma
goitre  Thyroid hormone
 Toxic adenoma resistance syndrome
 Functioning metastatic  Chorionic
thyroid carcinoma gonadotropin
 TSH receptor mutation secreting tumours
 Struma ovarii  Gestational
 Iodine excess thyrotoxicosis
hyperthyroidism
- Grave’s Disease

- tall stature, hyperactivity

- high rate of metabolism

- high body temp

- high heart rate

Thyroid function in pregnancy
Four factors alter thyroid function in pregnancy
 Transient increase in hcG during first trimester
which stimulates TSH-R
 The estrogen induced rise in TBG during the
first trimester which is sustained during
pregnancy
 Alterations in the immune system ,leading to
onset, exacerbation ,or amelioration of an
underlying auto immune thyroid disease
 Increased urinary iodide excretion ,which can
cause impaired thyroid hormone production
 Iodine supplementation is considered to be
important in women with precarious iodine
intake
 Maternal hypothyroidism occurs in 2 to 3% of
women of child bearing age & is associated
with increased risk of developmental delay in
the offspring
 Thyroid hormone requirements are increased
by 25 to 50µg/day during pregnancy
THYROID FUNCTION
TESTS
Thyroid function tests
Estimation of thyroid Estimates of free
hormones hormone
 Total T4 concentration
 Total T3  FT4E (T4 X %FT4)

Estimation of free  FT3E (T3 X % FT3)

hormone fraction  FT4I (T4 X THBR)

 Free T4 fraction  FT3I (T3 X THBR)

%FT4  T4: TBG ratio
 Free T3 fraction
%FT3
 THBR
Thyroid function tests
Serum binding  Anti TPO antibodies
proteins  TSH receptor anti
 Thyroxine binding bodies
globulin Other hormones &
 Thyroxine binding thyroid related
prealbumin proteins
Tests for auto immune  TRH

thyroid disease  Thyroglobulin

 Anti thyroglobulin  calcitonin
Abs
Measurement of T4,T3 &rT3
 METHOD
 Immunoassay
 Chemiluminiscence
 The major clinical role for T3 measurements are in
the diagnosis & monitoring of hyperthyroid pts
with suppressed TSH &normal FT4
 r T3 test is not always elevated with illness.It is
seldom used in pts with euthyroid sick syndrome
 Specifially,renal failure is associated with low r T3
conc.
Sandwich ELISA
Radioimmunoassay
Determination of free thyroid
hormones
 Direct assays – currently serve as reference
methods

 Indirect assays - more widely available for

general laboratory use
Direct methods
 Direct measurement of FT4&FT3 is a technical
challenge as free hormone conc. are low in serum
healthy individuals

 Assays for free thyroid hormones must be capable of

measuring sub picomole amounts

 Only minimal dilution of serum specimens is allowed

as dilution alters the binding of drugs, FFAs and
other substances to serum proteins
Methods
 Equilibrium dialysis

 Ultra filtration techniques

these techniques physically separate free hormone

from protein bound hormone (before direct
measurement of the free fraction with a sensitive T4
or T3 immunoassay)

These methods are unaffected by variations in SBPs

or thyroid hormone auto antibodies
Indirect methods
 More convenient & less expensive than direct

methods
 Automated immuno assay instuments

 Two step immunoassay

 One step immunoassay

 These methods estimate free hormone conc.

by using antibody extraction techniques

 FT4 is 0.03% of total serum T4
 FT3 is 0.3% of total serum T3
 Because T3 is less firmly bound by TBG than is
T4 the dialyzable fraction of T3 is appreciably
greater (by almost 10 times) than that of T4
Free hormone estimates
 FT4E = total T4 X %FT4
 The free hormone fraction as measured
dialysis or ultra filtration of diluted serum
containing tracer T4 or t3 is multiplied by the
respective total hormone concentration to
obtain indirect estimates

 THBR = %uptake(patient serum)/% uptake

(reference serum)
 Invitro I –T3resin uptake by Resin
 A known amount of I-T3 is added to a standard
volume of serum from a patient

 The amount of I-T3 which binds to the serum

proteins varies inversely with the endogenous
thyroid hormones already bound to serum
proteins(TBG)

 Residual free I-T3 then adsorbed by resin is

removed from the sample and then adsorbed/bound
I is measured
FT4 index

 Unlike direct free T4 methods , index methods

measure both the serum total T4 & the free T4
fraction
 They have an advtantage that they can define
whether an abnormal FT4 estimate is due to
abnormal hormone production or due to abnormal
protein binding
 An FT4 index is sometimes directly calculated
using the percentage T-uptake
 FT4I =total T4(µg/dl) x % thyroid uptake/ 100
Plasma TSH
Method- Immunoassay
-chemiluminiscence
Secretion of TSH occurs in a circadian fashion
Primary Hypothyroidism-TSH increased
Secondary hypothyroidism-TSH ,T3 ,T4 are low
Primary hyper thyroidism –TSH decreased
Secondary hyperthyroidism-TSH,T3,T4 high
TSH stimulation test
Measurement of serum T4 after TSH injection
 No response - primary

 Increase of T4- secondary

 Useful for distinguishing primary from

secondary hypothyroidism
TRH response test
 TRH administration will stimulate the
production of TSH
 Useful for differentiating hypothalamic from a
pituitary hypotyroidism
 There is increase of TSH after TRH in
hypothalamic disorder
If the hypothalamo pituitary axis is normal .the
T3 and T4 secretions will be increased
An abnormal response is seen in
Hyperthyroidism – T4 elevated

 Hypopituitarism- T4 Levels subnormal

 Primary hypothyroidism-exaggerated response

Determination of thyroid binding
globulin
 TBG is the thyroid binding globulin with the
greatest affinity for T4
 TBG is very important for regulating the conc. And
availability of the FT4 hormone.
 Method - immunoassay
- commercial kit methods available
- chemiluminiscence
 Estrogen induced TBG excess and congenital
TBG deficiency are important abnormalities that
affect the test results
Calculation of T4:TBG & T3:TBG
ratios
 These ratios correlate with FT4 or FT3 conc.
And are particularly useful in sera with altered
TBG conc.
 failures:They may fail however to compensate
for TBG variants with reduced T4 affinity & for
abnormal albumin binding
 Ref . Interval is 3.8 to 4.5
Determination of thyroglobulin
 Method –immunometric assay method
 These assays are based on the use of two or

more monoclonal antibodies directed to

different portions of the Tg molecule
 Difficulty: interference with anti-Tg antibodies
as seen in pts with thyroid cancer
Heterophilic antibody interference(HAMA)
 Ref interval is 3 to 42 μg/dl
 Thyroglobulin is used primarily as tumor marker in
pts carrying a diagnosis of differentiated thyroid
carcinoma

Tg levels are elevated in

Thyroid follicular &papillary carcinoma
Certain non neoplastic conditions like..,
 Thyroid adenoma

 Subacute thyroiditis

 hashimoto’s thyroiditis

 Grave’s disease
 Serum Tg conc. are  Tg determination is
not increased in pts used as an adjunct to
with medullary thyroid ultrasound and radio
carcinoma iodine scanning
 Serial measurements  Assessment of serum
of Tg is most useful in Tg also aids in
detecting recurrence management of
of diff. thyroid infants with congenital
carcinoma following hypo thyroidism
surgical resection  In hyperthyroidism-Tg

Low conc.-
thyrotoxicosis factita
Determination of antithyroid
antibodies
Anti thyroid antiodies are found in autoimmune
diseases and certain malignancies
These autoantibodies are directed against
several thyroid and thyroid hormone antigens
 Tg (Tg Ab)

 Thyroid peroxidase(TPO Ab)

 Thyroid receptor(TR Ab)

 TSH,T4,T3
 The presence of TPO antibodies is a risk
factor for autoimmune thyroid dysfunction
 However there is a high prevalence of anti-
TPO antibodies in the elderly
 With sensitive assays,low conc of TPO
antibodies may be detected in some healthy
individuals—they may have occult or
subclinical thyroid dysfunction
Method
 RIA

 CHEMILUMINISCENCE based immunometry

 Radioimmunometric technique

Reference value is ≤2U/ml(with sensitive

chemiluminiscence assay)
Detectable conc. Of TPO Ab are seen in
hashimoto’s thyroiditis,idiopathic myxedema,
grave’s disease, Type 1 IDDM
Determination of thyrotropin
receptor antibodies
Thyrotropin receptor antibodies are a group of related
immunoglobulins that bind to TSH receptors

Seen in pts with Graves disease & other auto immune

thyroid disorders

These Ab s demonstrate substantial heterogeneity

Some cause thyroid stimulation , where as others

have no effect or decrease thyroid secretion by
blocking the action of TSH
 Invitro bioassays assess the capacity of
immunoglobulins to stimulate functional activity of
thyroid gland such as..,

1.adenylatecyclase stimulation
2. c AMP formation
3.colloid mobilization
4.iodothyronine release
TSI s are present in 95% of pts with untreated
Grave’s disease
TSI measurement is also used for following the
course of therapy & predicting relapse & remission
Radio active iodine
uptake(RAIU)
 Radioactive iodine uptake by thyroid gland and
thyroid scanning with Tc 99 are of diagnostic
value.
calcitonin
 Calcitonin is secreted by the para follicular or
C cells ,which arise from the neural crest & are
distributed through out the thyroid gland
 A marker for medullary thyroid carcinoma
(tumor of C cells)
 Ref range ≤ 25pg/m L in men and ≤20 pg/m L
Normal ranges
 T3 :120-190 ng/dl
 r T3 : 10-25 ng/dl
 T4 : 5-12 µg/dl
 Thyroglobulin:3-5 µg/dl
 TRH :5-60 ng/L
 TSH :0.5-5 µU/ L
 Thyroxine binding globulin :1-2 mg/dl
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