THYROID HORMONES

M.Prasad NaiduMSc 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 diminished TSH response to TRH at the pituitary level.

T3 and T4 act at the hypothalamic level by inhibiting mRNA for TRH synthesis.

Only unbound fractions of hormone are metabolically active and only this free hormone has an inhibitory effect on the secretory activity of the thyroid.

dopamine physiologically inhibits TSH secretion

glucocorticoids have been shown to dull the response of the pituitary to TRH

oestrogens increase the sensitivity of thyrotrophs to TRH

Mechanism of thyroid hormone receptor action

Actions of thyroid hormones Brain----growth&development

of nervous system

Bone&tissue growth– linear growth & maturation of bones

CVS-- increased contractility,heart rate &cardiac output

GUT—increased absorption of nutrients, increased motility

Liver -increased gluconeogenesis&glyco genolysis

Adipose tissue –increased lipolysis

Muscle –increased protein catabolism in skeletal muscle

Kidney -increased erythropoietin synthesis

Respiration- increased central stimulation of respiration

Energy metabolism -increased 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 out TSH

Thyroid follicle with high TSH stimulation

High T3 or T4 gives

decreased TSH subunit synthesis

inactive thyrotrophs may lose the capacity to respond to reduced T3 or T4 levels

inhibits TSH release potentiates the

effect of thyroid hormones on thyrotrophs, ie thyroid hormone has inhibitory effects on TSH release

derives from the median eminence of the hypothalamus

thyrotropin releasing hormone, ie stimulates TSH release

somatostatin TRH

Iodine deficiency Hasimoto’s thyroiditis Thyroidectomy Radiation therapy Drugs-lithium,antithyroid

drugs and PAS Absent or ectopic thyroid

gland Dyshormonogenesis TSH receptor mutation

Hypopituitarism Tumors,pituitary

surgery, irradiation/infiltration, sheehan’s syndrome & isolated TSH deficiency

Hypothalamic disease Trauma & infiltration

Primary hypo thyroidism Secondary hypotyroidism

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

Grave’ disease Toxic multinodular

goitre Toxic adenoma Functioning metastatic

thyroid carcinoma TSH receptor mutation Struma ovarii Iodine excess

TSH secreting pituitary adenoma

Thyroid hormone resistance syndrome

Chorionic gonadotropin secreting tumours

Gestational thyrotoxicosis

Primary hyperthyroidism Secondary hyper thyroidism

hyperthyroidism- Grave’s Disease

- tall stature, hyperactivity

- high rate of metabolism

- high body temp

- high heart rate

Thyroid function in pregnancyFour 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 testsEstimation of thyroid

hormones Total T4 Total T3Estimation of free

hormone fraction Free T4 fraction %FT4 Free T3 fraction %FT3 THBR

Estimates of free hormone concentration

FT4E (T4 X %FT4) FT3E (T3 X % FT3) FT4I (T4 X THBR) FT3I (T3 X THBR) T4: TBG ratio

Thyroid function testsSerum binding proteins Thyroxine binding

globulin Thyroxine binding

prealbuminTests for auto immune

thyroid disease Anti thyroglobulin

Abs Anti microsomal Abs

Anti TPO antibodies TSH receptor anti

bodiesOther hormones &

thyroid related proteins

TRH Thyroglobulin calcitonin

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 -chemiluminiscenceSecretion of TSH occurs in a circadian fashionPrimary Hypothyroidism-TSH increasedSecondary hypothyroidism-TSH ,T3 ,T4 are lowPrimary hyper thyroidism –TSH decreasedSecondary hyperthyroidism-TSH,T3,T4 high

TSH stimulation testMeasurement 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 carcinomaCertain non neoplastic conditions like.., Thyroid adenoma Subacute thyroiditis hashimoto’s thyroiditis Grave’s disease

Serum Tg conc. are not increased in pts with medullary thyroid carcinoma

Serial measurements of Tg is most useful in detecting recurrence of diff. thyroid carcinoma following surgical resection

Tg determination is used as an adjunct to ultrasound and radio iodine scanning

Assessment of serum Tg also aids in management of infants with congenital hypo thyroidism

In hyperthyroidism-TgLow 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 techniqueReference 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 stimulation2. c AMP formation3.colloid mobilization4.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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