Biochemical Aspect of Endocrine Glands- I.

Biomedical Importance: Survival depends on- ability to adapt to a constantly changing environment. 2. Necessity of intercellular communication.
1.

Definition: Hormone: (Greek - arouse to activity). Is a substance that is synthesized in one organ and transported by the circulatory system to act on another tissue. (endocrine) Overview of Hormonal cascade & Important Endocrine glands:

Environmental/ internal signal

CNS

Limbic System

Hypothalamus

Ant. Pituitary

Target gland

Final hormone

Systemic effects

Nature of Hormones Hormones can be divided into five major classes: (1) amino acid derivatives- (dopamine, catecholamine, and thyroid hormone) (2) small neuropeptides- ( gonadotropin-releasing hormone (GnRH), thyrotropin-releasing hormone (TRH), somatostatin, and vasopressin) (3) large proteins- (luteinizing hormone (LH), and PTH) (4) steroid hormones- (cortisol and estrogen that are synthesized from cholesterol-based precursors) (5) vitamin derivatives- (retinoids (vitamin A) and vitamin D). A variety of peptide growth factors, most of which act locally, share actions with hormones. As a rule, amino acid derivatives and peptide hormones interact with cell-surface membrane receptors. Steroids, thyroid hormones, vitamin D, and retinoids are lipidsoluble and interact with intracellular nuclear receptors. Classification:

General Features of Hormone Classes

Group I Steroids, iodothyronines, calcitriol, retinoids Lipophilic Yes Long (hours to days) Intracellular Receptor-hormone complex Group II Polypeptides, proteins, glycoproteins, catecholamines Hydrophilic No Short (minutes) Plasma membrane cAMP, cGMP, Ca2+, metabolites of complex phosphinositols, kinase cascades

Types Solubility Transport proteins Plasma halflife Receptor Mediator

RECEPTOR SPECIFICITY & CROSS TALK Hormone Synthesis & Processing.

Secretion, Transport & Degradation.

Classification of Hormones by Mechanism of Action

I. Hormones that bind to intracellular receptors Steroid hormones Retinoic acid Thyroid hormones (T3 and T4 ) II. Hormones that bind to cell surface receptors A. The second messenger is cAMP Adrenergic catecholamines (2, ) TSH,FSH, LH, MSH, ACTH, PTH ADH Calcitonin CRH Glucagon Somatostatin

B. The second messenger is cGMP Atrial natriuretic factor Nitric oxide C. The second messenger is calcium or phosphatidylinositols (or both) Angiotensin II Antidiuretic hormone (vasopressin) Gastrin, CCK GnRH Oxytocin Thyrotropin-releasing hormone (TRH) D. The second messenger is a kinase or phosphatase cascade Adiponectin Chorionic somatomammotropin EGF, FGF,IGF 1 & 2, GH Erythropoietin Leptin Prolactin

1 -Adrenergic catecholamines

Membrane Receptor Families and Signaling Pathways

Receptors Effectors Signaling Pathways

G ProteinCoupled Seven-Transmembrane (GPCR) -Adrenergic, LH, FSH, TSH Glucagon PTH, PTHrP ACTH, MSH GHRH, CRH -Adrenergic, Somatostatin Gs, adenylate cyclase Ca2+ channels Gi Stimulation of cyclic AMP production, protein kinase A Calmodulin, Ca2+-dependent kinases Inhibition of cyclic AMP production Activation of K+, Ca2+ channels TRH, GnRH Gq, G11 Phospholipase C, diacylglycerol, IP3, protein kinase C, voltage-dependent Ca2+ channels

Receptor Tyrosine Kinase Insulin, IGF-I EGF, NGF GH, PRL Serine Kinase Activin, TGF-, MIS Serine kinase Smads Tyrosine kinases, IRS Tyrosine kinases, ras JAK, tyrosine kinases MAP kinases, PI 3-kinase; AKT, also known as protein kinase B, PKB Raf, MAP kinases, RSK STAT, MAP kinase, PI 3-kinase, IRS-1

Cytokine ReceptorLinked Kinase

Hormone Action & Signal transduction: a. A hormonereceptor interaction results in generation of an intracellular signal: b. Regulate the activity of a select set of genes, c. Affect the activity of specific proteins. d. The signal can influence the location of proteins in the cell and e. Affect general processes such as protein synthesis I. Intracellular Receptors:(Steroid and thyroid hormone("superfamily") Receptors) Composed of a single polypeptide chain that has, in the simplest analysis, three distinct domains: The amino-terminus: In most cases, this region is involved in activating or stimulating transcription by interacting with other

components of the transcriptional machinery. The sequence is highly variable among different receptors. DNA binding domain: Amino acids in this region are responsible for binding of the receptor to specific sequences of DNA. The carboxy-terminus or ligand-binding domain: This is the region that binds hormone. In addition to these three core domains, two other important regions of the receptor protein are a nuclear localization sequence, which targets the the protein to nucleus, and a dimerization domain, which is responsible for latching two receptors together in a form capable of binding DNA.

Hormone-Receptor Binding and Interactions with DNA When hormone binds to receptor, a characteristic series of events occurs:

Receptor activation is the term used to describe conformational changes in the receptor induced by binding hormone. The major consequence of activation is that the receptor becomes competent to bind DNA. Activated receptors bind to "hormone response elements", which are short specific sequences of DNA which are located in promoters of hormone-responsive genes. In most cases, hormone-receptor complexes bind DNA in pairs. Transcription from those genes to which the receptor is bound is affected. Most commonly, receptor binding stimulates transcription. The hormone-receptor complex thus functions as a transcription factor. I. Cell-Surface receptors:a. cAMP Mediated.

Synthesised from ATP by adenylyl cyclase located on the inner side of the plasma membrane. Adenylyl cyclase is activated by a range of signaling molecules through the activation of adenylyl cyclase stimulatory G (Gs)-protein-coupled receptors and inhibited by agonists of adenylyl cyclase inhibitory G (Gi)-proteincoupled receptors.

Liver adenylyl cyclase responds more strongly to glucagon, and muscle adenylyl cyclase responds more strongly to adrenaline. cAMP decomposition into AMP is catalyzed by the enzyme phosphodiesterase. cyclic AMP works by activating protein kinase A (PKA, a heterotetrameric molecule consisting of two regulatory subunits (R) and two catalytic subunits (C).)

a.

cGMP Mediated. Guanylate cyclase (GC) catalyzes cGMP synthesis. This enzyme converts GTP to cGMP. In turn, peptide hormones such as the atrial natriuretic factor activate membranebound GC, while soluble GC is typically activated by nitric oxide to stimulate cGMP synthesis. cGMP is a common regulator of ion channel conductance, glycogenolysis, and cellular apoptosis. It also relaxes smooth muscle tissues & causes vasodilation to increase blood flow. Calcium/ Phosphatidylinositol.

a.

b.

Kinase/ Phosphatase Cascade. (Insulin)

(IGFBP, insulin-like growth factor binding protein; IRS 14, insulin receptor substrate isoforms 14; PI-3 kinase, phosphatidylinositol 3-kinase; PTEN, phosphatase and tensin; PKD1, phosphoinositide-dependent kinase; PKB, protein kinase B; SGK, serum and glucocorticoid-regulated kinase; aPKC, atypical protein kinase C; p70S6K, p70 ribosomal protein S6 kinase; mTOR, mammalian target of rapamycin; GRB2, growth factor receptor binding protein 2; mSOS, mammalian son of sevenless; MEK, MAP kinase kinase and ERK kinase; MAP kinase, mitogen-activated protein kinase; mTOR, mammalian Target Of Rapamycin).

Pathologic Mechanisms of Endocrine disease: 1. Hormone Excess. 2. Hormone deficiency. 3. Resistance.

The Glands & Various Axes:

The Anterior Pituitary The Master Gland Produces 6 Hormones: PRL, GH, ACTH, TSH, FSH & LH Secreted in a pulsatile manner,(-Hypothalamic RF) . Specific responses in peripheral target tissues. Feedback control. Hormone deficiency may be inherited or acquired. Diagnoses are often elusive Performing the correct laboratory diagnostic tests.

Anterior Pituitary Hormone Expression and Regulation Cell Fetal appearance Hormone Protein Corticotrope 6 weeks POMC Polypeptide Somatotrope 8 weeks GH Polypeptide Lactotrope 12 weeks PRL Polypeptide Thyrotrope 12 weeks TSH Glycoprotein , , subunits 211 TRH Gonadotrope 12 weeks FSH LH Glycoprotein , , subunits 210, 204 GnRH, activins, estrogen Sex steroids, inhibin

Amino acids 266 (ACTH 1 191 39) Stimulators CRH, AVP, GHRH, ghrelin

199 Estrogen, TRH, VIP

Inhibitors

Glucocorticoids Somatostatin, IGF-I Dopamine

T3, T4, dopamine, somatostatin, glucocorticoids

Target gland Adrenal Trophic effect Steroid production

Liver, other tissues IGF-I production, growth induction, insulin antagonism

Breast, other Thyroid tissues Milk production T4 synthesis and secretion

Ovary, testis Sex steroid production, follicle growth, germ cell maturation M, 520 IU/L, F (basal), 520 IU/L

Normal range

ACTH, 422 pg/L

<0.5 g/L

M < 15; F <20 g/L

0.15 mU/L

Hormone Test Growth hormone Insulin tolerance test: Regular insulin (0.050.15 U/kg IV) GHRH test: 1 g/kg IV L-Arginine test: 30 g IV over 30 min L-Dopa test: 500 mg PO Prolactin ACTH TRH test: 200500 g IV Insulin tolerance test: regular insulin (0.050.15 U/kg IV)

Blood Samples 30, 0, 30, 60, 120 min for glucose and GH

Interpretation Glucose < 40 mg/dL; GH should be >3 g/L

0, 15, 30, 45, 60, 120 min for Normal response is GH >3 g/L GH 0, 30, 60, 120 min for GH 0, 30, 60, 120 min for GH 0, 20, and 60 min for TSH and PRL 30, 0, 30, 60, 90 min for glucose and cortisol Normal response is GH >3 g/L Normal response is GH >3 g/L Normal prolactin is >2 g/L and increase >200% of baseline Glucose <40 mg/dL Cortisol should increase by >7g/dL or to >20 g/dL

CRH test: 1g/kg ovine CRH IV at 8 A.M.

0, 15, 30, 60, 90, 120 min for Basal ACTH increases 2- to 4-fold and ACTH and cortisol peaks at 20100 pg/mL Cortisol levels >2025 g/dL Plasma 11-deoxycortisol and Plasma cortisol should be <4 g/dL to cortisol at 8 A.M.; ACTH assure an adequate response can also be measured Normal response is 11-deoxycortisol >7.5 g/dL or ACTH >75 pg/mL

Metyrapone test: Metyrapone (30 mg/kg) at midnight

Standard ACTH stimulation 0, 30, 60 min for cortisol and Normal response is cortisol >21 g/dL and test: ACTH 1-24 (cosyntropin), aldosterone aldosterone response of >4 ng/dL above 0.25 mg IM or IV baseline Low-dose ACTH test: ACTH 1-24 (cosyntropin), 1 g IV 3-day ACTH stimulation test consists of 0.25 mg ACTH given IV over 8 h each day TSH Basal thyroid function tests: T4, T3, TSH Basal measurements 0, 30, 60 min for cortisol Cortisol should be >21 g/dL Cortisol >21 g/dL

Low free thyroid hormone levels in the setting of TSH levels that are not appropriately increased indicate pituitary insufficiency TSH should increase by >5 mU/L unless thyroid hormone levels are increased Basal LH and FSH should be increased in postmenopausal women Low testosterone levels in the setting of low LH and FSH indicate pituitary insufficiency In most adults, LH should increase by 10 IU/L and FSH by 2 IU/L. Combined or individual releasing hormone responses must be elevated in the context of basal target gland hormone values and may not be uniformly diagnostic.

TRH test: 200500 g IV LH, FSH

0, 20, 60 min for TSH and PRL

LH, FSH, testosterone, estrogen Basal measurements

GnRH test: GnRH (100 g) IV 0, 30, 60 min for LH and FSH Multiple hormones Combined anterior pituitary 30, 0, 15, 30, 60, 90, 120 test: GHRH (1 g/kg), min for GH, ACTH, cortisol, CRH (1g/kg), GnRH (100 LH, FSH, and TSH g), TRH (200 g) are given IV

Pituitary Adenomas: (e.g., acromegaly, prolactinomas, or Cushing's syndrome) . When a pituitary adenoma is suspected based on MRI, initial hormonal evaluation usually includes (1) basal PRL; (2) insulin-like growth factor (IGF) I; (3) 24-h urinary free cortisol (UFC) and/or overnight oral dexamethasone (1 mg) suppression test; (4) subunit, FSH, and LH; and (5) thyroid function tests. Screening Tests for Functional Pituitary Adenomas Test Serum IGF-I Oral glucose tolerance test with GH obtained at 0, 30, and 60 min Serum PRL Comments Interpret IGF-I relative to age- and sex-matched controls Normal subjects should suppress growth hormone to <1 g/L Exclude medications MRI of the sella should be ordered if prolactin is elevated Ensure urine collection is total and accurate Normal subjects suppress to <5 g/dL

Acromegaly

Prolactinoma

Cushing's disease

24-h urinary free cortisol Dexamethasone (1 mg) at 11 P.M. and fasting plasma cortisol measured at 8
A.M.

ACTH assay

Distinguishes adrenal adenoma (ACTH suppressed) from ectopic ACTH or Cushing's disease (ACTH normal or elevated)

Posterior Pituitary: Formed by axons that originate in large cell bodies in the supraoptic and paraventricular nuclei of the hypothalamus. 1. Arginine Vasopressin (ADH) AVP is a nonapeptide composed of a six-member disulfide ring and a tripeptide tail. Average threshold:280 mosmol/L or 135 meq/L, respectively. Action is mediated via binding to G proteincoupled V2 receptors on the serosal surface of the cell, activation of adenyl cyclase, and insertion into the luminal surface of water channels composed of a protein known as aquaporin 2 (AQP2). Helps in maintaining BP by RAA cycle. Diseases: DI, SIADH.

At high concentrations, AVP also causes contraction of smooth muscle in blood vessels and in the gastrointestinal tract, induces glycogenolysis in the liver, and potentiates adrenocorticotropic hormone (ACTH) release by corticotropin-releasing factor. These effects are mediated by V1a or V1b receptors that are coupled to phospholipase C.

1.

Oxytocin.

Thyroid

Table:Characteristics of Circulating T4 and T3 Hormone Property Serum concentrations Total hormone Fraction of total hormone in the free form Free (unbound) hormone Serum half-life Fraction directly from the thyroid Production rate, including peripheral conversion Intracellular hormone fraction Relative metabolic potency Receptor binding T4 T3

8 g/dL 0.02% 21 1012M 7d 100% 90 g/d 20% 0.3 1010M

0.14 g/dL 0.3% 6 1012M 0.75 d 20% 32 g/d 70% 1 1011M

Investigation of primary hypothyroidism Serum TSH is the investigation of choice; a high TSH level confirms primary hypothyroidism. A low free T4 level confirms the hypothyroid state (and is also essential to exclude TSH deficiency if clinical hypothyroidism is strongly suspected and TSH is normal or low). Thyroid and other organ-specific antibodies may be present. Other abnormalities include the following: anaemia, which is usually normochromic and normocytic in type but may be macrocytic (sometimes this is due to associated pernicious anaemia) or microcytic (in women, due to menorrhagia) increased serum aspartate transferase levels, from muscle and/or liver increased serum creatine kinase levels, with associated myopathy hypercholesterolaemia and hypertriglyceridaemia hyponatraemia due to an increase in ADH and impaired free water clearance. Hyperthyroidism: Investigations Serum TSH is suppressed in hyperthyroidism (< 0.05 mU/L), except for the very rare instances of TSH hypersecretion. A raised free T4 or T3 confirms the diagnosis; T4 is almost always raised but T3 is more sensitive as there are occasional cases of isolated T3 toxicosis.