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Hormones and Signal Transduction

Dr.Thomas Nyambo
Department of Biochemistry
School of Medicine MUCHS
Hormone

Definition: An active regulatory chemical


substance formed in one part of the body
and carried by the blood to another part of
the body where it signals the coordination
of cellular functions.
Structure and Function of Hormones

The integration of body functions in higher


organisms such as humans, is carried out by
the nervous system, the immune system, and
the endocrine system.
The endocrine system is composed of a number
of tissues that secrete their products into the
circulatory system
They are then disseminated throughout the
body, regulating the function of distant tissues
and maintaining homeostasis.
Structure and Function of Hormones

Endocrine hormones are derived from amino acids,


peptides, or sterols and to act at sites distant from their
tissue of origin.
Exocrine tissues secrete their products into ducts and
then to the outside of the body or to the intestinal tract.
Some secreted substances act close to the cells that
secrete them and are referred to as paracrines.
Some secret products that act directly on the cell that
secreted them referred to as autocrines
Some secreted products behave as endocrine,
paracrine, and autocrine. A good example is the
Insulin-like growth factor-I (IGF-I)
Hormone targets
.
Broad classification of animal hormones

There are four (4) categories of endocrine


hormones.
1) Amines. (Amino acid derivatives)
2) oligopeptides
3) polypeptides proteins
4) lipid-like hormones
Classification of Hormones
Oligopeptide hormones:
All the hormones from the posterior
pituitary gland.
All the hormones from the hypothalamus
(except dopamine ).
Angiotensin I and II.
Classification of Hormones
Polypeptide hormones:
All the hormones from the anterior pituitary gland.
The anterior pituitary-like hormones from the
placenta.
Atrial natriuretic hormone (ANP).All the hormones
from the stomach, duodenum, pancreas and liver.
Parathyroid hormone (PTH) and calcitonin.
Erythropoietin.
Renin.
Interleukins (IL)
Classification of Hormones
AMINE HORMONES (all derived from the
amino acid: tyrosine):
Thyroid hormones (T3 and T4).
Adrenalin.
Noradrenalin.
Dopamine .
Classification of Hormones
STEROID HORMONES (all derived from
cholesterol)
Sex hormones from the gonads and
placenta.
All the adrenal cortex hormones.
1,25 dihydroxy cholecalciferol(Vit D3).
Mechanism of action
Protein and peptide hormones, catecholamines like
epinephrine, and eicosanoids such as prostaglandins find
their receptors on the plasma membrane of target cells.
Binding of hormone to receptor initiates a series of events
which leads to generation of so-called second messengers
within the cell (the hormone is the first messenger).
The second messengers then trigger a series of molecular
interactions that alter the physiologic state of the cell.
Another term used to describe this entire process is signal
transduction.
General mechanism of action of water soluble hormones

.
Mechanism of action
Hormones are normally present in the plasma and
interstitial tissue at concentrations in the range of 10-
7M to 10-10M.

The effect of this low concentrations, the need to


amplify the small signals has been achieved by
presence of sensitive protein receptors
There is also a systemic feedback mechanisms to
regulate the production of endocrine hormones.
There is also specific plasma protein carriers, which
complex with hormones and help their dissemination
to distant tissues.
Mechanism of action
Target tissues have two properties in common:
i. a receptor having very high affinity for hormone
ii. a receptor coupled to a process that regulates metabolism of the
target cells.
iii. receptors for most amino acid derived hormones and all peptide
hormones are located on the plasma membrane.
iv. activation of these plasma membrane receptors by hormones
referred to as the first messenger leads to the intracellular production
of a second messenger, such as cAMP, Ca2+, inositol triphosphate
(IP3) , and diacylglycerol (DAG) which initiates the intracellular
biological response.
v. Steroid and thyroid hormones are hydrophobic and diffuse from the
plasma, across the plasma membrane to intracellular receptors
Structure of Cell Surface Receptors
Cell surface receptors are integral membrane proteins and, as such, have regions
that contribute to three basic domains:
Extracellular domains: Some of the residues exposed to the outside of the cell
interact with and bind the hormone - another term for these regions is the ligand-
binding domain.

Transmembrane domains: Hydrophobic stretches of amino acids are


"comfortable" in the lipid bilayer and serve to anchor the receptor in the membrane.

Cytoplasmic or intracellular domains: Tails or loops of the receptor that are


within the cytoplasm react to hormone binding by interacting in some way with
other molecules, leading to generation of second messengers. Cytoplasmic residues
of the receptor are thus the effector region of the molecule.
Second Messengers
cAMP: Epinephrine and norepinephrine, glucagon, luteinizing
hormone, follicle stimulating hormone thyroid-stimulating hormone,
calcitonin, parathyroid hormone, antidiuretic hormone
Calcium and/or phosphoinositides(IP3):Epinephrine and
norepinephrine, angiotensin II, antidiuretic hormone, gonadotropin-
releasing hormone, thyroid-releasing hormone.
Cyclic GMP: Atrial naturetic hormone, nitric oxide
Protein kinase activity:Insulin, growth hormone, prolactin, oxytocin,
erythropoietin, several growth factors
Structure of Cell Surface Receptors

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The G protein cycle

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G proteins and Signal transduction

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cAMP

.
Metabolism of cAMP

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cAMP as a second messenger

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Activation of protein kinase A by cAMP
Signal amplification

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Signal transductional pathways of the
Upon binding of FSH to the FSH receptor, the Gs subunit
FSH
dissociates.
Together with GTP, this complex directly activates adenylyl cyclase,
thereby leading to cAMP synthesis.
PKA is activated by cAMP, which causes the dissociation of the
catalytic subunit (C) from the regulatory subunit (R).
The active catalytic site can activate proteins by phosphorylation.
Conversely, the production of cAMP leads to an intracellular rise of
Ca2+, presumably due to the gating of calcium channels.
In the nucleus the catalytic subunit of the PKA can phosphorylate
transcription factors which then bind to CREs preceding certain
genes.
Finally, mRNA synthesis of primary response genes of FSH action
starts.
Signal transductional pathways of the FSH

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An Overview of Calcium Regulation in Cells

.
The Action of Nitric Oxide on Blood
Vessels

.
G proteins
G proteins mediate signal transduction through G
protein-linked receptors.
When a messenger binds to its receptor, the Gs
protein is activated.
In the inactive state, the alpha, beta, and gamma
subunits are present as a complex, with GDP bound
to the alpha subunit.
(a) When a receptor is activated by binding of its
specific ligand on the outer surface of the plasma
membrane, the receptor-messenger complex
associates with the Gs protein, causing the
displacement of GDP by GTP and the dissociation of
the Gs(alpha)-GTP complex.
G proteins
(b) The GTP-Gs complex then binds tightly to a molecule of
membrane-bound adenyl cyclase, activating it for synthesis
of cAMP.
(c) Activation ends when the ligand leaves the receptor, the
GTP is hydrolyzed to GDP by the GTPase activity of the
Gs subunit, and the Gs dissociates from the adenylyl
cyclase.
(d) Adenylyl cyclase then reverts to the inactive form, the Gs
alpha reassociates with the Gs complex, and cAMP
molecules in the cytosol are hydrolyzed to AMP by the
enzyme phosphodiesterase.
G proteins and Signal transduction
The cascade ends with activation of the final target enzyme

.
Phosphoinositide Cascade
Not all G proteins signal through adenylate cyclase .
Certain G proteins, such as Gq, act by activating phospholipase C.
When a particular receptor such as the vasopressin receptor is
activated, it activates a G protein that then activates the membrane-
bound enzyme phospholipase C (PLC).
Phospholipase C cleaves PIP2, releasing inositol 1,4,5-trisphosphate
(IP3) and diacylglycerol (DAG).
These two messengers then cause the activation of other proteins
Different isoforms of PLC are activated by different receptors by a
variety of mechanisms.
Examples of processes regulated by the phosphoinositide cascade are:
1) histamine secretion by mast cells
2) aggregation of blood platelets and release of serotonin
3) insulin secretion by pancreatic beta cells
4) smooth muscle contraction
IP3 and DAG

.
IP3 and DAG linked cascade

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Insulin

.
Insulin

.
Insulin Receptor

.
Insulin signal transduction pathways

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GLUT4 translocation
Insulin promotes glucose uptake by muscle and
adipose tissue via stimulation of glucose transporter
(GLUT) 4 from intracellular sites to the plasma
membrane.
Attenuated GLUT4 translocation and glucose uptake
by muscle and fat cells following insulin stimulation
represent a prime defect in insulin resistance
The PI3 kinase/Akt pathway has been demonstrated
to be upstream of GLUT4 translocation.
The insulin receptor is a tyrosine kinase
One can say that it functions as an enzyme that
transfers phosphate groups from ATP to tyrosine
residues on intracellular target proteins.
Binding of insulin to the subunits causes the beta
subunits to autophosphorylate,thus activating the
catalytic activity of the receptor.
The activated receptor then phosphorylates a
number of intracellular proteins, which in turn alters
their activity, thereby generating a biological
response.
Actions of insulin on cell level
and global metabolism level

The actions of insulin on the global human metabolism level


include:
cellular intake of certain substances, most prominently
glucose
increase of DNA replication and protein synthesis
modification of the activity of numerous enzymes (allosteric
effect)
The actions of insulin on cells include:
increased glycogen synthesis
increased fatty acid synthesis
Actions of insulin on cell level and global metabolism level

increased esterification of fatty acids (to make fats)


decreased proteolysis
decreased lipolysis
decreased gluconeogenesis
increased amino acid uptake
increased potassium uptake
arterial muscle tone forces arterial wall muscle to
relax increasing blood flow, especially in micro
arteries.
Actions of insulin on cell level and global metabolism level

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Glucagon

Glucagon is a 29 amino acid peptide hormone


synthesized the -cells of the islets of Langerhans.
Glucagon is a hormone that opposes the action of
insulin in peripheral tissues, predominantly the liver,
where the insulin:glucagon ratio determines the
delicate control of gluconeogenesis and
glycogenolysis.
Meals generally suppress glucagon secretion from
the normal a cell.However,diabetic individuals
frequently exhibit disordered control of
glucagon secretion
Glucagon

Glucagon release is stimulated by hypoglycemia and


inhibited by hyperglycemia, insulin, and
somatostatin.
Glucagon generally functions as a counter
regulatory hormone, opposing the actions of insulin,
and maintaining the levels of blood glucose
In diabetics, excess glucagon secretion plays a
primary role in the metabolic perturbations
associated with diabetes, such as hyperglycemia.
Hypothalamo-pituitary axis

.
Hypothalamo-pituitary hormones

.
Hypothalamic/pituitary axis
Releasing factors produced by the hypothalamus are
Thyrotropin releasing hormone (TRH) releases thyrotropin (TSH)
Corticotropin releasing factor (CRF) releases adrenocorticotropin
(ACTH)
LH/FSH releasing hormone (GnRH) releases luteinizing hormone
(LH) and follicle-stimulating hormone (FSH)
Growth hormone releasing factor (GHRF) releases growth hormone
Prolactin releasing factor (PRF) releases prolactin
Melanocyte-stimulating hormone releasing factor releases
melanocyte- stimulating hormone (MSH)
Inhibitory factors produced by the hypothalamus are
Dopamine inhibits prolactin release
Somatostatin inhibits growth hormone and TSH release
Melanocyte-stimulating hormone inhibitory factor (MIF) inhibits
MSH release
Growth Hormone

Human placental lactogen (hPL), GH, and prolactin (Prl)


comprise the growth hormone family.
Each has its own receptors. They all possess growth-promoting
and lactogenic activity.
Growth hormone (22,000 daltons) is synthesized in acidophilic
pituitary somatotropes as a single polypeptide chain.
In humans, growth hormone promotes gluconeogenesis and is
consequently hyperglycemic. It promotes amino acid uptake by
cells, with the result that GH therapy puts an organism into
positive nitrogen balance, similar to that seen in growing
children.
Growth hormone is lipolytic, inducing the breakdown of tissue
lipids and thus providing energy supplies that are used to
support the stimulated protein synthesis.
Prolactin
Prolactin is produced by acidophilic
pituitary lactotropes.
Prolactin is constantly under negative

control by prolactin inhibiting


hormone(dopamine).
Prolactin initiates and maintains lactation in

mammals, but normally only in mammary


tissue that has been primed with estrogens.
Gonadotropins
All gonadotrophins are highly glycosylated. Each of the
glycoprotein hormones is an () heterodimer, with the
subunit being identical in all members of the family.
The biological activity of the hormone is determined by the -
subunit, which is not active in the absence of the subunit.
Molecular weights of the gonadotropins FSH, LH, and CG is
about 25,000, whereas that of the thyroid tropic hormone TSH
is about 30,000.
All members of the glycoprotein family transduce their
intracellular effects via the receptor, G-protein, adenylate
cyclase, second-messenger system.
The gonadotropins (LH, FSH and CG) bind to cells in the
ovaries and testes, stimulating the production of the steroid sex
hormones estrogen, testosterone , and dihydrotestosterone.
Gonadotropins
In males, luteinizing hormone (LH) binds to Leydig cells of the
testes to induce the secretion of testosterone, while follicle
stimulating hormone (FSH) binds to Sertoli cells and induces
the secretion of estosterone and dihydrotestosterone .
In females, LH induces thecal cells to secrete estradiol, and FSH
stimulates estrogen synthesis by granulosa cells.
Human chorionic gonadotropin (hCG) is a placental hormone.

The production of hCG increases markedly after implantation;


its appearance in the plasma and urine is one of the earliest
signals of pregnancy and the basis of many pregnancy tests.
Thyroid Stimulating Hormone (TSH)
Secretion of TSH is stimulated by TRH from the hypothalamus.
TSH receptors are coupled through a G-protein to adenylate cyclase.
Following ligand binding,there is an increases of thyrocyte cAMP and
PKA, leading in the short term to increased secretion of thyroxin (T4) and
triiodothyronine (T3).
Chronic stimulation of the receptor causes an increase in the synthesis of a
major thyroid hormone precursor, thyroglobulin.
Thyroglobulin has a molecular weight of 660,000. It is glycosylated and
contains more than 100 tyrosine residues, of which 4 to 5 become iodinated
and are used to synthesize T3 and T4.
A Na+/K+-ATPase-driven pump concentrates iodide (I-) in thyroid cells, and
the iodide is transported to the follicle lumen. There it is oxidized to I+ by a
thyroperoxidase . The addition of I+ to tyrosine residues of thyroglobulin is
catalyzed by the same enzyme, leading to the production of thyroglobulin
containing monoiodotyrosyl (MIT) and diiodotyrosyl (DIT) residues. The
thyronines, T3 and T4, are formed by combining MIT and DIT residues on
thyroglobulin.
Vasopressin and Oxytocin

Oxytocin and vasopressin are the principal hormones of the


posterior pituitary.
They are synthesized as prohormones in neural cell bodies of the
hypothalamus and mature as they pass down axons in association
with carrier proteins called neurophysins.
The axons terminate in the posterior pituitary, and the hormones
are secreted directly into the systemic circulation.
Vasopressin is also known as antidiuretic hormone (ADH), because
it is the main regulator of body fluid osmolarity.
The secretion of vasopressin is regulated in the hypothalamus by
osmoreceptors, which sense water concentration and stimulate
increased vasopressin secretion when plasma osmolarity increases.
The secreted vasopressin increases the reabsorption rate of water
in kidney tubule cells, causing the excretion of urine that is
concentrated in Na+.
Vasopressin and Oxytocin

Deficiency of vasopressin dilute urine and polydipsia, a


condition known as diabetes insipidus.

Vasopressin binds plasma membrane receptors and acts


through G-proteins to activate the cAMP/PKA regulatory
system.
. Oxytocin secretion in lactating women is stimulated by
neural feedback obtained by stimulation of the nipple during
suckling.
Oxytocin causes contraction of mammary gland
myoepithelial cells, which induces the ejection of milk from
mammary glands, and the stimulation of uterine smooth
muscle contraction leading to childbirth.
The Renin-Angiotensin System(RAS)

The RAS is responsible for regulation of blood pressure.


A fall in pressure results in the release of renin from the
juxtaglomerular cells of the kidneys.
Renin cleaves a 10-amino acid peptide from the N-terminal
end of angiotensinogen.
This decapeptide is called angiotensin I(AT1). AT1is then
cleaved by the action of angiotensin-converting enzyme,
ACE to the active hormone, angiotensin II(AT2).
ACE removes 2 amino acids from the C-terminal end of
(AT1)I.
It is one of the most potent naturally occuring
vasoconstrictors. The vasoconstrictive action of AT2 is
primarily exerted on the arterioles and leads to a rise in both
systolic and diastolic blood pressure.
The Renin-Angiotensin System
Other physiological responses to angiotensin
II include induction of adrenal cortex
synthesis and secretion of aldosterone.
Angiotensin II also acts on the brain leading
to increased blood pressure, vasopressin and
ACTH secretion and increased water intake.
Angiotensin II affects the contractility of the
mesangial cells of the kidney leading to
decreased glomerular filtration rate.
Angiotensin II also potentiates the release of
norepinephrine.
Parathyroid Hormone (PTH)
PTH is a 9.5kdprotein.It is synthesized and secreted by
chief cells of the parathyroid in response to systemic Ca
2+ levels.

The Ca2+receptor of the parathyroid gland responds to


Ca2+ by increasing intracellular levels of PKC, Ca2+ and
IP3.
The synthesis and secretion of PTH in chief cells is
constitutive, but Ca2+ regulates the level and secretion of
PTH in chief cells by increasing the rate of PTH
proteolysis when plasma Ca2+ levels rise and by
decreasing the proteolysis of PTH when Ca2+levels fall.
PTH is to regulates Ca2+ concentration in extracellular
fluids. PTH acts by binding to cAMP-coupled plasma
membrane receptors, initiating a cascade of reactions.
The response to PTH in all tissues results into increase
of Ca2+levels in extracellular fluids.
Parathyroid hormone
PTH induces the dissolution of bone by stimulating
osteoclast activity, which leads to elevated plasma Ca 2+
and phosphate.
In the kidney, PTH reduces renal Ca2+ clearance by
stimulating its reabsorption; at the same time, PTH
reduces the reabsorption of phosphate and thereby
increases its clearance.
PTH acts on the liver, kidney, and intestine to stimulate
the production of the steroid hormone 1,25-
dihydroxycholecalciferol (calcitriol), which is
responsible for Ca2+ absorption in the intestine.
Catecholamines

.
Synthesis of the catecholamines from tyrosine
The first step in the process requires tyrosine hydroxylase in the
precence of tetrahydrobiopterin as cofactor.
The hydroxylation reaction generates DOPA (3,4-
dihydrophenylalanine).
DOPA decarboxylase converts DOPA to dopamine, dopamine b-
hydroxylase converts dopamine to norepinephrine and
phenylethanolamine
N-methyltransferase converts norepinephrine to epinephrine.this
reaction uses SAM as a methyl donor generating S-
adenosylhomocysteine.
In the substantia nigra and some other regions of the brain, synthesis
proceeds only to dopamine.
In the adrenal medulla dopamine is converted to norepinephrine and
epinephrine.
Catabolism of the catecholemines occurs through the actions of
catecholamine-O-methyltransferase, (COMT) and monoamine
oxidase, (MAO). Both of these enzymes are widely distributed
throughout the body. However, COMT is not found in nerve endings
Synthesis of the catecholamines from tyrosine.

.
Steroid hormones
The steroid hormones are all derived from
cholesterol, with the exception of retinoic acid
They all contain the same cyclopentanophenanthrene
ring and atomic numbering system as cholesterol,
with the exception of vitamin D, .
The conversion of C27 cholesterol to the C18, C19,
and C21 involves the rate-limiting, irreversible
cleavage of a 6-carbon residue from cholesterol.
This cleavage produces a C21(pregnenolone) and an
isocaproaldehyde.
Steroid hormones
Steroid hormones can be roughly divided into 4 groups
according to their physiological behaviour:
i. Adrenal mineralocorticoids, which regulate the salt
balance and maintain blood pressure.
ii. Glucocorticoids, which regulate carbohydrate metabolism
and manage stress,
iii. Progestogens and estrogens, which are mainly produced
by the ovaries and regulate reproductive function and
secondary sex characteristics in the female.
iv. Androgens, which are mainly testicular in origin and are
essential for fertility and secondary sex characteristics in
the male
Steroidogenic hormones
Most of the steroidogenic enzymes belong to the cytochrome
P450 oxidation group, among which five enzymes are
involved in adrenal steroidogenesis:
i. P450scc (side chain cleavage)
ii. P450c11 (11--hydroxylase)
iii. P450c17 (17--hydroxylase)
iv. P450c21 (21-hydroxylase)
v. P450aldo (aldosterone synthase).
The first two of these are located in the mitochondria and the
last two in the endoplasmic reticulum. In addition,
P450aro mediates the aromatisation of androgens to
estrogens in the gonads
Steroid hormones
Steroids with 21 carbon atoms are known
systematically as pregnanes
Those containing 19 and 18 carbon atoms are
known as androstanes and estranes,
respectively.
As already mentioned,retinoic acid and
vitamin D are not derived from pregnenolone,
but from vitamin A and cholesterol
respectively.
Steroid hormones
Pregnenolone: is produced directly from cholesterol,
the precusor molecule for all C18, C19 and C21 steroids

.
Progesterone: is a progestin which is produced directly
from pregnenolone and secreted from the corpus luteum,
responsible for changes associatedwith luteral phase of
the menstral cycle

Aldosterone: is a mineralocorticoid, produced from


progesterone in the zona glomerulosa of adrenal cortex,
raises blood pressure and fluid volume, increases Na+
uptake
Steroid hormones

Testosterone: is an androgen, a male sex hormone synthesized from


progesterone in the testes ,responsible for secondary male sex
characteristics.
.

Estradiol: is an estrogen, the principal female sex hormone,


produced in the ovary, responsible for secondary female sex
characteristics

Cortisol: is a glucocorticoid , synthesized from progesterone in


the zona fasciculata of the adrenal cortex. Is involved in stress
adaptation, elevates blood pressure and Na+ uptake,influence of
the immune system
Steroidogenesis
.
Mechanisms of action
All steroid hormones and thyroid hormone exert their
action by passing through the plasma membrane and
binding to intracellular receptors.
Both the steroid and thyroid hormone-receptor
complexes exert their action by binding to specific
sequences in the DNA of target cell genes.
These DNA sequences are called hormone response
elements (HREs).
An interaction of steroid-receptor complexes with DNA
leads to altered rates of transcription of the associated
genes to produce the targeted Gene product.
Steroid Hormone Biosynthesis
A particular steroid hormone type is synthesized by a given
cell type depending on its complement of peptide hormone
receptors.
Indicates below is a list of peptide hormone responsible for
stimulating the synthesis of the respective steroid hormone:
Luteinizing Hormone (LH):
progesterone and testosterone
Adrenocorticotropic hormone (ACTH):
cortisol
Follicle Stimulating Hormone (FSH):
estradiol
Angiotensin II/III:
Aldosterone
Steroid Hormone Biosynthesis
The first reaction is the committed step and it involves
cleaving C6 carbon group from cholesterol into C18, C19 and
C21 steroids.
This reaction is catalyzed by a mitochondrial enzyme system
known as P450-linked side chain cleaving enzyme (P450ssc),
or desmolase.
Mitochondrial desmolase is a complex enzyme system
consisting of cytochrome P450, and a P450 reductant called
adrenadoxin.
The activity of each of these components is increased by two
cAMP- and PKA-dependent processes:
i. cAMP stimulates PKA, leading to the phosphorylation of a
cholesteryl-ester esterase and generating increased
concentrations of cholesterol.
ii. A long-term regulation is effected at the level the gene for
desmolase.
Steroid Hormone Biosynthesis
Control at the level of cholesterol synthesis is by a
negative feedback regulator of HMG CoA reductase
activity.
That is,when cytosolic cholesterol is depleted, de
novo cholesterol synthesis is stimulated by activation
of HMG CoA reductase .
Following the desmolase activity, pregnenolone
moves to the cytosol, where further processing
depends on the tissue under consideration.
Various hydroxylases are involved in the synthesis of
the steroid hormones.They have a nomenclature that
indicates the site of hydroxylation (e.g. 21-
introduces a hydroxyl group to carbon C21).
Steroids of the Adrenal Cortex
The adrenal cortex is composed of three tissue regions:
i. zona glomerulosa
ii. zona fasciculata
iii. zona reticularis.
The pathway to pregnenolone synthesis is the same in all
zones of the cortex. However, the zones are morphologically
enzymatically distinct and the exact steroid hormone product
dependent on the enzymes present in the cells of each zone.
Adrenal cortex is responsible for production of three major
classes of steroid hormones:
i. Glucocorticoids
ii. mineralocorticoids
iii. androgens
The Adrenal Cortex

.
Reactions of the zona glomerulosa
The zona glomerulosa cells lack the P450c17 that converts
pregnenolone and progesterone to their C17 hydroxylated
analogs.
In these cells the pathways to the glucocorticoids
(deoxycortisol and cortisol) and the androgens
(dehydroepiandosterone [DHEA] and androstenedione) are
blocked.
Zona glomerulosa cells contain the enzyme responsible for
converting corticosterone to aldosterone, the most potent
mineralocorticoid.
This enzyme is P450c18 (or 18a-hydroxylase), also called
aldosterone synthase.
Thus, the zona glomerulosa is responsible for the conversion
of cholesterol to corticosterone and aldosterone.
Reactions of zona fasciculata and zona reticularis
The cells of the zona fasciculata and zona reticularis
lack aldosterone synthase (P450c18) that converts
corticosterone to aldosterone, and thus these tissues
produce only corticosterone.
These two zones contain P450c17 and thus produce
the major glucocorticoid, cortisol.
Zona fasciculata and zona reticularis cells also
contain the C17,20 lyase, whose activity is
responsible for producing the androgens,
dehydroepiandosterone (DHEA) and
androstenedione.
Steroid Hormone Biosynthesis
Regulation of Adrenal Steroid Synthesis

Activities of zona fasciculata and zona reticularis are


controlled by Adrenocorticotropic hormone (ACTH)
from the hypothalamu
ACTH has a feedback loop responsible for
regulating the circulating levels of corticotropin
releasing hormone, (CRH), ACTH itself , and
cortisol.
Mineralocorticoid from the zona glomerulosa Is
controlled by angiotensins
Angiotensins II and III are derived from the action of
the kidney protease renin on liver-derived
protein,angiotensinogen.
Angiotensins
Thus, angiotensin II and III binding to their receptor leads to the
activation of PKC and elevated intracellular Ca2+ levels.
This leads to increased P450ssc activity and increased production of
aldosterone.
In the kidney, aldosterone regulates sodium retention by stimulating
gene expression of mRNA for the Na+/K+-ATPase responsible for the
reaccumulation of sodium from the urine.
The interplay between renin from the kidney and plasma
angiotensinogen is important in regulating plasma aldosterone levels,
sodium and potassium levels, and ultimately blood pressure.
Some drugs that are employed to lower blood pressure are in the class
of the angiotensin converting enzyme (ACE) inhibitors.
Angiotensins
These compounds are potent competitive inhibitors of
the enzyme that converts angiotensin I to the physiologically
active angiotensins II and III.
Although fasciculata and reticularis cells each have the capability
of synthesizing androgens and glucocorticoids, the main pathway
normally followed is that leading to glucocorticoid production.
When a genetic defect occurs in the 3 enzyme complexes leading
to glucocorticoid production, large amounts of a potent androgen,
dehydroepiandrosterone (DHEA), are produced. These lead to
hirsutism and other masculinizing changes in secondary sex
characteristics.
Gonadal Steroid Hormones
Two most important steroids from the testes and the ovary are
testosterone and estradiol.
These hormones are under a very tight control, with a short
and a long negative feedback loops that regulate the secretion
of follicle stimulating hormone (FSH) and leutinizing hormone
(LH) by the pituitary.
The pituitary is inturn under control by gonadotropin releasing
hormone (GnRH) of the the hypothalamus.
Low levels of circulating testosterone and estradiol reduce
feedback inhibition on GnRH synthesis leading to elevated
FSH and LH.
The follicle stimulating hormone (FSH) and leutinizing
hormone (LH) bind to gonadal tissue and stimulate P450ssc
activity, resulting in sex hormone production via cAMP and
PKA mediated pathways.
Gonadal Steroid Hormones
Testes and ovaries contain 17b-hydroxysteroid dehydrogenase,
whichenables androgens to be converted to testosterone
In males, LH binds to Leydig cells, stimulating production of
the principal Leydig cell hormone, testosterone. Testosterone
is secreted to the plasma and also carried to Sertoli cells by
androgen binding protein (ABP).
In Sertoli cells the -4 double bond of testosterone is reduced,
producing dihydrotestosterone. Testosterone and
dihydrotestosterone are carried in the plasma, and delivered to
target tissue, by a specific gonadal-steroid binding globulin
(GBG).
In a number of target tissues, testosterone can be converted to
dihydrotestosterone (DHT).
DHT is extremely potent, with an activity that is 10 times that
of testosterone.
Gonadal Steroid Hormones
Testosterone is also produced by Sertoli cells but in these
cells it is regulated by FSH.
FSH also stimulates Sertoli cells to secrete androgen-
binding protein (ABP), which transports testosterone and
DHT from Leydig cells to sites of spermatogenesis.
In females, LH binds to thecal cells of the ovary, where it
stimulates the synthesis of androstenedione and
testosterone.
Females have an additional enzyme complex, aromatase
responsible for the final conversion to estrogens.
Aromatase is a complex endoplasmic reticulum enzyme
found in the ovary and in numerous other tissues in both
males and females.
Synthesis of the male sex hormones in Leydig cells of the testis

.
Synthesis of the major female sex hormones in the ovary

.
Adrenal genital syndrome
Deficiency of 21-hydroxylase inhibits the synthesis of
glucocorticoids and mineralocorticoids .
This leadsto overproduction of testosterone in the
adrenal glands and underproduction of cortisol.
Underproduction of cortisol effectively increases
adrenocorticotropic hormone (ACTH), which
stimulates the adrenals to grow and synthesize
steroids, exacerbating the testosterone
overproduction.
This leads to masculinization of females.
Synthetic steroids
Synthetic steroid hormones include the
following:
Diethylstilbestrol, a synthetic estrogen
previously used to promote growth of beef
cattle, until it was found to be potentially
carcinogenic at the levels found in meat from
treated cattle.
Oral contraceptives, such as norethynodrel
and mestranol.
Anabolic Steroids
Were first developed in the 1930's by the Germans in the
second world war.
Then in the 1950's many athletes began to find that anabolic
steriods were beneficial to their physical strength
characteristics.
Pure Testosterone was the available drug, resulting in severe
side-affects which soon became apparent after prolonged use.
There after that, other choices like Dianabol or
Methandrostenolone were developed, and a decade later
anabolic steriods were available on the market.
Latest products are so called designer steroids such as
tetrahydrogestrinone (THG), which have been chemically
engineered to deceive anti doping testers.
.

.
Congenital adrenal hyperplasia (CAH)
CAH is al term used to describe a group of inherited
disorders in which a defect in cortisol biosynthesis is
present.
An enzymatic defect in 11--hydroxylase is the
second most common variant of CAH.
Individuals with 11--hydroxylase deficiency present
with features of androgen excess, including
masculinization of female newborns and precocious
puberty in male children.
Approximately two thirds of cases also have
hypertension, which may or may not be associated
with mineralocorticoid excess, hypokalemia, and
metabolic alkalosis.
17-hydroxylase deficiency

Individuals with 17-hydroxylase deficiency have alterations


in their CYP17 gene that encodes the P450C17 enzyme. This
enzyme plays a central role in steroidogenesis .
17-hydroxylase is essential for the production of cortisol
and sex steroids. Thus, individuals with 17-hydroxylase
deficiency have reduced secretion of cortisol, androgen, and
estrogen, with both adrenal and gonadal steroidogenesis
impairment.
Although these individuals with 17-hydroxylase deficiency
have decreased cortisol production, they do not have signs
or symptoms of adrenal insufficiency due to elevations of
corticosterone and glucocorticoids
Addison disease

Is an adrenocortical insufficiency due to the


destruction or dysfunction of the entire
adrenal cortex.
Destruction or dysfunction is caused by a
number of pathologies such as
hemorrhage,adrenal tuberculosis etc.
It affects both glucocorticoid and
mineralocorticoid function.
Cushing syndrome

Cushing syndrome is caused by prolonged


exposure to elevated levels of either
endogenous or exogenous glucocorticoids.
Individuals with Cushing syndrome can
develop moon facies, buffalo torso, truncal
obesity, and skin striae.
Conn syndrome

Conn syndrome is characterized by increased


aldosterone secretion from the adrenal glands,
suppressed plasma renin activity, hypertension, and
hypokalemia.
Because there are a number hyperaldosteronism
conditions that were discovered after the 1955 1st
Conns syndrome, the term primary
hyperaldosteronism is used to describe Conn
syndrome and other etiologies of primary
hypersecretion of aldosterone (eg, adrenal
hyperplasia).
Steroid Hormone Receptors
Steroid hormones bind to are ligand-activated proteins that regulate
transcription of selected genes.
Unlike peptide hormone receptors, steroid hormone receptors are
found in the cytosol and the nucleus.
The steroid hormone receptors belong to the steroid and thyroid
hormone receptor super-family of proteins, that includes receptors for
steroid hormones, thyroid hormones, vitamin D and retinoic acid.
These receptors bind ligands and undergo a conformational change
that renders them activated to recognize and bind to specific
nucleotide sequences in the DNA referred to as hormone-response
elements (HREs).
When ligand-receptor complexes interact with DNA they alter the rate
of transcription which can be either activating or repressing the
associated gene.
Thyroid Hormones

TRH (thyrotropin releasing hormone) made by


the hypothalamus stimulates the release of
TSH (thyroid stimulating hormone) from the
anterior pituitary which regulates the
production and release of thyroid hormones.
They increase protein synthesis in virtually
every body tissue and increase oxygen
consumption dependent upon Na+ -K+ ATPase
(Na pump).
Hypothyroidism

Hypothyroidism in Adults
Low serum thyroid hormone resulting from hypothalmic, pituitary, or
thyroid insufficiency. Untreated, can lead to life-threatening myxedema
coma (early signs of fatigue, forgetfulness, sensitivity to cold,
unexplained weight gain, constipation progress to decreasing mental
stability, dry, flaky inelastic skin; puffy face, hands, and feet; hoarseness;
periorbital edema; upper eyelid droop; dry, sparse hair; and thick, brittle
nails.
Hypothyroidism in Children (Cretinism)
Deficiency of thyroid hormone secretion during fetal development or
early infancy results in infantile cretinism (congenital hypothyridism).
Respiratory difficulties, persistent jaundice, and hoarse crying in infants;
stunted growth (dwarfism), bone and muscle dystrophy, and mental
deficiency in older children.
Thyroid Hormones

The thyroid produces two hormones:


i. thyroxine (T4, a tetra-iodinated tyrosine derivative)
ii. triiodothyronine (T3).
They are derived from the amino acid tyrosine.
T4 is the major active hormone.
In tissues outside the thyroid, particularly in the liver and kidney, T4 is
converted to T3, an active metabolite, by mono-deiodination of the
outer phenol ring.
Reverse T3, or rT3, is a minimally active metabolite which derives
from mono-deiodination of the inner ring of T4.
rT3 is used to detect fetal hypothyrodism at as early as the 15wk of
pregnancy.
Thyroid Hormones

T4
T3
rT3
Actions of T3 and T4.
T3 and T4 regulate growth and development, especially
critical for the fetus, with requirements lasting past the time of
birth.
T3 and T4 impact upon numerous systems including muscle,
bones, CNS development, myelination, dendritic formation,
and synapse formation.
T3 and T4 act as metabolic stimulants (calorigenic effects) to
increase the rate of oxygen consumption by the heart, skeletal
muscle, liver, and kidney.
In adults, brain, spleen, and gonad metabolism is less
susceptible to the effects of T3 and T4. CNS input through the
hypothalamus (TRH) in cold temperatures increase T3 and T4
secretion to increase metabolic rates. T4 and T3 have
cardiovascular effects, increasing the heart rate and force of
contraction to achieve increased cardiac output.
Biosynthesis of thyroid hormones
The thyroid hormones are synthesized in the
follicular cells of the thyroid
The first step to hormone synthesis is the import of
iodide into the follicular cells a process which
depends an ATP-dependent pump which pumps K+ in
and Na+ out of the cell.
The iodide itself is pumped in with Na+ by a
cotransporter also callad sodium/iodide symporter
Active transport of iodide is required into the thyroid
cell since there is a greater than 50 fold higher
concentration of iodide inside the follicular cell than
outside.
Biosynthesis of thyroid hormones
Once in the follicular cell, iodide is converted into iodide by the enzyme thyroid
peroxidase, which uses hydrogen peroxide (H2O2) as a cofactor.
Thyroid peroxidase catalyzes the incorporation of iodide molecule onto both the 3
and/or 5 positions of the phenol rings of tyrosines found in a glycoprotein called
thyroglobulin.
Thyroglobulin has a molecular weight of about 660,000.
Thyroid peroxidase also appears to couple iodinated tyrosine rings to iodinated
phenol rings which it obtains from other iodinated tyrosine residues within the
protein.
Thyroglobulin contains approximately 300 carbohydrate residues and 5500 amino
acids, including 140 tyrosine residues.
Of these 140 tyrosines, but only two to five of these tyrosines are converted into
either T4 or T3! The entire thyroglobulin protein with its thyroid hormones is stored
in the lumen of the thyroid follicle cell.
Thyroid stimulatory hormone (TSH) stimulates enzymatic degradation of
thyroglobulin to effect release of the thyroid hormones.
T4 is formed exclusively in the thyroid and secreted. In contrast, 80% of T3 found
circulating in the blood is derived from metabolism of T4 in peripheral tissues,
especially the liver and kidney
Biosynthesis of thyroid hormones

.
Thyroid hormone receptors
Thyroid hormone receptors occur in the cytoplasm and nucleus.
The thyroid hormones appear to modulate the dimerization of two
proteins which, when dimerized properly, turn on the transcription of
genes which presumably encode for proteins which mediate the
effects observed for the thyroid hormones.
As part of a negative feedback mechanism, the anterior pituitary has
one of the highest densities of thyroid hormone receptors. T4 and T3
cause a decrease in the levels of TSH.
Heart and skeletal muscle also have high amounts of thyroid hormone
receptors, though there are few in the gonads.
The thyroid hormone receptor in the nucleus is a nonbasic protein
with a MW of 60-65,000. The receptor, influenced by the bound
hormone, binds to DNA and regulates mRNA transcription.
Agonists
Agonists are used to treat hypothyroidism (i.e.
myxedema, infant cretinism, or simple goiter).
T4 and T3 are used to replace natural production of
the hormones if loss is due to disease or medical
intervention.
T4 has a 7 day plasma half-life while T3 has a 1.3 day
half-life.
Since most of T3 comes from T4 through metabolism
in the peripheral tissues, particularly the liver (i.e.
thyroid independent T3 formation),
T4 is often prescribed because of its longer half-life.
Liver dysfunction can effect T3 levels. T3 is
considered the active thyroid hormone.
Inhibitors

Inhibitors are used to treat hyperthyroidism.


The antagonists block the thyroid peroxidase
which catalyzes both the incorporation of
iodine into the tyrosine residues and the
coupling of the outer phenol ring to the inner.
Presently, two kinds of thioureylenes are used
clinically to inhibit the thyroid peroxidase.
One is Propylthiouracil (PTU), the other is a
mercaptoimidazole.
Enzyme immunoassays (EIAs) for Antigen
Detection
Direct EIAs have four steps:
Antigen-specific antibody is attached to a solid-
phase surface (plastic beads)
Test specimen is added, which may or may not
contain the antigen
An enzyme-labeled antibody specific to the antigen
is added (conjugate)
Chromogenic substrate is added, which in the
presence of the enzyme, changes color. The amount
of color that develops is proportional to the amount
of antigen in the test specimen.
Direct EIA

.
ELISA (Enzyme-Linked Immunosorbent
Assay)

Is a useful and powerful method in


estimating ng/ml to pg/ml ordered materials
in the solution, such as serum, urine and
culture supernatant.
An ELISA plate

.
ELISA

Purified,(Hormone) antigens pre-coated onto an ELISA


platePatient serum which contains antibodies.
Antibodies raised against the hormone will bind to the
antigens on the plate.
Anti-immunoglobulin coupled to an enzyme is introduced.
This is the second antibody, and it binds to first antibodies.
Chromogen or substrate which changes color when cleaved
by the enzyme attached to the second antibody
Radioimmunassay (RIA),
In radioimmunassay (RIA), a fixed concentration of labeled tracer
antigen is incubated with a constant amount of antiserum such that the
concentration of antigen binding sites on the antibody is limiting, for
example, only 50% of the total tracer concentration may be bound by
antibody.
If unlabeled antigen is added to this system, there is competition
between labeled tracer and unlabeled antigen for the limited and
constant number of binding sites on the antibody, and thus the amount
of tracer bound to antibody will decrease as the concentration of
unlabeled antigen increases.
This can be measured after separating antibody-bound from free tracer
and counting either the bound fraction, the free fraction or both.
A calibration or standard curve is set up with increasing amounts of
known antigen, and from this curve the amount of antigen in the
unknown samples can be calculated.
RIA