Endocrinology

Question 1

What is endocrinology?

Answer 1

• Endocrinology is the branch of physiology and medicine concerned with endocrine glands and hormones
• Endocrine glands, are specialised organs that secrete chemical mediators (hormones) into the circulation.
• Hormones(a regulatory substance of cellular activity) act at sites remote from the site of production
• Target tissues/organs dictated by expression of receptors for the hormone

Question 2

What is the difference between endocrine, autocrine and paracrine?

Answer 2

Endocrine - act on remote tissues (main focus in clinical biochem)
Autocrine - mediators act on the cell where they are produced
Paracrine - mediators act on adjacent cells

Question 3

What are the major functions of hormones?

Answer 3

• Homeostatic mechanisms in the body
• Energy production, utilisation and storage
• Growth and development
• Reproduction

Basically every major mechanism in the body.
- anything involving many of the body analytes, such as Calcium, used
for signalling.
- or anything involving energy production for example insulin regulation
of glucose.

Question 4

How are hormones classified?

Answer 4

They are classified based on structure.

Peptides: chains of among acids varying in length - e.g. ACTH, Prolactin, Insulin

Glycoproteins: AA with an attached sugar moiety throughout the length of the chain – e.g. LH, FSH, TSH

Steroids: three 6 sided rings and one 5 sided ring.- e.g. Cortisol, Testosterone, Vitamin D

Catechol amines: different analogous involving a Tryptophan with a benzene ring attached- Adrenaline, + Noradrenaline

Question 5

What is the mechanism of hormone action for proteins and amines?

Answer 5

Proteins and amines, cant enter the lipid membrane, so bind to cell surface receptors, which in turn triggers intracellular secondary messenger systems which propagate the signal into and throughout a cell e.g. Adrenaline

Intracellular 2nd Messenger Systems:
Cyclic AMP
Phospholipid systems, eg insulin
Intracellular calcium

Question 6

What is the mechanism of hormone action for steroids?

Answer 6

Steroid hormones can diffuse directly into cells and bind intracellular receptors. The steroid-receptor complex then moves into the nucleus and activates or suppresses specific genes.

Eg Vitamin D or testosterone are lipid soluble so can diffuse across the membrane into cells. Once inside they will bind to a steroid receptor, causing a conformational change that allows entry into the nucleus where it can bind to DNA and alter the expression of genes.

Question 7

What are the different kinetic rates of each hormone mechanism?

Answer 7

Steroid hormones can take hours to activate a response but by altering gene expression, steroids produce more long lasting effects. Whereas protein/amine hormones that act via a cell surface receptor can act very quickly (30s – 2mins) but have shorter effects.

The two different mechanisms have different kinetic rates. The peptide/amine mechanism has a very rapid action, but the effect is less long lived. Whereas the steroid mechanism can take take several ours to affect a gene in gene regulation, but they last a much longer time.

Question 8

How are endocrine responses modulated?

Answer 8

Sets of endocrine glands are usually organized into hierarchical loops that allow feed forward and feedback to regulate responses.

Can be short or long loops

Vast majority of endocrine pathways are self limiting – incorporate some form of negative feedback loop

Example:
Hypothalamus

Question 9

What are the major endocrine pathways in biochemistry?

Answer 9

1. Hypothalamus – Pituitary – Adrenal (HPA) axis
2. Hypothalamus – Pituitary – Gonadal axis
3. Hypothalamus – Pituitary – Thyroid axis
4. Anti Diuretic Hormone (ADH) – water balance

Many others including, Insulin Signalling Pathway, gastrointestinal tract, PTH/Calcium, Renin/Aldosterone, Catecholamines.

Question 10

What are the features of the hypothalamus and the pituitary?

Answer 10

At the core is the hypothalamus, involved in sensing and control mechanisms. The pituitary gland is directly below the hypothalamus, and is made up of two anatomically and endocrinologically distinct and different lobes.

At the front of the gland is the anterior lobe, linked to the hypothalamus via a network of capillary vessels. So when the hypothalamus wants to talk to the anterior pituitary it does so by secreting hormones into the capillary blood stream, that travel down and activate the lobe.

In contrast the posterior pituitary is connected to the hypothalamus by a series of nerves.

So the hypothalamus can interact with each lobe in different ways, this allows different levels of regulation. Patients with pituitary tumours can affect either the neuronal or capillary connections.

Summary:
Hypothalamus: key area of the brain involved in a wide range of homeostatic and metabolic processes.
- Exerts many of its effects through stimulation of the Pituitary gland.
- Stimulates the Anterior pituitary using peptides carried by capillaries.
- Stimulates the Posterior pituitary using neuronal signals

Question 11

What are examples of anterior pituitary hormones?

Answer 11

Multiple effects:
Adrenocorticotrophic hormone (ACTH) 
Growth hormone (GH)

Reproductive effects:
Follicle stimulating hormone (FSH)
Luteinising hormone (LH)
Prolactin

Metabolic effects:
Thyroid stimulating hormone (TSH)

Question 12

What are examples of posterior pituitary hormones?

Answer 12

Water balance effects:
Antidiuretic hormone (ADH)
AKA Arginine vasopressin (AVP)

Oxytocin (induced during labour)

Question 13

What hormones are involved in the HP Adrenal Major Endocrine Axis?

Answer 13

• Cortisol is a key stress response steroid hormone
• Stimulates glucose production and modulates the immune
  system
• stress causes CRH to be released from the hypothalamus, this travels down capillaries inducing ACTH release from the pituitary.
• ACTH reaches the adrenal glands and induces cortisol release.
• Cortisol can inhibit CRH release and to a lesser extent ACTH - regulates
  its own feedback loop

Hypothalamus: CRH

Question 14

What hormones are involved in the HP Gonadal Major Endocrine Axis?

Answer 14

• Different in men and women.
• GnRH (Gonadotrophin releasing hormone) release by the hypothalamus is pulsatile - hormones are released most in puberty, but women also have varied release during menstruated life cycle.

IN MEN:
- Testosterone regulates feedback loop.
Hypothalamus: GnRH

Question 15

What hormones are involved in the HP Thyroid Major Endocrine Axis?

Answer 15

• Thyroid Hormones T4 and T3 are key regulators of metabolic
  rate 
- TRH released from the Hypothalamus stimulates TSH 
- In turn stimulates thyroid hormone synthesis and release from the
  thyroid gland
• Neg feedback loop

Hypothalamus: TRH

Question 16

What hormones are involved in the ADH and Water balance Axis?

Answer 16

• Major axis regulating circulatory volume
• More complex pathway.
• Dehydration leads to high plasma osmolality, detected by
  hypothalamus, triggering thirst response and signalling to POSTERIOR
  pituitary to release ADH through neuronal connection (contrast
  to others).
• ADH goes round body and binds to collecting duct of kidneys
  stimulating greater reabsorption and less urine production, reducing
  plasma osmolality.
• Problem is lots of other stimuli can cause this, for example, drop
  of blood pressure, stress pain etc. These stimulants can override
  the work of the hypothalamus, so that unnecessary amounts of ADH
  are released from the pituitary, leading to water imbalance
  disorders.
• Parallel thirst pathway

                             Water intake \_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_                 
                                   \➕                                             |  
                                   thirst                                         ↓ ➖
                                       \➕                                         | Hypothalamus:             ⬆️plasma osmolality \_\_\_|

Question 17

What physiological factors affect hormone production?

Answer 17

Time of day
Time of month
Time of year
Age
Sex
Stress
Diet
Illness
Drugs

• Essential to consider these factors when investigating and monitoring
  disease - always check that the required info is supplied with the test
  request/on request form.

Question 18

How is immunoassay used to measure of hormones in Clin Biochemistry?

Answer 18

Uses a labelled antibody designed to specifically recognise a specific region (epitope) present on the target protein or molecule (antigen). Often only feasible if there is a unique antigen epitope

• competitive
• sandwich

Question 19

What are measurement considerations that must be accounted for when measuring steroids?

Answer 19

1. Are you confident your assay is measuring what you think?

   Many steroid hormones exists with very similar chemical structures, causing interference/mis-recognition in many commercial assays - falsely high or low levels
   - This is why steroid tend to be measured by a chemical detection
   approach, e.g. HPLC-UV – catecholamines, or mass spec (Vit D)
2. Are you measuring the correct ‘form’ of the hormone?

Many hormones exists in circulation as both protein-bound and unbound ‘free’ forms. Often only the ‘free’ form is biologically active and is the one we want to measure, however measuring ‘free’ hormones can be difficult and some assays may only measure the ‘Total’:
- ‘Total’ amount = [Free] + [bound]
- e.g. ‘bound forms’ of testosterone may be biologically inactive but all
assays commonly measure total testosterone. So in females we
estimate the free fraction using the Free Androgen Index (FAI)
calculation: FAI = 100 x [Total Testosterone]/SHBG
- For thyroid hormones we commonly measure and clinically interpret
(Total) TSH and Free T4 (FT4)
- Will often measure the binding proteins to help interpret results of free
hormone assays – e.g. SHBG (as above) or Thyroid Binding Globulin
(TBG)

3. Are you measuring the correct sample type ?

• Serum (most common)- but some compounds are very lability and
  quickly degraded in blood
• Lithium Heparin – ACTH, Insulin, Aldosterone
• EDTA – Gut hormones
• Urine – 24hr urine ‘free’ cortisol, metadrenalines, 5-HIAA
• Also – saliva e.g. cortisol (equivalent to the ‘free form’)

4. Is the patient on treatment ?

Some medications use the same or similar hormone to that produced endogenously. So results can be very misleading if not aware!
- e.g. Thyroid medication (Thyroxine) is the same as endogenous T4
- Commonly used steroids are derivatives of Cortisol, or of a similar
enough structure to give a falsely raised Cortisol level when measured
by immunoassay.

5. What is the timing of sample? -

Many physiological factors can affect the level – see previous slide.
- Stabilty. We don’t typically measure the Hypothalmic hormones as they are very unstable(short half lifes)!

Question 20

How are hormone results interpreted?

Answer 20

Often need to consider the entire hormone axis, as a single analyte result in isolation can be misleading.

 Pit Hormone End Hormone
End organ failure ↑ A ↓ B
Pituitary failure ↓ A ↓ B
End organ over activity ↓ A ↑ B
Pituitary over activity ↑ A ↑ B


A = Pituitary Hormone
B = End Organ Hormone

Question 21

What are dynamic function tests?

Answer 21

Investigation of hormone excess and deficient states, involves doing something to the axis and see how it responds.

? Hypofunction → perform a stimulation test (End organ isn’t producing the desired hormone effectively. Give (synthetic?) pituitary hormone and see if the end organ responds and produces its hormone.

? Hyperfunction → perform a suppression test (End organ is responding too much by overproducing end hormone. Give a stimulus that in a normal person would shipwreck off the end hormone, see if you get this appropriate response)

Why perform?
- Low hormone levels alone are not diagnostic.
- “Normal” hormone levels alone do not exclude disease.
- Pulsatile secretion and diurnal variations confuse interpretation of
baseline values - no static background for comparison.
- Can aid in differential diagnosis in hormone excess states.

→ Remember to consider results in the context of the entire axis

Question 22

How are dynamic function tests used to assess the HP Adrenal Major Endocrine Axis?

Answer 22

Suppression Test:
Give Dexamethasone in a Dexamethasone Suppression Test (DST) - investigation of XS Cortisol (Cushing’s)
Acts on hypothalamus to assess its endocrine function

Stimulation Test:
Give Synacthen (ACTH) in a Short Synacthen Test (SST) - investigation of low Cortisol (Addison’s)
Acts on adrenals to assess their endocrine function

Question 23

What are causes of water balance disorders?

Answer 23

ADH binds to collecting duct of kidneys in order to stimulate greater water reabsorption and less urine production, hence reducing plasma osmolality.

Water imbalance disorders can be caused when other stimuli result in inappropriate ADH production; for example blood pressure, stress, pain etc. These stimulants override the feedback loop involving the hypothalamus and pituitary.