Clinical Endocrinology (Gunn)

Question 1

What are the features of diabetes?

Answer 1

• key sign of diabetes = extreme wasting
• diabetes = variety of disorders characterised by polyuria
• pathologically an autoimmune disease
• over time, there is infiltration of entire islet, which is progressively destroyed

Question 2

Describe the Time Course of T1D

Answer 2

• non-linear decline of β-cell mass over time + development of autoantibodies associated with hyperglycaemia
• time course of T1DM is slow
• at time of patient presentation, it is well advanced for a long period of time (80-90% of β cells lost)
• honeymoon phase = period of time (~9 months) following diabetes diagnosis when b cells can still produce enough insulin for blood glucose control (no hyperglycaemia)
• immunological response to T1D is cyclic
• ↑ autoreactive effector T cells are controlled by ↑ regulatory T cells
• over time gradual disequilibrium, leading to number of autoreactive T cells surpassing regulatory T cells
• therefore, regulatory T cells can no longer control autoreactive effector T cell responses
• this leads to declined pancreatic islet function
• relative β-cell proliferation ↑ in a cyclical fashion over time
• inflammatory process of pancreatic islets may enhance b-cell proliferation and antigenic presentation, ultimately leading to generation of more effector and regulatory T cells
• in addition, as b-cell mass ↓, pressure on each b-cell to produce insulin ↑, which may be sufficient to alter recognition of b-cells by immune system and to alter their ability to regenerate and ↑ insulin production

Question 3

Describe the Carbohydrate Metabolism

Answer 3

• glucose and lipid metabolism are regulated by:
  
  • insulin (glucose uptake)
• And Counter-regulatory hormones
  
  • glucagon,
  • catecholamines,
  • cortisol,
  • growth hormone (glucose release)

Question 4

What are the actions of insulin?

Answer 4

Stimulates

• glucose uptake in muscle and adipose tissue
• glycolysis
• glycogen synthesis
• protein synthesis (muscle wasting in DM)
• uptake of ions (especially K+ and PO₄^3-)

Inhibits

• gluconeogenesis (liver and kidneys)
• glucogenolysis
• lipolysis
• ketogenesis
• proteolysis

Question 5

Describe the Blood Glucose and Insuline Concentrnation over 24 hours

Answer 5

• Rise and fall of insulin concentration is tightly matched with rise and fall of glucose concentration.
  • After meal, there is increased blood glucose, followed immediately by increased insulin in blood.
  • Insulin is slightly elevated between meals.
• After dinner (last meal of night), there is a steady decline in glucose concentration, which is coupled with decreased insulin.
  • Cortisol also decrease overnight due to diurnal rhythm associated with cortisol
  • This is mirrored by diurnal rhythm associated with insulin.

Question 6

Describe the Effect Of Insulin On Glucose Uptake/Metabolism

Answer 6

Insulin binds to insulin receptor (1), which in turn starts many protein activation cascades (2). These include:

• Translocation of GLUT4 transporter to plasma membrane and influx of glucose (3)
• Glycogen synthesis (4)
• Glycolysis (5)
  
  • metabolic pathway that converts glucose into pyruvate,
• Fatty acid synthesis (6)

This o_ccurs in liver and skeletal muscle,_ but not in brain, since insulin cannot cross BBB.

• This is a mechanism in which brain protects itself from hypoglycaemia.
• Glucose is constantly delivered to brain, and brain glucose only decreases when blood glucose decreases.

Question 7

How do we make Ketons?

Answer 7

2 Step process (1st is in the Fat)

1) Fat and Liver

• in fat, production of ketones = mobilising fat and stimulating β-oxidation (stimulated by glucagon)
• in adipose tissue, there are triglycerides stored in the adipocytes
• lipolysis breaks down triglycerides into non-esterified FAs + glycerol
  
  • via hormone-sensitive lipase
  • lipase inhibitor is insulin
  • lipase stimulators = glucagon, corticosteroid, ACTH, catecholamine, GH
  • after lipolysis, NEFA + glycerol released into blood, taken up by hepatocytes for ketone oxidation
  • NEFAs are esterified into esterified fatty acids (EFA) in liver
  • esterified fatty acids (EFA) can either:
  • EFA + glycerol as triglycerides is stored in hepatocyte, or exported to bloodstream via VLDL
  • or EFAs are converted into acetyl-CoA via β-oxidation in mitochondria, acetyl-CoA can enter Kreb’s cycle or become ketone
• once ketones are produced, they are exported from hepatocyte to enter blood there they can be oxidised by peripheral tissues

2) Liver

• ↑ hepatic fatty acid oxidation to accelerate ketone production when:
• insulin deficiency (activates lipolysis, ↑ plasma FFA then ↑ liver FA)
• and excess counter-regulatory hormones (glucagon ↑ liver carnitine and ↓ malonyl-CoA, which leads to activation of carnitine acetyltransferase)

3) Mitochondria

• FFAs are converted into acetyl-CoA
• acetyl-CoA is converted into acetoacetyl-CoA via β-ketothiolase
• acetoacetyl-CoA is → into β-hydroxy-β-methylglutaryl-CoA (HMG-CoA) via HMG-cosynthase
• HMG-CoA is converted into acetoacetate via HMG-CoA lyase
• acetoacetate (acid) is in equilibrium with β-hydroxybutyrate (acid) and acetone (not acid)
• acidosis (more H+) balance towards β-hydroxybutyrate
• β-hydroxybutyric acid and acetoacetic acid dissociate completely (form excess H+)

Excess H⁺ + HCO₃^- ® CO₂ + H₂O, therefore ¯ [HCO₃^-]

• ketone bodies circulate as anions, which ↑ anion gap

Question 8

Why does Diabetic Ketoacidosis result in an increased anion gap?

Answer 8

• β-hydroxybutyric acid and acetoacetic acid dissociate completely (form excess H+)

Excess H⁺ + HCO₃^- ® CO₂ + H₂O, therefore ¯ [HCO₃^-]

• ketone bodies circulate as anions, which ↑ anion gap
  
  • anion gap is Na⁺ – (Cl^– + HCO₃^-)
  • normal anion gap is 12 2mmol/L
• in diabetic ketoacidosis, bicarbonate is replaced by ketone bodies (β-hydroxybutyric acid and acetoacetic acid),
• ↓ HCO3- approximates ↑ anion gap

Question 9

Describe the Sodium and Chloride Balance in Diabetic Ketoacidosis

Answer 9

• if β-hydroxybutyric acid is dissociated, and H+ → to CO₂ and H₂O,
  
  • Na-β-hydroxybutyrate and Cl- remains.
• Na-β-hydroxybutyrate lost in urine as salt, proportional to GFR (diagnosis = urine dipstick for ketones)
• for chloride:
  
  • in short term, net loss of Na+ but no loss of Cl- in ECF
  • after clinical treatment of DKA, volume replacement with 0.9% NaCl -> salt loading to replace lost Na+, large increase in Cl- -> normal Na and hyperchloremia (not problematic corrected over time)

Question 10

Describe the issues with Urine Ketone for Acetoacetate test

Answer 10

Test: Urine Ketones for Acetoacetate (ACAC) – urine dipstick

• ketone strips use sodium nitroprusside reaction to produce a purple colour in presence of acetoacetate (ACAC)
  
  • if reagent contains glycine in addition to sodium nitroprusside, acetone can be detected
  • it doesn’t detect ß-hydroxybutyrate (BOHB)
• normally, serum BOHB:ACAC ~ 1:1
• in acidosis, serum _BOHB:ACAC = 1.3:1 to 5.5:1! (_in favour of BOHB)
• plasma or urine ACAC concentration alone underestimate severity of ketonemia, especially in DKA!

Question 11

Comapre the bedside tools vs Lab assay for b-OH-Butyrate (BOHB)

Answer 11

There are bedside kits that can measure [BOHB] and [glucose]. These are screening but not diagnostic tool. Measurement of [BOHB] in bedside measure is a lot less than laboratory assay.

• For high [BOHB], there is steady ¯[BOHB] in laboratory assay after insulin infusion. However, bedside kit doesn’t show ¯[BOHB] until ~8hr after treatment is initiated.
• For low blood [BOHB] within physiological limits, there is steady fall with both measures. This highlights that bedside kit is screening but not diagnostic tool. It works well with low [BOHB].

Pure infusion of insulin results in reduced formation of ketones over course of several hours.

Question 12

What are the current vs suggested criteria to diagnose DKA?

Answer 12

Test: Criteria to Diagnose DKA (All Three Needed)

Current

1. Hyperglycemia (blood glucose >11mmol/L);
2. Venous pH <7.3 or bicarbonate <15mmol/L;
3. Presence of ketonemia or ketonuria

Suggested

1. Hyperglycemia (blood glucose >11mmol/L);
2. Venous pH <7.3
3. Serum BOHB >3mmol/L

Question 13

Diabetic ketoacidosis requires both ………….

Answer 13

Diabetic ketoacidosis requires both relative lack of insulin and physiological stress.

Question 14

Describe the relationship between Hormones and DKA

Answer 14

• Insulin (anabolic) is responsible for
  
  • (1) glucose used for energy substrate or stored as glycogen;
  • (2) protein formation;
  • (3) fats stored as triglycerides
• Counterregulatory hormones (catabolic) is responsible for
  
  • (1) glycogenolysis;
  • (2) proteolysis®gluconeogenesis;
  • (3) lipolysis -> FFA + ketone bodies

Question 15

If DKA is bad, how is a girl with extreme wasting alive?

Answer 15

Reasons of Extremely Wasted Girl Being Alive (If DKA Is Bad)

DKA has the following three requirements:

• Lack of insulin
• Physiological stress (to active counter-regulatory system)
• Fat (if no TGs, no NEFAs, no ketone bodies can be made)

This girl has lack of insulin, but no stress or no fat. Therefore, there are no ketone production for DKA (i.e. starvation diet before discovery of insulin). She is starving to death but not DKA!

Question 16

What happens when you have dietary intake of Potassium via the gut?

Answer 16

Roughly 2% K+ is in ECF (70mEq). Majority 98% K+ is in ICF (3300mEq), which are stored in muscle and non-muscle cells.

Dietary intake of potassium is 80mEq via gut, which is more than total K+ in ECF.

• There is immediate shift in/out of ECF (highly regulated).
  
  • ECF serum concentration of K+ is 4.2mmol/L (3.5-5mmol/L).
    
    • Extracellular K+ pool is 14L (4.2 14 = 59mmol/L)
    • One meal of fruit/meat ~ 40 to 70mmol! (70mmol/14L = 5mM!)
    • If not regulated, s[K] rise from 4.2 to 9.2 mmol/L via single meal (lethal dose of cardiac arrest)
  • ICF concentration of K+ is ~140mmol/L.
• There is late excretion via stool (9mEq) and urine (70mEq) daily.

Feedback vs Feedforward

Feedback is effective, but s_low and overshoot_ (K+ intake, takes time for high ECF s[K+] before responding, can be lethal)

Feedforward is r_apid and anticipator_y (K+ intake, gut sensors bypass ECF to trigger K+ secretion, keep K+ stable)!

Question 17

What controls K after a meal?

Answer 17

Feedback vs Feedforward

Feedback is effective, but slow and overshoot (K+ intake, takes time for high ECF s[K+] before responding, can be lethal)

Feedforward is rapid and anticipatory (K+ intake, gut sensors bypass ECF to trigger K+ secretion, keep K+ stable)!

Regulations of Potassium

Feedforward Control of K+ After Meal

K+ control is mainly via insulin -> Na/K ATPase -> K⁺ uptake

• This is not linked to glucose uptake (GLUT4 does not affect K⁺ uptake!)
• Consumption of a meal containing glucose or amino acids release incretin from group, which facilitate insulin release from pancreas even before absorption.
• This stimulates K⁺ shift, independent of [K⁺].

There is some evidence of K⁺ control via glucagon and cAMP

• Protein rich meal -> increased [glucagon] + [cAMP]
• Glucagon portal vein infusion -> increased transtubular [K⁺] gradient + increased GFR -> Twice K⁺ excretion

There is possibility of addition gut factor?

Feedback Control To Increased [K+]

Feedback mechanism is backup to prevent major changes (for unanticipated changes).

• Aldosterone -> stimulates Na/K ATPase -> ­K⁺ uptake by cells (longer term shift)
• Aldosterone -> inserts K⁺ channel of apical membrane in principal cells -> ­renal K⁺ excretion
  
  • Aldosterone also affects muscle cells

Other Controls For K+ Gradient

In sympathetic system via b2 adrenergic receptors:

• b2 stimulation (NA/A) -> shift K⁺ from ECF to ICF -> ¯s[K⁺]
• Beta blockers tend to increase s[K⁺] by inhibiting b2 adrenergic stimulation

Acidosis -> ?inhibit Na/K ATPase? (does not work via Na/H exchanger) -> increases K⁺ loss from cells -> ¯s[K⁺]

Cell lysis (e.g. muscle, red cells) -> shift K⁺ from ICF to ECF -> ­s[K⁺]

Other Shifts Increasing s[K+]

Exercise will lead to increased serum potassium:

• Physiological response of contraction leads to K⁺ release to ECF -> ­s[K⁺]
  
  • Normally balanced by adrenalin
• Combined with beta-blocker or low insulin -> ­­­s[K⁺] -> cardiac arrest?

I_ncreased ECF osmolarity,_ e.g. diabetes mellitus, will lead to increased serum potassium:

• Hyperglycaemia -> increased ECF osmolartiy -> water out of cells (dehydration) -> increased ICF [K⁺] -> (drives K⁺ out to ECF via gradient) -> increased s[K⁺]
• The reverse is also correct (tonicity of cells)

Question 18

Describe the K+ within the glomerular filtration

Answer 18

Potassium Regulation in Kidney

Glomerular Filtration

K+ in kidney is all about early reabsorption and late secretion.

Amount of K+ that is filtered by glomerulus is determined by GFR

• GFR is in part determined by size of the patient.
• Because GFR is normally constant for a patient, the K+ filtered into the tubular part of the nephron is also constant.

Early Reabsorption

Proximal tubule reabsorbs ~65% K+

L_oop of Henle reabsorbs ~25-30% of K+_

• On apical membrane lining loop of Henle, there is Na/K/2Cl transporter for active reabsorption,
• On apical membrane lining loop of Henle, there is also K+ channels that allowed K+ to leak back out into tubular lumen.

Late Secretion

• K+ delivered to distal convoluted tubule is 4% (~31mmol/day). However, we excrete ~92mmol/day.
• Therefore, difference is the amount of active K+ secretion by distal convoluted tubule and collecting duct. Most variation in K+ excretion happens in the DCT and CD.

Reabsorption vs Secretion

Renal excretion is faster and greater than retention

• Kidney is able to excrete very large amounts of K+ (maximum secretion can be >GFR in healthy individuals)
• Kidney is slow to conserve K+. Fractional K+ excretion can be reduced to ~2% of filtered load (significant!). Therefore, it can result in K+ depletion (hypokalemia) if K intake is restricted.

Question 19

Describe the effects of Aldosterone

Answer 19

Principal Cells of _Collecting Ducts (_Aldosterone)

Aldosterone is a steroid hormone that _binds to receptor in principal cell nucleus to stimulate affec_t. Its effects are relatively slow (6-8 hours).

• Early effects are thru upregulating existing ENaC and ROMK channels on apical/luminal membrane, also NKA on basolateral membrane (create electrochemical gradient).
• Later/longer effects are synthesis of new pumps and channels (ENaC, ROMK, NKA).

There are three key factors determine K+ secretion:

• Activity of Na/K ATPase on basolateral membrane (Na+ in, K+ out)
• Permeability of apical/luminal membrane
• The electrochemical gradient from lumen to blood

Question 20

What are the three key factors determine K+ secretion?

Answer 20

• Activity of Na/K ATPase on basolateral membrane (Na+ in, K+ out)
• Permeability of apical/luminal membrane
• The electrochemical gradient from lumen to blood

Question 21

What Controls Distal Secretion of K+?

Answer 21

Summary

There is increased distal K+ secretion if:

• Increased s[K+]
• Increased distal tubular flow rate
• Increased aldosterone

Note that alkalosis increases K+ secretion (but clinically uncommon), and acidosis decreases K+ secretion.

Increased s[K+] (Increased K+ Secretion)

As there is increased s[K+], there is a proportional increase in aldosterone secretion, thus excreting more K+ in urine.

• The relationship between aldosterone and urinary K+ excretion is linear.
• The relationship between extracellular/serum potassium concentration (s[K+]) and urinary K+ excretion is not linear. There is non-linear increased urinary K+ excretion when s[K+]>4.1mM (i.e. greater than mean [K]). This is probably caused by:
  
  • Direct stimulation of Na/K ATPase (NKA), which generates a higher gradient (with less back-leak of K+ into cells).
  • And stimulation of aldosterone secretion (caused increased s[K+])

Increased Distal flow (Increased K+ Secretion)

• _Increased flow (_due to diuretics, high urine osmalarity in diabetic and hyperglycaemic patient) -> decreased [K+] in tubular fluid (washout) -> increased K+ gradient between lumen and blood -> more K+ secreted -> ¯s[K⁺]
• Decreased flow -> ­s[K⁺] in severe renal impairment

Question 22

There is increased distal K+ secretion if:

Answer 22

Increased s[K+]

Increased distal tubular flow rate

Increased aldosterone

Question 23

Describe how Increased s[K+] results in Increased K+ Secretion

Answer 23

Increased s[K+] (Increased K+ Secretion)

As there is increased s[K+], there is a proportional i_ncrease in aldosterone secretion,_ thus excreting more K+ in urine.

• The relationship between aldosterone and urinary K+ excretion is linear.
• The relationship between extracellular/serum potassium concentration (s[K+]) and urinary K+ excretion is not linear. There is non-linear increased urinary K+ excretion when s[K+]>4.1mM (i.e. greater than mean [K]). This is probably caused by:
  
  • Direct stimulation of Na/K ATPase (NKA), which generates a higher gradient (with less back-leak of K+ into cells).
  • And stimulation of aldosterone secretion (caused increased s[K+])

Question 24

Describe how Increased Distal flow results in Increased K+ Secretion

Answer 24

• Increased flow (due to diuretics, high urine osmalarity in diabetic and hyperglycaemic patient) -> decreased [K+] in tubular fluid (washout) -> increased K+ gradient between lumen and blood (osmotic diuresis?)-> more K+ secreted -> ¯s[K⁺]
• Decreased flow -> ­s[K⁺] in severe renal impairment