Insulin

1. INSULIN : SECRETION AND
ITS ROLE

2. History
◦ On July 27, 1921, Dr. Frederick Banting, a
Canadian surgeon and Charles Best, a
medical student, successfully isolated the
hormone insulin for the first time.
◦ On January 11, 1922, 14-year-old Leonard
Thompson became the first person to receive
an insulin injection as treatment for diabetes.
◦ The Nobel Prize in Physiology or Medicine
1923 was awarded jointly to Frederick Grant
Banting and JJR Macleod “for the discovery
of insulin.”
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3. Introduction
◦ Insulin is a polypeptide containing 51 Amino
acid .
◦ Two chains of amino acids, A chain ( 21
Amino acids ) and B chain (30 Amino acids),
linked by disulphide bridges.
◦ It is secreted from beta cells of Pancreas (
Molecular weight of 5808Da).
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4. ◦ Pancreatic beta cells co-secrete islet amyloid
polypeptide (IAPP) or amylin, a 37-amino-acid
peptide.
◦ The IAPP is the major component of the amyloid
fibrils found in the islets of patients with type 2
diabetes, and an analogue is sometimes used in
treating type 1 and type 2 DM.
◦ Human insulin is produced by recombinant
DNA technology.
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5. • Insulin circulates almost entirely in an unbound form.
• It has a plasma half- life of about 6 minutes, it is mainly cleared from
the circulation within 10 to 15 minutes.
• It is degraded by the enzyme insulinase mainly in the liver, to a lesser
extent in the kidneys and muscles.
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6. Insulin- Chemistry and Synthesis
• Synthesized in beta cells , by translation of
RNA ( in ribosomes attached to the
endoplasmic reticulum) to form
preproinsulin , (86-AA ),( molecular weight :
11,500 Da).
• Processed by removing the amino-terminal
peptide, in the endoplasmic reticulum to form
a proinsulin (molecular weight : 9000 Da ),
consisting of 3chains of peptides, A, B, and
C.
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7. ◦ Proinsulin : cleaved in the Golgi apparatus to
form insulin, : A and B chains connected by
disulfide linkages, and the C chain peptide, called
connecting peptide (C peptide).
◦ The insulin and C peptide are packaged in
secretory granules.
◦ The proinsulin and C peptide have virtually no
insulin activity.
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8. Significance in Diagnosis and
Assessment
◦ C-peptide is cleared more slowly than insulin, measured by
radioimmunoassay in insulin- treated diabetic patients.
◦ Useful marker of insulin secretion : discrimination of endogenous
and exogenous sources of insulin
◦ Patients with type 1 diabetes will have decreased levels of C
peptide.
◦ Elevated levels of serum proinsulin are indicative of beta cell
dysfunction.
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9. Insulin secretion
◦ Glucose : key regulator of insulin secretion by the pancreatic beta cell .
◦ Glucose levels >3.9 mmol/L (70 mg/dL) stimulate insulin synthesis.
◦ Begins with glucose transport into the beta cell by a facilitative glucose
transporter .
◦ Glucose phosphorylation by glucokinase : rate-limiting step that controls
glucose-regulated insulin secretion.
◦ Further metabolism via glycolysis generates ATP, which inhibits the
activity of an ATP-sensitive K+ channel.
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10. ◦ This channel consists of two
separate proteins:one is the binding
site for certain oral hypoglycemics
(e.g., sulfonylureas, meglitinides);
the other is an inwardly rectifying
K+ channel protein (Kir6.2).
◦ Inhibition of this K+ channel
induces beta cell membrane
depolarization, which opens
voltage-dependent calcium channels
(leading to an influx of calcium) and
stimulates insulin secretion.
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11. ◦ pulsatile pattern of hormone release,
◦ Small secretory bursts: every 10 min,
Greater amplitude oscillations of about
80–150 min.
◦ Insulin is produced @ 0.2 – 0.5 U/ kg /
day .
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12. ◦ Glucagon-like peptide-1 (GLP-1) and glucose-dependent
insulinotropic peptide (GIP) are incretin hormones .
◦ Bind specific receptors on the beta cell to stimulate insulin
secretion through cyclic AMP production, but have this
effect only when the blood glucose is above the fasting level.
◦ Incretin hormones also suppress glucagon production and
secretion.
◦ Incretin analogues or pharmacologic agents that prolong the
activity of endogenous GLP-1 are used therapeutically in type
2 DM.
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14. Activation of target cell receptors by insulin and
the resulting cellular effects
◦ Insulin must first bind with and activate a
membrane receptor protein (molecular weight
300,000 Da).
◦ The insulin receptor is a combination of four
subunits held together by disulfide linkages:
2 alpha subunits :( outside membrane )
2 beta subunits : that penetrate through into the
cell cytoplasm.
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15. ◦ Insulin binds with the alpha subunits, causing beta subunits to become
autophosphorylated.
◦ Insulin receptor is an enzyme-linked receptor.
◦ Autophosphorylation of the beta subunits activates tyrosine
kinase, which causes phosphorylation of multiple other intracellular
enzymes, including a group called insulin- receptor substrates (IRS).
◦ In this way, insulin directs the intracellular metabolic machinery to
produce the desired effects on carbohydrate, fat, and protein
metabolism.
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16. Effects of Insulin stimulation-
◦ Within seconds after insulin binds with its membrane receptors, cells
markedly increase their uptake of glucose.
◦ This action is especially true of muscle cells and adipose cells, it is not
true of most neurons in the brain.
◦ The increased glucose transported into the cells is immediately
phosphorylated and becomes a substrate for all the usual carbohydrate
metabolic functions.
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17. The cell membrane becomes more permeable to many of the amino acids,
potassium ions, and phosphate ions, causing increased transport of these
substances into the cell.
1. Slower effects (10 to 15 minutes ) to change the activity of many more
intracellular metabolic enzymes.
2. Much slower effects occur for hours and even several days. These result
from changed rates of translation of messenger RNAs to form new
proteins .
3. Still slower effects from changed rates of transcription of DNA in the
cell nucleus.
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18. Effect of Insulin on Carbohydrate Metabolism-
◦ Immediately after a high- carbohydrate meal, there is rapid secretion of
insulin.
Insulin Promotes Muscle Glucose Uptake and Metabolism .
◦ During much of the day, muscle tissue depends on fatty acids for its
energy.
◦ This is because the normal resting muscle membrane is only slightly
permeable to glucose, except when the muscle fiber is stimulated by
insulin.
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19. ◦ However, under two conditions the muscles do use large amounts of glucose.
1. During moderate or heavy exercise.
This does not require large amounts of insulin because muscle contraction
increases translocation of glucose transporter 4 (GLUT 4) to the cell membrane,
which, facilitates diffusion of glucose into the cell.
2. During the few hours after a meal.
At this time the blood glucose concentration is high and the pancreas is
secreting large quantities of insulin. It causes the muscle cell to use glucose
preferentially over fatty acids.
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20. Quantitative Effect of Insulin to Facilitate Glucose
Transport Through the Muscle Cell Membrane
◦ Curve “control” shows that the
intracellular glucose concentration
remained almost zero despite increased
extracellular glucose concentration up to as
high as 750 mg/100 ml.
◦ Curve labeled “insulin” show that the
intracellular glucose concentration rose to
as high as 400 mg/100 ml when insulin was
added.
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21. ◦ Thus, insulin can increase the rate of transport of glucose by at
least 15- fold.
◦ One of the most important effects of insulin is to store absorbed
glucose in the liver in the form of glycogen.
◦ Then, between meals, when the blood glucose concentration begins
to fall, insulin secretion decreases rapidly and the liver glycogen is
split back into glucose, which is released back into the blood to
keep the glucose concentration from falling too low.
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22. The mechanism by which insulin causes glucose uptake and storage in
the liver ( during well fed state ) includes several almost simultaneous
steps:
1. Insulin inactivates liver phosphorylase, the principal enzyme that
causes liver glycogen to split into glucose. This inactivation prevents
breakdown of the glycogen that has been stored in liver cells.
2. Insulin enhances the enzyme glucokinase, that causes the initial
phosphorylation of glucose after it diffuses into the liver cells.
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23. 3 .Once phosphorylated ,glucose cannot diffuse back through the cell
membrane.
4 . Insulin increases the activities of enzymes that promote glycogen
synthesis, including especially glycogen synthase.
The net effect of all these actions is to increase the amount of glycogen
in the liver.
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24. Glucose is released from the liver between meals ,into the circulating
blood.
1. The decreasing blood glucose causes the pancreas to decrease its
insulin secretion.
2. The lack of insulin then reverses all the effects listed earlier for
glycogen storage
3. The lack of insulin activates the enzyme phosphorylase, which causes
the splitting of glycogen into glucose phosphate.
During fasting
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25. 4. The enzyme glucose phosphatase, now becomes activated by the lack
of insulin and causes the phosphate radical to split away from the
glucose, allowing the free glucose to diffuse back into the blood.
Thus, the liver removes glucose from the blood when it is present in
excess after a meal and returns it to the blood when the blood glucose
concentration falls between meals.
Ordinarily, about 60% of the glucose in the meal is stored in this way in
the liver and then returned later.
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26. Insulin Promotes Conversion of Excess Glucose Into
Fatty Acids and Inhibits Gluconeogenesis in the Liver
◦ Excess glucose entering the liver cells is converted into fatty acids.
◦ These fatty acids are subsequently packaged as triglycerides in very low
density lipoproteins( VLDL ) which are transported in the blood to
adipose tissue, and deposited as fat.
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27. Lack of Effect of Insulin on Glucose Uptake and
Usage by the Brain
◦ Insulin has little effect on uptake or use of glucose by brain.
◦ Most of the brain cells are permeable to glucose and can use glucose
without the insulin.
◦ The brain cells normally use only glucose for energy.
◦ When the blood glucose level falls too low, into the range of 20 to 50
mg/dl, symptoms of hypoglycemic shock develop, characterized by
progressive nervous irritability that leads to fainting, seizures, and even
coma.
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28. Effect of Insulin on Fat Metabolism
◦ The long- term insulin deficiency causes atherosclerosis, often leading
to heart attacks, cerebral strokes, and other vascular accidents .
◦ Insulin promotes fat synthesis and storage.
Insulin increases glucose utilization by most of the body’s tissues, which
automatically decreases fat utilization, thus functioning as a fat sparer.
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29. ◦ The following factors lead to increased fatty acid synthesis in the liver:
1. Insulin increases glucose transport into the liver cells. After the liver
glycogen concentration reaches 5% to 6%, further glycogen synthesis is
inhibited.
2. All the additional glucose entering the liver cells then becomes
available to form fat. The glucose is first split to pyruvate and the
pyruvate subsequently is converted to acetyl coenzyme A (acetyl-
CoA), the substrate from which fatty acids are synthesized.
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30. ,
3. Most of the fatty acids are then synthesized within the liver and used
to form triglycerides, the usual form of storage fat. They are
released from the liver cells to the blood in the lipoproteins.
4. Insulin activates lipoprotein lipase in the capillary walls of the
adipose tissue, which splits the triglycerides again into fatty acids, a
requirement for them to be absorbed into adipose cells, where they
are again converted to triglycerides and stored.
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31. Role of Insulin in Storage of Fat in the Adipose Cells
Insulin has two essential effects:
1. Insulin inhibits the action of hormone- sensitive lipase. Therefore,
release of fatty acids from adipose tissue into the circulating blood is
inhibited.
2. Insulin promotes glucose transport through cell membranes into fat
cells in the same way that it promotes glucose transport into muscle cells.
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32. ◦ Some of this glucose, also forms α- glycerol phosphate. This
substance combines with fatty acids to form triglycerides, the storage
form of fat in adipose cells.
◦ Therefore, when insulin is not available, even storage of the large
amounts of fatty acids transported from the liver in lipoproteins is
almost blocked.
◦ All aspects of fat breakdown and its use for providing energy are
greatly enhanced in the absence of insulin.
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33. Lipolysis of Stored Fat and Release of Free Fatty Acids
◦ In the absence of insulin, all the effects of
insulin noted earlier that cause storage of fat
are reversed.
◦ The enzyme hormone-sensitive lipase in the fat
cells gets activated.
◦ Hydrolysis of triglycerides, releasing fatty acids
and glycerol into the circulating blood.
◦ These free fatty acids then become the main
energy substrate to all tissues of the body except
the brain.
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34. Effect on Plasma Cholesterol and Phospholipid Concentrations
◦ Excess of fatty acids are converted into phospholipids and
cholesterol, two of the major products of fat metabolism.
◦ These two substances, are then discharged into the blood in the
lipoproteins.
◦ This high lipid concentration—especially the high concentration of
cholesterol—promotes development of atherosclerosis in people
with severe diabetes.
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35. Ketosis
◦ Insulin deficiency also causes excessive amounts of acetoacetic acid to
be formed in liver cells.
◦ In the mitochondria, beta oxidation of the fatty acids proceeds rapidly,
releasing extreme amounts of acetyl- CoA.
◦ This excess acetyl- CoA is then condensed to form acetoacetic acid,
which is then released into the circulating blood.
◦ At the same time, the absence of insulin also depresses utilization of
acetoacetic acid in peripheral tissues.
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36. Effect of Insulin on Protein Metabolism and Growth
◦ Insulin Promotes Protein Synthesis and Storage.
◦ It stimulates transport of many of the amino acids into the cells.
◦ It “turns on” the ribosomal machinery.
◦ It inhibits catabolism of proteins, thus decreasing the rate of
amino acid release from the cells, especially from muscle cells.
◦ It promotes formation of protein and prevents degradation of
proteins.
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37. Administration of either growth
hormone or insulin one at a time causes
almost no growth. However, a
combination of these hormones causes
dramatic growth.
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38. Feedback Relation Between Blood Glucose Concentration and
the Insulin Secretion Rate
◦ As blood glucose concentration rises above 100
mg/dl of blood, secretion of insulin rises rapidly,
reaching a peak some 10 to 25 times the basal
level at blood glucose concentrations between
400 and 600 mg/100 ml.
◦ Furthermore, the turnoff of insulin secretion is
almost equally as rapid, occurring within 3 to 5
minutes after a reduction in blood glucose
concentration back to the fasting level.
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39. Role of Insulin in switching between carbohydrate and
lipid metabolism
◦ The signal that controls this switching mechanism is principally the
blood glucose concentration.
◦ When glucose concentration is low, insulin secretion is suppressed, and
fat is used almost exclusively for energy everywhere except in the
brain.
◦ When the glucose concentration is high, insulin secretion is stimulated,
and carbohydrate is used instead of fat.
◦
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40. ◦ Growth hormone and cortisol are secreted in response to
hypoglycemia, and both inhibit cellular utilization of glucose while
promoting fat utilization.
◦ Epinephrine -increases plasma glucose concentration during
periods of stress when the sympathetic nervous system is excited.
◦ Epinephrine -increases plasma fatty acid concentration at the same
time.
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41. Effects of Anesthesia on insulin secretion and action -
◦ Surgical stress with inhalation anesthesia is associated with
increased circulating catecholamines, hyperglycemia, and
impaired insulin secretion.
◦ Inhalation anesthesia - suppress basal insulin levels and the
insulin response to intravenous glucose.
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42. ◦ In postoperative period, suppressed insulin secretion was found
to be correlated with elevated plasma epinephrine concentrations
mediated by adrenergic mechanisms.
◦ Impaired insulin secretion have two etiologies-
1. the type of anesthesia used
2. adrenomedullary stimulation due to pain.
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43. Summary
◦ Insulin is a polypeptide linked by disulphide bonds secreted by Beta
cells of Pancreas in the form of pre -pro insulin which is cleaved to
pro insulin releasing c -peptide which is useful in diagnostic tests.
◦ Glucose induces insulin secretion from pancreas wherein blockage
of K+ channels induce opening of voltage gated Ca+2 channels.
◦ GLP1 and GIP are incretin hormones involved in insulin production
during fasting.
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44. ◦ Insulin has an enzyme linked receptor, by autophosphorylation of
beta subunits forms Insulin Receptor Substrate ( IRS )
phosphorylation and further cellular effects.
◦ Insulin – increases permeability of cell to glucose,
-glucose uptake and storage in liver,
- promotes conversion of excess glucose into fatty acid,
- promotes protein synthesis and storage.
◦ Deficiency of insulin increases plasma cholesterol and phospholipid
concentration causing atherosclerosis, may cause acidosis and
ketosis due to excess use of fat.
◦ Type of anesthesia and surgical stress affect the secretion of Insulin
which is to be managed accordingly.
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45. References -
◦ Guyton’s And Hall textbook of Medical Physiology - 2nd – South Asia Edition
◦ Harrison’s Principles of Internal Medicine - 20th edition
◦ Ganong’s review of Medical Physiology – 26th edition
◦ Research publications from https://www.ncbi.nlm.nih.gov.in
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