Articles, Blog


October 12, 2019

Insulin is a peptide hormone, produced by
beta cells in the pancreas, and is central to regulating carbohydrate and fat metabolism
in the body. It causes cells in the skeletal muscles, and fat tissue to absorb glucose
from the blood. Insulin is a very old protein that may have
originated more than one billion years ago. The molecular origins of insulin go at least
as far back as the simplest unicellular eukaryotes. Apart from animals, insulin-like proteins
are also known to exist in Fungi and Protista kingdoms.
Insulin stops the use of fat as an energy source by inhibiting the release of glucagon.
Except in the presence of the metabolic disorder diabetes mellitus and metabolic syndrome,
insulin is provided within the body in a constant proportion to remove excess glucose from the
blood, which otherwise would be toxic. When blood glucose levels fall below a certain
level, the body begins to use stored sugar as an energy source through glycogenolysis,
which breaks down the glycogen stored in the liver and muscles into glucose, which can
then be utilized as an energy source. As a central metabolic control mechanism, its status
is also used as a control signal to other body systems. In addition, it has several
other anabolic effects throughout the body. When control of insulin levels fails, diabetes
mellitus can result. As a consequence, insulin is used medically to treat some forms of diabetes
mellitus. Patients with type 1 diabetes depend on external insulin for their survival because
the hormone is no longer produced internally. Patients with type 2 diabetes are often insulin
resistant and, because of such resistance, may suffer from a “relative” insulin deficiency.
Some patients with type 2 diabetes may eventually require insulin if dietary modifications or
other medications fail to control blood glucose levels adequately. Over 40% of those with
Type 2 diabetes require insulin as part of their diabetes management plan.
The human insulin protein is composed of 51 amino acids, and has a molecular weight of
5808 Da. It is a dimer of an A-chain and a B-chain, which are linked together by disulfide
bonds. Insulin’s name is derived from the Latin insula
for “island”. Insulin’s structure varies slightly between species of animals. Insulin from animal
sources differs somewhat in “strength” from that in humans because of those variations.
Porcine insulin is especially close to the human version. Gene
The preproinsulin precursor of insulin is encoded by the INS gene.
Alleles A variety of mutant alleles with changes in
the coding region have been identified. A read-through gene, INS-IGF2, overlaps with
this gene at the 5′ region and with the IGF2 gene at the 3′ region.
Regulation Several regulatory sequences in the promoter
region of the human insulin gene bind to transcription factors. In general, the A-boxes bind to Pdx1
factors, E-boxes bind to NeuroD, C-boxes bind to MafA, and cAMP response elements to CREB.
There are also silencers that inhibit transcription. Protein structure Within vertebrates, the amino acid sequence
of insulin is strongly conserved. Bovine insulin differs from human in only three amino acid
residues, and porcine insulin in one. Even insulin from some species of fish is similar
enough to human to be clinically effective in humans. Insulin in some invertebrates is
quite similar in sequence to human insulin, and has similar physiological effects. The
strong homology seen in the insulin sequence of diverse species suggests that it has been
conserved across much of animal evolutionary history. The C-peptide of proinsulin, however,
differs much more among species; it is also a hormone, but a secondary one. The primary structure of bovine insulin was
first determined by Frederick Sanger in 1951. After that, this polypeptide was synthesized
independently by several groups. The 3-dimensional structure of insulin was determined by X-ray
crystallography in Dorothy Hodgkin’s laboratory in 1969.
Insulin is produced and stored in the body as a hexamer, while the active form is the
monomer. The hexamer is an inactive form with long-term stability, which serves as a way
to keep the highly reactive insulin protected, yet readily available. The hexamer-monomer
conversion is one of the central aspects of insulin formulations for injection. The hexamer
is far more stable than the monomer, which is desirable for practical reasons; however,
the monomer is a much faster-reacting drug because diffusion rate is inversely related
to particle size. A fast-reacting drug means insulin injections do not have to precede
mealtimes by hours, which in turn gives diabetics more flexibility in their daily schedules.
Insulin can aggregate and form fibrillar interdigitated beta-sheets. This can cause injection amyloidosis,
and prevents the storage of insulin for long periods.
Synthesis, physiological effects, and degradation Synthesis
Insulin is produced in the pancreas and released when any of several stimuli are detected.
These stimuli include ingested protein and glucose in the blood produced from digested
food. Carbohydrates can be polymers of simple sugars or the simple sugars themselves. If
the carbohydrates include glucose, then that glucose will be absorbed into the bloodstream
and blood glucose level will begin to rise. In target cells, insulin initiates a signal
transduction, which has the effect of increasing glucose uptake and storage. Finally, insulin
is degraded, terminating the response. In mammals, insulin is synthesized in the
pancreas within the β-cells of the islets of Langerhans. One million to three million
islets of Langerhans form the endocrine part of the pancreas, which is primarily an exocrine
gland. The endocrine portion accounts for only 2% of the total mass of the pancreas.
Within the islets of Langerhans, beta cells constitute 65–80% of all the cells.
Insulin consists of two polypeptide chains, the A- and B- chains, linked together by disulfide
bonds. It is however first synthesized as a single polypeptide called preproinsulin
in pancreatic β-cells. Preproinsulin contains a 24-residue signal peptide which directs
the nascent polypeptide chain to the rough endoplasmic reticulum. The signal peptide
is cleaved as the polypeptide is translocated into lumen of the RER, forming proinsulin.
In the RER the proinsulin folds into the correct conformation and 3 disulfide bonds are formed.
About 5–10 min after its assembly in the endoplasmic reticulum, proinsulin is transported
to the trans-Golgi network where immature granules are formed. Transport to the TGN
may take about 30 min. Proinsulin undergoes maturation into active
insulin through the action of cellular endopeptidases known as prohormone convertases, as well as
the exoprotease carboxypeptidase E. The endopeptidases cleave at 2 positions, releasing a fragment
called the C-peptide, and leaving 2 peptide chains, the B- and A- chains, linked by 2
disulfide bonds. The cleavage sites are each located after a pair of basic residues. After
cleavage of the C-peptide, these 2 pairs of basic residues are removed by the carboxypeptidase.
The C-peptide is the central portion of proinsulin, and the primary sequence of proinsulin goes
in the order “B-C-A”. The resulting mature insulin is packaged inside
mature granules waiting for metabolic signals and vagal nerve stimulation to be exocytosed
from the cell into the circulation. The endogenous production of insulin is regulated
in several steps along the synthesis pathway: At transcription from the insulin gene
In mRNA stability At the mRNA translation
In the posttranslational modifications Insulin and its related proteins have been
shown to be produced inside the brain, and reduced levels of these proteins are linked
to Alzheimer’s disease. Release Beta cells in the islets of Langerhans release
insulin in two phases. The first phase release is rapidly triggered in response to increased
blood glucose levels. The second phase is a sustained, slow release of newly formed
vesicles triggered independently of sugar. The description of first phase release is
as follows: Glucose enters the β-cells through the glucose
transporter, GLUT2. Glucose goes into glycolysis and the Krebs
cycle, where multiple, high-energy ATP molecules are produced by oxidation, leading to a rise
in the ATP:ADP ratio within the cell. An increased intracellular ATP:ADP ratio closes
the ATP-sensitive SUR1/Kir6.2 potassium channel. This prevents potassium ions from leaving
the cell by facilitated diffusion, leading to a buildup of potassium ions. As a result,
the inside of the cell becomes more positive with respect to the outside, leading to the
depolarisation of the cell surface membrane. On depolarisation, voltage-gated calcium ion
channels open which allows calcium ions to move into the cells by facilitated diffusion.
An increased intracellular calcium ion concentration causes the activation of phospholipase C,
which cleaves the membrane phospholipid phosphatidyl inositol 4,5-bisphosphate into inositol 1,4,5-trisphosphate
and diacylglycerol. Inositol 1,4,5-trisphosphate binds to receptor
proteins in the plasma membrane of the endoplasmic reticulum. This allows the release of Ca2+
ions from the ER via IP3-gated channels, and further raises the intracellular concentration
of calcium ions. Significantly increased amounts of calcium
ions in the cells causes the release of previously synthesized insulin, which has been stored
in secretory vesicles. This is the primary mechanism for release
of insulin. Other substances known to stimulate insulin release include the amino acids arginine
and leucine, parasympathetic release of acetylcholine, sulfonylurea, cholecystokinin, and the gastrointestinally
derived incretins glucagon-like peptide-1 and glucose-dependent insulinotropic peptide.
Release of insulin is strongly inhibited by the stress hormone norepinephrine, which leads
to increased blood glucose levels during stress. It appears that release of catecholamines
by the sympathetic nervous system has conflicting influences on insulin release by beta cells,
because insulin release is inhibited by α2-adrenergic receptors and stimulated by β2-adrenergic
receptors. The net effect of norepinephrine from sympathetic nerves and epinephrine from
adrenal glands on insulin release is inhibition due to dominance of the α-adrenergic receptors.
When the glucose level comes down to the usual physiologic value, insulin release from the
β-cells slows or stops. If blood glucose levels drop lower than this, especially to
dangerously low levels, release of hyperglycemic hormones forces release of glucose into the
blood from cellular stores, primarily liver cell stores of glycogen. By increasing blood
glucose, the hyperglycemic hormones prevent or correct life-threatening hypoglycemia.
Evidence of impaired first-phase insulin release can be seen in the glucose tolerance test,
demonstrated by a substantially elevated blood glucose level at 30 minutes, a marked drop
by 60 minutes, and a steady climb back to baseline levels over the following hourly
time points. Oscillations Even during the digestion, in general, one
or two hours following a meal, insulin release from the pancreas is not continuous, but oscillates
with a period of 3–6 minutes, changing from generating a blood insulin concentration
more than about 800 pmol/l to less than 100 pmol/l. This is thought to avoid downregulation
of insulin receptors in target cells, and to assist the liver in extracting insulin
from the blood. This oscillation is important to consider when administering insulin-stimulating
medication, since it is the oscillating blood concentration of insulin release, which should,
ideally, be achieved, not a constant high concentration. This may be achieved by delivering
insulin rhythmically to the portal vein or by islet cell transplantation to the liver.
It is hoped that future insulin pumps will address this characteristic.
Blood content The blood content of insulin can be measured
in international units, such as µIU/mL or in molar concentration, such as pmol/L, where
1 µIU/mL equals 6.945 pmol/L. A typical blood level between meals is 8–11 μIU/mL.
Signal transduction Special transporter proteins in cell membranes
allow glucose from the blood to enter a cell. These transporters are, indirectly, under
blood insulin’s control in certain body cell types. Low levels of circulating insulin,
or its absence, will prevent glucose from entering those cells. More commonly, however,
there is a decrease in the sensitivity of cells to insulin, resulting in decreased glucose
absorption. In either case, there is ‘cell starvation’ and weight loss, sometimes extreme.
In a few cases, there is a defect in the release of insulin from the pancreas. Either way,
the effect is the same: elevated blood glucose levels.
Activation of insulin receptors leads to internal cellular mechanisms that directly affect glucose
uptake by regulating the number and operation of protein molecules in the cell membrane
that transport glucose into the cell. The genes that specify the proteins that make
up the insulin receptor in cell membranes have been identified, and the structures of
the interior, transmembrane section, and the extra-membrane section of receptor have been
solved. Two types of tissues are most strongly influenced
by insulin, as far as the stimulation of glucose uptake is concerned: muscle cells and fat
cells. The former are important because of their central role in movement, breathing,
circulation, etc., and the latter because they accumulate excess food energy against
future needs. Together, they account for about two-thirds of all cells in a typical human
body. Insulin binds to the extracellular portion
of the alpha subunits of the insulin receptor. This, in turn, causes a conformational change
in the insulin receptor that activates the kinase domain residing on the intracellular
portion of the beta subunits. The activated kinase domain autophosphorylates tyrosine
residues on the C-terminus of the receptor as well as tyrosine residues in the IRS-1
protein. phosphorylated IRS-1, in turn, binds to and
activates phosphoinositol 3 kinase PI3K catalyzes the reaction PIP2 + ATP → PIP3
+ ADP PIP3 activates protein kinase B
PKB phosphorylates glycogen synthase kinase and thereby inactivates GSK
GSK can no longer phosphorylate glycogen synthase unphosphorylated GS makes more glycogen
PKB also facilitates vesicle fusion, resulting in an increase in GLUT4 transporters in the
plasma membrane After the signal has been produced, termination
of signaling is then needed. As mentioned below in the section on degradation, endocytosis
and degradation of the receptor bound to insulin is a main mechanism to end signaling. In addition,
signaling can be terminated by dephosphorylation of the tyrosine residues by tyrosine phosphatases.
Serine/Threonine kinases are also known to reduce the activity of insulin. Finally, with
insulin action being associated with the number of receptors on the plasma membrane, a decrease
in the amount of receptors also leads to termination of insulin signaling.
The structure of the insulin–insulin receptor complex has been determined using the techniques
of X-ray crystallography. Physiological effects The actions of insulin on the global human
metabolism level include: Control of cellular intake of certain substances,
most prominently glucose in muscle and adipose tissue
Increase of DNA replication and protein synthesis via control of amino acid uptake
Modification of the activity of numerous enzymes. The actions of insulin on cells include:
Increased glycogen synthesis – insulin forces storage of glucose in liver cells in the form
of glycogen; lowered levels of insulin cause liver cells to convert glycogen to glucose
and excrete it into the blood. This is the clinical action of insulin, which is directly
useful in reducing high blood glucose levels as in diabetes.
Increased lipid synthesis – insulin forces fat cells to take in blood lipids, which are
converted to triglycerides; lack of insulin causes the reverse.
Increased esterification of fatty acids – forces adipose tissue to make fats from fatty acid
esters; lack of insulin causes the reverse. Decreased proteolysis – decreasing the breakdown
of protein Decreased lipolysis – forces reduction in
conversion of fat cell lipid stores into blood fatty acids; lack of insulin causes the reverse.
Decreased gluconeogenesis – decreases production of glucose from nonsugar substrates, primarily
in the liver; lack of insulin causes glucose production from assorted substrates in the
liver and elsewhere. Decreased autophagy – decreased level of degradation
of damaged organelles. Postprandial levels inhibit autophagy completely.
Increased amino acid uptake – forces cells to absorb circulating amino acids; lack of
insulin inhibits absorption. Increased potassium uptake – forces cells
to absorb serum potassium; lack of insulin inhibits absorption. Insulin’s increase in
cellular potassium uptake lowers potassium levels in blood. This possibly occurs via
insulin-induced translocation of the Na+/K+-ATPase to the surface of skeletal muscle cells.
Arterial muscle tone – forces arterial wall muscle to relax, increasing blood flow, especially
in microarteries; lack of insulin reduces flow by allowing these muscles to contract.
Increase in the secretion of hydrochloric acid by parietal cells in the stomach
Decreased renal sodium excretion. Insulin also influences other body functions,
such as vascular compliance and cognition. Once insulin enters the human brain, it enhances
learning and memory and benefits verbal memory in particular. Enhancing brain insulin signaling
by means of intranasal insulin administration also enhances the acute thermoregulatory and
glucoregulatory response to food intake, suggesting that central nervous insulin contributes to
the control of whole-body energy homeostasis in humans. Insulin also has stimulatory effects
on gonadotropin-releasing hormone from the hypothalamus, thus favoring fertility.
Degradation Once an insulin molecule has docked onto the
receptor and effected its action, it may be released back into the extracellular environment,
or it may be degraded by the cell. The two primary sites for insulin clearance are the
liver and the kidney. The liver clears most insulin during first-pass transit, whereas
the kidney clears most of the insulin in systemic circulation. Degradation normally involves
endocytosis of the insulin-receptor complex, followed by the action of insulin-degrading
enzyme. An insulin molecule produced endogenously by the pancreatic beta cells is estimated
to be degraded within about one hour after its initial release into circulation.
Hypoglycemia Although other cells can use other fuels,
neurons depend on glucose as a source of energy in the nonstarving human. They do not require
insulin to absorb glucose, unlike muscle and adipose tissue, and they have very small internal
stores of glycogen. Glycogen stored in liver cells can be converted to glucose, and released
into the blood, when glucose from digestion is low or absent, and the glycerol backbone
in triglycerides can also be used to produce blood glucose.
Sufficient lack of glucose and scarcity of these sources of glucose can dramatically
make itself manifest in the impaired functioning of the central nervous system: dizziness,
speech problems, and even loss of consciousness. Low blood glucose level is known as hypoglycemia
or, in cases producing unconsciousness, “hypoglycemic coma”. Endogenous causes of insulin excess
are very rare, and the overwhelming majority of insulin excess-induced hypoglycemia cases
are iatrogenic and usually accidental. A few cases of murder, attempted murder, or suicide
using insulin overdoses have been reported, but most insulin shocks appear to be due to
errors in dosage of insulin or other unanticipated factors.
Possible causes of hypoglycemia include: External insulin
Oral hypoglycemic agents Ingestion of low-carbohydrate sugar substitutes
in people without diabetes or with type 2 diabetes. Animal studies show these can trigger
insulin release, albeit in much smaller quantities than sugar, according to a report in Discover
magazine, August 2004, p 18. Diseases and syndromes
There are several conditions in which insulin disturbance is pathologic:
Diabetes mellitus – general term referring to all states characterized by hyperglycemia
Type 1 – autoimmune-mediated destruction of insulin-producing β-cells in the pancreas,
resulting in absolute insulin deficiency Type 2 – multifactoral syndrome with combined
influence of genetic susceptibility and influence of environmental factors, the best known being
obesity, age, and physical inactivity, resulting in insulin resistance in cells requiring insulin
for glucose absorption. Other types of impaired glucose tolerance Insulinoma – a tumor of pancreatic β-cells
producing excess insulin or reactive hypoglycemia. Metabolic syndrome – a poorly understood
condition first called Syndrome X by Gerald Reaven, Reaven’s Syndrome after Reaven, CHAOS
in Australia. It is currently not clear whether these signs have a single, treatable cause,
or are the result of body changes leading to type 2 diabetes. It is characterized by
elevated blood pressure, dyslipidemia, and increased waist circumference. The basic underlying
cause may be the insulin resistance that precedes type 2 diabetes, which is a diminished capacity
for insulin response in some tissues. It is common that morbidities, such as essential
hypertension, obesity, type 2 diabetes, and cardiovascular disease develop.
Polycystic ovary syndrome – a complex syndrome in women in the reproductive years where anovulation
and androgen excess are commonly displayed as hirsutism. In many cases of PCOS, insulin
resistance is present. Medication uses Biosynthetic human insulinfor clinical use
is manufactured by recombinant DNA technology. Biosynthetic human insulin has increased purity
when compared with extractive animal insulins, enhanced purity reducing antibody formation.
Researchers have succeeded in introducing the gene for human insulin into plants as
another method of producing insulin in safflower. This technique is anticipated to reduce production
costs. Several analogs of human insulin are available
for clinical therapy. These insulin analogs are closely related to the human insulin structure,
they were developed for specific aspects of glycemic control in terms of fast action and
long action. The first biosynthetic insulin analog was developed for clinical use at mealtime,
Humalog,it is more rapidly absorbed after subcutaneous injection than regular insulin,
with an effect 15 minutes after injection. Other rapid-acting analogues are NovoRapid
and Apidra, with similar profiles. All are rapidly absorbed due to sequence that will
reduce formation of dimers and hexamers. Fast acting insulins do not require the injection-to-meal
interval previously recommended for human insulin and animal insulins. The other type
is long acting insulin; the first of these was Lantus. These have a steady effect for
an extended period from 18 to 24 hours. Likewise, another protracted insulin analogue is based
on a fatty acid acylation approach. A myristyric acid molecule is attached to this analogue,
which in turn associates the insulin molecule to the abundant serum albumin, which in turn
extends the effect and reduces the risk of hypoglycemia. Both protracted analogues need
to be taken only once daily, and are used for type 1 diabetics as the basal insulin.
A combination of a rapid acting and a protracted insulin is also available, making it more
likely for patients to achieve an insulin profile that mimics that of the body´s own
insulin release. Insulin is usually taken as subcutaneous injections
by single-use syringes with needles, via an insulin pump, or by repeated-use insulin pens
with disposable needles. Unlike many medicines, insulin currently cannot
be taken orally because, like nearly all other proteins introduced into the gastrointestinal
tract, it is reduced to fragments, whereupon all activity is lost. There has been some
research into ways to protect insulin from the digestive tract, so that it can be administered
orally or sublingually. While experimental, several companies now have various formulations
in human clinical trials, and one, the India-based Biocon, has formed an agreement with BMS to
produce an oral-insulin alternative. History
Discovery In 1869, while studying the structure of the
pancreas under a microscope, Paul Langerhans, a medical student in Berlin, identified some
previously unnoticed tissue clumps scattered throughout the bulk of the pancreas. The function
of the “little heaps of cells”, later known as the islets of Langerhans, initially remained
unknown, but Edouard Laguesse later suggested they might produce secretions that play a
regulatory role in digestion. Paul Langerhans’ son, Archibald, also helped to understand
this regulatory role. The term “insulin” originates from insula, the Latin word for islet/island.
In 1889, the Polish-German physician Oscar Minkowski, in collaboration with Joseph von
Mering, removed the pancreas from a healthy dog to test its assumed role in digestion.
Several days after the removal of the dog’s pancreas, Minkowski’s animal-keeper noticed
a swarm of flies feeding on the dog’s urine. On testing the urine, they found sugar, establishing
for the first time a relationship between the pancreas and diabetes. In 1901 Eugene
Lindsay Opie took another major step forward when he clearly established the link between
the islets of Langerhans and diabetes: “Diabetes mellitus . . . is caused by destruction of
the islets of Langerhans and occurs only when these bodies are in part or wholly destroyed”.
Before Opie’s work, medical science had clearly established the link between the pancreas
and diabetes, but not the specific role of the islets. Over the next two decades researchers made
several attempts to isolate – as a potential treatment – whatever the islets produced.
In 1906 George Ludwig Zuelzer achieved partial success in treating dogs with pancreatic extract,
but he was unable to continue his work. Between 1911 and 1912, E.L. Scott at the University
of Chicago used aqueous pancreatic extracts, and noted “a slight diminution of glycosuria”,
but was unable to convince his director of his work’s value; it was shut down. Israel
Kleiner demonstrated similar effects at Rockefeller University in 1915, but World War I interrupted
his work and he did not return to it. In 1916 Nicolae Paulescu, a Romanian professor
of physiology at the University of Medicine and Pharmacy in Bucharest, developed an aqueous
pancreatic extract which, when injected into a diabetic dog, had a normalizing effect on
blood-sugar levels. He had to interrupt his experiments because of World War I, and in
1921 he wrote four papers about his work carried out in Bucharest and his tests on a diabetic
dog. Later that year, he published “Research on the Role of the Pancreas in Food Assimilation”.
Extraction and purification In October 1920, Canadian Frederick Banting
concluded that it was the very digestive secretions that Minkowski had originally studied that
were breaking down the islet secretion(s), thereby making it impossible to extract successfully.
He jotted a note to himself: “Ligate pancreatic ducts of the dog. Keep dogs alive till acini
degenerate leaving islets. Try to isolate internal secretion of these and relieve glycosurea.”
The idea was the pancreas’s internal secretion, which, it was supposed, regulates sugar in
the bloodstream, might hold the key to the treatment of diabetes. A surgeon by training,
Banting knew certain arteries could be tied off that would lead to atrophy of most of
the pancreas, while leaving the islets of Langerhans intact. He theorized a relatively
pure extract could be made from the islets once most of the rest of the pancreas was
gone. In the spring of 1921, Banting traveled to
Toronto to explain his idea to J.J.R. Macleod, who was Professor of Physiology at the University
of Toronto, and asked Macleod if he could use his lab space to test the idea. Macleod
was initially skeptical, but eventually agreed to let Banting use his lab space while he
was on holiday for the summer. He also supplied Banting with ten dogs on which to experiment,
and two medical students, Charles Best and Clark Noble, to use as lab assistants, before
leaving for Scotland. Since Banting required only one lab assistant, Best and Noble flipped
a coin to see which would assist Banting for the first half of the summer. Best won the
coin toss, and took the first shift as Banting’s assistant. Loss of the coin toss may have
proved unfortunate for Noble, given that Banting decided to keep Best for the entire summer,
and eventually shared half his Nobel Prize money and a large part of the credit for the
discovery of insulin with the winner of the toss. Had Noble won the toss, his career might
have taken a different path. Banting’s method was to tie a ligature around the pancreatic
duct; when examined several weeks later, the pancreatic digestive cells had died and been
absorbed by the immune system, leaving thousands of islets. They then isolated an extract from
these islets, producing what they called “isletin”, and tested this extract on the dogs starting
July 27. Banting and Best were then able to keep a pancreatectomized dog named Marjorie
alive for the rest of the summer by injecting her with the crude extract they had prepared.
Removal of the pancreas in test animals in essence mimics diabetes, leading to elevated
blood glucose levels. Marjorie was able to remain alive because the extracts, containing
isletin, were able to lower her blood glucose levels.
Banting and Best presented their results to Macleod on his return to Toronto in the fall
of 1921, but Macleod pointed out flaws with the experimental design, and suggested the
experiments be repeated with more dogs and better equipment. He then supplied Banting
and Best with a better laboratory, and began paying Banting a salary from his research
grants. Several weeks later, the second round of experiments was also a success; and Macleod
helped publish their results privately in Toronto that November. However, they needed
six weeks to extract the isletin, which forced considerable delays. Banting suggested they
try to use fetal calf pancreas, which had not yet developed digestive glands; he was
relieved to find this method worked well. With the supply problem solved, the next major
effort was to purify the extract. In December 1921, Macleod invited the biochemist James
Collip to help with this task, and, within a month, the team felt ready for a clinical
test. On January 11, 1922, Leonard Thompson, a 14-year-old
diabetic who lay dying at the Toronto General Hospital, was given the first injection of
insulin. However, the extract was so impure, Thompson suffered a severe allergic reaction,
and further injections were canceled. Over the next 12 days, Collip worked day and night
to improve the ox-pancreas extract, and a second dose was injected on January 23. This
was completely successful, not only in having no obvious side-effects but also in completely
eliminating the glycosuria sign of diabetes. The first American patient was Elizabeth Hughes
Gossett, the daughter of the governor of New York. The first patient treated in the U.S.
was future woodcut artist James D. Havens; Dr. John Ralston Williams imported insulin
from Toronto to Rochester, New York, to treat Havens.
Children dying from diabetic ketoacidosis were kept in large wards, often with 50 or
more patients in a ward, mostly comatose. Grieving family members were often in attendance,
awaiting the death. In one of medicine’s more dramatic moments,
Banting, Best, and Collip went from bed to bed, injecting an entire ward with the new
purified extract. Before they had reached the last dying child, the first few were awakening
from their coma, to the joyous exclamations of their families.
Banting and Best never worked well with Collip, regarding him as something of an interloper,
and Collip left the project soon after. Over the spring of 1922, Best managed to improve
his techniques to the point where large quantities of insulin could be extracted on demand, but
the preparation remained impure. The drug firm Eli Lilly and Company had offered assistance
not long after the first publications in 1921, and they took Lilly up on the offer in April.
In November, Lilly made a major breakthrough and was able to produce large quantities of
highly refined insulin. Insulin was offered for sale shortly thereafter.
Synthesis Purified animal-sourced insulin was the only
type of insulin available to diabetics until genetic advances occurred later with medical
research. The amino acid structure of insulin was characterized in the early 1950s by Frederick
Sanger, and the first synthetic insulin was produced simultaneously in the labs of Panayotis
Katsoyannis at the University of Pittsburgh and Helmut Zahn at RWTH Aachen University
in the early 1960s. The first genetically engineered, synthetic
“human” insulin was produced using E. coli in 1978 by Arthur Riggs and Keiichi Itakura
at the Beckman Research Institute of the City of Hope in collaboration with Herbert Boyer
at Genentech. Genentech, founded by Swanson, Boyer and Eli Lilly and Company, went on in
1982 to sell the first commercially available biosynthetic human insulin under the brand
name Humulin. The vast majority of insulin currently used worldwide is now biosynthetic
recombinant “human” insulin or its analogues. Recombinant insulin is produced either in
yeast or E. coli. In yeast, insulin may be engineered as a single-chain protein with
a KexII endoprotease site that separates the insulin A chain from a c-terminally truncated
insulin B chain. A chemically synthesized c-terminal tail is then grafted onto insulin
by reverse proteolysis using the inexpensive protease trypsin; typically the lysine on
the c-terminal tail is protected with a chemical protecting group to prevent proteolysis. The
ease of modular synthesis and the relative safety of modifications in that region accounts
for common insulin analogs with c-terminal modifications. The Genentech synthesis and
completely chemical synthesis such as that by Bruce Merrifield are not preferred because
the efficiency of recombining the two insulin chains is low, primarily due to competition
with the precipitation of insulin B chain. Nobel Prizes The Nobel Prize committee in 1923 credited
the practical extraction of insulin to a team at the University of Toronto and awarded the
Nobel Prize to two men: Frederick Banting and J.J.R. Macleod. They were awarded the
Nobel Prize in Physiology or Medicine in 1923 for the discovery of insulin. Banting, insulted
that Best was not mentioned, shared his prize with him, and Macleod immediately shared his
with James Collip. The patent for insulin was sold to the University of Toronto for
one half-dollar. The primary structure of insulin was determined
by British molecular biologist Frederick Sanger. It was the first protein to have its sequence
be determined. He was awarded the 1958 Nobel Prize in Chemistry for this work.
In 1969, after decades of work, Dorothy Hodgkin determined the spatial conformation of the
molecule, the so-called tertiary structure, by means of X-ray diffraction studies. She
had been awarded a Nobel Prize in Chemistry in 1964 for the development of crystallography.
Rosalyn Sussman Yalow received the 1977 Nobel Prize in Medicine for the development of the
radioimmunoassay for insulin. George Minot, co-recipient of the 1934 Nobel
Prize for the development of the first effective treatment for pernicious anemia, had diabetes
mellitus. Dr. William Castle observed that the 1921 discovery of insulin, arriving in
time to keep Minot alive, was therefore also responsible for the discovery of a cure for
pernicious anemia. Nobel Prize controversy The work published by Banting, Best, Collip
and Macleod represented the preparation of purified insulin extract suitable for use
on human patients. Although Paulescu discovered the principles of the treatment his saline
extract could not be used on humans, and he was not mentioned in the 1923 Nobel Prize.
Professor Ian Murray was particularly active in working to correct “the historical wrong”
against Nicolae Paulescu. Murray was a professor of physiology at the Anderson College of Medicine
in Glasgow, Scotland, the head of the department of Metabolic Diseases at a leading Glasgow
hospital, vice-president of the British Association of Diabetes, and a founding member of the
International Diabetes Federation. Murray wrote: Insufficient recognition has been given to
Paulesco, the distinguished Roumanian scientist, who at the time when the Toronto team were
commencing their research had already succeeded in extracting the antidiabetic hormone of
the pancreas and proving its efficacy in reducing the hyperglycaemia in diabetic dogs. In a recent private communication Professor
Tiselius, head of the Nobel Institute, has expressed his personal opinion that Paulescu
was equally worthy of the award in 1923. See also
Insulin analog Anatomy and physiolology
Pancreas Islets of Langerhans
Endocrinology Leptin. Forms of diabetes mellitus
Diabetes mellitus Diabetes mellitus type 1
Diabetes mellitus type 2 Treatment
Diabetic coma Insulin therapy
Intensive insulinotherapy Insulin pump
Conventional insulinotherapy Other medical / diagnostic uses
Insulin tolerance test Triple bolus test References Further reading External links

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