Insulin

Background

Insulin is a hormone that regulates the amount of glucose (sugar) in the blood and is required for the body to function normally. Insulin is produced by cells in the pancreas, called the islets of Langerhans. These cells continuously release a small amount of insulin into the body, but they release surges of the hormone in response to a rise in the blood glucose level.

Certain cells in the body change the food ingested into energy, or blood glucose, that cells can use. Every time a person eats, the blood glucose rises. Raised blood glucose triggers the cells in the islets of Langerhans to release the necessary amount of insulin. Insulin allows the blood glucose to be transported from the blood into the cells. Cells have an outer wall, called a membrane, that controls what enters and exits the cell. Researchers do not yet know exactly how insulin works, but they do know insulin binds to receptors on the cell's membrane. This activates a set of transport molecules so that glucose and proteins can enter the cell. The cells can then use the glucose as energy to carry out its functions. Once transported into the cell, the blood glucose level is returned to normal within hours.

Without insulin, the blood glucose builds up in the blood and the cells are starved of their energy source. Some of the symptoms that may occur include fatigue, constant infections, blurred eye sight, numbness, tingling in the hands or legs, increased thirst, and slowed healing of bruises or cuts. The cells will begin to use fat, the energy source stored for emergencies. When this happens for too long a time the body produces ketones, chemicals produced by the liver. Ketones can poison and kill cells if they build up in the body over an extended period of time. This can lead to serious illness and coma.

People who do not produce the necessary amount of insulin have diabetes. There are two general types of diabetes. The most severe type, known as Type I or juvenile-onset diabetes, is when the body does not produce any insulin. Type I diabetics usually inject themselves with different types of insulin three to four times daily. Dosage is taken based on the person's blood glucose reading, taken from a glucose meter. Type II diabetics produce some insulin, but it is either not enough or their cells do not respond normally to insulin. This usually occurs in obese or middle aged and older people. Type II diabetics do not necessarily need to take insulin, but they may inject insulin once or twice a day.

There are four main types of insulin manufactured based upon how soon the insulin starts working, when it peaks, and how long it lasts in the body. According to the American Diabetes Association, rapid-acting insulin reaches the blood within 15 minutes, peaks at 30-90 minutes, and may last five hours. Short-acting insulin reaches the blood within 30 minutes, it peaks about two to four hours later and stays in the blood for four to eight hours. Intermediate-acting insulin reaches the blood two to six hours after injection, peaks four to 14 hours later, and can last in the blood for 14-20 hours. And long-acting insulin takes six to 14 hours to start working, it has a small peak soon after, and stays in the blood for 20-24 hours. Diabetics each have different responses to and needs for insulin so there is no one type that works best for everyone. Some insulin is sold with two of the types mixed together in one bottle.

History

If the body does not produce any or enough insulin, people need to take a manufactured version of it. The major use of producing insulin is for diabetics who do not make enough or any insulin naturally.

Before researchers discovered how to produce insulin, people who suffered from Type I diabetes had no chance for a healthy life. Then in 1921, Canadian scientists Frederick G. Banting and Charles H. Best successfully purified insulin from a dog's pancreas. Over the years scientists made continual improvements in producing insulin. In 1936, researchers found a way to make insulin with a slower release in the blood. They added a protein found in fish sperm, protamine, which the body breaks down slowly. One injection lasted 36 hours. Another breakthrough came in 1950 when researchers produced a type of insulin that acted slightly faster and does not remain in the bloodstream as long. In the 1970s, researchers began to try and produce an insulin that more mimicked how the body's natural insulin worked: releasing a small amount of insulin all day with surges occurring at mealtimes.

Researchers continued to improve insulin but the basic production method remained the same for decades. Insulin was extracted from the pancreas of cattle and pigs and purified. The chemical structure of insulin in these animals is only slightly different than human insulin, which is why it functions so well in the human body. (Although some people had negative immune system or allergic reactions.) Then in the early 1980s biotechnology revolutionized insulin synthesis. Researchers had already decoded the chemical structure of insulin in the mid1950s. They soon determined the exact location of the insulin gene at the top of chromosome 11. By 1977, a research team had spliced a rat insulin gene into a bacterium that then produced insulin.

In 1891, Frederick Banting was born in Alliston, Ontario. He graduated in 1916 from the University of Toronto medical school. After Medical Corps service in World War I, Banting became interested in diabetes and studied the disease at the University of Western Ontario.

In 1919, Moses Barron, a researcher at the University of Minnesota, showed blockage of the duct connecting the two major parts of the pancreas caused shriveling of a second cell type, the acinar. Banting believed that by tying off the pancreatic duct to destroy the acinar cells, he could preserve the hormone and extract it from islet cells. Banting proposed this to the head of the University of Toronto's Physiology Department, John Macleod. Macleod rejected Banting's proposal, but supplied laboratory space, 10 dogs, and a medical student, Charles Best

Begining in May 1921, Banting and Best tied off pancreatic ducts in dogs so the acinar cells would atrophy, then removed the pancreases to extract fluid from islet cells. Meanwhile, they removed pancreases from other dogs to cause diabetes, then injected the islet cell fluid. In January 1922, 14 year-old Leonard Thompson became the first human to be successfully treat-ed for diabetes using insulin.

Best received his medical degree in 1925. Banting insisted Best also be credited, and almost turned down his Nobel Prize because Best was not included. Best became head of the University of Toronto's physiology department in 1929 and director of the university's Banting and Best Department of Medical Research after Banting's death in 1941.

In the 1980s, researchers used genetic engineering to manufacture a human insulin. In 1982, the Eli Lilly Corporation produced a human insulin that became the first approved genetically engineered pharmaceutical product. Without needing to depend on animals, researchers could produce genetically engineered insulin in unlimited supplies. It also did not contain any of the animal contaminants. Using human insulin also took away any concerns about transferring any potential animal diseases into the insulin. While companies still sell a small amount of insulin produced from animals—mostly porcine—from the 1980s onwards, insulin users increasingly moved to a form of human insulin created through recombinant DNA technology. According to the Eli Lilly Corporation, in 2001 95% of insulin users in most parts of the world take some form of human insulin. Some companies have stopped producing animal insulin completely. Companies are focusing on synthesizing human insulin and insulin analogs, a modification of the insulin molecule in some way.

Raw Materials

Human insulin is grown in the lab inside common bacteria. Escherichia coli is by far the most widely used type of bacterium, but yeast is also used.

Researchers need the human protein that produces insulin. Manufacturers get this through an amino-acid sequencing machine that synthesizes the DNA. Manufacturers know the exact order of insulin's amino acids (the nitrogen-based molecules that line up to make up proteins). There are 20 common amino acids. Manufacturers input insulin's amino acids, and the sequencing machine connects the amino acids together. Also necessary to synthesize insulin are large tanks to grow the bacteria, and nutrients are needed for the bacteria to grow. Several instruments are necessary to separate and purify the DNA such as a centrifuge, along with various chromatography and x-ray crystallography instruments.

The Manufacturing
Process

Synthesizing human insulin is a multi-step biochemical process that depends on basic recombinant DNA techniques and an understanding of the insulin gene. DNA carries the instructions for how the body works and one small segment of the DNA, the insulin gene, codes for the protein insulin. Manufacturers manipulate the biological precursor to insulin so that it grows inside simple bacteria. While manufacturers each have their own variations, there are two basic methods to manufacture human insulin.

Working with human insulin

• 1 The insulin gene is a protein consisting of two separate chains of amino acids, an A above a B chain, that are held together with bonds. Amino acids are the basic units that build all proteins. The insulin A chain consists of 21 amino acids and the B chain has 30.
• 2 Before becoming an active insulin protein, insulin is first produced as preproinsulin. This is one single long protein chain with the A and B chains not yet separated, a section in the middle linking the chains together and a signal sequence at one end telling the protein when to start secreting outside the cell. After preproinsulin, the chain evolves into proinsulin, still a single chain but without the signaling sequence. Then comes the active protein insulin, the protein without the section linking the A and B chains. At each step, the protein needs specific enzymes (proteins that carry out chemical reactions) to produce the next form of insulin.

STARTING WITH A AND B

• 3 One method of manufacturing insulin is to grow the two insulin chains separately. This will avoid manufacturing each of the specific enzymes needed. Manufacturers need the two mini-genes: one that produces the A chain and one for the B chain. Since the exact DNA sequence of each chain is known, they synthesize each mini-gene's DNA in an amino acid sequencing machine.
• 4 These two DNA molecules are then inserted into plasmids, small circular pieces of DNA that are more readily taken up by the host's DNA.
• 5 Manufacturers first insert the plasmids into a non-harmful type of the bacterium E. coli. They insert it next to the lacZ gene. LacZ encodes for 8-galactosidase, a gene widely used in recombinant DNA procedures because it is easy to find and cut, allowing the insulin to be readily removed so that it does not get lost in the bacterium's DNA. Next to this gene is the amino acid methionine, which starts the protein formation.
• 6 The recombinant, newly formed, plasmids are mixed up with the bacterial cells. Plasmids enter the bacteria in a process called transfection. Manufacturers can add to the cells DNA ligase, an enzyme that acts like glue to help the plasmid stick to the bacterium's DNA.
• 7 The bacteria synthesizing the insulin then undergo a fermentation process. They are grown at optimal temperatures in large tanks in manufacturing plants. The millions of bacteria replicate roughly every 20 minutes through cell mitosis, and each expresses the insulin gene.
• 8 After multiplying, the cells are taken out of the tanks and broken open to extract the DNA. One common way this is done is by first adding a mixture of lysozome that digest the outer layer of the cell wall, then adding a detergent mixture that separates the fatty cell wall membrane. The bacterium's DNA is then treated with cyanogen bromide, a reagent that splits protein chains at the methionine residues. This separates the insulin chains from the rest of the DNA.
• 9 The two chains are then mixed together and joined by disulfide bonds through the reduction-reoxidation reaction. An oxidizing agent (a material that causes oxidization or the transfer of an electron) is added. The batch is then placed in a centrifuge, a mechanical device that spins quickly to separate cell components by size and density.
• 10 The DNA mixture is then purified so that only the insulin chains remain. Manufacturers can purify the mixture through several chromatography, or separation, techniques that exploit differences in the molecule's charge, size, and affinity to water. Procedures used include an ion-exchange column, reverse-phase high performance liquid chromatography, and a gel filtration chromatography column. Manufacturers can test insulin batches to ensure none of the bacteria's E. coli proteins are mixed in with the insulin. They use a marker protein that lets them detect E. coli DNA. They can then determine that the purification process removes the E. coli bacteria.

PROINSULIN PROCESS

• 11 Starting in 1986, manufacturers began to use another method to synthesize human insulin. They started with the direct precursor to the insulin gene, proinsulin. Many of the steps are the same as when producing insulin with the A and B chains, except in this method the amino acid machine synthesizes the proinsulin gene.
• 12 The sequence that codes for proinsulin is inserted into the non-pathogenic E. coli bacteria. The bacteria go through the fermentation process where it reproduces and produces proinsulin. Then the connecting sequence between the A and B chains is spliced away with an enzyme and the resulting insulin is purified.
• 13 At the end of the manufacturing process ingredients are added to insulin to prevent bacteria and help maintain a neutral balance between acids and bases. Ingredients are also added to intermediate and long-acting insulin to produce the desired duration type of insulin. This is the traditional method of producing longer-acting insulin. Manufacturers add ingredients to the purified insulin that prolong their actions, such as zinc oxide. These additives delay absorption in the body. Additives vary among different brands of the same type of insulin.

Analog insulin

In the mid 1990s, researchers began to improve the way human insulin works in the body by changing its amino acid sequence and creating an analog, a chemical substance that mimics another substance well enough that it fools the cell. Analog insulin clumps less and disperses more readily into the blood, allowing the insulin to start working in the body minutes after an injection. There are several different analog insulin. Humulin insulin does not have strong bonds with other insulin and thus, is absorbed quickly. Another insulin analog, called Glargine, changes the chemical structure of the protein to make it have a relatively constant release over 24 hours with no pronounced peaks.

Instead of synthesizing the exact DNA sequence for insulin, manufacturers synthesize an insulin gene where the sequence is slightly altered. The change causes the resulting proteins to repel each other, which causes less clumping. Using this changed DNA sequence, the manufacturing process is similar to the recombinant DNA process described.

Quality Control

After synthesizing the human insulin, the structure and purity of the insulin batches are tested through several different methods. High performance liquid chromatography is used to determine if there are any impurities in the insulin. Other separation techniques, such as X-ray crystallography, gel filtration, and amino acid sequencing, are also performed. Manufacturers also test the vial's packaging to ensure it is sealed properly.

Manufacturing for human insulin must comply with National Institutes of Health procedures for large-scale operations. The United States Food and Drug Administration must approve all manufactured insulin.

The Future

The future of insulin holds many possibilities. Since insulin was first synthesized, diabetics needed to regularly inject the liquid insulin with a syringe directly into their bloodstream. This allows the insulin to enter the blood immediately. For many years it was the only way known to move the intact insulin protein into the body. In the 1990s, researchers began to make inroads in synthesizing various devices and forms of insulin that diabetics can use in an alternate drug delivery system.

Manufacturers are currently producing several relatively new drug delivery devices. Insulin pens look like a writing pen. A cartridge holds the insulin and the tip is the needle. The user set a dose, inserts the needle into the skin, and presses a button to inject the insulin. With pens there is no need to use a vial of insulin. However, pens require inserting separate tips before each injection. Another downside is that the pen does not allow users to mix insulin types, and not all insulin is available.

For people who hate needles an alternate to the pen is the jet-injector. Looking similar to the pens, jet injectors use pressure to propel a tiny stream of insulin through the skin. These devices are not as widely used as the pen, and they can cause bruising at the input point.

The insulin pump allows a controlled release in the body. This is a computerized pump, about the size of a beeper, that diabetics can wear on their belt or in their pocket. The pump has a small flexible tube that is inserted just under the surface of the diabetic's skin. The diabetic sets the pump to deliver a steady, measured dose of insulin throughout the day, increasing the amount right before eating. This mimics the body's normal release of insulin. Manufacturers have produced insulin pumps since the 1980s but advances in the late 1990s and early twenty-first century have made them increasingly easier to use and more popular. Researchers are exploring the possibility of implantable insulin pumps. Diabetics would control these devices through an external remote control.

Researchers are exploring other drug-delivery options. Ingesting insulin through pills is one possibility. The challenge with edible insulin is that the stomach's high acidic environment destroys the protein before it can move into the blood. Researchers are working on coating insulin with plastic the width of a few human hairs. The coverings would protect the drugs from the stomach's acid.

In 2001 promising tests are occurring on inhaled insulin devices and manufacturers could begin producing the products within the next few years. Since insulin is a relatively large protein, it does not permeate into the lungs. Researchers of inhaled insulin are working to create insulin particles that are small enough to reach the deep lung. The particles can then pass into the bloodstream. Researchers are testing several inhalation devices much like that of an asthma inhaler.

Another form of aerosol device undergoing tests will administer insulin to the inner cheek. Known as buccal (cheek) insulin, diabetics will spray the insulin onto the inside of their cheek. It is then absorbed through the inner cheek wall.

Insulin patches are another drug delivery system in development. Patches would release insulin continuously into the bloodstream. Users would pull a tab on the patch to release more insulin before meals. The challenge is finding a way to have insulin pass through the skin. Ultrasound is one method researchers are investigating. These low frequency sound waves could change the skin's permeability and allow insulin to pass.

Other research has the potential to discontinue the need for manufacturers to synthesize insulin. Researchers are working on creating the cells that produce insulin in the laboratory. The thought is that physicians can someday replace the non-working pancreas cells with insulin-producing cells. Another hope for diabetics is gene therapy. Scientists are working on correcting the insulin gene's mutation so that diabetics would be able to produce insulin on their own.

Where to Learn More

Books

Clark, David P, and Lonnie D. Russell. Molecular Biology Made Simple and Fun. 2nd ed. Vienna, IL: Cache River Press, 2000.

Considine, Douglas M., ed. Van Nostrand's Scientific Encyclopedia. 8th ed. New York: International Thomson Publishing Inc., 1995.

Periodicals

Dinsmoor, Robert S. "Insulin: A Never-ending Evolution." Countdown (Spring 2001).

Other

Diabetes Digest Web Page. 15 November 2001. <http://www.diabetesdigest.com>.

Discovery of Insulin Web Page. 16 November 2001. <http://web.idirect.com/~discover>.

Eli Lilly Corporation. Humulin and Humalog Development. CD-ROM, 2001.

Eli Lilly Diabetes Web Page. 16 November 2001. <http://www.lillydiabetes.com>.

Novo Nordisk Diabetes Web Page. 15 November 2001. <http://www.novonet.co.nz>.

M.RaeNelson

insulin Glucose, dissolved in the blood (blood sugar), is one of the main sources of energy for the body. Different organs and tissues use other fuels to varying extents, but the brain uses glucose exclusively. To protect vital functions mammals have evolved a mechanism for keeping the concentration of glucose in the blood fairly constant — an example of homeostasis. This includes diverse hormonal responses that increase the blood glucose concentration. Yet, in what seems a remarkable oversight of nature, the body relies almost entirely upon just one hormone, the protein insulin, to bring about a decrease in blood glucose. Insulin facilitates the entry of glucose from the blood into the tissues of the body.

It has been known since 1899 that removal of the pancreas from dogs led to diabetes, with its characteristic persistent increase in blood glucose (hyperglycaemia) and presence of sugar in the urine (glycosuria). The fascinating saga of the eventual discovery of insulin by Banting and Best in 1921 is well known. They were able to show that injection of an extract from the pancreas of a healthy dog led consistently to a decrease in the amount of sugar in the blood and urine of diabetic dogs. They published their account, entitled ‘The Internal Secretion of the Pancreas’, in 1922. Their experiments, which were to prove life-saving, assured the insulin molecule a key place in medical history, and won a Nobel Prize for Banting and Macleod (in whose Canadian laboratory the work was done) as well as earning the grateful thanks of diabetic people in their millions around the world. It was some 30 years later that Frederick Sanger embarked on his painstaking but highly successful molecular dissection of insulin, which unravelled its precise amino acid sequence. This was a landmark achievement, representing as it did the first successful sequencing of any protein molecule, and it earned Sanger his first Nobel Prize. With subsequent establishment of its three-dimensional structure, insulin was revealed as a vital protein of classic polypeptide design. Despite 300 million years of divergent evolution, the molecular form and function of insulin has been remarkably well conserved across the entire zoological spectrum.

The dynamic glucose–insulin system on which the body's metabolism so critically depends is controlled and modulated by various factors impinging on the ‘b-cells’, which are found in the pancreas in cellular nodules, the Islets of Langerhans — named after the German pathologist who described them in 1869, long before their function was known. These ‘b-cells’ detect glucose in the blood and secrete insulin in appropriate amounts in response to the meal-induced tidal changes in blood glucose level.

Insulin is normally quite rapidly removed from the blood and survives in the circulation for only 5–15 min, thus placing a continuing demand on the b-cells for the release of more insulin in order to establish an effective feedback control of blood glucose concentration. This moment-by-moment process requires, in the b-cells, mechanisms for the manufacture, storage, and rapid release of insulin. To replenish its insulin supply, the genes of a pancreatic b-cell switch on their protein manufacturing machinery, producing a much larger single chain precursor molecule, called pro-insulin, which contains the amino acid sequence of insulin. Successive and controlled proteolysis (breakdown of this protein molecule) finally leaves the 51 amino acid sequence of insulin itself, and ensures its correct folding to create the three-dimensional shape of the molecule. Once formed in the b-cell, insulin is stored in granules as a symmetrical hexagonal array of 6 insulin molecules combined in a stable crystalline structure with 2 atoms of zinc. When released into the circulation at effective concentrations, insulin is transported as, and normally acts as, a single molecule.

To exert its effects on target cells — muscle, liver, or fat cells — the insulin molecule must first be recognized by specific insulin-receptor protein molecules in the cell membranes. Part of the insulin receptor spans the membrane, so that, when an insulin molecule binds to the external part of the receptor, a signal is transmitted across the membrane to other molecules, leading to a cascade of enzyme activity in the target cells.

Insulin resistance may occur when the blood glucose level is not well controlled, as in a type of diabetes which begins in adult life, where the pancreatic b-cells do not produce enough insulin. Not only does this lead to the appearance of the symptoms of diabetes, but the high level of glucose in the blood decreases the sensitivity of the target cell receptors for insulin and so makes the situation worse. It is possible to treat this type of diabetes by mouth with agents that boost the output of insulin from any viable b-cells that are present, or reduce the blood sugar by other means. If, on the other hand, pancreatic b-cells have all been destroyed, as in juvenile diabetes, then insulin must be injected daily as replacement therapy.

Unfortunately insulin cannot be given orally because it is a peptide and is therefore rapidly broken down by enzymes in the gut. Different preparations of insulin are available for injection, depending on the duration of action required. Insulin was originally extracted on a massive scale from the pancreas of animals (cows or pigs). It can now be obtained by genetic engineering of bacterial cells, causing them to express human insulin. It is noteworthy that insulin was the first protein to be commercially produced by such recombinant technology. Although this allows large scale production and isolation, pig pancreas remains the main source of insulin for human treatment: pig insulin differs from human insulin by only one amino acid.

Various insulin formulations may combine rapid-, medium-, or long-acting forms of crystalline insulin so that individual requirements for insulin can be matched to blood glucose levels following meals. Insulin has thus become firmly established in modern medicine as a remarkably effective therapeutic agent, but a whole life-time of constant injection is an unwelcome hazard for anyone suffering from juvenile diabetes. The design and production of a non-peptide, orally-active insulin analogue therefore remains a major goal of pharmaceutical research.

E. K. Matthews

INSULIN

Insulin

Diabetes mellitus, also known as "sugar disease," is a disease that killed thousands every year until the discovery of insulin in 1921. Diabetes is often seen in children, and before the treatment existed, the disease was essentially a death sentence. It is caused by a defect in the pancreas, which is then unable to produce the hormone insulin needed by muscle cells to utilize glucose. Without glucose the tissues are deprived of their main energy and are forced to produce energy from fat. High blood levels of toxic ketone bodies (acetone) result.

Symptoms

The diabetic shows a high level of glucose in blood and urine. Symptoms of the disease include increased thirst and hunger, increased urination, weakness, and a loss of weight. If left untreated, the acetone accumulates in the blood, brain function ceases, and the patient may slip into a coma and die.

Pancreas Defect Responsible for Disease

Although a disease thought to be diabetes was recognized by the Egyptians as early as 1500 B.C., it was not until 1899 that the causative factor in the disease was discovered to be a defect in the pancreas. This was a major step forward because scientists were then able to produce the disease in laboratory animals. However, even with these advances, no scientist was able to isolate the exact substance that was causing the pancreatic malfunction.

Banting Resolves to Find Cure

Many experiments were conducted in the early 1900s in the search for the elusive pancreatic substance, but no real progress was made until the 1920s when Frederick Grant Banting was allowed access to a proper laboratory. Banting was a young Canadian physician whose life had been altered by diabetes at the age of fifteen when his sweetheart and his best friend died of the disease. Banting served as a pall-bearer for both, and this experience influenced his resolve in later life to find a cure for diabetes.

Banting Proposes Research

After studying medicine at the University of Toronto and serving as a battlefield surgeon in World War I, Banting set up practice as an orthopedic surgeon in London, Ontario. He also took a parttime teaching assignment in the School of Medicine at the University of Western Ontario. While preparing a lecture on physiology, he became interested in the pancreas and began to wonder how it might have influenced the death of his friends years earlier. He proposed to research the subject at Western Ontario, but the faculty refused him with the recommendation that he discuss his ideas with professor John J. R. MacLeod at the University of Toronto, an authority on diabetes research. Banting met with MacLeod in 1920, but the elder physician contemptuously questioned how a young nonscientist could hope to find a cure for diabetes when the best physiologists in the world had been trying without success for thirty years. The door of opportunity remained closed.

A Determination to Succeed

However, Banting learned that MacLeod was going to Scotland for twelve weeks in the summer of 1921. Banting asked if he could use one of MacLeod's laboratories while he was away. He would need ten dogs and a helper who could run blood-sugar determinations. MacLeod agreed to his request if he would work without compensation and without standing at the university. Charles Best, age twenty-one, who was just graduating with a degree in biochemistry, agreed to help. Best was concerned with a recent diabetic death in his family. Between them, Banting and Best had less than $500. They cooked on a Bunsen burner in the lab and frequently slept there. By the end of July they were able to induce diabetic coma by removing the pancreas of a dog and restore the animal to health using a substance they had isolated from the pancreas of another dog, a substance they called the "X Factor."

"X Factor" Isolated and Purified

The animal's recovery was short-lived, and it became evident that an additional injection of the X Factor would be necessary every day. Banting and Best would have to find another method of producing the substance in quantity. When MacLeod returned from Europe, he insisted that the researchers repeat the experiments with dogs before trying to work with humans. After the second experiment worked, MacLeod offered Banting and Best sixty dollars a month each and dedicated the entire efforts of his department to the project. James Collip, who had a doctorate in chemistry, worked on purifying the X Factor, which MacLeod named "insulin."

"RESCUED BY INSULIN"

"Will I die, daddy?' asked my small daughter Elizabeth as she slipped her hand into mine as we were leaving the office of the specialist. His verdict had been—diabetes.

'No,' I assured her; 'thanks to Dr. Banting and his wonderful discovery of insulin, you will live/

'But Eloise had diabetes and she died/ reasoned my daughter. Eloise was a little neighbor.

'Yes/ I answered, 'but that was before insulin had been discovered.' "

Source:

"Rescued by Insulin," Hygeia, 6 (December 1928): 672.

First Successful Treatment of Diabetes

In January 1922 Banting and Best first tested insulin on themselves to show that it was not harmful, then administered an injection to Leonard Thompson, a twelve-year-old boy dying of diabetes. His recovery seemed nothing less than miraculous, and other diabetics quickly came forward for treatment. There was soon difficulty in producing the substance in the quantities needed.

Banting and MacLeod Honored

While Banting and Best worked on problems of mass production, MacLeod traveled around reading papers on "his" discovery. The 1923 Nobel Prize for medical research was awarded to MacLeod and Banting as "codiscoverers" of insulin. Banting gave half of his money and half of the credit to Best, after which MacLeod decided to give half of his money to Collip.

Insulin Promises New Life

The discovery of insulin has been called one of the most revolutionary events in the history of medicine. Although not a cure, insulin changed the devastating effects of a previously deadly disease. Insulin promised sufferers of diabetes a full and healthy life. Today, there are an estimated fifteen million diabetics alive due to insulin.

Sources:

Michael Bliss, Banting: A Biography (Toronto: McClelland 6c Stewart, 1984);

Bliss, The Discovery of Insulin (Toronto: McClelland & Stewart, 1982);

Wayne Martin, Medical Heroes and Heretics (Old Greenwich, Conn.: Devon-Adair, 1977), pp. 2-19;

Insulin

Insulin is a small peptide (protein) consisting of fifty-one amino acids synthesized and stored within the pancreas, an organ situated behind the stomach. The protein itself consists of two chains, denoted A and B, linked by disulfide (sulfur-sulfur) bridges between cysteine residues (see Figure 1).

Insulin is a hormone, a chemical transported in the blood that controls and regulates the activity of certain cells or organs in the body. When blood sugar levels rise following a meal, the pancreas is stimulated to release insulin into the bloodstream. In order for tissues to absorb glucose from the blood, they must first bind insulin. Glucose metabolism is necessary for cell growth and energy needs associated with cell function. When insulin binds to receptors on cell membranes, glucose transporter proteins are released from within the cell to the surface of the cell membrane. Once on the exterior surface of cells, glucose transporters can carry sugar from the blood into the tissue where it is metabolized. Without insulin, cells cannot absorb glucose and effectively starve.

A deficiency in insulin production results in a condition called diabetes mellitus. Approximately 6.2 percent of the population in the United States is affected with diabetes. Type 1 diabetics account for 10 percent of those individuals suffering from diabetes mellitus. It is also known as juvenile diabetes and generally develops in young people, typically between the ages of ten and fifteen years, as a result of an autoimmune disorder. Why the body's immune system turns on itself, attacking and destroying beta cells, the pancreatic cells in which insulin is synthesized, is not clear. The unfortunate consequence is insulin deficiency.

The majority of individuals afflicted with diabetes mellitus suffer from type 2 diabetes. The onset of type 2 diabetes occurs much later than for type 1 and typically in people over the age of fifty. The pancreas of type 2 diabetics continues to produce and release insulin. However, cells do not respond appropriately to insulin levels in the blood. This condition is known as insulin-resistance and is associated with obesity and high blood pressure. Children who are obese can also develop type 2 diabetes.

Gestational diabetes affects 3 to 5 percent of pregnant women. Onset generally occurs in the fifth or sixth month of pregnancy. In pregnant women, the placenta produces hormones to support the growing fetus. Some of these hormones (e.g., estrogen and cortisol) interfere with the actions of insulin. Insulin-resistance develops despite adequate blood insulin levels. Gestational diabetes does not last beyond pregnancy and the condition disappears after delivery.

Before the discovery of insulin, type 1 diabetics usually died within a few years of onset of the disease. During the early 1920s, a young Canadian physician, Frederick Grant Banting, working with John James Rickard Macleod, professor of physiology at the University of Toronto, and Charles Best, a medical student, discovered insulin while performing investigations on extracts acquired from dog pancreas. Soon after, Banting and Macleod were awarded the 1923 Nobel Prize in physiology or medicine for their discovery (Banting reportedly split his share of the prize money with Best).

Insulin was rapidly put into clinical use, chiefly through the efforts of August Krogh, a Danish scientist who cofounded the Nordic Insulin Laboratory in Copenhagen, Denmark, for the production of insulin. Large quantities of insulin were initially acquired from the pancreatic tissues of slaughtered animals, typically cows and pigs. Although bovine (cow) and porcine (pig) insulin are still the major components of commercially available insulin in the United States, the use of human insulin preparations is rapidly growing. Since the 1980s, recombinant DNA techniques have made human insulin available for clinical use. Genetically modified strains of Escherichia coli bacteria or Saccharomyces cerevisiae (baker's yeast) containing human genes coding for insulin have been developed for the mass production of human insulin.

Insulin is not a cure for diabetes. It does, however, allow diabetics, especially those with type 1 diabetes, to gain some control over their condition. Insulin is typically administered by subcutaneous (under the skin) injection. There are four principal types of insulin preparations commercially available. Insulin lispro (ultra-short-acting insulin) is rapidly absorbed into the blood and lasts only 3 to 4 hours. Regular insulin, on the other hand, takes 30 minutes to become effective, and lasts 5 to 7 hours. Lente and NPH (neutral protamine Hagedorn) insulin preparations are longeracting formulations typically administered every 12 hours. Finally, mixtures of insulin preparations are also commercially available. Typical mixtures consist of 70 percent NPH and 30 percent insulin lispro.

see also Genetic Engineering; Glycolysis.

Nanette M. Wachter

Bibliography

Dewitt, Dawn E. and Hirsh, Irl B. (2003). "Outpatient Insulin Therapy in Type 1 and Type 2 Diabetes Mellitus: Scientific Review." Journal of the American Medical Association 289(17):2254–2264.

Katzung, Bertram G. (1998). Basic & Clinical Pharmacology, 7th edition. Stamford, CT: Appleton & Lange.

Internet Resources

National Institute of Diabetes & Digestive & Kidney Diseases of the National Institutes of Health. Available from <http://www.niddk.nih.gov>.

Nobel Foundation E-Museum. "The Nobel Prize in Physiology or Medicine 1923." Available from <http://www.nobel.se>.

Insulin

Insulin is a hormone produced by the pancreas (a gland that releases a digestive juice into the intestine). The pancreas is composed of acinar cells, which produce digestive enzymes, and the islet cells of Langerhans, which produce hormones.

What Insulin Does

Four hormones are produced by the Langerhans islet cells. Insulin is produced in the B cells, glucagon in the A cells, somatostatin in the D cells, and pancreatic polypeptide in the F cells. Insulin promotes anabolism (building up of tissues) and inhibits catabolism (breaking down of tissues) in muscle, liver, and fat cells. It increases the rate of synthesis (blending) of glycogen, fatty acids, and proteins. Lack of insulin causes diabetes mellitus (a disease characterized by excess sugar in the blood and other body fluids).

Insulin's most important feature is its ability to increase the rate of glucose (a crystalline sugar) absorption by cells. Glucose is the most efficient fuel used by and found in almost all cells. Insulin causes a decreased concentration of glucose in the blood and causes the cells to store glycogen (a starchlike substance), mostly in the liver. It also promotes the entry of other sugars and amino acids into the muscle and fat cells. Insulin is therefore responsible for promoting fat storage in fat cells and for the total quantity of protein in the body.

Insulin Production

Insulin production is stimulated by high levels of glucose and inhibited (limited) by lower levels of glucose. Insulin regulates glucose with glucagon. Glucagon catabolizes (changes into a product of simpler composition) glycogen to glucose and also raises the blood sugar. Glucagon can be given to increase the blood sugar when intravenous (by needle) glucose cannot be given. Glucagon takes about twenty minutes to raise the blood sugar. Intravenous glucose raises it instantaneously, which is why it is preferred in treatment. Together insulin and glucagon ensure that the body stores and maintains the proper level of glucose for its energy needs.

Diabetes

Diabetes is from the Greek word meaning "siphon," and "mellitus" comes from melliferous, meaning "of or relating to honey." Diabetes has been recognized for centuries and was originally diagnosed by tasting the urine and finding it sweet (melliferous). The high sugar also causes the kidneys to excrete (or siphon) large amounts of water. In 1815, French chemist Michel Eugene Chevreul discovered that the sweetness came from grape sugar or glucose. Later discoveries showed how the body makes, stores, and uses glucose.

Injury to the pancreas was linked to diabetes beginning in the seventeenth century and confirmed by animal experiments, particularly those of the German physiologist Joseph von Mehring (1849-1908) and a Russian pathologist, Oscar Minkowski (1858-1931). The acinus cells were found in the seventeenth century by the Dutch anatomist Regnier de Graaf and the islet cells in 1869 by a German pathologist Paul Langerhans (1847-1888).

Hormones

In 1905 English physiologists Ernest Starling and William Bayliss discovered hormones. Hormones are substances secreted (released) by glands and carried in the blood to control cell activity elsewhere. In 1916 an English physiologist named Edward Sharpey-Schafer proposed that a hormone produced by the pancreas lowered the level of glucose in the blood. He called the hormone "insuline," the Latin word for "island," because he believed it came from the islet cells of the pancreas.

Credit for discovering insulin is given to Canadian surgeon Frederick Grant Banting (1891-1941) and Canadian physiologist Charles Herbert Best (1899-1978). Banting and Scottish physiologist and professor John James Rikard Macleod (1876-1935) were jointly awarded the Nobel Prize for medicine in 1923. Banting gave half of his share to Best, and Macleod gave half of his share to James Bertram Collip, because of the men had contributed to the discovery.

The First Insulin Patient

Collip, a professor at the University of Alberta, had experience in the chemistry of hormones. Prior to January 1922, he had prepared an insulin pure enough to be used on human patients. The first patient to receive insulin was 14-year-old Leonard Thompson. Thompson was admitted to Toronto General Hospital with a high blood glucose level; he also was urinating between three and five liters of fluid per day. Despite his rigid diet of only 450 calories (the only known treatment at this time was a diet low in carbohydrates), Thompson continued to excrete (get rid of through bodily waste) large amounts of glucose. On January 11, 1922, he was given insulin. Within a fairly short time, his blood sugar level came down and he stopped urinating large amounts of liquid.

Humulin

In 1982 insulin became available as a genetically-engineered product called Humulin. Humulin's structure is identical with human insulin. The A and B chains are produced separately in different strains of E. coli bacteria. The E. coli have been genetically encoded to produce each of these strains. The strains are separated from the bacteria and purified. The purified chains are combined chemically and repurified.

Insulin

Insulin is a hormone produced by specialized cells in the pancreas. Secreted into the bloodstream at each meal, insulin helps the body use and store glucose (sugar) produced during the digestion of food. In people with diabetes , the pancreas either does not produce enough insulin or the body cannot use the insulin that is produced in an efficient manner.

Treatment for diabetes requires the delivery of insulin into the bloodstream by either an insulin pen, needle and syringe, or pump. An insulin pen is a device that looks like a pen but contains an insulin cartridge. Both the syringe and pen methods require injection of the insulin into the arm, thigh, or abdomen. Pump therapy, however, continuously administers insulin according to a programmed plan unique to the pump wearer. Several types of insulin exist, and they differ in when the insulin begins working after it is injected, when the insulin is working hardest, and how long the insulin lasts in the body.

Insulin release and glucose absorption depend on a number of factors, including the glycemic index of food and the co-ingestion of fat and protein . Consumption of high-glycemic foods causes hyperglycemia which results in the release of too much insulin. On the other hand, low-glycemic foods or the ingestion of fat and protein in a meal provide steady glucose absorption and release of insulin.

Exercise lowers blood glucose levels and increases the amount of insulin in the bloodstream, along with improving the body's use of insulin. A balance must exist between the sugar used for energy , the sugar available from food, and the insulin used in lowering blood sugar. Consequently, changes may have to be made to insulin, or food intake, or both, prior to and after exercise.

see also Diabetes Mellitus; Glycemic Index; Hyperglycemia; Hypoglycemia.

Julie Lager

Bibliography

Bode, Bruce W.; Sabbah, Hassan T.; Gross, Todd M.; Fredrickson, Linda P.; and Davidson, Paul C. (2002). "Diabetes Management in the New Millennium Using Insulin Pump Therapy." Diabetes/Metabolism Research and Reviews 18 (Suppl. 1):S14–S20.

DeWitt, Dawn E. and Hirsch, Irl B. (2003). "Outpatient Insulin Therapy in Type 1 and Type 2 Diabetes Mellitus: A Scientific Review." Journal of the American Medical Association 289(17):2254–2264.

Parmet, Sharon; Cassio, Lynm; and Glass, Richard M. (2003). "Insulin." Journal of the American Medical Association 289(17):2314.

Internet Resources

American Diabetes Association. "About Insulin." Available from <http://www.diabetes.org/>

National Library of Medicine. "Diabetes." Updated July 2, 2003. Available from <http://medlineplus.gov/>

insulinhormone secreted by the β cells of the islets of Langerhans, specific groups of cells in the pancreas . Insufficiency of insulin in the body results in diabetes . Insulin was one of the first products to be manufactured using genetic engineering .

Action

In general, insulin acts to reduce extracellular (including blood plasma) levels of glucose by interacting in some way yet unknown with various cell membranes. In adipose (fatty) tissue it facilitates the cellular uptake of glucose and its subsequent conversion to fatty acids , and it inhibits the breakdown of fatty acids to simpler compounds. In muscle it again facilitates the transport of glucose into cells and in addition stimulates its conversion to glycogen . It also increases protein synthesis in muscle. In the liver, insulin facilitates glucose catabolism and its conversion to glycogen and inhibits its synthesis from simpler compounds.

Isolation and Structure

Canadians Frederick G. Banting and Charles H. Best were the first to obtain, from extracts of pancreas (1921–22), a preparation of insulin that could serve to replace a deficiency of the hormone in the human body. The complete amino acid sequence of the insulin molecule was described in the early 1950s; insulin was the first protein to be sequenced entirely. This pioneering work was confirmed from 1963 to 1966, when several groups reported laboratory synthesis of biologically active insulin. The three-dimensional structure of the crystalline hormone was published in 1969.

Insulin has been shown to be a protein consisting of two polypeptide chains (see peptide ), one of 21 amino acid residues and the other of 30, joined by two disulfide bridges (see cysteine ). The two chains are synthesized in the β cells as part of one continuous polypeptide chain called proinsulin; a 32-amino acid sequence (the connecting peptide) is subsequently split out of the proinsulin molecule by an enzyme resembling trypsin to yield active insulin.

Insulin in Diabetes Treatment

Many, but not all, of the symptoms of diabetes can be controlled by the administration of insulin. The forms of insulin available early in the 20th cent. had to be injected frequently because they were quick-acting. Later modifications gave the insulin solution a more prolonged action so that hypodermic injections could be made less frequently. Some now control their insulin levels via a small, portable insulin pump. In certain cases of mild diabetes, oral medications that stimulate production of insulin can be taken in lieu of insulin. See glucagon .

insulin (ins-yoo-lin) n. a protein hormone, produced in the pancreas by the beta cells of the islets of Langerhans, that is important for regulating the amount of sugar (glucose) in the blood. Lack of this hormone gives rise to diabetes mellitus, in which large amounts of sugar are present in the blood and urine. This condition may be treated successfully by subcutaneous injections or continuous subcutaneous infusion of insulin; insulin is now also available in a form that can be inhaled. isophane i. insulin combined with protamine, which reduces its rate of absorption from the injection site and hence prolongs its action in the bloodstream. i. analogue one of a group of insulin medications that are either rapidly absorbed from the injection site and have a short duration of action or are slowly absorbed at a constant rate. i. pen a penlike device to inject a measured dose of insulin. i. shock see shock. i. stress test a test of anterior pituitary gland function involving the induction of a hypoglycaemic episode with injected insulin and subsequent measurements of plasma cortisol and growth hormone.

insulin A protein hormone, secreted by the β (or B) cells of the islets of Langerhans in the pancreas, that promotes the uptake of glucose by body cells, particularly in the liver and muscles, and thereby controls its concentration in the blood. Insulin was the first protein whose amino-acid sequence was fully determined (in 1955). Underproduction of insulin results in the accumulation of large amounts of glucose in the blood (hyperglycaemia) and its subsequent excretion in the urine in abnormally high concentrations (glycosuria). This condition, known as diabetes mellitus, can be treated successfully by insulin injections.

insulin Hormone secreted by the islets of Langherhans in the pancreas and concerned with the control of blood-glucose levels. Insulin lowers the blood-glucose level by helping the uptake of glucose into cells, and by causing the liver to convert glucose to glycogen. In the absence of insulin, glucose accumulates in the blood and urine, resulting in diabetes. Insulin was isolated in 1921 by Canadian physician Frederick Banting and Canadian physiologist Charles Best. Its structure was discovered in the 1940s by English biochemist Frederick Sanger.

in·su·lin / ˈinsələn/ • n. Biochem. a hormone produced in the pancreas by the islets of Langerhans that regulates the amount of glucose in the blood. The lack of insulin causes a form of diabetes. ∎  an animal-derived or synthetic form of this substance used to treat diabetes.

insulin Hormone secreted by the β‐cells of the pancreas which controls carbohydrate metabolism. Diabetes mellitus is the result of an inadequate supply of insulin or failure of its function. Since insulin is a protein it would be digested if taken by mouth so must be injected. See also diabetes; diet, diabetic; glucose tolerance.

insulin A protein hormone that is secreted by the beta-cells of the islets of Langerhans in the pancreas. It stimulates the utilization of glucose by the cells and thereby lowers the blood-sugar level. A deficiency of this hormone results in the condition known as diabetes mellitus.

insulin hormone extracted from the islets of Langerhans in the pancreas of animals. XX. f. L. insula island; see -IN1.

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