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My term paper on "Insulin - Production,Biochemistry & Transport" gives an brief description on how function of insulin how insulin is secreted by the human body and is transported to our blood via various mechanisms.It also explains the biochemistry & role of various enzymes which take part in insulin production and release.The production of insulin by recombinant Dna technology which has proved to be of extreme importance for patients with diabetis has also been discussed.Lastly it discusses about the various effects of Insulin and with my references at the back I end my paper.
The Discovery of Insulin
Â Before insulin was discovered Diabetis was a fearfull disease.Doctors knew that due to sugar increase in blood it was caused so doctors gave a sugat free diet for the patients. This type of treatment could only give the patients a few extra years to live but it could not save them.At times patients died because of harsh diets given to them due to starvarion
During the 19th century, observations on the patients who died of diabetis showed that their pancreas were damaged. In 1869, a German medical student, Paul Langerhans, found the pancreatic tissue which produces digestive juices had a clusters of cells whose function was unknown. Some of these cells were shown to be the insulin-producing beta cells. Later, in honor of the person who discovered them, the cell clusters were named the islets of Langerhans.
In 1889 in Germany, physiologist Oskar Minkowski and physician Joseph von Mering, showed that if the pancreas was removed from a dog, the animal got diabetes. But if the duct through which the pancreatic juices flow to the intestine was ligated - surgically tied off so the juices couldn't reach the intestine - the dog developed minor digestive problems but no diabetes. So it seemed that the pancreas must have at least two functions:
To produce some digestive juices
Production of a substance which regulates glucose.
THE BEGEINING OF EXPERIMENT:
Banting and Best started their experiments by removing the pancreas from dog. This results are as follows:
It's blood sugar increased.
It became thirsty, drank lots of water, and urinated frequently..
It became weaker.
The dog developed diabetes.
Experimenting on another dog, Banting and Best surgically ligated the pancreas, stopping the flow of nourishment, so that the pancreas got degenerated.
After a while, they removed the pancreas & sliced it up & froze the pieces in a mixture of salts & water. When the pieces were half frozen, they were ground up and filtered. The isolated substance were named "isletin.
When the extract was injected into the dog the glucose levels of dog decreased and even it seemed healthier. By giving the diabetic dog a few injections a day, Banting and Best could keep it healthy and free of symptoms. Banting and Best showed their result to Macleod, who was impressed, but he wanted more tests to prove that their pancreatic extract really worked.
TESTING ON HUMANS:
The team was eager to start testing on humans. But on whom should they test? Banting and Best began by injecting themselves with the extract. They felt weak and dizzy, but they were not harmed. In January 1922 in Toronto, Canada, a 14-year-old boy, Leonard Thompson, was chosen as the first person with diabetes to receive insulin. The test was a success. Leonard, who before the insulin shots was near death, rapidly regained his strength and appetite. The team now expanded their testing to other volunteer diabetics, who reacted just as positively as Leonard to the insulin extract.
THE NOBEL PRIZE:
The news of the successful treatment of diabetes with insulin rapidly spread outside of Toronto, and in 1923 the Nobel Committee decided to award Banting and Macleod the Nobel Prize in Physiology or Medicine.
The decision of the Nobel Committee made Banting furious. He felt that the prize should have been shared between him and Best, and not between him and Macleod. To give credit to Best, Banting decided to share his cash award with him. Macleod, in turn, shared his cash award with Collip.
The Nobel Prize in Physiology or Medicine for insulin has been much debated. It has been questioned why Macleod received the prize instead of Best and Collip. However, Macleod played a central role in the discovery of insulin. It was he who supported the project from the beginning. He supervised the work and it is also most likely that Macleod's contacts in the scientific world helped the team in getting a speedy recognition of their discovery.
Insulin is chemically a hormone and is produced by the Islets present in the pancreas present in our stomach.
Insulins main function is to control the levels of blood glucose in our body.
Human insulin is composed of a dimer comprising of one chain of 21 amino acids(A chain) & the other 30 amino acids (B chain).
Insulin allows the cell to use glucose for energy as because the cells cannot utilize glucose without insulin.insulin helps the body cells to ta glucose from blood stream which the cells use for production of energy or else it is sent to the liver to preserve it.the main functions of insulin are:
It helps in initiation of lipogenesis( a process by which acetyl COA is converted to fats)
It reduces lipolysis(a process of breaking down of lipids)
It increases amino acid carry into cells
It modulate transcription
It shifts the cell content of many mRNAs
It Stimulates growth.
Once insulin enters the human brain it stimulates an increase in memory and benefits verbal memory in particular(7).
Insulin helps in regulating the levels of blood glucose in our body
FUNCTIONS OF THE ORGANS & CELLS RESPONSIBLE FOR INSULIN PRODUCTION:
PANCREAS:Pancreas are the organs which are situated inside the stomach
ISLETS: Islets are those cells which produce insulin.they are present within the
Human pancreas in the langerhan cells.
BETA CELLS: Beta cells are those cells which are present in the islet of langerhans,
This Beta cells finally produce Insulin.
IMAGE SHOWING PRODUCTION OF INSULIN:
Fig1:showing production of insulin
The simple systematic process showing insulin production are as follows:
Pancreas have within them cells called the islet cells
This islets have small cells attached with it called the beta cells.
This beta cells helps present in the islets produces insulin.
Now after this insulin gets produced than this insulin granules are transported after certain modifications to the blood vessels and released with glucose
Initially insulin is released as pre-pro insulin in the pancreatic beta cells which are than modified by the organ system to functional pro insulin.
SYNTHESIS IN HUMAN BODY:
Mature insulin is packaged inside mature granules waiting for metabolic signals (such as leucine, arginine, glucose and mannose) and vagal nerve stimulation to be exocytosed from the cell into the circulation(4).
Initially the process begins with transcription of the gene,production of mRNA,then synthesis of insulin proteins.
The ER(endoplasmic reticulum) export golgi transport & vesicle packaging
In Î²-cells, insulin is synthesized from the proinsulin precursor molecule by the action of proteolytic enzymes, known as prohormone convertase (PC1 & PC2) well as the exoprotease carboxypeptidase cpE.(shown in figure 3)
This modifications of insulin removes the centre peptide that is the C peptide from the C & N terminal ends at the specific site which is breaking the bonds between lysine 64 and arginine 31 & 32.
After that pro insulin is released into the blood(1,2,3)
Fig 3:Synthesis of insulin
BIOCHEMISTRY OF INSULIN PRODUCTION
Glucose enters the Î²-cells all the way through the glucose transporter 2(Glut 2)
Glucose go into glyclolysis and the respiratory cycle, where multiple high-energy ATP molecules are formed by oxidation
Increased intracellular ATP:ADP ratio closes the ATP-dependent potassium channels
The cell membrane potential depolarizes.
On the depolarization, voltage-controlled calcium channels (Ca)2+ open and calcium flows into the cells .
An increased calcium level causes activation of phospholipase c, which cleaves the membrane phospholipid phosphatidyl inositol into inositol 1,4,5 triphosphate and diacylglycerol.
Inositol 1,4,5-triphosphate (IP3) binds to receptor proteins in the membrane of (ER). This allows the release of Ca2+ from the ER via IP3-gated channels, and further raises the cell concentration of calcium.
8.Significantly improved amounts of calcium in the cells causes release of previously synthesized insulin, which has been stored in secretory vessels.
Fig 4:Metabolic regulation of insulin secretion.
PRODUCTION OF INSULIN BY RECOMBINANT DNA TECHNOLOGY:
Fig 5:showing insulin production by recombinant DNA technology
Firstly the required DNA of interest is ligated out from the host organism where the gene is present.
Then vectors especially the plasmid of bacterial cell are isolated and the insulin producing gene along with moleqular markers are are inserted into the vector.
And then further the prepared vector is integrated into the bacterial cell such as
E. coli and put into a suitable culture medium for its growth.
The bacteria is engineered in a fermentation tank to produce insulin
Then the prepared insulin is injected into patient.
In this process the trinucleotides representing each codon were first synthesized,then they were joined in a correct order to represent the amino acid sequence of the chains A & B.
The genes for A & B were integrated separately In a vector called pBR322 that had lac Z alpha sequence driven by a strong lac promoter region.this strategy of integration was called as translational fusion.
A & B chains were designed in such a manner that chains A & B were separated by a methionine residue from the beta galactosidase sequence encoded by the lac Zalpha .The insulin sequences were separated from Beta galactosidase by treating the fusion proteins with cyanogens bromide.s
The purified chains of A & B chains were attached to each other by di sulphide bonds induced in vitro and finally insulin could be derived from E.coli cells
Fig 6:A mature insulin protein.
EFFECTS OF INSULIN:
1.ACTION ON PROTEINS: Insulin increases the uptake of amino acids by tissues, which leads to the decrease of their plasma concentration. except alanine, because of its formation starting from pyruvate, and tryptophan whose relative concentration rises because being more bound to plasma albumin, its concentration drops less than that of the other amino acidS
2.LIPID METABOLISM: Insulin induces the lipogenesis and inhibits lipolysis in the liver, adipose tissues and striated muscles. In absence of, fatty acid catabolism ß-oxidationn increases, which induces excessive production of acetyl-CoA at the origin of ketogenesis which is the production of acetone and ß-hydroxybutyrate.
3.POTASSIUM TRANSPORT:Insulin induces potassium uptake bycells which tends to induce hypokalemia.A potassium deficiency decreases the hypoglycemic action of insulin.it also has the same effect as on magnesium.
4.CENTRAL EFFECTS: By activation cerebral receptors, insulin modulates the food behavior. An insulin deficiency elicits release of neuropeptide Y, responsible of the increase in appetite and a decrease of the release by adipocytes of leptine which, by acting on the hypothalamus, reduces appetite and increase thermogenesis.
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 glucose is known as hypoglycemia or, in cases producing unconsciousness, "hypoglycemic coma" (sometimes termed "insulin shock" from the most common causative agent).
Endogenous causes of insulin excess (such as an insulinoma) 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 (e.g., 20 units instead of 2) or other unanticipated factors (did not eat as much as anticipated, or exercised more than expected, or unpredicted kinetics of the subcutaneously injected insulin itself).
Possible causes of hypoglycemia include:
External insulin (usually injected subcutaneously)
Oral hypoglycemic agents (e.g., any of the sulfonylureas, or similar drugs, which increase insulin release from Î²-cells in response to a particular blood glucose level)
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. (This can never be a cause of hypoglycemia in patients with type 1 diabetes, since there is no endogenous insulin production to stimulate.)
Glucagon is a hyperglycemic polypeptic hormone secreted by the alpha cells of the pancreas and intestinal tract, discovered little time after insulin.
Glucagon contains a single-chain of 29 amino acid residues whose secretion is inhibited by glucose and somatostatin and is stimulated by amino acids. Its plasma half-life is a few minutes because it is hydrolyzed in many tissues, particularly the liver.
An excess of glucagon secretion is seen in certain diabetics.
Glucagon stimulates the receptors coupled to Gs proteins and activation of adenylcyclase which leads to an increase of cyclic AMP, responsible of its pharmacological effects.
Glucagon induces an hyperglycemic action by transforming glucagon into glucose.
It can be used in the emergency treatment in of hypoglycemic comas while waiting for the administration of glucose.
Glucagon has an positive inotropic effect which increases cardiac output. It induces an arterial and venous vasodilator effect and decreases the peripheral resistance. It can be used as an inotropic agent in the event of acute myocardial failure and in cases of beta-blocker overdosage..
EFFECT IN THE DIGESTIVE TRACT:
the motility of the digestive tract is reduced by glucagon and can be used to facilitate endoscopical and radiological examinations of the digestive tract.
Glucagon is indicated primarily in the treatment of severe hypoglycaemia when the patient can not take glucose by mouth and when intravenous administration of glucose is not possible.. It is administered by intravenous, intramuscular or subcutaneous routes. Its effect is fast and of short duration, less than one half an hour, thus the need for supplementing its administration by that of glucose.
Glucagon has adverse effects: nausea, vomiting, reactional hypoglycemia after the phase of hyperglycemia, catecholamine release, rarely hypokalemia.