Insulin is the small protein which is required for the regulation of the glucose levels in the body. It is produced by the beta cells in the pancreas. Insulin is composed of two peptide chains referred to as the A chain and B chain. Although the amino acid sequence of insulin varies among species, certain segments of the molecule are highly conserved. Insulin act on various insulin receptors to exhibit its actions. Insulin signaling also controls embryonic growth and development, reproduction, and appetite regulation in the body. Insulin binding to the receptors activates various pathways inside the cell as Ras/MAPK pathway to control the glucose levels. Nowadays more research is going on the action of insulin receptors in cancer therapy as insulin potentiation theory to combine the chemotherapy with insulin for the selective action on the cancer cell growth. Insulin receptors are also presents in CNS to regulate various functions. So insulin receptors are the targeting agents for the cure of various diseases as energy is the main source of growth in case of any cell. It is a rather small protein, with a molecular weight of about 6000 Daltons. It is composed of two chains held together by disulphide bonds. Insulin is composed of two peptide chains referred to as the A chain and B chain. A and B chains are linked together by two disulphide bonds, and an additional disulphide is formed within the A chain. In most species, the A chain consists of 21 amino acids and the B chain of 30 amino acids.
Although the amino acid sequence of insulin varies among species, certain segments of the molecule are highly conserved, including the positions of the three disulphide bonds, both ends of the A chain and the C-terminal residues of the B chain. These similarities in the amino acid sequence of insulin lead to a three dimensional conformation of insulin that is very similar among species, and insulin from one animal is very likely biologically active in other species. Indeed, pig insulin has been widely used to treat human patients. Insulin molecules have a tendency to form dimers in solution due to hydrogen-bonding between the C-termini of B chains. Additionally, in the presence of zinc ions, insulin dimers associate into hexamers.
Biosynthesis of insulin
Insulin is synthesized in significant quantities only in B cells in the pancreas. The insulin mRNA is translated as a single chain precursor called preproinsulin, and removal of its signal peptide during insertion into the endoplasmic reticulum generates proinsulin. Proinsulin consists of three domains: an amino-terminal B chain, a carboxy-terminal A chain and a connecting peptide in the middle known as the C peptide. Within the endoplasmic reticulum, proinsulin is exposed to several specific endopeptidases which excise the C peptide, thereby generating the mature form of insulin. Insulin and free C peptide are packaged in the Golgi into secretory granules which accumulate in the cytoplasm.
Control of insulin secretion
Insulin is secreted in primarily in response to elevated blood concentrations of glucose. This makes sense because insulin is "in charge" of facilitating glucose entry into cells. Some neural stimuli (e.g. site and taste of food) and increased blood concentrations of other fuel molecules, including amino acids and fatty acids, also promote insulin secretion. Understandings of the mechanisms behind insulin secretion remain somewhat fragmentary. Nonetheless, certain features of this process have been clearly and repeatedly demonstrated, yielding the following model:
Glucose is transported into the B cell by facilitated diffusion through a glucose transporter; elevated concentrations of glucose in extracellular fluid lead to elevated concentrations of glucose within the B cell.
An elevated concentration of glucose within the B cell ultimately leads to membrane depolarization and an influx of extracellular calcium. The resulting increase in intracellular calcium is thought to be one of the primary triggers for exocytosis of insulin-containing secretory granules. The mechanisms by which elevated glucose levels within the B cell cause depolarization is not clearly established, but seems to result from metabolism of glucose and other fuel molecules within the cell, perhaps sensed as an alteration of ATP:ADP ratio and transduced into alterations in membrane conductance.
An increased level of glucose within B cells also appears to activate calcium-independent pathways that participate in insulin secretion.
Structure of insulin receptor
Insulin Receptors are areas on the outer part of a cell that allow the cell to join or bind with insulin that is in the blood. When the cell and insulin bind together, the cell can take glucose (sugar) from the blood and use it for energy
The structure of InR is similar to the mammalian insulin receptor (Inr) and the IGF1 receptor (IGF1R). It is a tetramer composed of two subunits containing the putative ligand binding domains and two transmembrane subunits containing the cytoplasmic tyrosine kinase domains. In contrast to human receptors, InR possesses extensions at the amino and carboxy termini. The C-terminal extension contains binding sites for Downstream components similar to those found in insulin receptor substrates (IRS), and has been shown to be able to signal in the absence of IRS proteins. For example, they identified proteins mediating insulin signaling, which are known as insulin receptor substrate proteins—IRS1 and IRS2. IRS1 controls body growth and peripheral insulin action, whereas IRS2 regulates brain growth, body-weight control, glucose homeostasis and female fertility, researchers have found.
Insulin binding to its receptor results in receptor autophosphorylation on tyrosine residues and the tyrosine phosphorylation of insulin receptor substrates (e.g. IRS and Shc) by the insulin receptor tyrosine kinase. This allows association of IRSs with downstream effectors such as PI-3K via its Src homology 2 (SH2) domains leading to end point events such as Glut4 translocation. Shc when tyrosine phosphorylated associates with Grb2 and can thus activate the Ras/MAPK pathway independent of the IRSs.
Signal transduction by the insulin receptor is not limited to its activation at the cell surface. The activated ligand-receptor complex initially at the cell surface, is internalised into endosomes itself a process which is dependent on tyrosine autophosphorylation. Endocytosis of activated receptors has the dual effect of concentrating receptors within endosomes and allows the insulin receptor tyrosine kinase to phosphorylate substrates that are spatially distinct from those accessible at the plasma membrane. Acidification of the endosomal lumen, due to the presence of proton pumps, results in dissociation of insulin from its receptor. (The endosome constitutes the major site of insulin degradation). This loss of the ligand-receptor complex attenuates any further insulin-driven receptor re-phosphorylation events and leads to receptor dephosphorylation by extra-lumenal endosomally-associated protein tyrosine phosphatases (PTPs). The identity of these PTPs is not clearly established yet. A discussion of candidates will be added in the near future.
Insulin potentiation therapy for cancer
Chemotherapy drugs are powerful cell-killing agents in case of cancer but very high doses of these drugs are required in the treatment to force them across the cell membrane of cancer cells. Dose related side effects are produced because these drugs can not discriminate between cancer cells and other normal cells in the patient's body. They kill both kinds of cells, therefore there are side effects. But insulin potentiation theory states that cancer cells also need glucose for their functions. So cancer cells actually manufacture and secrete their own insulin as cancer cells know their requirements for large amounts of glucose to fuel their processes of uncontrolled growth. cancer cells have ten times more insulin receptors per cell than any of the normal cells in the body. This fact creates a valuable opportunity for the chemotherapy of cancer because it significantly differentiates normal cells from the cancerous ones.
The mechanisms that cancer cells use to kill people are the same ones manipulated in IPT to selectively potentiate chemotherapy effects in them, and to more safely and effectively kill the cancer cells. A published article about cancer cells in tissue culture reported that the addition of insulin to the culture medium enhanced the cell-killing effect of methotrexate - a commonly used chemotherapy drug - by a factor of up to ten thousand. This striking result was attributed to two effects on the cancer cells.
■ An effect of insulin to increase the trans-membrane transport of the methotrexate into the cell.
■ "Metabolic modification by insulin" within the cancer cells. There is yet another wonderful and powerful coincidence of cancer cell biology involved in this factor of "metabolic modification" - one that fits right in with the workings of Insulin Potentiation Therapy.
■ The metabolic modification by insulin mentioned above results from the fact that not only can it join up with its own specific receptors on cell membranes, but insulin is also able to join up with the receptors for insulin-like growth-factor, and to communicate messages about growth to these cells. While it may seem highly undesirable for a cancer therapy to actually promote cancer cell growth, this is in fact a valuable effect of insulin here.
Chemotherapy side-effects result from actions on the cells of patient's hair follicles, their bone marrow, and the cells lining the stomach and intestines. This is what causes the hair loss, low blood cell counts, and the nausea and vomiting. What these different cell types all have in common - along with cancer cells - is that they are all rapidly dividing cells.
Chemotherapy drugs like to attack rapidly dividing cells, indiscriminately. In a tumor, not all the cancer cells are in this rapidly dividing stage all at once. They take turns. When insulin joined up with the excess of insulin-like growth-factor receptors on those cancer cells in the tissue culture, it stimulated growth in many of the cells that were not in this growth phase. This "metabolic modification by insulin" rendered more of these cells susceptible to chemotherapy attack, contributing to their increased death rate as observed in the experiment
In Insulin Potentiation Therapy, insulin administration does cause the blood glucose to go down. This is called hypoglycemia. This hypoglycemia is an anticipated side-effect of the insulin, one rapidly and effectively controllable with intravenous glucose infusions at an appropriate time, according to the IPT protocol. The principal role insulin plays in IPT is that of a biologic response modifier. It modifies the biologic response of cancer cells in such a way that lowered doses of anticancer drugs, administered in conjunction with insulin, will kill the cancer cells more effectively. Insulin modifies the cell membrane allowing more anticancer drugs into the cell. It also modifies the growth characteristics in tumors making more of the cancer cells vulnerable to anticancer drug effects. Due to the great excess of insulin-sensitive receptors on cancer cell membranes, we are now able to create a clear differentiation between cancer and normal cells using insulin.
Role of insulin in muscle contraction
The potential mechanisms are: A general mitochondrial dysfunction, reducing ATP production; An impairment in the central nervous system (CNS) in the glucose- or insulin-induced excitation of muscle efferents, leading to reduced ß-adrenergic activation of the muscle; A reduced increase in mitochondrial ATP synthesis in response to activation of the ß-adrenergic receptor (ßAR); A defect in insulin-induced opening of the terminal arterioles controlling blood flow through muscle fibre capillaries and, thus, preventing increases in the insulin concentration in the interstitial fluid and in binding of insulin to the insulin receptor (IR) in the muscle membrane; and a molecular defect in the insulin signalling cascade in the muscle, leading to reduction in the insulin-induced stimulation of muscle glucose uptake, glycogen synthesis, and protein synthesis. ADP, adenosine diphosphate, nitric oxide.
Indirect actions of insulin on liver
The indirect actions of insulin on HGP are diverse. Glucagon secretion from the cell of the pancreas is diminished by insulin, which in turn causes a reduction in HGP .Likewise, nonesterified fatty acid (NEFA) release from the adipocyte is reduced by insulin, and a reduction in the supply of NEFAs to the liver causes an increase in hepatic glycolytic flux, resulting in glucose-6-phosphate exiting the liver after being converted to lactate rather than glucose.
Direct actions of insulin on liver
In the absence of any change in arterial plasma glucagon or insulin levels, plasma NEFA level, gluconeogenic precursor load reaching the liver, or insulinization of the brain, hepatic glucose output rose 3-fold when portal vein insulin fell by 75%, and decreased by 50% when portal vein insulin rose by 75% (7, 2). These data leave no doubt that the liver responds directly, rapidly, and sensitively to the insulin in the plasma perfusing it.
Effects of insulin receptors in CNS
Proteins called receptors straddle cell membranes and pass on messages from hormones like insulin to the rest of the cell. The researchers used genetic tricks to deactivate genes that encode receptors in many worm tissues, creating a group of long-lived individuals. Next, they put the genes back in, but only in certain areas of the body. In separate experiments, they turned insulin receptor genes back on in the intestine but not the brain, then in the brain but not the muscle, and so on. The results were striking: While putting insulin receptors back in muscle and intestinal cells had only a small effect on longevity, replacing the receptors in the nervous system restored the worms’ normal life span. Since the worm insulin receptor seems to function in the nervous system, it seemed likely that these proteins would be active in nervous tissues. To see if this is the case, the researchers altered the genes for about a dozen of the insulin family members so that they produced a glowing green protein instead of their normal insulin-like product. Looking through the transparent worms to see where the protein was produced revealed that they are expressed primarily in sensory neurons and in the nerve ring that functions as the roundworm’s brain. Based on this information, the researchers proposed that members of the insulin family might help neurons direct the organism to go into a dormant state in adverse environmental conditions or to continue growth and development in good conditions.
The new picture of a more diverse insulin family with direct functions in the nervous system provides impetus for investigations into the mysteries of human insulin. Caloric restriction, which effects insulin level, has known effects on human longevity, which may be related to the direct effects of roundworm insulin on lifespan. The discovery that the insulin receptor family is more diverse than previously thought will provide immediate impetus for a search for new insulin relatives in humans. The strong role that insulin plays in the fruitfly and roundworm brain will spur on a search for the functions that insulin family proteins - both known and yet to be discovered - play in the human nervous system.
Insulin receptors antibodies
Auto antibodies to the insulin receptor have been demonstrated to antagonize the physiologic actions of insulin, most often resulting in hyperglycemia unresponsive to massive doses of insulin (type B insulin resistance). Patients with these anti-insulin receptor antibodies typically have a coexistent autoimmune disorder, most commonly systemic lupus erythematosus (SLE) or undifferentiated autoimmune syndromes. Attempting to determine the prevalence and significance of anti-insulin receptor antibodies, sera from consecutive patients with SLE and early undifferentiated connective tissue disease (UCTD) were tested for the presence of anti-insulin receptor antibodies by radio-immuno assay. Thirty-eight patients participated in the study.
(The author is with MCOPS, Manipal University, Manipal, Karnataka, India)