Essay: Diabetes and Metabolic Abnormalities: An Overview of Carbohydrate, Protein, and Lipid Metabolism

CHAPTER 1
Diabetes and Metabolic abnormalities
Diabetes mellitus is an endocrine disorder and is a major source of morbidity in the developed countries. It is not only associated with carbohydrate metabolism but also with lipid and protein metabolism.
Diabetes and Carbohydrate metabolism
Carbohydrate from various sources is the primary exogenous source of glucose. Glucose is the main fuel for energy requirement of the body. Therefore a continuous supply of glucose is necessary to ensure proper function and survival of all organs. The capacity of the liver in the glucose metabolism by glycogen synthesis and glycolysis in the absorptive state is sufficient to account for the clearance of more than one third of the dietary glucose that is absorbed from the gut (Agius, 2007). In normal subjects fasting blood glucose is maintained constant by control of hepatic glucose output. After an overnight fasting approximately 75% of hepatic glucose output is utilized by glycogenolysis and rest by gluconeogenesis from lactate, alanine, glycerol and pyruvate in decreasing order of importance( Hers and Hue, 1983).Alteration in carbohydrate metabolism in diabetes are frequently accompanied by changes in the activity of the enzymes that control glycolysis and gluconeogenesis in liver and muscle( Prince and Kannan,2006).Alteration in glucogenic and glucolytic pathways results in increased rate of hepatic glucose production which lead to the development of overt hyperglycemia especially fasting hyperglycemia in patients with type-2 diabetes(Abdollahi et al.,2013).There are several enzymic checkpoints to control glycolysis (hexokinase, glucokinase), glycogenesis (glycogen synthase kinase -3), glycogenolysis ( glycogen phosphorylase) and glyconeogenesis, fructose1,6-bisphospatase,glucose-6-phosphatase).The activity of certain enzymes is directly controled by insulin via phosphsrylation and dephosphorelation mechanisms( Zhang,2002).
Diabetes and protein metabolism
Insulin acts on protein metabolism by increasing the rate of protein synthesis and decreasing the rate of protein degradation. Proteins are an important targets for oxidative challenge. ROS modify amino acid side chains of proteins such as arginine, lysine, threonine and proline residues to form protein carbonyls (Chevion et al.,2000).Oxidation of cystein residues may lead to the reversible formation of mixed disulphide between protein thiol groups(-SH) and particular GSH (S-glutathiolation) (Dalledonne et al.,2005).The increased rate of proteolysis leads to elevated concentration of amino acids serve in plasma. These aminoacids serve as precursors for hepatic and renal gluconeogenesis. In Livert he increased gluconeogenesis further contributed to the hyperglycemia seen in NIDDM ( Gastal delli et al.,2004)
Diabetes and lipid metabolism
Lipid abnormality is a major problem in patients with diabetes. Among individuals with diabetes 97% had at least one lipid abnormality( Fagot et al.,2000). Patients with diabetes mellitus have a higher total mortality than the general population. This mortality rate can be attributed to cardiovascular disease. Prolonged hyperglycemia may directly injure coronary arteries and there by promote atherogenesis. Furthermore since diabetic patients are frequently hyperlipidemic and are at higher risk even when apparently normolipidemic factors associated with diabetes ( Yashino et al.1996).Patients with diabetes had a two- to three-fold increase of cardiovascular disease as compared to non-diabetic subjects. Patients with diabetes do not have concentrations of LDL-C different from non-diabetic patients. However the LDL-C in diabetic patients is more atherogenic secondary to altered composition, being smaller, denser and highly oxidized (Lamarche et al., 1997). In addition patients with type 2 diabetes also have elevated triglycerides, LDL-C and decreased HDL-C level which is additional risk factors for cardiovascular disease (Henry, 2001).Hence diabetes mellitus (DM) is a major risk factor for cardiovascular disease (CVD) (Grundy et al., 1999). Incidence and mortality due to cardiovascular disease is two to five times higher in patients with diabetes than in nondiabetic subjects of similar age (Laakso and Lehto, 1997).
MANAGEMENT OF DIABETES MELLITUS
Elements of the management plan include short-term and long-term goals, diet, lifestyle changes (exercise, smoking cessation) and medications (e.g., oral antidiabetic agents and insulin).
Diet and life style changes
Medical nutrition therapy is an essential component of diabetes management unfortunately; patient adherence to nutrition principles is one of the most challenging aspects of diabetes care. A goal of medical nutrition therapy is to achieve and maintain blood glucose concentrations as close to normal as possible by balancing food intake with antidiabetic drug therapy and physical activity levels. No more than 30% of the total daily caloric intake should come from fats, 10% to 20% from proteins and the balance of daily calories from carbohydrates. Exercise improves insulin sensitivity and glycaemic control, especially in patients with mild diabetes or a high degree of insulin resistance (ADA, 2012).
Oral hypoglycemic agents
The oral antidiabetic agents that are used in the treatment of type 2 diabetes fall into four categories: beta cell stimulators (Sulfonylureas , Meglitinide), biguanides (Metformin), α-glucosidase inhibitors and thiazolidinediones Fig 8 (Vaaler, 2000). The -cell stimulators act at the level of the pancreatic beta cells to stimulate insulin release. They require the presence of functioning -cells and used only in the treatment of type 2 diabetes and have the potential for producing hypoglycaemia . The sulfonylureas reduce blood glucose by stimulating the release of insulin from beta cells in the pancreas and increasing the sensitivity of peripheral tissues to insulin. Chronic treatment with sulphonylureas may desensitize the -cells and high sulphonylurea concentration may inhibit insulin biosynthesis in vivo and in vitro (Stenman et al., 1990; Andersson and Borg, 1980). Moreover, these agents can produce a series of side effects including hematological, gastro-intestinal disturbances, hypoglycaemic coma. In addition, they are not suitable for use during pregnancy (Larner, 1985). Repaglinide and nateglinide are nonsulfonylurea beta cell stimulators. These agents which are rapidly absorbed from the gastrointestinal tract are taken shortly before meals. Both repaglinide and nateglinide can produce hypoglycaemia, thus proper timing of meals in relation to drug administration is important.
Figure 1. Action sites of oral hypoglycemic agents and mechanisms of lowering blood glucose in type 2 diabetes mellitus (Stumvoll et al., 2005)
Metformin, the only currently available biguanide inhibits hepatic glucose production and increases the sensitivity of peripheral tissues to the actions of insulin. Secondary benefits of metformin therapy include weight loss and improved lipid profiles. Unlike the sulfonylureas whose primary action is to increase insulin secretion, metformin exerts its beneficial effects on glycemic control through decreased hepatic glucose production (main effect) and increased peripheral use of glucose. This medication does not stimulate insulin secretion; therefore, it does not produce hypoglycemia. Because of the risk for lactic acidosis, metformin is contraindicated in people with elevated serum creatinine levels, clinical and laboratory evidence of liver disease or conditions associated with hypoxemia or dehydration.
The α-glucosidase inhibitors block the action of the brush border enzymes in the small intestine that break down complex carbohydrates. By delaying the breakdown of complex carbohydrates, the α-glucosidase inhibitors delay the absorption of carbohydrates from the gut and blunt the postprandial increase in plasma glucose and insulin levels. The postprandial hyperglycemia probably accounts for sustained increases in HbA1c levels.
The thiazolidinediones (TZDs)or glitazones are the only class of drugs that directly target insulin resistance, a fundamental defect in the pathophysiology of type 2 diabetes. The TZDs improve glycemic control by increasing insulin sensitivity in the insulin-responsive tissues liver, skeletal muscle and fat allowing the tissues to respond to endogenous insulin more efficiently without increased output from already dysfunctional beta cells. A secondary effect is the suppression of hepatic glucose production. The mechanism of action of the TZDs is complex and not fully understood but is believed to be associated with binding of the drug to a nuclear receptor that plays a role in the regulation of genes involved in lipid and glucose metabolism (Schoonjans and Auwerx, 2000). Because of a potential problem with liver toxicity, liver enzymes should be measured when using these drugs.
Combination therapy
Using a combination of oral agents with different mechanism of action provides additive efficacy in reducing haemoglobin A1c levels. Adding a second agent will generally lower haemoglobin A1c levels by an additional 0.5% to 2%, depending on the class of oral agents used. Effective Food and Drug Administration-approved oral combination therapies include sulfonylureas (glyburide, glipizde, glimepriride) and metformin (DeFronzo and Goodman, 1995), nateglinide and metformin (Horton et al., 2000), repaglinide and metformin (Moses et al., 1999), metformin and thiazolidinediones (Einhorn et al., 2000) sulfonyureas and acarbose (Willms and Ruge, 1999), metformin and acarbose (Phillips et al., 2003), and sulfonylureas and thiazolidinediones (Horton et al., 1998).
DPP-4 inhibitors
Inhibitors of dipeptidyl peptidase 4 also DPP-4 inhibitors, are a class of oral hypoglycemics that block DPP-4. They can be used to treat type 2 diabetes mellitus. The first agent of the class – sitagliptin – was approved by the FDA in 2006. Their mechanism of action is thought to result from increased incretin levels (GLP-1 and Gastric inhibitory polypeptide) which inhibit glucagon release, the effect of which, in turn, decreases blood glucose, but, more significant, increases insulin secretion and decreases gastric emptying. Drugs belonging to this class are: sitagliptin, vildagliptin,, saxagliptin, linagliptin and alogliptin.
Insulin therapy
Insulin is an important hormone needed by the human body to utilize carbohydrates, protein and fats. However, in type 1 diabetes the pancreas does not produce insulin and replacement therapy is required with exogenous insulin. Type 2 diabetics on the other hand have a problem with either the secretion of insulin or have become insulin-resistant; thus, the common name for the condition is non insulin dependent diabetes mellitus. Insulin injections are a necessary daily component of therapy for type 1 diabetics. Insulin injections however, are not always necessary for treatment and control of diabetes in type 2 diabetics (Buse, 1999).
Method of insulin injection
There are a number of different methods of getting insulin into the body, including the use of injections and pumps. Depending on the type and severity of patients the mode of insulin injection will be recommended.
Syringe
Syringes are a common method of quickly delivering insulin directly to the blood stream. In order to use a syringe, diabetics need to be sure that the injection site is clean and sterile before they can inject the needle into their skin. Often, Diabetics will inject themselves with insulin in the thigh or stomach where there is an increased amount of fat to absorb the needle.
Insulin pen
An insulin pen looks like a writing pen, but instead of being filled with ink, the pen is filled with insulin. The delivery method is similar to that of the syringe, except that users can control the amount of insulin by turning a dial on the pen itself. The pen is ideal for diabetics that are quite mobile, as it can easily slip into a bag or pocket and can be used without drawing too much attention. in increased doses after a meal. Note: This method is not recommended if you will be playing contact sports or participating in other physical activities.
Jet injectors
A jet injector is a less common method of delivering insulin, but is ideal for diabetics that do not want to use needles. The jet injector is a device that sends insulin into the body using a high-pressure spray of insulin across the skin. The insulin gets absorbed by the skin, thereby eliminating the need for needles and other devices.
Insulin pump
An insulin pump is an especially useful insulin delivery method for someone that requires careful insulin regulation and has proven to be an effective method of treatment for Type 1 diabetes. The insulin pump is a small device that is attached to the body through a catheter that is located under the skin of the abdomen. The pump sends the right amount of insulin into your body through flexible tubing continuously throughout the day
OXIDATIVE STRESS AND DIABETES
Oxidative stress is the common pathomechanism that greatly influences the progression of both cardiovascular and metabolic diseases (Boudina and Abel, 2007).The sustained hyperglycemia has been identified as a principle mediator of increased reactive oxygen species (ROS) generation in diabetes (Kumar, Banu, & Murugesan, 2008; Maritim, Sanders & Watkins, 2003).Over production and insufficient removal of free radicals in diabetes mellitus results oxidative stress. Under chronic hyperglycemic condition oxygen radical species (ROS) are accelerated through multiple sources including enzymatic, on enzymatic and mitochondrial pathways thus causes cellular damage through the oxidation of protein, lipid and DNA leading to the disease complication ( Kaneto, Katakami, Matsuhisa, & Matsuoka, 2010).
FREE RADICALS
Free radicals are the molecular fragments or byproducts of the endogenous metabolism having one or more unpaired electrons in atomic or molecular orbital’s plays a significant role in cell signaling. (Vinay et al., 2013).There are other reactive molecules particularly derived from oxygen which are not radicals. Hydrogen peroxide (H2O2), Singlet oxygen (O21) is characterised by the anti-parallel spin of its two unpaired electrons, which is highly reactive as compared to normal triplet oxygen. These uncoupled electrons are very reactive with adjacent molecules lipids, proteins and carbohydrates and can cause cellular damage (Kuhn, 2003). NO is an endothelial relaxing factor and neurotransmitter produced through nitric oxide synthase enzyme. NO and O21 radicals are converted to powerful oxidizing radicals like hydroxyl radicals (OH•), alkoxyl (RO•) and peroxyl (ROO•),singlet oxygen (O21) by complex transformation reactions. Some of the radical species are converted to molecular oxidants like hydrogen peroxide (H2O2), peroxy nitrite (ONOO-), Hypochlorous acid (HOCl). (Amit Kumar and K.L. Priyadarsini., 2011).
LIPID PEROXIDATION AND ANTIOXIDANTS
Lipid peroxidation is the oxidative deterioration of lipids containing a number of carbon-carbon double bonds. Among biological molecules lipids are the most susceptible to oxidative damage or peroxidation. Oxidative deterioration of polyunsaturated fatty acids (PUFAs) which are present in abundance in cell membranes initiates a self-perpetuating chain reaction that yields a wide range of cytotoxic products such as malondialdehyde (MDA), 4-hydroxynonenal etc. Increased lipid peroxidation is generally believed to be an important underlying cause of the initiation of oxidative stress related various tissue injury and cell death and further progression of many acute and chronic diseases (Halliwell and Gutteridge, 1999).
ANTIOXIDANTS
Nature has endoured with adequate protective mechanism against harmful effects of free radicals. Antioxidants are subatances that neutralize free radicals or their actions and can act as different stages (Van Toan and Hanh, 2013). Cells have developed several enzymic antioxidants, superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx), glutathione S-transferase (GST)) and non enzymic antioxidants (vitamin C, vitamin E, β-carotene, reduced glutathione (GSH)) to prevent or limit ROS-associated damage (de Haan et al., 2003). Free radicals scavenging enzymes such as SOD, CAT, Gpx and GST are first line of cellular defense against oxidative injury. The equilibrium between these enzymes is an important process for the removal of oxidative stress in intracellular organelles (Rakhshandehroo,2013). Superoxide dismutase is known to catalyse the dismutase of superoxide to H20 and 02, since SOD is present in all aerobic organisms and in most subcellular compartments that generate activated oxygen, it has been assumed that SOD has a central role in defense against oxidative stress (Alzoghaibi et al.,2013). There are three different types of SOD classified on the basis of their metal cofactor: the Copper/Zinc(Cu/Zn-SOD, The manganese (Mn-SOD) and the Iron (Fe-SOD) isoezymes. Most common are the Cu/Zn containing SOD present in the cytosol in the mitochondrial (Kocot et al.,2013).
Catalase is a hydrogen peroxide decomposing enzyme mainly localized to peroxisomes or microperoxisomes (Tesfamariam, 1994). GSH is a tripeptide (Glu-Cys-Gly), whose antioxidant function is a facilitated by sulfhydral groups of cysteine (Rennenberg, 1982).GSH has a redox potential of 340 mv that enables GSH to reduce dehydroascarbate to ascorbate or to reduce the disulphide bonds of proteins. It can react chemically with singlet oxygen superoxide and hydroxyl radicals and therefore function directly as a free radical scavenger.GSH may stabilize membrane structure by removing acy-peroxides formed by LPO reactions (Jing et al.,2007). GSH-metabolizing enzymes, GPx and GST work in concert with glutathione in the decomposition of hydrogen peroxide and other organic hydroperoxides to non-toxic products, respectively at the expense of reduced glutathione (Halliwell and Gutteridge, 1999). GSH plays a key role in the liver in detoxification reactions and in regulating the thiosulphide status of the cell (Chavan et al., 2005).
Ascorbic acid is well known water soluble antioxidant vitamin that scavenger O2 and other ROS (Hoffer, 2013). Vitamin C, in addition to directly scavenging the free radicals in the cytoplasm also participates in the regeneration of vitamin E and antioxidant peptide glutathione (Bendich, 1997). By reacting with activated oxygen more readily than any other aqueous component, ascorbic protects certain macromolecules from oxidative damage. Ascorbic acid has a lower redox potential that α-tocopherol and acts as an efficient co- antioxidant in vitro for the generation of α-tocopherol from the tocopheroxyl radical( Jablonnowska-Lietz, 2013).
Vitamin E is a fat soluble vitamin that exists in eight different forms α-tocopherol is the most active form of vitamin E, in human and is considered to be more powerful membrane bound antioxidant by the cell (Jeon,2013). The main principle of vitamin E is to break and terminate the free radical chain reaction in most tissues (Halliwell and Gutteridge, 1999).So Vitamin E functions as a chain breaking antioxidant that prevents the propagation of free radical reactions.It quenches the lipid peroxidants and protects the cell structure from attach(Pryor,2000).
PATHWAYS LEADING TO HYPERGLYCEMIA-INDUCED CELL DAMAGE
A large amount of data emphasize four key metabolic pathways as being major contributors to hyperglycemia induced cell damage (Nishikawa et al., 2000; Brownlee,2001;Robertson,2004): (1) increased polyol pathway flux; (2) increased advanced glycation end product (AGE) formation; (3) activation of protein kinase C (PKC) isoforms; and (4) increased hexosamine pathway flux.
Polyol pathway
In the polyol pathway, glucose is converted to sorbitol, then to fructose; and it involves the oxidation of nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) to NADP+. Increased activity of this metabolic pathway soon depletes the NADPH needed to regenerate the antioxidant glutathione and without adequate glutathione, nerves have a diminished ability to scavenge ROS, promoting oxidative stress (Vincent and Feldman, 2004). In addition, as glucose is metabolized via the polyol pathway, it causes sorbitol to accumulate. Excessive amounts of sorbitol can lead to cellular osmotic stress (Obrosova et al., 2003): this alters the antioxidant potential of the cell; further increasing ROS accumulation (Figure 2). Excessive fructose produced in the polyol pathway also leads to nonenzymatic glycation/glycoxidation that accelerates ROS-mediated damage of cellular proteins and lipids (Feldman et al., 2003).
Figure 2. Polyol pathway
The AGE pathway
Intracellular hyperglycemia appears to be the primary initiating event in the formation of AGEs (Brownlee, 2001) via the advanced glycation end product pathway (Figure 3). Glycation/glycosylation occurs as glucose combines with proteins (Schiff bases) forming early glycation products at a rate proportional to glucose concentration. These Schiff bases rearrange, becoming more stable Amadori-type early glycation products. These reactions are reversible and there is no evidence that shows these early products correlate to diabetes complications (Brownlee et al., 1988). However, some of these early glycation products undergo a slow, complex series of chemical reactions, becoming AGEs. Because AGEs are irreversible, they do not normalize when hyperglycemia is corrected, but rather accumulate overtime. AGEs bind to a number of receptor proteins including the receptor for advanced glycation end products (RAGE) (Lander et al., 1997). In mesangial and endothelial cells, activation of RAGE by AGEs results in a burst of ROS production.
Figure 3. Advanced glycation end product pathway
Protein kinase C (PKC) pathway
The PKC family of serine-threonine kinases is among the integrin targeted/activated intracellular signaling molecules clustered at focal adhesions (Schlaepfer and Hunter, 1998). Eleven isotypes of PKC with different tissue distribution and biological functions have been reported (Nishizuka, 1992). High glucose concentration stimulates the PKC gene expression, especially of beta iso-form (Koya and King, 1998). Normally, the PKC activation is involved in the regulation of vascular permeability, contractility, extracellular matrix prodution, cell growth, angiogenesis, cytokine action, and leukocyte adhesion, all of which are involved in the pathophysiology of diabetic complications (Way et al., 2001). The process of PKC activation is preceded by diacylglycerol (DAG) formation generated from the phosphoinositides.
Figure 4. Protein kinase C pathway
An enhanced PKC activity increases the production of cytokines and of extracellular matrix as well as the contractility and permeability of the vascular wall (Figure 4). All these consequences of the PKC activation may be blocked by specific PKC inhibitors and by alpha tocopherol (Gabriele and King, 2001). The role of ROS in the DAG-PKC pathway is suggested. Additionally, sorbitol is metabolized to fructose by sorbitol dehydrogenase, increasing the ratio of NADH/NAD+. This results in oxidized triose phosphates with de novo synthesis of diacylglycerol (DAG). Increased DAG content activates protein kinase C, and leads to several pathologies of diabetic complications (Brownlee, 2001).
Hexosamine pathway
The hexosamine pathway is activated when excessive metabolites of glycolysis accumulate (Feldman et al, 2003; Leinninger et al., 2004). Intermediates in the pathway lead to changes in gene expression and protein function that contribute to the pathogenesis of diabetic complications (Brownlee, 2001). For example, many of the acylglycoslated proteins that are produced in the pathway are transcription factors that increase proteins associated with complications of diabetes. These proteins are often inflammatory intermediates and include transforming growth factor-β1 that promotes nephropathy and plasminogen-activator inhibitor that inhibits normal blood clotting, increasing vascular complications (Leinninger et al., 2004). Thus, the activation of this pathway increases nerve oxidative stress through vascular disease leading to microvascular occlusion that produces ROS (Figure 5). The hexosamine pathway appears to be particularly important in type 2 diabetes through 2 major mechanisms (Leinninger et al., 2004). The rate-limiting enzyme glutamine: fructose-6-phosphate amidotransferase (GFAT) is specifically increased in the muscles of diabetic mice. The overexpression of GFAT promotes insulin resistance, and hyperinsulinemia. In addition, activation of the hexosamine pathway induces oxidative stress via the generation of intracellular hydrogen peroxide. Several hexosamine pathway-mediated changes can be suppressed by treatment with antioxidants (Kaneto et al., 2001).
Figure 5. Hexoseamine pathway
DIABETOGENIC AGENTS
Many chemicals are used for the induction of diabetes mellitus in the animal models for testing new antihyperglycemic drugs. The most commonly used chemicals for the induction of diabetes in the experimental animals are alloxan and streptozotocin. The most disadvantage of alloxan diabetes model is multiorgan damage; hence this diabetogen is not widely employed to study the anti-diabetic effect of newer agents (Grussner et al., 1993).
Streptozotocin (STZ)
It is now well established that streptozotocin selectively destroys the pancreatic cells and produces hyperglycaemia (Gilman, 1990). Hence, streptozotocin is commonly employed for the induction of diabetes mellitus in experimental rats (Tomlinson et al., 1992). Streptozotocin is produced by a strain of Streptomyces achromogens, is used for induction of diabetes mellitus in experimental animals (Szkudelski, 2001). STZ, a glucosamine-nitrosourea compound, has a chemical name of 2-deoxy-2-(3-methyl-3-nitrosoureido)-D-glucopyranose (C8H15N3O7). The structure is composed of a nitrosourea moiety with a methyl group attached at one end and a glucose molecule at the other as shown in Figure (Weiss, 1982).
GLUCOSE TRANSPORTERS
Glucose is the main source of energy in eukaryotic organisms and plays a central role in metabolism and cellular homeostasis. Most mammalian cells are dependent on a continuous supply of glucose which acts as primary source for the generation of adenosine-5’-triphosphate (ATP) (Joost et al., 1994). Glucose homeostasis is maintained by the coordinated regulation of 3 processes. First glucose absorption via the small intestine; second glucose production in the liver and third consumption of glucose by nearly all tissues (Kahn 1992). As glucose is the main regulator of insulin secretion and production excessive amounts of glucose over a prolonged period have negative effects on pancreatic β-cell function resulting in increased sensitivity to glucose, increased basal insulin release, reduced maximal secretory response, and a gradual depletion of insulin stores. Thus blood glucose concentrations need to be maintained with narrow limits. Because the lipid bilayer of the eukaryotic plasma membrane is impermeable for hydrophilic molecules, glucose is a hydrophilic compound; it cannot pass through the lipid bilayer by simple diffusion and therefore required specific carrier proteins to mediate its specific transport into the cytosol. All mammalian cells contain one or more members of the facilitative glucose transporter gene family named GLUT. These transporters have a high degree of stereoselectivity providing for the bidirectional transport of substrate with passive diffusion down its concentration gradient. The GLUTs are intrinsic membrane proteins which differ in tissue-specific expression and response to metabolic and hormonal regulation (James et al., 1994; Mueckler, 1994; Stephens & Pilch, 1995). The family of facilitative glucose transporter (GLUT) proteins is responsible for the entry of glucose into cells throughout the periphery and the brain (Shepherd and Kahn, 1999; Vannucci et al., 1997b). The expression, regulation and activity of glucose transporters play an essential role in neuronal homeostasis, since glucose represents the primary energy source for the brain (Pardridge, 1983).
Tissue-specific expression of the GLUT family members.
Protein Expression Function Reference
GLUT-1 All tissues
(abundant in brain and erythrocytes) Basal uptake Mueckler et al, 1985
GLUT-2 Liver, pancreatic islet cells,
retina Glucose sensing Fukumoto et al., 1989
Watanabe et al, 1999
GLUT-3 Brain Supplements GLUT1 in
tissues with
high energy demand Kayano et al., 1998
GLUT-4 Muscle, adipose tissue, heart Insulin responsive Fukumoto et al., 1989
GLUT-5 Intestine, testis, kidney,
erythrocytes Fructose transport Kayano et al., 1990,
Concha et al., 1997
GLUT-6 Spleen, leukocytes, brain Doege et al., 2000
Protein Expression Function Reference
GLUT-7 Liver Mediates glucose flux across endoplasmic reticulum membrane Joost & Thorens, 2001
GLUT-8 Testis, brain Glucose transport Doege et al., 2000a
GLUT-9 Liver, kidney Regulator of urate homeostasis Phay et al., 2000
GLUT-10 Liver, pancreas Glucose transport McVie-Wylie et al., 2001
GLUT-11 Heart, muscle Fructose transport Doege et al., 2001
GLUT-12 Heart, prostate Rogers et al., 1998
Diabetes and abnormalities in glucose-stimulated insulin secretion.
Glucose stimulation of insulin secretion begins with its transport into the beta cell by the GLUT-2 glucose transporter. Glucose is the key regulator of insulin secretion by the pancreatic beta cells although amino acids, ketones, various nutrients, gastrointestinal peptides and neurotransmitters also influence insulin secretion. Glucose level >3.9 mmol/L (70 mg/dL) stimulate insulin synthesis primarily by enhancing protein translocation and processing. Glucose stimulation of insulin secretion begins with its transport into beta cell by the Glut-2 glucose transporter. Glucose phosphorylation by glucokinase is the rate-limiting step that controls glucose-regulated insulin secretion. Further metabolism of glucose-6-phosphate via glycolysis generates ATP which inhibits the activity of an ATP-sensitive K+ channel. Inhibition of this K+ channel induces beta cell membrane depolarization which opens voltage-dependent calcium channels (leading to an influx of calcium), and stimulates insulin secretion.
Insulin signaling pathways that regulate glucose metabolism in muscle cells and adipocytes
Skeletal muscle is the primary tissue responsible for the postprandial uptake of glucose from the blood. The two major transporters expressed in skeletal muscle are the muscle/fat specific glucose transporter GLUT 4 and GLUT 1 (Pessin and Bell, 1992). Insulin-stimulated intracellular movement of GLUT 4 is initiated by the binding of insulin to the extracellular portion of the transmembrane insulin receptor. Its binding activates tyrosine kinase phosphorylation at the intracellular portion of the receptor. The chief substrates for this tyrosine kinase include insulin-receptor–substrate molecules (IRS-1, IRS-2, IRS-3, and IRS-4), Grb-2 (growth factor receptor–bound protein-2) and associated binder-1 (Gab-1), and SHC (Src and collagen-homologous protein) (Holman and Holman, 1997, Shepherd et al., 1998) and other proteins. These substrates form complexes with docking proteins such as phosphoinositide-3 kinase at its 85-kD subunit (p85) by means of SH2 (Scr homology region 2) domains. Then p85 is constitutively bound to the catalytic subunit (p110). Activation of phosphoinositide-3 kinase is a major pathway in the mediation of insulin stimulated glucose transport and metabolism. The function of the activated phosphoinositol-3 kinase is to phosphorylate the phosphotidyl inositol 3, 4, 5–triphosphate (PIP3) and to lesser extent phosphotidyl inositol 4-phosphate to phosphoinositol 4, 5 bisphosphate. PI-3 kinase activates phosphoinositide-dependent kinases that participate in the activation of protein kinase B (also known as Akt) and atypical forms of protein kinase C (PKC). Over expression of constitutively active forms of PKC ζ or PKC λ increases transporters activity and GLUT 4 translocation by 50-100% of the extent observed in response to maximal insulin. Exercise stimulates glucose transport by pathways that are independent of phosphoinositide-3 kinase and that may involve adenosine monophosphate (5′-AMP) activated kinase .Thus the insulin-stimulated acute activation of glucose transport mainly occurs by one of two mechanisms: translocation of GLUT 4 and GLUT 1 from intracellular vesicles to the plasma membrane and augmentation of the intrinsic catalytic activities of the transporters (Mitsumoto et al., 1991, Mitsumoto et al., 1992).
INSULIN SIGNALING AND MECHANISM OF INSULIN SIGNALING
Insulin is a peptide secreted by β-cells from pancreas and it has a central role in the regulation of fuel homeostasis. It is particular effective in keeping glucose level within narrow limits. After a meal circulating levels of insulin rise in response to the increased level of plasma glucose promoting its uptake and storage. It is also responsible for the activation of anabolic pathways such as glycogen, lipids, and protein synthesis while it inhibits the oxidation of these metabolites. All these effects are mediated by the binding of this hormone to the insulin receptor (IR) a process characterised by high specificity and affinity while prompts an intra-cellular cascade of the events that signals the presence of plasma insulin. Insulin signaling is initiated by the binding of insulin to the extra cellular α subunit of the IR on the cell surface resulting in the auto phosphorylation of a number of residues in the Trans membrane of β-subunits (Gammel and Van abberghen, 1986).The active tyrosine kinase phosphorylates insulin receptor substrate (IRS) proteins. The IRS proteins function as docking proteins for other signalling proteins containing Src homology 2 domains. IRS-1 and IRS-2 best characterized and are the main docking proteins for the binding of the p85 regulatory subunit of p13K which leads to the activation of the catalytic subunitp110(Thirone et al.,2006).WhenactivatedP13Kcatalysisthephosphorylationofph sphatidylinositol4,5bisphosphate[(PI(4,5)P2)]tophospatidylinostal-3,4,5-triphosphate [PI(3,4,5)P3)].[PI (3, 4, 5) P3)].In turn activates phosphoinasitide dependent kinase 1(PDK-1) promoting the phosphorylation and activation of the Akt complex (Interaction between the Akt with PDK-1) to regulate phenomena like glucose uptake, glycogen synthesis and gene expression.
INSULIN RECEPTOR SUBSTRATE
Insulin signal transduction is highly conserved pathway which regulates multiple of cellular physiology including the regulation of cellular growth and glucose uptake and utilization (Luo et al., 2007). Cell insulin signaling starts when insulin binds to its specific receptors in several tissues including a complex intracellular cascade. The mainly of insulin receptor substrates (IRS, s) is an important regulatory key in the insulin signaling pathways and includes at least four distinct IRS proteins among IRS-1 and IRS-2 which are found in several human tissues (Cleaseby et al., 2007).IRS proteins link various membrane receptors to intracellular signalling pathways (While, 1997, Kaburagi et al., 1999; Johnston et al., 2003).IRS play an key role in the arrangement of the complex hepatic metabolic responses. IRS are unique docking molecules whose actions are very lightly regulated by the phosphorylation at various sites (While, 2002, Zick, 2005; Taniguchi et al., 2006).The IRS bind to the activated insulin receptor become phosphorylated thereby providing docking sites for the multitude of signaling molecules essential for the diversification and modulation of insulin action and hence for the light regulation of the hepatic glucose and lipid metabolism .Diversification of this complex system leads to impaired signal transduction resulting in pathological states. Reduced IRS protein levels in the liver and hyper phosphorylation of IRS on serine/threonine residues are hallmarks in the development of type-2 diabetes mellitus (Zick, 2004; Gval et al., 2005).Type-2 diabetes is caused by the impairment action of insulin an essential hormone for maintaining glucose homeostasis. Insulin entities a wide variety of growth promoting and metabolic effects by binding to the insulin receptor and activating its intrinsic tyrosine kinase (While, 1997) .Binding of insulin to its receptor and subsequently phosphorylation of tyrosine residues on the IRS proteins (Bertrand et al., 2008).These proteins have an important regulatory role providing an interface between insulin receptors and downstream effector molecules. Considerable evidence demonstrates that insulin receptor tyrosine kinase activity is essential for many if not all, of the biological effects of insulin (Saad et al., 1992).These adaptor proteins recognise phosphorylated tyrosine residues on the receptor leading to a stable association between the receptor and the IRS proteins. IRS proteins do not contain kinase activities but they play a key role as branching point by transferring the extra cellular signal( e.g. binding of insulin to its receptor) into multiple metabolic and anti-apoptosis signaling pathways. Insulin is the major physiological regulator of the metabolism
ETHNOMEDICAL IMPORTANCE OF NATURAL REMEDIES
Herbal medicine also called phytomedicine refers to the use of any plant seeds, berries, roots, leaves ,bark or flowers for medicinal purposes (Ang-Lee et al.,2000).The hypoglycemic activity of a large number of plants has been evaluated and confirmed in different animal models (Jouad et al., 2001). The ethnobotinical information reports about 1200 plant species that may possess antidiabetic potential (Grover et al.,2002, Jung et al.,2006).Oral hypoglycemic drugs or medicines, biguanides, sulfonylureas and thiozolidiesdiones are available for the treatment of diabetes along with undesirable side effects (Herrera et al.,2011). Alternatively development process in antidiabetic drug discovery has shifted focus on plant derived drug due to their safety, efficacy, cultural acceptability and lesser side effect ( Veerapur et al.,2010). Hundrends of herbs and traditional Chinese herbal formulas reported to have been used to treat diabetes (Xu et al., 2013). Many plant derived phytomedicines such as β-carotene and α-lipoic acid have reported to the anti-diabetic effects (Chen et al.,2013).Treating diabetes mellitus with phytochemicals which are accessible and do not require laborious pharmaceutical synthesis seems highly attractive. (Venkata Raman et al., 2012). Phytotherapeutic agents are normally marketed as standardized preparations in the form of liquid, solid (Powdered extract) or viscous preparations. Some phytotherapeutic agents are greatly concentrated in order to improve their therapeutic efficacy (Calixto,2000).Hence we are interested to study the antidiabetic effect of Aconitum hetrophyllum wall.x.Royle on STZ- induced diabetic rats.
ACONITUM HETEROPHYLLUM
Many unknown and lesser-known plants are used in folk and tribal medicinal practice in India. The medicinal values of these plants are not much known to the scientific world. Aconitum heterophyllum wall x.Royle (Family Renunculaceae) is one such medicinal plant popularly used for the anti-obesity and anti-diabetes in traditional practices .Aconitum heterophyllum is a temperate medicinal plant distributed 2400 and 4500 m in alpine regions of the Himalaya. Distribution and morphology of this species has been extensively documented by various workers from time to time (Hooker and Thomson, 1987; Collett, 1902; Prajapati et al., 2007and Nautial et al., 2009).The plants are usually perennial or biennial herbs, often with stout leafy stems, bulbs or creeping rhizomes. Leaves are mostly cauline, lobed, rarely divided and dentate. Flowers are simple or branched recemes. It comprises of over 300 species, including some medicinal and ornamental plants (Utelli et al., 2000).
Nomenclature
Family – Renunculaceae
Genus – Aconitum
Species – Aconitum heterophyllum wallx Royle.
Vernacular names
English -Indian Atees
Persian -Vajjcturki
Malayalam-Atividayam
Telugu -Ati vasa
Tamil -Atividayam
.PHARMACOLOGICAL PROPERTIES- HYPOGLYCAEMIC ACTION
Aconitum heterophyllum wall commonly known as Atis or Patris belonging to family renunculace is a perennial herb distributed over temperate parts of western Himalaya extending from Kashmir to Kumaonh.(Uniyal et al.,2002). The plant has shown to contain alkaloids. heteratisine,heterophyllisine, heterophyllidine,atidine,isoatisine,hetidine,benzonyheteroatisine. (Zhaobong et al., 2005).Three widely occurring alkaloids in the aconite roots mesaconitine, aconitine, and hypaconitine showed analgesic activities (Hikino et al., 1979). The monograph of dried roots of Aconitum heterophyllum is listed in the Ayurvedic Pharmacopoeia of India. The roots of Aconitum heterophyllum are considered relatively less toxic and they are used for treatment of emesis, inflammation, fever, cough, diarrhea and helminthiasis. The roots are also included in formulations like Rodhrasava (infusion), Mahavisagarbha Taila (medicated oils), and Siva Guika (pills) as listed in the Ayurvedic formulary of India (Anonymous, 1977, 1987). The paste of A. heterohyllum dried tubers mixed with water and sugar is taken orally to treat body ache. It is also used as an aphrodisiac and tonic (Semwal et al., 2009). Kutajghan Vati is a classical Ayurvedic anti-dysentery lozenge prepared from Aconitum heterophyllum and other medicinal plants (Lather et al., 2010). Recent studies have shown that its roots are used for curing arthritis (Subramanian et al., 2013) as well as in the preparation of Caspa Drops a polyherbal formulation for improving digestion and preventing abdominal distension (Sojitra et al., 2013). Aconitum has also shown to exhibit antipyretic,analgesic,anti-fungal,anti-bacterial,insecticidal,brime shrimp cytotoxicity, antiviral , anti-diarrheal and immunostimulant properties.( Anwar et al., 2003) A. heterophyllum is traditionally used to control obesity and included in “lekhaneyagana,” a pharmacological classification mentioned in Charaka samhita (Sharma, 2000). The development of anti-diabetic agents that are devoid of adverse effects is still a challenge to the health care systems globally .Thus medicinal plants are constantly being explored with the hope of developing a relatively safe antidiabetic plant based product alone or in combination with other agents (Tamiru et al., 2012).Medicinal plants are widely used in the management of diseases all over the world (Adewunmi and Ojewole,2004; Aliyu et al., 2007).Mechanism behind the hypolipidemic, antioxidant and antilipidimic effect of A. heterophyllum is unknown as so far in wistar diabetic rats. Thus the present study was under taken to explore hypolipidimic, antioxidant and lipidperoxidation effect of Aconitum heterophyllium on STZ induced wistar diabetic rats.
Fig. 6 Antidiabetic herbs – various mechanisms of actions and persisting models of its therapeutic evaluation.
RATIONALE FOR CHOOSING ACONITUM HETEROPHYLLIUM FOR THE STUDY
The research in the development of plant derived newer antidiabetic drugs is going on relentlessly throughout the world. No drug has been proved to be cheap and safe without any contraindications.
1. Protective effect of Aconitum heterophyllum against oxidative stress and hyperlipidemia, the important factors for the redressal of diabetic complications.
2. Easy availability of Aconitum hetrophyllum through out some parts of Kashmir
3. Relatively non-toxic nature of Aconitum heterophyllum among their other species.
No attempt has been made to isolate and characterize scientifically antidiabetic principle(s) from Aconitum heterophyllum root and to study its effect on STZ – diabetic rats.