Insulin and C-peptide Date – 8.1.2013. Time – 9.30 – 10.30 Dr. Jeevan Shetty MBBS MD Associate Professor Reference Book: Text book of Biochemistry By DM Vasudevan 5th edition Lippincott’s biochemistry – illustrated reviews – 4th edition.

INSULIN and C- PEPTIDE

Pancreas A triangular gland, which has both exocrine and endocrine cells, located behind the stomach Strategic location Acinar cells produce an enzyme-rich juice used for digestion (exocrine product) Pancreatic islets (islets of Langerhans) produce hormones involved in regulating fuel storage and use.

Islets of Langerhans 1 million islets 1-2% of the pancreatic mass Beta (β) cells produce insulin Alpha (α) cells produce glucagon Delta (δ) cells produce somatostatin F cells produce pancreatic polypeptide

Islets of Langerhans

Insulin Hormone of nutrient abundance. 4 major organs that play a dominant role in fuel metabolism: liver, adipose, muscle, and brain Tissues work together and communication between them is mediated through several ways Integration of energy metabolism controlled mainly by hormones such as insulin, glucagon, catecholamine's: epinephrine and norepinephrine. Insulin and glucagon profoundly affect energy metabolism

Insulin Structure

Insulin Structure Synthesized as part of proinsulin(86AA) and then excised by enzymes, releasing functional insulin (51 AA) and C peptide (31 AA). Insulin is a polypeptide hormone of 51 amino acids.(MW-6000) Composed of 2 polypeptide Chains A (21 AA) Chain B (30 AA) Held together by disulfide bonds.

Species variation Porcine insulin is more similar in sequence to human insulin except at terminal amino acid of B chain (Thr is replaced by Ala) Bovine insulin in A chain ( 8-Thr-Ala,10-Ile-val) B chain at 30 (Thr-Ala)

Insulin Synthesis Insulin gene encodes a large precursor of insulin (preproinsulin – 110 AA) During translation, the signal peptide is cleaved (proinsulin – 86 AA) During packaging in granules by Golgi, proinsulin is cleaved into insulin and C peptide

Insulin Insulin is released when there is a stimulus ( high blood glucose ) and exerts its function. Insulin has half life of 6 mins. Insulin is degraded by enzyme insulinase present in the liver and kidneys C- peptide C-peptide is essential for proper folding of insulin C – peptide has longer half life compared to insulin. So quantity of C- peptide in the serum indicator of amount of insulin production and secretion.

Protein and Polypeptide Synthesis and Release

Insulin Synthesis Insulin synthesis is stimulated by glucose or feeding and decreased by fasting. Threshold of glucose- stimulated insulin secretion is 100 mg/dl. Glucose rapidly increase the translation of the insulin mRNA and slowly increases transcription of the insulin gene. Insulin has got half life of 6 minutes.

Regulation of insulin secretion Glucose is the primary stimulator of insulin secretion. Ingestion of glucose or carbohydrate rich meal leads to rise in blood glucose, which is signal for increased insulin secretion and decreased glucagon synthesis and release. Amino acids- ingestion of protein causes transient rise in plasma amino acid levels, which, in turn, induces immediate secretion of insulin.

Secretin and gastrointestinal hormones stimulate insulin secretion. Inhibition of Insulin secretion Epinephrine – inhibit insulin release and epinephrine levels are high during Starvation Stress

Regulation of insulin secretion – When glucose low

Regulation of insulin secretion – When glucose high

Regulation of Insulin Secretion

Regulation of Insulin Secretion No insulin is produced when plasma glucose below 50 mg/dl Half-maximal insulin response occurs at 150 mg/dl A maximum insulin response occurs at 300 mg/dl Insulin secretion is biphasic: Upon glucose stimulation– an initial burst of secretion (5-15 min.) Then a second phase of gradual increment that lasts as long as blood glucose is high.

Mechanism of action of insulin Insulin binds to specific , high affinity receptors in the cell membrane of the most tissues, including liver, muscle, adipose tissue. This is first step followed by cascade of events. Insulin receptor – tetramer with 2 alpha (extracellular) and 2 beta subunits (cytosolic), which are linked by disulfide bridges. Alpha subunit contains insulin binding site and beta subunit is a tyrosine kinase, which is activated by insulin.

Signal Transduction Binding of insulin to alpha subunits of the insulin receptor induces conformational changes that are transduced to beta subunits. This promotes a rapid auto phosphorylation of specific tyrosine residues on each beta subunit. Auto phosphorylation initiates a cascade of cell signaling responses, including phosphorylation of a family of proteins called insulin receptor substrate proteins. (IRS). Actions of insulin are terminated by dephosphorylating of the receptor.

Membrane effects of insulin Glucose transport in some tissues, such as skeletal muscle and adipocytes, increases in the presence of insulin. Insulin promotes the recruitment of insulin sensitive glucose transporters (GLUT-4) from pool located in intracellular vesicles. Entry of glucose is independent of insulin - Hepatocytes, erythrocytes, nerve cells, intestinal mucosa, cornea, renal tubule, cornea

Membrane effects of insulin

Receptor regulation – binding of insulin is followed by internalization of the hormone – receptor complex. In the cell insulin is degraded by lysosomes. Receptors are recycled to the cell surface. Action of insulin can be minutes to hours.

Insulin Action on Carbohydrate Metabolism: Liver Stimulates glucose oxidation - Glycolysis Promotes glucose storage as glycogen - Glycogenesis Inhibits glycogenolysis (break down of glucose) Inhibits gluconeogenesis (synthesis of glucose from non carbohydrate sources.

Insulin Action on Carbohydrate Metabolism Muscle Stimulates glucose uptake (GLUT4) Promotes glucose storage as glycogen(Glycogenes is)

Adipose Tissue: Stimulates glucose transport into adipocytes Promotes the conversion of glucose into triglycerides and fatty acids

Glucose Transport GLUT2 (liver, pancreas) GLUT4, insulin sensitive transporter (muscle, adipose tissue) GLUT3 (brain)

Action of insulin on Glycogen Synthesis Short term storage of glucose Activates glycogen synthase Inhibit glycogen phosphorylase Glycolysis is also stimulated by insulin

Lipogenic and antilipolytic Insulin promotes lipogenesis and inhibits lipolysis Promotes formation of α-glycerol phosphate and fatty acid synthesis Stimulates fatty acid synthase (FAS) Inhibits hormone sensitive lipase (HSL) Activates lipoprotein lipase (LPL) Protein Synthesis and Degradation Insulin promotes protein accumulation: Stimulates amino acid uptake Increases the activity of protein synthesis Inhibits protein degradation

Insulin action (summary): Dominates in Fed State Metabolism  glucose uptake in most cells  glucose use & storage  protein synthesis  fat synthesis

Insulin & Glucagon Regulate Metabolism

The Regulation of Blood Glucose Concentrations

Diabetes Mellitus (DM) A disorder of carbohydrate metabolism Results from hypo secretion or hypo activity of insulin The three cardinal signs of DM are: Polyuria – increased urine output Polydipsia – excessive thirst Polyphagia – excessive hunger and food consumption

Diabetes Mellitus Type I Type 1: beta cells destroyed- no insulin producedchronic fasted state, "melting flesh", ketosis, acidosis, glycosuria, diuresis & coma

Diabetes Mellitus: Type II a Group of Diseases Relative deficiency of insulin or Insulin resistance keeps blood glucose too high Chronic complications: atherosclerosis, renal failure& blindness

Symptoms of Diabetes Mellitus

Diabetes Mellitus (DM)

Glucagon A 29-amino-acid polypeptide hormone that is a potent hyperglycemic agent Produced by α cells in the pancreas Its major target is the liver, where it promotes: Glycogenolysis – the breakdown of glycogen to glucose Gluconeogenesis – synthesis of glucose from lactic acid and noncarbohydrates Release of glucose to the blood from liver cells

Glucagon Signaling

DNA in α cells mRNA Preproglucagon Proglucagon Glucagon Synthesis DNA in α cells mRNA Preproglucagon Proglucagon Glucagon

Factors Affecting Glucagon Secretion:

Glucagon Action on Cells: