Insulin

•	Insulin hormone.
•	Endocrine effects of insulin.
•	Sources of insulin.
•	Preparation of insulin.
•	Choices of preparation.

Insulin hormone

1.	Chemistry:

a.	contains 51 amino acids arranged in two chains (A and B) linked by disulfide bridges.

b.	within the B cell, insulin precursor is produced by DNA- or RNA-directed synthesis.

c.	proinsulin, a long single-chain protein molecule, is processed within the Golgi apparatus and packaged into granules, where it is hydrolyzed into insulin and a residual connecting segment, the C peptide by removal of 4 amino acids.

d.	insulin and C-peptide are secreted in equimolar amounts in response to all insulin secretagogues.

e.	granules within the B cell store the insulin in the form of crystals containing two atoms of zinc and six molecules.

f.	the entire human pancreas contains up to 8mg of insulin, representing about 200 biological units.

2.	Secretion:

a.	insulin is released from pancreatic B cells at a low basal rate and at a much higher stimulated rate in response to a variety of stimuli, especially glucose.

b.	if glucose concentration rises, ATP production increases, potassium channels close, and depolarization of the cell results.

c.	voltage-gated calcium channels open in response to depolarization, allowing more calcium to enter the cell.

d.	increased intracellular calcium results in increased insulin secretion.

3.	Degradation:

a.	the liver and kidney are the two main organs that remove insulin from the circulation by hydrolysis of the disulfide connections between the A and B chains.

b.	after this reductive cleavage, further degradation by proteolysis occurs.

4.	Insulin receptor:

a.	the receptors bind insulin with high specificity and affinity in the picomolar range.

b.	the full insulin receptor consists of two heterodimers, each containing an alpha subunit, which is entirely extracellular and constitutes the recognition site, and a beta subunit that spans the membrane.

c.	the beta subunit contains a tyrosine kinase.

d.	when insulin binds to the alpha subunit at the outside surface of the cell, tyrosine kinase activity is stimulated in the beta portion.

Endocrine effects of insulin

1.	Effects on liver:

a.	inhibits glycogenolysis.

b.	inhibits conversion of fatty acids and amino acids to keto acids.

c.	inhibits conversion of amino acids to glucose.

d.	promotes glucose storage as glycogen.

e.	increases triglyceride synthesis and very low density lipoprotein formation.

2.	Effects on muscle:

a.	increases amino acid transport.

b.	increases ribosomal protein synthesis.

c.	increases glucose transport.

d.	induces glycogen synthase and inhibits phosphorylase.

3.	Effects on adipose tissue:

a.	lipoprotein lipase is induced and activated by insulin to hydrolyze triglycerides from lipoproteins.

b.	glucose transport into cell is enhanced.

c.	intracellular lipase inhibited by insulin.

Sources of insulin

1.	Commercial insulin preparations differ in:

a.	animal species from which they are obtained.

b.	their purity, concentration and solubility.

c.	the time of onset and duration of their biological action.

2.	Bovine insulin differs from human insulin by 3 amino acids and is more antigenic to man than is porcine insulin, which differs from human by only 1 amino acid.

3.	Human insulin:

a.	made either by enzyme modification of porcine insulin, or by using recombinant DNA to synthesize the proinsulin.

b.	this is done by artificially introducing the DNA into either Escherichia coli or yeast.

4.	Pharmacokinetics:

a.	insulin naturally secreted by the pancreas enters the portal vein and passes straight to the liver which takes up half of it.

b.	the rest enters the systemic circulation where its concentration is only about 15%.

c.	injected insulin is absorbed into the blood and is inactivated in the liver and kidney.

d.	the half-life is 5 min.

5.	Differences between human and animal insulin:

a.	human insulin is absorbed from subcutaneous tissues slightly more rapidly than animal insulin and it has a slightly shorter duration of action.

b.	human insulin is less immunogenic than bovine, but not porcine, insulin.

c.	lessened awareness of hypoglycaemia with human insulin.

Preparations of Insulin

1.	Three major factors:

a.	strength.

b.	source.

c.	formulation.

2.	Formulation:

a.	short-acting solution of insulin for use s.c., i.m., or i.v..

b.	intermediate and longer acting (sustained release) preparations in which the insulin has been physically modified by combination with protamine or zinc to give an amorphous or crystalline suspension; this is given s.c. and slowly dissociates to release insulin in its soluble form.

3.	Dosage is measured in international units now standardized by chemical assay.

4.	Short acting insulin:

a.	regular (soluble, neutral) insulin is a short-acting soluble crystalline zinc insulin whose effect appears within 15 minutes of s.c. injection and generally lasts 5-7 hours.

b.	short-acting soluble insulin is the only type of insulin that can be administered intravenously or by infusion pumps.

c.	it is useful for intravenous therapy in the management of diabetic ketoacidosis and when the insulin requirement is changing rapidly, such as after surgery or during acute infections.

d.	insulin lispro (humalog) is a modified human insulin in which the reversing of two amino acids has resulted in a very rapid onset of action, within 15 minutes of injection.

5.	Lente and ultralente insulin:

a.	lente insulin is a mixture of 30% semilente (an amorphous precipitate of insulin with zinc ions in acetate buffer that has a rapid onset of action) with 70% ultralente insulin (a poorly soluble crystal of zinc insulin that has a delayed onset and prolonged duration of action).

b.	these two components provide a combination of rapid absorption, with sustained long action making lente insulin a useful therapeutic agent.

c.	lente insulin is the most widely used of the lente series of insulins, particularly in combination with regular insulin, which has a more rapid onset of action.

6.	Isophane (NPH) insulin:

a.	NPH insulin is an intermediate-acting insulin wherein the onset of action is delayed by combining appropriate amounts of insulin and protamine so that neither is present in an uncomplexed form.

b.	to form an isophane complex, about a 1:10 ratio by weight of protamine to insulin is required: 0.3 – 0.4mg of protamine to 4mg of insulin.

c.	NPH insulin is usually mixed with regular insulin and given at least twice daily for insulin replacement in patients with insulin-dependent diabetes.

7.	Mixtures of insulin:

a.	since intermediate-acting insulins require several hours to reach adequate therapeutic levels, their use in IDDM patients requires supplements of regular insulin preprandially.

b.	for convenience, these are often mixed together in the same syringe and injected subcutaneously in split dosage before breakfast and supper.

c.	the excess zinc in lente and ultralente insulin can precipitate some of the soluble regular insulin when mixed in vitro.

8.	Allergy:

a.	to addictives (protamine), preservative or to insulin itself.

b.	it may take the form of local reactions or insulin resistance.

9.	Antibodies to insulin:

a.	they act as a carrier or store, binding insulin after injection and releasing it slowly as the free insulin in the plasma declines.

b.	in this way they smooth and prolong insulin action.

c.	too high concentrations cause insulin resistance.

10.	Intravenous insulin:

a.	only soluble insulin injection should be used.

b.	the standard strength of insulin preparations is 100 units per ml.

Choice of preparation

1.	Soluble (regular, neutral) insulin:

a.	an aqueous solution of insulin.

b.	it is given s.c. 2-3 times a day, 30 min before meals.

c.	there is little risk of serious hypoglycaemic reaction.

d.	biggest disadvantages of soluble insulin for long term use are the need for frequent injections, and the occurrence of high blood glucose before breakfast.

2.	Intravenous soluble insulin:

a.	used in diabetic ketoacidosis.

b.	it may be given intermittently but continuous infusion is preferred.

c.	use of a slow-infusion pump with am ore concentrated solution (insulin 1.0unit/ml) is preferred.

3.	Insulin zinc suspensions and isophane insulin: sustained-release formulations in which rate of release is controlled by modifying particle size.

Dose of Insulin

1.	The total daily output of endogenous insulin from pancreatic islet cells is 30-40 units.

2.	Most insulin-deficient diabetics will need 30-50 unit/day (0.5-0.8 units/kg) of insulin.

3.	Initial treatment of Type I patient is with two injections of intermediate-acting insulin, or a mixed insulin.

4.	Initial daily dose requirements:

a.	0.3 units/kg (16-20 units daily).

b.	increasing to 0.5 units/kg.

5.	Giving one injection per day: 10 – 14 units of an intermediate acting isophane suspension may be given.

6.	Excessive dose of insulin leads to overeating and obesity, hypoglycaemia.

Treatment of a hypoglycaemic attack

1.	Prevention depends largely upon patient education, but it is an unavoidable aspect of intensive glycaemic control.

2.	Patients should never miss meals, must know the early symptoms of an attack, and always carry glucose with them.

3.	Treatment is to give sugar, either by mouth if the patient can still swallow or glucose (dextrose) i.v. (20-50ml of 50% solution).

4.	The patient should be given a meal to avoid relapse.

5.	If the patient does not respond within 30 minutes, it may be because of cerebral edema, which recovers slowly and may require treatment with i.v. dexamethasone and perhaps mannitol.

6.	If the patient has been severely hypoglycaemic, large amounts of 20% glucose may be given by i.v. infusion for several days.