diabetes mellitus - biochemistry

Diabetes mellitus is the 3rd leading cause of

death in many developed countries.

Diabetes is a major cause of blindness, renal

failure, amputation, heart attacks and stroke.

Diabetes mellitus is a characterized by

increased blood glucose level

(hyperglycemia) due to insufficient or

inefficient (incompetent) insulin.

Insulin is a polypeptide hormone produced

by the β-cells of islets of Langerhans of

pancreas.

It influences the metabolism of

carbohydrate, fat & protein.

It is an anabolic hormone, promotes the

synthesis of glycogen, triacylglycerols &

proteins.

Human insulin (mol. wt. 5,7341) contains 51

amino acids, arranged in 2 polypeptide chains.

A chain – 21 amino acids & B chain – 30 amino

acids.

Both are held together by 2 interchain disulfide

bridges, connecting A7 to B7 & A20 to B19.

There is an intrachain disulfide link in chain A

between the amino acids 6 & 11.

The gene for insulin synthesis is located on

chromosome 11.

The synthesis of insulin involves two

precursors, namely preproinsulin with 108

amino acids (mol. wt. 11,500) & proinsulin with

86 amino acids (mol. wt. 9,000).

They are sequentially degraded to form the

active hormone insulin & a connecting

peptide (C-peptide).

Insulin & C-peptide are produced in

equimolar concentration.

C-peptide is biologically inactive.

Its estimation is useful index for the

endogenous production of insulin.

In the β-cells, insulin (also proinsulin)

combines with zinc to form complexes.

In this complex form, insulin is stored in the

granules of the cytosol which is released in

response to various stimuli by exocytosis.

Factors stimulating insulin secretion:

Glucose & amino acids

Gastrointestinal hormones – secretin, gastrin,

pancreozymin increase the secretion.

Factors inhibiting insulin secretion:

Epinephrine from adrenal medulla is most

potent inhibitor of insulin secretion.

Insulin has a half-life of 4-5 minutes.

About 40-50 units of insulin is secreted daily

human pancreas.

The normal range of insulin: 20-30 μU/ml.

A protease enzyme – insulinase degrades

insulin.

Insulinase is mainly present in liver & kidney.

Effects on carbohydrate metabolism:

Insulin lowers blood glucose level by

promoting its utilization & storage & by

inhibiting its production.

Effect on glucose uptake by tissues:

Insulin is required for uptake of glucose by

muscle (skeletal, cardiac & smooth), adipose

tissue, leukocytes & mammary glands.

About 80% of glucose uptake in the body is

not dependent on insulin.

Effect on glucose utilization:

Insulin increases glycolysis in muscle & liver.

Insulin activates key enzymes of glycolysis –

glucokinase, PFK & pyruvate kinase.

Glycogen production is increased, due to

increased activity of glycogen synthase by

insulin.

Effect on glucose production:

Insulin decreases gluconeogenesis by

suppressing the enzymes pyruvate

carboxylase, phosphoenol pyruvate

carboxykinase & glucose 6- phosphatase.

Insulin also inhibits glycogenolysis by

inactivating the enzyme glycogen

phosphorylase.

Effects on lipid metabolism:

The net effect of insulin on lipid metabolism

is to reduce the release of fatty acids from

the stored fat & decrease the production of

ketone bodies.

Adipose tissue is the most sensitive to the

action of insulin.

Effect on lipogenesis:

Insulin favours the synthesis of

triacylglycerols from glucose by providing

more glycerol 3-phosphate & NADPH.

Insulin increases the activity of acetyl CoA

carboxylase, a key enzyme in fatty acid

synthesis.

Effect on lipolysis:

Insulin decreases the activity of hormone-

sensitive lipase & reduces the release of fatty

acids from stored fat.

The mobilization of fatty acids from liver is

also decreased by insulin.

Effect on ketogenesis:

Insulin reduces ketogenesis by decreasing

the activity of HMG CoA synthetase.

Effects on protein metabolism:

It stimulates the entry of amino acids into the

cells, increases protein synthesis & reduces

protein degradation.

Insulin promotes cell growth & replication.

This is mediated through certain factors such

as epidermal growth factor (EGF), platelet

derived growth factor & prostaglandins.

Insulin receptor mediated signal transduction:

Insulin receptor:

It is a tetramer consisting of 4 subunits – α2β2.

The subunits are in the glycosylated form.

They are held together by disulfide linkages.

α – subunit (mol. wt. 135,000) is extracellular &

it contains insulin binding site.

β – subunit (mw. 95,000) is a transmembrane

protein which is activated by insulin.

The cytoplasmic domain of β – subunit has

tyrosine kinase activity.

The insulin receptor is synthesized as a single

polypeptide & cleaved to α & β subunits which

are then assembled.

Insulin receptor has a half-life of 6-12 hours.

About 20,000 receptors/cell in mammals.

Signal transduction:

Insulin binds to the receptor, a conformational

change is induced in the α-subunits of insulin

receptor.

This results in the generation of signals which

are transduced to β-subunits.

The net effect is that insulin binding activates

tyrosine kinase activity of intracellular β-

subunit of insulin receptor.

This causes the autophosphorylation of

tyrosine residues on β-subunit.

Receptor tyrosine kinase also phosphorylates

insulin receptor substrate (IRS).

The phosphorylated IRS, in turn, promotes

activation of other protein kinases &

phosphatases, finally leading to biological

action.

Insulin-mediated glucose transport:

The binding of insulin to insulin receptor signals the

translocation of vesicles containing glucose

transporters from intracellular pool to the plasma

membrane.

The vesicles fuse with the membrane recruiting the

glucose transporters.

The glucose transporters are responsible for the

insulin-mediated uptake of glucose by the cells.

As the insulin level falls, the glucose

transporters move away from the membrane

to the intracellular pool for storage & recycle.

Insulin mediated enzyme synthesis:

Insulin promotes the synthesis of enzymes

such as glucokinase, PFK & pyruvate kinase.

This is brought about by increased

transcription & translation.

Glucagon, secreted by α-cells of the pancreas.

It is a polypeptide hormone composed of 29

amino acids (mol. wt. 3,500) in a single chain.

It is synthesized as proglucagon, on sequential

degradation releases active glucagon.

Its amino acid sequence is the same in all

mammalian species & half-life. i.e. about 5

minutes.

The secretion of glucagon is stimulated by

low blood glucose concentration, amino

acids derived from dietary protein & low

levels of epinephrine.

Increased blood glucose level markedly

inhibits glucagon secretion.

Glucagon enhances the blood glucose level

(hyperglycemic).

Primarily, glucagon acts on liver to cause

increased synthesis of glucose & enhanced

degradation of glycogen.

Effects on lipid metabolism:

Glucagon promotes fatty acid oxidation

resulting in energy production & ketone

body synthesis.

Effects on protein metabolism:

Glucagon increases the amino acid uptake

by liver & promotes gluconeogenesis.

The maintenance of glucose level in blood

within narrow limits is a very finely &

efficiently regulated system.

It is essential to have continuous supply of

glucose to the brain.

Following a meal, glucose is absorbed from

the intestine and enters the blood.

The rise in blood glucose level stimulates the

secretion of insulin.

The uptake of glucose by most extrahepatic

tissues, except brain is dependent on insulin.

Insulin helps in the storage of glucose as

glycogen or its conversion to fat.

Normally, 2 to 2½ hours after a meal, blood

glucose level falls to near fasting levels.

It may go down further; but this is prevented

by processes that contribute glucose to the

blood.

For another 3 hours, hepatic glycogenolysis

will take care of the blood glucose level.

Thereafter gluconeogenesis will take charge

of the situation.

Liver is the major organ that supplies the

glucose for maintaining blood glucose level.

Glucagon, epinephrine, glucocorticoids,

growth hormone, ACTH & thyroxine will keep

the blood glucose level from falling.

They are referred to as antiinsulin hormones

or hyperglycemic hormones.

Random blood sugar

Fasting blood sugar

Post-prandial blood sugar

Hyperglycemia

Hypogycemia

Glucose is estimated by GOD/POD or

hexokinase method.

Insulin:

It is produced in response to hyperglycemia.

Some amino acids, free fatty acids, ketone

bodies, drugs such as tolbutamide also cause

the secretion of insulin.

It is hypoglycemic hormone that lowers in

blood glucose level.

Glucagon:

Hypoglycemia stimulates its production.

It increases blood glucose concentration.

It enhances gluconeogenesis & glycogenolysis.

Epinephrine:

It is secreted by adrenal medulla.

It acts on muscle & liver to bring about

glycogenolysis by increasing phosphorylase

activity.

Thyroxine:

It is a hormone of thyroid gland.

It elevates blood glucose level by stimulating

hepatic glycogenolysis & gluconeogenesis.

Glucocorticoids:

Glucocorticoids increases gluconeogenesis.

The glucose utilization by extrahepatic tissues is

inhibited by glucocorticoids.

The overall effect of glucocorticoids is to elevate

blood glucose concentration.

GH & ACTH also increases blood glucose.

A fall in plasma glucose less than 50 mg/dl is

called as hypoglycemia.

Hypoglycemia is life-threatening.

The manifestations include headache,

anxiety, confusion, sweating, slurred speech,

seizures & coma, and, if not corrected, death.

Post-prandial hypoglycemia:

This is also called reactive hypoglycemia & is

observed in subjects with an elevated insulin

secretion following a meal.

This causes transient hypoglycemia & is

associated with mild symptoms.

The patient is advised to eat frequently rather

than the 3 usual meals.

Fasting hypoglycemia:

Fasting hypoglycemia is not very common.

It is observed in patients with pancreatic β-

cell tumor & hepatocellular damage.

Hypoglycemia due to alcohol intake:

Alcohol consumption causes hypoglycemia

This is due to the accumulation of NADH,

which diverts pyruvate & oxaloacetate to

form lactate & malate.

Finally gluconeogenesis is reduced due to

alcohol consumption.

Hypoglycemia due to insulin overdose:

The most common complication of insulin

therapy in diabetic patients is hypoglycemia.

This is particularly observed in patients who

are on intensive treatment.

Diabetes mellitus (DM) is a metabolic disease

due to absolute or relative insulin deficiency.

DM is a common clinical condition.

It is a major cause for morbidity & mortality.

Mainly two types.

Type 1 diabetes mellitus (T1DM).

Type 2 diabetes mellitus (T2DM).

Also known as IDDM or (less frequently)

juvenile onset diabetes, mainly occurs in

childhood (between 12 -15 years age).

IDDM accounts for about 10 to 20% of the

known diabetics.

Characterized by almost total deficiency of

insulin due to destruction of β-cells.

The β-cell destruction may be caused by drugs,

viruses or autoimmunity.

Due to certain genetic variation, the β-cells are

destroyed by immune mediated injury.

Symptoms of diabetes appear when 80-90% of

the - β cells have been destroyed.

The pancreas ultimately fails to secrete insulin

The patients of IDDM require insulin therapy.

Also called as non-insulin dependent diabetes

mellitus (NIDDM).

Accounting for 80 to 90% of diabetic population.

NIDDM occurs in adults (above 35 years) & is

less severe than IDDM.

The causative factors of NIDDM include genetic

& environmental.

NIDDM commonly occurs in obese individuals.

Gestational diabetes mellitus (GDM):

This term is used when carbohydrate intolerance is

noticed, for the first time, during a pregnancy.

A known diabetic patient, who becomes pregnant, is

not included in this category.

Glucose challenge test (GCT) is done between 22 & 24

weeks of pregnancy by giving an oral glucose load

of 50 g of glucose regardless of the time.

If the 2-hour post-glucose value is >140 mg/dl, the test

is positive.

Impaired glucose tolerance (IGT):

Also called as Impaired Glucose Regulation (IGR).

Plasma glucose values are above the normal level, but

below the diabetic levels.

In IGT, the FBS value is 110 & 126 mg/dl & PPBS value is

between 140 & 200 mg/dl.

Requires careful follow-up because IGT progresses to

frank diabetes at the rate of 2% patients per year.

Impaired fasting glycemia (IFG):

In this condition, fasting plasma glucose is

above normal (between 110 & 126 mg/dl); but

the 2 hour post-glucose value is within

normal limits (less than 140 mg/dl).

These persons need no immediate treatment;

but are to be kept under constant check up.

Secondary to other known causes:

Endocrinopathies (Cushing's disease,

thyrotoxicosis, acromegaly)

Drug induced (steroids, beta blockers, etc.)

Pancreatic diseases (chronic pancreatitis,

fibrocalculus pancreatitis, hemochromatosis,

cystic fibrosis).

The diagnosis of diabetes can be made on

the basis of individual's response to the oral

glucose load, commonly referred to as oral

glucose tolerance test (OGTT).

Preparation of the subject:

Carbohydrate-rich diet for at least 3 days

prior to the test.

All drugs known to influence carbohydrate

metabolism should be discontinued (2 days).

The subject should avoid strenuous exercise

on the previous day of the test.

Person should be in an overnight fasting

state.

During the course of GTT, the person should

be comfortably seated & should refrain from

smoking & exercise.

Glucose tolerance test should be conducted

preferably in the morning (ideal 9 to 11 AM).

A fasting blood sample is drawn and urine

collected.

The subject is given 75 g glucose orally,

dissolved in about 300 ml of water, to be

drunk in about 5 minutes.

Blood & urine samples are collected at 30

minute intervals for at least 2 hours.

All blood samples are subjected to glucose

estimation while urine samples are

qualitatively tested for glucose.

The fasting plasma glucose level is 75-110 mg/dl

in normal persons.

On oral glucose load, concentration increases

& peak value (140 mg/dl) is reached in less

than an hour which returns to normal by 2

hours.

Glucose is not detected in any of the urine

samples

In individuals with impaired glucose

tolerance, the fasting (110-126 mg/dl) as well as

2 hour (140-200 mg/dl) plasma glucose levels

are elevated.

These subjects slowly develop frank diabetes.

Dietary restriction & exercise are advocated

for the treatment of impaired glucose

tolerance.

Condition Plasma glucose concentration as mmol/l (mg/dl)

Normal IGT Diabetes

Fasting <6.1

(<110)<7.0

(<126)>7.0

(>126)

2 hours after glucose

<7.8(<140)

<11.1(<200)

>11.1(>200)

For conducting GTT in children, oral glucose is

given on the basis of weight (1.5 to 1.75 g/kg).

In case of pregnant women, 100 g oral

glucose is recommended.

Mini GTT carried out in some laboratories,

fasting and 2 hrs. sample (instead of 1/2 hr.

intervals) of blood & urine are collected.

To evaluate the glucose handling of the body

under physiological conditions, fasting blood

sample is drawn, the subject is allowed to

take heavy breakfast, blood samples are

collected at 1 hour & 2 hrs (post-prandial-

meaning after food).

Urine samples are also collected.

This type of test is commonly employed in

established diabetic patients for monitoring

the control.

For individuals with suspected malabsorption,

intravenous GTT is carried out.

Corticosteroid stressed GTT is employed to detect

latent diabetes.

Glycosuria:

The commonest cause of glucose excretion in urine

(glycosuria) is diabetes mellitus.

Glycosuria is the first line screening test for diabetes.

Normally, glucose does not appear in urine until the

plasma glucose concentration exceeds renal

threshold (180 mg/dl).

Renal glycosuria:

Renal glycosuria is a benign condition due to

a reduced renal threshold for glucose.

It is unrelated to diabetes & should not be

mistaken as diabetes.

Further, it is not accompanied by the classical

symptoms of diabetes.

Alimentary glycosuria:

In certain individuals, blood glucose level rises

rapidly after meals resulting in its spill over

into urine.

This condition is referred to as alimentary

glycosuria.

It is observed in some normal people & in

patients of hepatic diseases, hyperthyroidism

& peptic ulcer.

Hyperglycemia:

Elevation of blood glucose concentration is the

hallmark of uncontrolled diabetes.

Hyperglycemia is primarily due to reduced

glucose uptake by tissues & its increased

production via gluconeogenesis &

glycogenolys.

Glucose is excreted into urine (glycosuria).

Ketoacidosis:

Increased mobilization of fatty acids results

in overproduction of ketone bodies which

often leads to ketoacidosis.

Hypertriglyceridemia:

Conversion of fatty acids to TAGs & secretion

of VLDL & chylomicrons is higher in diabetics.

Plasma levels of VLDL, chylomicrans, TAGs &

cholesterol are increased.

Glycosuria – glucose excretion in urine.

Due to osmotic effect, more water

accompanies the glucose (polyuria).

To compensate for this loss of water, thirst

center is activated & more water is taken

(polydypsia).

To compensate the loss of glucose & protein,

patient will take more food (polyphagia).

Diabetic keto acidosis (DKA):

DKA more common in T1DM.

Normally the blood level of ketone bodies is

<1 mg/dl & only traces are excreted in urine.

Increased synthesis causes the accumulation

of ketone bodies in blood.

It causes ketonemia, ketonuria & smell of

acetone in breath.

Together constitute ketosis.

Detected by Rothera's test.

Supportive evidence may be derived from

estimation of serum electrolytes, acid–base

parameters & glucose estimation.

The urine of a patient with diabetic keto

acidosis will give positive Benedict's test as

well as Rothera's test.

But in starvation ketosis, Benedict's test is

negative, but Rothera's test will be positive.

Diabetes Mellitus:

The combination of hyperglycemia,

glucosuria, ketonuria & ketonemia is called

diabetic ketoacidosis (DKA).

Untreated diabetes mellitus is the most

common cause for ketosis.

Deficiency of insulin causes accelerated

lipolysis & more fatty acids are released into

circulation.

Oxidation of these fatty acids increases the

acetyl CoA pool.

Enhanced gluconeogenesis restricts the

oxidation of acetyl CoA by TCA cycle, since

availability of oxaloacetate is less.

In starvation, dietary supply of glucose is

decreased.

Available oxaloacetate is channelled to

gluconeogenesis.

The increased rate of lipolysis provides

excess acetyl CoA which is channeled to

ketone bodies.

The high glucagon favors ketogenesis.

Hyperemesis (vomiting) in early pregnancy may

also lead to starvation-like condition & may lead to

ketosis.

In both diabetes mellitus & starvation, the

oxaloacetate is channelled to gluconeogenesis.

Acetyl CoA cannot be fully oxidized in TCA cycle.

This excess acetyl CoA is channelled into ketogenic

pathway.

Metabolic acidosis:

Acetoacetate & β-hydroxy butyrate are

accumulated, causes metabolic acidosis.

There will be increased anion gap.

Reduced buffers:

The plasma bicarbonate is used up for

buffering of these acids.

Kussmaul's respiration:

Patients will have typical acidotic breathing due to

compensatory hyperventilation.

Smell of acetone in patient's breath.

Osmotic diuresis induced by ketonuria may lead to

dehydration.

Sodium loss:

The ketone bodies are excreted in urine as their

sodium salt, leading to loss of cations from the body.

High potassium:

Due to lowered uptake of potassium by cells

in the absence of insulin.

Dehydration:

Sodium loss further aggravates dehydration.

Coma:

Hypokalemia, dehydration & acidosis

contribute to the lethal effect of ketosis.

Parenteral administration of insulin & glucose.

Intravenous bicarbonate to correct acidosis.

Correction of water imbalance by normal

saline.

Correction of electrolyte imbalance.

Hyperglycemia is directly or indirectly

associated with several complications.

These include

Atherosclerosis

Retinopathy

Nephropathy

Neuropathy.

Dietary management:

A diabetic patient is advised to consume low

calories (i.e. low carbohydrate & fat), high

protein & fiber rich diet.

Diet control & exercise will help to a large

extent obese NIDDM patients.

Hypoglycemic drugs:

The oral hypoglycemic drugs are broadly of

two categories-sulfonylureas & biguanides.

Sulfonylurea such as acetohexamide,

tolbutamide & gibenclamide are frequently

used.

They promote the secretion of endogenous

insulin & help in reducing blood glucose level.

Management with insulin:

Two types of insulin preparations are

commercially available – short acting & long

acting.

The short acting insulins are unmodified &

their action lasts for about 6 hours.

The long acting insulins are modified ones &

act for several hours, which depends on the

type of preparation.

Glycated hemoglobin:

Refers to the glucose derived products of normal adult

hemoglobin (HbA).

Glycation is a post-translational, non-enzymatic

addition of sugar residue to amino acids of proteins.

Among the glycated hemoglobins, the most abundant

form is HbA1c.

HbA1c is produced by the condensation of glucose

with N-terminal valine of each β-chain of HbA.

The rate of synthesis of HbA1c is directly related to

the exposure of RBC to glucose.

The concentration of HbA1c serves as an indication of

the blood glucose concentration over a period.

HbA1c concentration is about 3-5%.

In diabetic patients, HbA1c is elevated (15%).

HbA1c reflects the mean blood glucose level over 2

months period prior to its measurement.

Other proteins in the blood are glycated.

Glycated serum proteins (fructosamine) can

also be measured in diabetics.

Albumin is the most abundant plasma

protein, glycated albumin largely contributes

to plasma fructosamine measurements.

Albumin has shorter half-life than Hb.

Glycated albumin represents glucose status

over 3 weeks prior to its determination.

Microalbuminuria is defined as the excretion of 30-

300 mg of albumin in urine per day.

Microalbuminuria represents an intermediary stage

between normal albumin excretion (2.5-30 mg/d) &

macroalbuminuria (>300 mg/d).

The small increase in albumin excretion predicts

impairment in renal function in diabetic patients.

It indicates reversible renal damage.

Textbook of Biochemistry – U Satyanarayana

Textbook of Biochemistry – DM Vasudevan