1. Alloxan & Streptozotocin-induced diabetes is a form of
insulin-dependent diabetes mellitus that occurs as a result of alloxan
administration or injection to animals. It has been successfully induced in a
variety of animal species; rabbits, mice, rats, monkeys, cats and dogs. Alloxan
& Streptozotocin has been administered in single or multiple doses, through
different routes (intraperitoneal, intravenous and subcutaneous); with single
intraperitoneal administration apparently the most employed mode.
2.
Alloxan is a toxic glucose analogue,
which selectively destroys insulin-producing cells in the pancreas (that is,
beta cells) when administered to rodents. Alloxan is a classical
diabetogenic chemical that exerts selective cytotoxic influences on pancreatic
β-cells, resulting in the destruction of β-cells and type 1 diabetes.
3.
Alloxan selectively inhibits
glucose-induced insulin secretion through specific inhibition of glucokinase,
the glucose sensor of the beta-cell which facilitates phosphorylation of
glucose to glucose-6-phosphate. Inhibition of glucokinase reduces glucose
oxidation and ATP generation, thereby suppressing the ATP signal that triggers
insulin secretion. Inhibition of glucokinase is achieved within 1 min of
exposure to alloxan.
4.
It causes insulin-dependent diabetes by
inducing ROS formation in a cyclic reaction with its reduction
product, dialuric acid, resulting in the selective necrosis of beta cells.
5. After administration of Alloxan, in the first 2 hours, blood glucose rises. This transient hyperglycaemia is thought to be due to sudden glycogen breakdown in the liver. The second phase is a hypoglycaemic phase which may be severe enough to lead to death if it is not prevented or treated with supplemental glucose. Hypoglycaemia is due to a sudden outpouring of insulin from dying beta cells. Hypoglycaemia generally appears after 6 hours. Hypoglycaemia is more pronounced in fasted animals.
6.
Streptozotocin is a naturally
occurring alkylating antineoplastic agent that is particularly toxic to the
insulin-producing beta cells of the pancreas in mammals.
7.
Streptozotocin is selectively
accumulated in pancreatic beta cells via the low-affinity GLUT2 glucose
transporter in the plasma membrane. Thus, insulin-producing cells that do not
express this glucose transporter are resistant to Streptozotocin.
8.
Toxicity of Streptozotocin is dependent
upon the DNA alkylating activity of its methylni-trosourea moiety. The transfer
of the methyl group from streptozotocin to the DNA molecule causes damage,
which along a defined chain of events, results in the fragmentation of the DNA.
DNA methylation is ultimately responsible for beta cell death, but it is likely
that protein methylation contributes to the functional defects of the beta
cells after exposure to streptozotocin.
9.
Both alloxan and streptozotocin induce
insulin deficiency. While the mechanisms of beta cell-selective action through
uptake via the GLUT2 glucose transporter and beta-cell death via necrosis are
identical, ROS in the case of alloxan and DNA alkylation in the case of
streptozotocin mediate the toxic action of these glucose analogues.
10.
Due to its chemical properties, in
particular the greater stability, streptozotocin is the agent of choice for
reproducible induction of a diabetic metabolic state in experimental
animals. Alloxan, on the other hand, as a model compound of
ROS mediated beta-cell toxicity, is the agent with the greater impact upon
the understanding of ROS mediated mechanisms of beta-cell death in type 1
and type 2 diabetes mellitus.
Key Points for Streptozotocin &
Alloxan preparation and induction in laboratory animals
·
Streptozotocin and
Alloxan are available in powder form. As both chemicals are very sensitive to
light, temperature and humidity, they should be kept under tightly closed containers and should be protected from light.
·
For the preparation
of a solution of these chemicals, weigh the appropriate amount of compound and place it in an aluminium foil wrapped container under an icebox. Ice cold citrate buffer
prepared in autoclaved water should be used for the preparation of the solution.
·
All steps of the
solution preparation should be performed in a dark room to protect the compound
from light.
·
When you add the
buffer in powdered chemicals, it started to degrade. So, you should use the
solution at the earliest.
·
Solution should be
filtered through 0.22 uM nylon syringe to remove any particulate matter
from solution.
·
Streptozotocin and
alloxan should be induced by the intraperitoneal route. The dose of Streptozotocin and Alloxan is 50 to 60 mg/kg.
· Before induction of diabetes, animals should be kept on fasting for 12 to 15 hours. After induction,
blood glucose level (BGL) should be measured after 4, 24, 72 hours of induction
and insulin injection should be given when BGL is too high. If diabetes is not
induced in any animal, re-induce in that individual animal.
·
After diabetes-inducing
agent induction, provide 10% dextrose water solution to the animals for 24 hours
to protect them from hypoglycaemia.
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