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Amandeep Dhillon
M-Pharm 2nd Sem
Pathogenesis of Type 1 & 2 Diabetes mellitus.
Diabetes mellitus is a chronic metabolic disorder characterised by a high blood glucose concentration Hyperglycemia due to insulin deficiency and/or insulin resistance.
Hyperglycemia occurs because of uncontrolled hepatic glucose output & reduced uptake of glucose by skeletal muscles with reduced glycogen synthesis.
Hypoglycemic effects causes “Blood sugar lowering” effects (Assay of insulin & insulin like activity).
Cataract Formation
Diabetic patients are known to have a higher incidence of cataract formation. Which produce changes in the transparency or the refractory index of the lens.
Aldose reductase catalyst the reduction of aldoses like “Glucose & Galactose” to corresponding polyols.
Aldose reductase inhibitors have been shown to prevent sugar induced cataract fomation.
Pathogenesis
GLUT Glucose transporter IAPP Islet amyloid poly-peptide
ClassificationA. Enhance Insulin secretion
Sulfonylureas (KATP channel blockers)
1st Gen:- Tolbutamide
2nd Gen:-Glimepiride
B. Overcome Insulin resistance
Biguanide (AMPK Activator)
Thiazolidinediones (PPAR Activator)
C. Miscellaneous anti-diabetic drugs
α- glucosidase inhibitors Bromocriptin, Pramlintide
In-Vivo Models
1. Streptozotocin induced DM
2. Blood sugar lowering effects in rabbits/mice.
3. Cataract formation in streptozotocin Diabetic rats.
4. Galactose induced cataract in rats.
5. Naphthalene induced cataract in rats.
6. Hypoglycemic seizures in mice.
7. Assay of glucose transport.
In-Vitro Models1. Assays of insulin & of insulin like activity
2. Isolated organs, cell & membranes
3. Insulin receptor Binding assay
4. Assays of other glucose regulating peptide hormones
5. Inhibition of polysaccharide degrading enzyme
6. Effect on secondary diabetes symptoms
7. Glucose uptake by the isolated diaphragm from mice & rats
8. Insulin receptor binding assays
9. Isolated pancreas of rat
10. Rat diaphragm test.
Streptozotocin induced DMStreptozotocin [60 mg/kg body weight] is prepared in citrated buffer [ph 4.5]
Albino rats of either sex weighing 150-200 g are injected i.p with above solution.
Animals showing fasting blood glucose levels > 140mg/dl after 48 hours ofstreptozotocin administration are considered diabetic.
After six weeks of treatment blood samples are collected from 6 hour fastinganimal
Serum is separated by centrifuge (3000 rpm) under cooling (2-4 °C) for tenminutes
Serum glucose level is estimated by glucose- peroxidase method [GOD-PODkit] using autoanalyser
Broad spectrum anti biotic – 200 mg/kg i.p.
MOA
β cell damage by free radical injury
Fragmentation of DNA
Nitric oxide generation
Induces diabetes in all species
Advantages
Greater selectivity towards beta cells
Low mortality
Longer and irreversible diabetes
Guinea pigs and rabbits are resistant
Insulin antibody induced diabetesPrinciple: - A transient diabetic syndrome can be induced by injecting guinea pigs with anti insulin serum.
Preparation of antibody
Bovine insulin, dissolved in acidified water [ph 3.0] at a doseof 1mg /ml
Anti insulin sera is collected after two weeks of antigenic challenge
Injected into guinea pigs
ProcedureAdult albino rats are injectedwith 0.25-1.0 ml of guinea piganti- insulin serum.
increase of blood glucose level up to 300mg/ dl.
The drug sample to bescreened is given andblood glucose level isanalyzed to determinethe activity.
Blood sugar lowering effecs in rabbits/mice.PURPOSE & RATIONALE :- A biological assay of
insulin preparations in comparison with a stable standard using the blood sugar lowering effects in rabbits.
EVALUATION :-The result of the assay are calculated by standard methods.
Rabbits 150mg/kg
Procedure
Rabbits 3-4.5kg div into groups of 4-5
• Insulin and insulin like compound-
• Food stopped a day earlier
• Hypoglycemic agents-diet till experiment
Test drug –orally 1ml/kg in0.4%starch suspension
• Control receives only the vehicle
• Different doses are tried on other groups
Blood is withdrawn before &1,2,3,4,5,24,48 and 72 hr
• Blood glucose is determined
• Blood sugar values are plotted against time
RAT MODEL Male Wistar rats -250 gms
Divided into 4 groups of 7 each
Test dose is given as per dose in starch suspension
Blood is drawn from the tip of the tail at 1,2,3,5,24 hours
IN VITRO METHODS ON ISOLATED ORGANS AND CELLS PRINCIPLE:- To study the effect of the drug on
insulin, glucagon & somatostatin secretion without interference from other organs.
INSULIN ASSAY:-
Four groups of six rabbits weighing at least 1.8 kg
Two standard solutions of insulin containing one unit and two units
respectively
Two dilutions of sample whose potency is being examined are prepared
After one hour and 2.5 h of each injection, a suitable blood sample is taken from the ear vein of each rabbit.
blood sugar determined by glucose oxidase method
Summary