Clinical diagnostic biochemistry - 2 Dr. Maha Al-Sedik 2015 CLS 334

Oral Glucose Tolerance Test Serial measurement of plasma and urine glucose before and after a specific amount of glucose given orally. Indications : 1- Diagnosis of impaired glucose tolerance. 2- Diagnosis of gestational D.M. 3- Diagnosis of unexplained glucosuria. 4- Unexplained nephropathy, retinopathy or neuropathy.

Precautions before test : 1- Unrestricted diet 3 days before. 2- No drugs affecting glucose level. 3- No smoking or excessive exercise in the day of the test.

How to do the test : 1- Fasting 8 – 12 hours. Measure blood and urine glucose. 2- Give glucose load (50 – 100 g glucose or 1 g/kg body weight) dissolved in 250 – 300 ml water. 3- Blood and urine samples are taken every 1 hr for 2or 3 hours. Blood glucose level is plotted against time to draw glucose tolerance curve.

Normal curve : Fasting: 70 – 100 mg / dl Peak: 130 – 155 mg / dl after 1 hour Less than 140 mg l dl after 2 hours All urine samples contain no glucose.

Abnormalities in OGTT 1.Diabetic curve: Fasting: more than 126. After 2 hours: more than 200. One or more urine samples contain glucose.

2. lag storage response: It characterized by a sharp raise in blood glucose in the first hour with peak value exceeding the renal threshold with appearance of glucose in urine and return to normal fasting or below fasting occur in the second hour. This is due to rapid absorption of glucose from intestine.

It occurs in: Post-gastrectomy. Hyperthyrodisim & stress.

3- Flat response There is an ↑ in glucose tolerance or failure of the plasma glucose level to raise significantly after an oral glucose load and return the fasting level is rapid this is due to:  Insulinoma.  intestinal malabsortion syndrome.

Diabetes mellitus is a group of metabolic disorders of carbohydrate metabolism in which glucose is underused, producing hyperglycemia. Some patients may experience acute life threatening hyperglycemic episodes, such as ketoacidosis. Diabetes Mellitus

Classification: Type 1 diabetes. Type 2 diabetes. Gestational diabetes mellitus (GDM). Impaired glucose tolerance (IGT).

Criteria for the Diagnosis of Diabetes Mellitus Diabetes Mellitus Diabetes Mellitus Classic symptoms of diabetes. Fasting plasma glucose > 126 mg/dL (7mmol/L). 2-hour post prandial plasma glucose concentration > 200 mg/dL (11.1 mmol/L).

1- Type I D.M.  Old names: Insulin dependent (IDDM).  Usually starts acutely in young people <30 years but may start at any age.  There defective insulin secretion or destruction of B-cells.  Most individuals have antibodies that identify autoimmune process.

Pathogenesis of Type 1 Diabetes Mellitus Antibodies Islet cell cytoplasmic antibodies (ICAs) Insulin autoantibodies (IAAs) Genetics HLA-DR3 or HLA-DR4 histocompatibility antigens. Environment

2- Type II D.M. :  Old names : Non-insulin dependent D.M. (NIDDM), adult onset or maturity onset D.M.  Insulin secretion is slightly affected but there is impaired insulin action due to insulin resistance.  Usually starts over 40 years.

 Decrease of β-Cell Function.  Insulin Resistance Insulin resistance is defined as a decreased biological response to normal concentrations of circulating insulin, is found in both obese, non-diabetic individuals and patients with type 2 diabetes.  Environment: Diet and exercise.  Diabetogenes: Insulin-Resistance Genes. Body gain genes. Pathogenesis of Type 2 Diabetes Mellitus

Gestational Diabetes Mellitus Screening  Perform between 24 and 28 week of gestation on all average- and high-risk pregnant women not identified as having glucose intolerance.  Give 50 g oral glucose load without regard to time of day or time of last meal.  Measure venous plasma glucose at 1 hr.  If glucose is >140mg/dL, perform glucose tolerance test for pregnant womem.

Diagnosis  Perform in the morning after a 8 hr fast.  Measure fasting venous plasma glucose.  Give 75 or l00g of glucose orally.  Measure plasma glucose hourly for 3hr if you give 100 g (or 2hr if 75 g of glucose given).  At least two values must meet or exceed the following:

75-g load100-g load 95mg/dL Fasting 180mg/dL 1 hr 155mg/dL 2 hr mg/dL 3 hr

Prediabetes is when blood glucose levels are higher than normal but not high enough for a diagnosis of diabetes. Prediabetes means a person is at increased risk for developing type 2 diabetes, as well as for heart disease and stroke. Many people with prediabetes develop type 2 diabetes within 10 years. Pre diabetes

DiabetesPre - diabetesNormal More than 126 mg / dl100 – 126 mg / dlLess than 100 mg/dl Fasting More than 200 mg / dl140 – 199 mg / dlLess than 140 mg /dl Post - prandial

Keton bodies acetoacetic acid  -hydroxy butyric acid acetone

Ketogenesis  It is the formation of ketone bodies.  Site of ketogenesis: liver mitochondria.  The ketone bodies are acetoacetate, b-hydroxybutyrate, and acetone.

Importance of keton bodies: 1.When carbohydrates are low, the keton bodies are used as fuel. 2.Acetoacetate and beta-hydroxy buterate are important in biosynthesis of neonatal cerebral lipids. 3.In early stage of starvation, the heart and skeletal muscles use keton bodies as source of energy to keep glucose enough for brain as long as possible.

1.Two molecules of acetyl CoA react with each other in the presence of thiolase enzyme to form acetoacetyl CoA. Steps of synthesis of Ketone bodies:

2. Condensation of acetoacetyl CoA with acetyl CoA to form HMG CoA (3 hydroxyl- 3 methyl glutaryl CoA) catalyzed by HMG CoA synthetase,

3.HMG-CoA lyase enzyme catalyzes the cleavage of HMG-CoA to acetoacetate and acetyl CoA.

4. Acetoacetate produces β-hydroxybutyrate in a reaction catalyzed by β-hydroxybutyrate dehydrogenase in the present NADH.

5. Both acetoacetate and β-hydroxybutyrate can be transported across the mitochondrial membrane and the plasma membrane of the liver cells, enter to the blood stream to be used as a fuel by other cells of the body.

6. In the blood stream, small amounts of acetoacetate are spontaneously (non- enzymatically) decarboxyated to acetone.

 Acetone is volatile and can not be detected in the blood.  The odor of acetone may be detected in the breath and also in the urine of a person who has high level of ketone bodies in the blood.  e.g. in severe diabetic ketoacidosis, while under normal conditions, acetone formation is negligible.

Ketolysis Ketolysis is the complete oxidation of ketone bodies to C02 and water. Site: Mitochondria of extrahepatic tissues but not in the liver due to deficiency of the enzymes needed for ketolysis.

ketonemia : Excessive formation of ketone bodies results in increased blood concentrations. ketonuria: increased excretion of ketone bodies in the urine.

Diabetic ketoacidosis Hyperglycemia Ketosis Acidosis ↑ K

Signs and symptoms: 1-Classic symptoms of hyperglycemia: polyuria, polydipsia, polyphagia and dry skin. 2-Symptoms of hyperkalemia: arrhythmia 3- Symptoms of acidosis: Acidotic breath ( deep and rapid), confusion, arrhythmia and may lead to coma.

4- Symptoms due to increase in keton bodies: -Nausea, vomiting and abdominal pain. - fruit smell of mouth. - Disturbed consciousness and confusion. 5- Symptoms of dehydration: -Skin: Dry, hot and flushed skin. -Tongue: Dry (sometimes woody tongue). -Eyes: Sunken eyes and dark circles under the eyes.

LAB VALUES IN DKA: Blood glucose is > 300 mg/dl Serum ketones are positive. Arterial pH is < 7.3 Potassium is high.

Reference: Burtis and Ashwood Saunders, Teitz fundamentals of Clinical Chemistry, 4th edition, 2000.