الدكتور الصيدالني احمد يحيى دالل باشي أستاذ في الكيمياء الحياتية الطبية رئيس فرع الكيمياء الحياتية كلية الطب/جامعة الموصل مدير وحدة التعليم الطبي في زاخو Disorders of Carbohydrates metabolism, Hyperglycemia, Diabetes Mellitus & Hypoglycemia PATHWAYS IN GLUCOSE METABOLISM The major energy pathways involved with glucose metabolism in the body includes the followings:. Glycolysis: Metabolism of glucose molecule to pyruvate or lactate for production of energy. Gluconeogenesis: Formation of glucose-6-phosphate from non carbohydrate sources. Glycogenolysis: Breakdown of glycogen to glucose for use as energy. Glycogenesis: Conversion of glucose to glycogen for storage. Lipogenesis: Conversion of carbohydrates to fatty acids. Lipolysis: Decomposition of fat. Regulation of Carbohydrate Metabolism The liver, pancreas, and other endocrine glands are all involved in controlling the blood glucose concentrations within a narrow range. Control of blood glucose is under two major hormones: insulin and glucagon both produced by the pancreas. Their actions oppose each other. Other hormones also exert some control over blood glucose concentrations. Insulin is the primary hormone responsible for the entry of glucose into the cell. It is synthesized by the cells of islets of Langerhans in the pancreas. When these cells detect an increase in body glucose, they release insulin. The release of insulin causes an increased movement of glucose into the cells and increased glucose metabolism. Insulin is normally released when glucose levels are high and is not released when glucose levels are decreased. It decreases plasma glucose levels by increasing the entry of glucose in muscle and adipose tissue. It also regulates glucose by increasing glycogenesis, lipogenesis, and glycolysis and inhibiting glycogenolysis. Insulin is the only hormone that decreases glucose levels and can be referred to as a hypoglycemic agent . Glucagon is the primary hormone responsible for increasing glucose levels. It is synthesized by the β cells of islets of Langerhans in the pancreas and released during stress and fasting states. When these cells detect a decrease in body glucose, they release glucagon. Glucagon acts by increasing plasma glucose levels by glycogenolysis in the liver and an increase in gluconeogenesis. It can be referred to as a hyperglycemic agent Two hormones produced by the adrenal gland affect carbohydrate metabolism. Epinephrine, produced by the adrenal medulla, increases plasma glucose by inhibiting insulin secretion, increasing glycogenolysis, and promoting lipolysis. Epinephrine is released during times of stress. Glucocorticoids, primarily cortisol, are released from the adrenal cortex on stimulation by adrenocorticotropic hormone (ACTH). Cortisol increases plasma glucose by decreasing intestinal entry into the cell and increasing gluconeogenesis, glycogenolysis, and lipolysis. Two anterior pituitary hormones, growth hormone and ACTH, promote increased plasma glucose. Growth hormone increases plasma glucose by decreasing the entry of glucose into the cells and decreasing glycolysis. Its release from the pituitary is stimulated by decreased glucose levels and inhibited by increased glucose. Decreased levels of cortisol stimulate the anterior pituitary to release ACTH. ACTH, in turn, stimulates the adrenal cortex to release cortisol and increases plasma glucose levels by converting liver glycogen to glucose and promoting gluconeogenesis. Two other hormones affect glucose levels: Thyroxine and somatostatin. The thyroid gland is stimulated by the production of thyroid-stimulating hormone (TSH) to release thyroxine that increases plasma glucose levels by increasing glycogenolysis, gluconeogenesis, and intestinal absorption of glucose. Somatostatin, produced by the β cells of the islets of Langerhans of the pancreas, increases plasma glucose levels by the inhibition of insulin, glucagon, growth hormone, and other endocrine hormones. HYPERGLYCEMIA Hyperglycemia is an increase in plasma glucose levels. In healthy patients, during a hyperglycemia state, insulin is secreted by the β cells of the pancreatic islets of Langerhans. Insulin enhances membrane permeability to cells in the liver, muscle, and adipose tissue. Diabetes Mellitus Diabetes mellitus is actually a group of metabolic diseases characterized by hyperglycemia resulting from defects in insulin secretion, insulin action, or both. In 1979, the National Diabetes Data Group developed a classification and diagnosis scheme for diabetes mellitus. This scheme included dividing diabetes into two broad categories: type 1, insulin-dependent diabetes mellitus (IDDM); and type 2, non–insulin-dependent diabetes mellitus (NIDDM). In 1995, the International Expert Committee and the American Diabetes Association, describe the classification as follow (Table 13-3). Therefore, the ADA/World Health Organization (WHO) guidelines recommend the following categories of diabetes: ■ Type 1 diabetes ■ Type 2 diabetes ■ Other specific types of diabetes ■ Gestational diabetes mellitus (GDM) Type 1 diabetes is characterized by inappropriate hyperglycemia primarily a result of pancreatic islet β-cell destruction and cellularmediated autoimmune destruction of the β cells of the pancreas, causing an absolute deficiency of insulin secretion and a tendency to ketoacidosis. Type 2 diabetes, in contrast, includes hyperglycemia cases that result from insulin resistance with an insulin secretory defect. An intermediate stage, in which the fasting glucose is increased abovenormal limits but not to the level of diabetes has been named, impaired fasting glucose. Use of the term impaired glucose tolerance to indicate glucose tolerance values above normal but below diabetes levels was retained. Also, the term gestational diabetes mellitus was retained for women who develop glucose intolerance during pregnancy. Upper limit of 110 mg/dL on the fasting plasma glucose is designated as the upper limit of normal blood glucose. Type 1 constitutes only 10 - 20% of all cases of diabetes and commonly occurs in childhood and adolescence. Characteristics of type 1 diabetes include abrupt onset, insulin dependence, and ketosis tendency. Signs and symptoms include polydipsia (excessive thirst), polyphagia (increased food intake), polyuria (excessive urine production), rapid weight loss, hyperventilation, mental confusion, and possible loss of consciousness (due to increased glucose to brain). Complications include microvascular problems such as nephropathy, neuropathy, and retinopathy. Increased heart disease is also found in patients with diabetes. Type 2 DM is characterized by hyperglycemia as a result of an individual’s resistance to insulin with an insulin secretory defect. This resistance results in a relative, not an absolute, insulin deficiency. Type 2 constitutes the majority of the diabetes cases. Most patients in this type are obese or have an increased percentage of body fat distribution in the abdominal region. This type of diabetes often goes undiagnosed for many years and at increased risk with an increase in age, obesity, and lack of physical exercise. Characteristics usually include adult onset of the disease and milder symptoms than in type 1, with ketoacidosis seldom occurring. However, these patients are more likely to go into a hyperosmolar coma and are at an increased risk of developing macrovascular and microvascular complications. Other specific types of diabetes are associated with certain conditions (secondary), including genetic defects of β-cell function or insulin action, pancreatic disease, diseases of endocrine origin, drug- or chemical-induced insulin receptor abnormalities, and certain genetic syndromes. The characteristics and prognosis of this form of diabetes depend on the primary disorder. GDM is “any degree of glucose intolerance with onset or first recognition during pregnancy.” Causes of GDM include metabolic and hormonal changes. Patients with GDM frequently return to normal postpartum. However, this disease is associated with increased perinatal complications and an increased risk for development of diabetes in later years. Infants born to mothers with diabetes are at increased risk for respiratory distress syndrome, hypocalcemia, and hyperbilirubinemia. Pathophysiology of Diabetes Mellitus In both type 1 and type 2 diabetes, the individual will be hyperglycemic, which can be severe. Glucosuria can also occur after the renal tubular transporter system for glucose becomes saturated. This happens when the glucose concentration of plasma exceeds roughly 180 mg/dL in an individual with normal renal function and urine output. As hepatic glucose overproduction continues, the plasma glucose concentration reaches around 300 to 500 mg/dL (17–28 mmol/L). The difference in glucagon and insulin concentrations in these two groups appears to be responsible for the generation of ketones through increased β-oxidation. In type 1, there is an absence of insulin with an excess of glucagon. This permits gluconeogenesis and lipolysis to occur. The laboratory findings of a patient with diabetes with ketoacidosis tend to reflect dehydration, electrolyte disturbances, and acidosis. Acetoacetate, β-hydroxybutyrate, and acetone are produced from the oxidation of fatty acids. The two former ketone bodies contribute to the acidosis, normal or elevated plasma sodium and potassium, slightly decreased bicarbonate, elevated blood urea nitrogen (BUN) and creatinine. Criteria for Testing Prediabetes and Diabetes The testing criteria for asymptomatic adults for type 2 diabetes mellitus were modified by the ADA Expert Committee to allow for earlier detection of the disease. According to ADA recommendations, all adults older than 45 years should have a measurement of fasting blood glucose every 3 years. Testing should be carried out at an earlier age or more frequently in individuals who display overweight tendencies (i.e., body mass index [BMI] ≥25 kg/m2) and have additional risk factors, as follows: ■ Habitually physically inactive ■ Family history of diabetes in a first-degree relative ■ History of GDM ■ Hypertension (blood pressure ≥140/90 mm Hg) ■ Low (HDL) high-density lipoprotein cholesterol concentrations (<35 mg/dL [0.90 mmol/L]) ■ Elevated triglyceride concentrations >250 mg/dL (2.82 mmol/L) ■ History of impaired fasting glucose/impaired glucose tolerance ■ Women with polycystic ovarian syndrome (PCOS) ■ Other clinical conditions associated with insulin resistance (e.g., severe obesity) ■ History of cardiovascular disease In the absence of the above criteria, testing for prediabetes and diabetes should begin at age 45 years. If results are normal, testing should be repeated at least at 3-years intervals. As the incidence of adolescent type 2 diabetes has raised dramatically in the past few years, criteria for the testing for type 2 diabetes in asymptomatic children have been developed. These criteria include initiation of testing at the age 10 years or at onset of puberty, with follow-up testing every 2 years for, overweight plus any two of the following risk factors: ■ Family history of type 2 diabetes in first or second degree relative ■ Race/ethnicity (e.g., Native American, African American, Latino, Asian American, and Pacific Islander) ■ Signs of insulin resistance or conditions associated with insulin resistance (e.g., hypertension, dyslipidemia) ■ Maternal history of diabetes or GDM Criteria for the Diagnosis of Diabetes Mellitus Three methods of diagnosis are suggested: (1) symptoms of diabetes plus a random plasma glucose level of ≥200 mg/dL, (2) a fasting plasma glucose of ≥126 mg/dL, or (3) an oral glucose tolerance test (OGTT) with a 2-hour postload (75-g glucose load) level ≥200 mg/dL, each of which must be confirmed on a subsequent day by any one of the three methods. An intermediate group who did not meet the criteria of diabetes mellitus but who had glucose levels above normal was defined by two methods. First, those patients with fasting glucose levels ≥100 mg/dL but <126 mg/dL were called the impaired fasting glucose group. Another set of patients who had 2-hour OGTT levels of ≥140 mg/dL but <200 mg/dL was defined as having impaired glucose tolerance. Patients with impaired fasting glucose and/or impaired glucose tolerance are referred to as having “prediabetes,” indicating the relatively high risk for the development of diabetes in these patients. The preferred test for diagnosing diabetes is measurement of the fasting plasma glucose level. HYPOGLYCEMIA Hypoglycemia involves decreased plasma glucose levels and can have many causes—some are transient and relatively insignificant, but others can be life threatening. The plasma glucose concentration in Hypoglycemia is usully less than 65 mg/dL (3.6 mmol/L); at about 50 to 55 mg/dL (2.8–3.0 mmol/L), observable symptoms of hypoglycemia appear. The warning signs and symptoms of hypoglycemia are all related to the central nervous system. The release of epinephrine into the systemic circulation and of norepinephrine at nerve endings of specific neurons act with glucagon to increase plasma glucose. Symptoms of hypoglycemia are increased hunger, sweating, nausea and vomiting, dizziness, nervousness and shaking, blurring of speech and sight, and mental confusion. Laboratory findings include decreased plasma glucose levels during hypoglycemic episode . Extremely elevated insulin levels may be found in patients with pancreatic β-cell tumors (insulinoma). Genetic Defects in Carbohydrate Metabolism Glycogen storage diseases are result of the deficiency of a specific enzyme that causes an alternation of glycogen metabolism. The most common congenital form of glycogen storage disease is glucose6-phosphatase deficiency type 1, which is also called von Gierke disease, an autosomal recessive disease. This disease is characterized by severe hypoglycemia that coincides with metabolic acidosis, ketonemia, and elevated lactate and alanine. Hypoglycemia occurs because glycogen cannot be converted back to glucose by way of hepatic glycogenolysis. A glycogen buildup is found in the liver; causing hepatomegaly. The patients usually have severe hypoglycemia, hyperlipidemia, uricemia, and growth retardation. A liver biopsy will show a positive glycogen stain. Although the glycogen accumulation is irreversible, the disease can be kept under control by avoiding the development of hypoglycemia. Liver transplantation corrects the hypoglycemic condition. Other enzyme defects or deficiencies that cause hypoglycemia include glycogen synthase, fructose-1,6-bisphosphatase, phosphoenolpyruvate carboxykinase, and pyruvate carboxylase. . Galactosemia, a cause of failure to thrive syndrome in infants, is a congenital deficiency of one of two enzymes involved in galactose metabolism, resulting in increased levels of galactose in plasma. The most common enzyme deficiency is galactose-1-phosphate uridyl transferase. Galactosemia occurs because of the inhibition of glycogenolysis and is accompanied by diarrhea and vomiting. Galactose must be removed from the diet to prevent the development of irreversible complications. If left untreated, the patient will develop mental retardation and cataracts. The disorder can be identified by measuring erythrocyte galactose-1-phosphate uridyltransferase activity. Laboratory findings include hypoglycemia, hyperbilirubinemia, and galactose accumulation in the blood, tissue, and urine following milk ingestion. Another enzyme deficiency, fructose-1-phosphate aldolase deficiency, causes nausea and hypoglycemia after fructose ingestion. There are also alimentary hypoglycemias, appears to be caused by an increase in the release of insulin in response to rapid absorption of nutrients after a meal ROLE OF LABORATORY IN DIFFERENTIAL DIAGNOSIS AND MANAGEMENT OF PATIENTS WITH GLUCOSE METABOLIC ALTERATIONS Laboratory tests are used for demonstration of hyperglycemia or hypoglycemia to diagnose diabetes mellitus and hypoglycemic conditions. Other laboratory tests have been developed to identify insulinomas and to monitor glycemic control and the development of renal complications. Methods of Glucose Measurement Glucose can be measured from serum, plasma, or whole blood. Today, most glucose measurements are performed on serum or plasma. The glucose concentration in whole blood is approximately 11% lower than the glucose concentration plasma. Serum or plasma must be separated from the cells within 1 h to prevent substantial loss of glucose by the cellular fraction and can be refrigerated. Sodium fluoride (gray-top tubes) are often used as an anticoagulant and preservative of whole blood. The fluoride inhibits glycolytic enzymes. However, although fluoride maintains long-term glucose stability, the rates of decline of glucose in the first hour after sample collection in tubes with and without fluoride are virtually identical. Therefore, the plasma should be separated from the cells as soon as possible. Fasting blood glucose (FBG) should be obtained in the morning after an approximately 8- to 10-hours fast (not longer than 16 hours). Fasting plasma glucose values have a diurnal variation with the mean FBG higher in the morning than in the afternoon. Diabetes in patients tested in the afternoon may be missed because of this variation. Cerebrospinal fluid and urine can also be analyzed. Urine glucose measurement is not used in diabetes diagnosis; however, some patients use this measurement for monitoring purposes. Self-Monitoring of Blood Glucose The ADA has recommended that individuals with diabetes monitor their blood glucose levels in an effort to maintain levels as close to normal as possible. For persons with type 1 diabetes, the recommendation is 3 to 4 times/day. It is important that patients be learned how to use the testing instrument to ensure the accuracy of their results. Urine glucose testing should be replaced by self-monitoring of blood glucose; however, urine ketone testing will remain for type 1 and gestational diabetes. Glucose Tolerance and 2-Hour Postprandial Tests A solution containing 75 g of glucose is administered, and a specimen for plasma glucose measurement is drawn 2 hours later. Children receive 1.75 g/kg of glucose to a maximum dose of 75 g. The oral glucose tolerance test (OGTT) is not recommended for routine use under the ADA guidelines. This procedure is inconvenient to patients and is not being used by physicians for diagnosing diabetes. However, if the OGTT is used, WHO recommends the criteria listed in Table 13-7. It is important that proper patient preparation be given before this test is performed. The patient should be ambulatory and on a normal-to high carbohydrate intake for 3 days before the test. The patient should be fasting for at least 10 hours and not longer than 16 hours, and the test should be performed in the morning because of the hormonal diurnal effect on glucose. Just before tolerance and while the test is in progress, patients should be prevented from exercise, eating, drinking (except that the patient may drink water), and smoking. Factors that affect the tolerance results include medications such as large doses of salicylates, diuretics, anticonvulsants, oral contraceptives, and corticosteroids. Also, gastrointestinal problems, including malabsorption problems, gastrointestinal surgery, vomiting and endocrine dysfunctions, can affect the OGTT results. Glycosylated Hemoglobin/Hemoglobin A1c Glycosylated hemoglobin is the term used to describe the formation of a hemoglobin compound produced when glucose (a reducing sugar) reacts with the amino group of hemoglobin (a protein). The glucose molecule attaches non-enzymatically to the hemoglobin molecule to form a ketoamine. The rate of formation is directly proportional to the plasma glucose concentrations. Because the average red blood cell lives approximately 120 days, the glycosylated hemoglobin level at any one time reflects the average blood glucose level over the previous 2 to 3 months. Therefore, measuring the glycosylated hemoglobin provides the clinician with a time-averaged picture of the patient’s blood glucose concentration over the past 3 months. Hemoglobin A1c (HbA1c), the most commonly detected glycosylated hemoglobin, is a glucose molecule attached to one or both N-terminal valines of the β-polypeptide chains of normal adult hemoglobin. HbA1c is a more reliable method of monitoring long-term diabetes control than random plasma glucose. Normal values range from 4.5 to 8.0. It is determined that for every 1% change in the HbA1c value, there is a 35 mg/dL (2 mmol/L) change in the mean plasma glucose (Table 13-10). It is important to remember that two factors determine the glycosylated hemoglobin levels: (1) the average glucose concentration and (2) the red blood cell life span. If the red blood cell life span is decreased because of another disease state such as hemoglobinopathies, the hemoglobin will have less time to become glycosylated and the glycosylated hemoglobin level will be lower. Current ADA guidelines recommend that an HbA1c test be performed at least two times a year with patients who are meeting treatment goals and who have stable glycemic control. For patients whose therapy has changed or who are not meeting glycemic goals, a quarterly HbA1c test is recommend. Lowering HbA1c to an average of less than 7% has clearly been shown to reduce the microvascular, retinopathic, and neuropathic complications of diabetes. Ketones The ketone bodies are produced by the liver through metabolism of fatty acids to provide a ready energy source from stored lipids at times of low carbohydrate availability. The three ketone bodies are acetone (2%), acetoacetic acid (20%), and β-hydroxybutyric acid (78%). A low level of ketone bodies are present in the body at all times. However, in cases of carbohydrate deprivation or decreased carbohydrate use such as diabetes mellitus, starvation/fasting, high-fat diets, prolonged vomiting, and glycogen storage disease, blood levels of ketone bodies increase due to lincreased lipolysis to meet energy needs. The term ketonemia refers to the accumulation of ketones in blood, and the term ketonuria refers to accumulation of ketones in urine. The measurement of ketones is recommended for patients with type1 diabetes during acute illness, stress, pregnancy, or elevated blood glucose levels above 300 mg/dL or when the patient has signs of ketoacidosis. The specimen requirement is fresh serum or urine; the sample should be tightly stoppered and analyzed immediately. Microalbuminuria Diabetes mellitus causes progressive changes to the kidneys and ultimately results in diabetic renal nephropathy. This complication progresses over years and may be delayed by aggressive glycemic control. An early sign that nephropathy is occurring is an increase in urinary albumin. Microalbumin measurements are useful to assist in diagnosis at an early stage and before the development of proteinuria. An annual assessment of kidney function by the determination of urinary albumin excretion is recommended for diabetic patients. Microalbuminuria is defined as persistent albuminuria in the range of 30 to 299 mg/24 h or an albumin-creatinine ratio of 30 to 300 g/mg. Clinical proteinuria or macroalbuminuria is established with an albuminuria ≥300 mg/24 h or an albumin-creatinine ratio of ≥300 g/mg. Insulin measurements are not required for the diagnosis of diabetes mellitus, but in certain hypoglycemic states, it is important to know the concentration of insulin in relation to the plasma glucose concentration.