34 GLUCOSE

1 Why does delayed processing or analysis of a blood sample result in a lower glucose concentration?

Prolonged exposure of serum or plasma to leukocytes, platelets, and erythrocytes allows the cells to consume glucose and lower the glucose concentration. The blood cells use the plasma glucose for their glycolytic pathways. The decline of plasma or serum glucose concentration is generally 5% to 10% per hour. This usually does not interfere with the interpretation of glucose concentrations because the processing of patient samples and the samples used for reference interval determination should be the same (i.e., removing serum or plasma from cells within 30 to 60 minutes after blood collection). The rate of glucose consumption can increase when there are more cells in the blood (marked leukocytosis, marked thrombocytosis, or possibly erythrocytosis). The rate of glucose consumption can be decreased by placing the blood sample in a cool environment, but clot formation and contraction are also reduced.

Serum separator tubes (tubes with gold or red/black stoppers) contain an activator and gel that enhance clot formation and allow easier separation of a blood clot and serum, respectively. After centrifugation, the serum glucose concentration remains stable in a refrigerator for at least 48 hours if the gel barrier is intact.

Erythrocytes from pigs are unique in that they lack a functional glucose transporter and therefore utilize glucose at a much lower rate than erythrocytes from other species. Inosine is the major energy substrate in pig erythrocytes, whereas glucose is the primary substrate in other domestic species.

2 If blood samples cannot be processed within an hour of collection, how can the glucose concentration in the sample be maintained?

Blood can be collected into evacuated tubes that contain sodium fluoride (NaF); these tubes have a gray stopper or cap. NaF tubes may contain potassium oxalate, sodium EDTA, or no anticoagulant. The fluoride ion enters the blood cells and reduces glycolysis by complexing with magnesium ion (Mg²⁺), a cofactor for some enzymes (e.g., phosphopyruvate hydratase, or enolase). The oxalate or EDTA in some tubes binds free calcium ions (Ca²⁺) to prevent the formation of fibrin through the coagulation pathways.

If NaF is not available, keeping the sample cool (not frozen) will reduce glycolysis. The blood sample should not be frozen because freezing will lyse blood cells and thus add their contents to the plasma.

3 What are the disadvantages of using sodium fluoride tubes?

Disadvantages of using NaF tubes include falsely lowering plasma glucose concentration, erythrocyte lysis, and limited use of the samples for other assays. Studies involving people, cats, and marine mammals have shown that plasma glucose concentrations decreased by 5% to 10% during the first hour after blood collection into NaF tubes. The decrease may result from the osmotic movement of water from erythrocytes to plasma when the blood is mixed with the hyperosmotic salts (NaF, potassium oxalate). Other data indicate that it takes about 1 hour for the fluoride to inhibit glycolysis completely. The antiglycolytic effect of fluoride depends on the fluoride concentration in the blood sample.

Erythrocytes are prone to lysis when exposed to the NaF and potassium oxalate, especially if an optimal amount of blood is not drawn into the evacuated blood tube. The release of erythrocyte contents (e.g., hemoglobin, phosphorus, potassium) may result in erroneous results in some clinical chemistry assays.

Besides inhibiting enolase in the glycolytic pathway, fluoride inhibits other enzymes, including the glucose oxidase of some glucose assays. Thus the NaF sample may not be an acceptable sample for determining glucose concentration.

4 Why does a blood glucose concentration not equal a plasma/serum glucose concentration?

The differences between blood glucose and plasma or serum glucose concentration typically will be slight, but differences can be caused by erythrocytosis, sample processing errors, or differences in assay methods. Erythrocytosis will increase the difference because the glucose concentration in erythrocytes is lower than in plasma/serum. The concentration of glucose in the plasma H₂O and erythrocyte H₂O is the same, but there is less H₂O in erythrocytes (most volume is occupied by hemoglobin) and thus less glucose in erythrocytes per unit volume. The greater the erythrocyte concentration in blood, the greater will be the difference between blood glucose concentration and plasma glucose concentration (blood glucose concentration will be lower). Plasma concentrations were calculated in Table 34-1 using the following conversion formula:

Table 34-1 Calculated Plasma Glucose Concentrations in Blood Samples with Given Whole-Blood Glucose Concentrations (WBG) and Hematocrit (Hct) Values^*

5 How do point-of-care glucose instruments measure glucose concentrations in whole blood?

The analytical methods vary and include reactions catalyzed by glucose oxidase, glucose dehydrogenase, or hexokinase. Each type of assay method has it advantages and disadvantages.

Some blood glucose analyzers measure the blood glucose concentration but then convert it to a plasma glucose concentration assuming a normal hematocrit percentage or fraction. If the hematocrit value is either increased or decreased, the calculated concentration would not equal the true plasma concentration. In the blood glucose assays, erythrocytes are lysed so that a whole-blood glucose (plasma glucose + erythrocyte glucose) concentration is measured. In other assays, a barrier excludes erythrocytes from the reacting reagents, and thus the assays measure plasma glucose concentrations.

Other blood glucose analyzers determine glucose concentration by assessing plasma molality. Displacement of plasma water by proteins or lipids can result in incorrect values. For some glucose oxidase methods, high arterial oxygen tension (high PaO₂, as may occur during gas anesthesia) can produce a positive interference.

Because there are at least 12 point-of-care glucose instruments and each has its unique features, the user should closely follow the manufacturer’s recommendations and know the limitations as described by the manufacturer or other resources.

6 How do most serum glucose assays measure glucose concentrations?

Most current serum glucose assays use hexokinase, glucokinase, glucose dehydrogenase, or glucose oxidase to catalyze the breakdown of glucose and the generation of products that can be measured by spectrophotometry, oxygen detection, or electron transfer. The glucose oxidase reaction is specific for glucose, but other factors can interfere with later stages of the assay (e.g., isopropyl alcohol can produce positive interference in reagent pad methods). Hexokinase can catalyze reactions with other hexoses, but rarely will there be another hexose in blood at sufficient concentration to increase the measured glucose concentration falsely.

7 How do most urine glucose assays estimate glucose concentrations?

In the reagent pad method, glucose reacts with glucose oxidase (which is glucose specific) to produce hydrogen peroxide (H₂O₂), which is detected when it reacts with a color indicator. The glucose concentration is proportional to color change in the reagent pad. Contamination of urine with H₂O₂ and sodium hypochlorite will produce false-positive reactions. False-negative reactions are caused by ascorbic acid, ketones, very concentrated urine samples, and possibly cold urine.

In the copper-reduction method (Clinitest), copper ion (Cu²⁺) reacts with a reducing substance (e.g., a hexose) to produce cuprous (Cu⁺) oxide and cuprous hydroxide and thus a color change. Cephalosporin and ascorbic acid may cause false-positive reactions.

8 In health, how similar are plasma/serum glucose concentrations and cerebrospinal fluid glucose concentrations?

Cerebrospinal fluid (CSF) glucose concentrations in health are approximately 60% to 80% of serum or plasma concentrations. CSF glucose concentrations may take 30 to 90 minutes to equilibrate with sudden changes in plasma/serum glucose concentration. In humans a decreased glucose concentration in the CSF (hypoglycorrachia) has been attributed to increased glucose metabolism by microorganisms or inflammatory cells (as in bacterial meningitis). However, this relationship is not a consistent finding in dogs.

9 How much does the plasma/serum glucose concentration contribute to plasma/serum osmolality?

The easiest way to understand the contribution of glucose to osmolality is to recognize that a glucose concentration of 1 mmol/L contributes 1 mOsm/L to osmolarity, which is approximately 1 mOsm/kg H₂O. Thus in healthy dogs and cats, a euglycemia of 5 mmol/L contributes about 5 mOsm/kg H₂O to the total plasma osmolality (almost 300 mOsm/kg H₂O). A marked hyperglycemia increases the plasma or serum osmolality; for example, increasing the glucose concentration from 5 to 25 mmol/L will increase the plasma osmolality about 20 mOsm/kg H₂O. Conversely, a marked hypoglycemia decreases the plasma or serum osmolality, but the minor changes (e.g., from 5 to 3 mmol/L) reduces the osmolality about 2 mmol/kg H₂O, a change of no clinical significance.

If glucose concentrations are reported in non-SI units (e.g., mg/dl), the glucose concentration is converted to mmol/L with the formula mg/dl × 0.05551 = mmol/L. The conversion is based on relative molecular weight of glucose (180 g = 1 mole) and the conversion of deciliters to liters.

10 What are the three major changes or alterations in physiologic processes that produce a hyperglycemia?

Hyperglycemia can be created by (a) increased glucose intake, (b) increased glucose production, and/or (c) decreased glucose uptake by peripheral tissues.

Increased glucose intake can result from ingestion of carbohydrate meal or the infusion of a glucose-containing fluid (e.g., 5% dextrose). Strictly speaking, increased glucose production is limited to increased gluconeogenesis within hepatocytes. However, hepatic glycogenolysis can also result in the release of glucose into blood. Decreased glucose uptake by peripheral tissues can result in a mild hyperglycemia by itself, but increased glucose uptake or increased glucose production can enhance the severity of the hyperglycemia. Figure 34-1 illustrates how these processes are influenced by activities of hormones and other factors.

Figure 34-1 Physiologic factors that influence blood glucose concentration.

Intestine: In monogastric animals, dietary carbohydrates (CHO) are broken down to monosaccharides (including glucose) that are absorbed in the small intestine, from which they enter portal blood and then systemic blood if not removed by hepatocytes.

Pancreas: Insulin and glucagon are released from pancreatic islet cells (beta cells and alpha cells, respectively). Insulin secretion is stimulated by increased blood concentrations of glucose, growth hormone (GH), glucagon, and amino acids. Glucagon secretion is stimulated by increased blood concentrations of amino acids and cortisol or by decreased blood glucose concentrations.

Liver: Hepatocytes are the primary source of blood glucose during fasting. Glucose can be obtained from glycogenolysis (stimulated by epinephrine and glucagon but inhibited by insulin) or gluconeogenesis (stimulated by glucagon and cortisol but inhibited by insulin). Insulin also promotes glycolysis. Increased glucose release from hepatocytes is promoted by increased glucagon, cortisol, or epinephrine. Insulin promotes the uptake of glucose by promoting glucokinase activity.

Muscle: Glucose uptake by myocytes is promoted by insulin through specific insulin receptors and glucose transporters; GH and cortisol inhibit the uptake of glucose. Insulin promotes glycogen synthesis in myocytes, whereas GH, glucagon (in cardiac muscle), and epinephrine promote glycogenolysis.

Adipose tissue: Insulin promotes the uptake of glucose by adipocytes through specific insulin receptors and glucose transporters.

Kidney: If the renal threshold is exceeded, hyperglycemic glucosuria will develop.

Pituitary: GH release from the pituitary is stimulated by growth hormone–releasing hormone (GHRH), which is released from the hypothalamus during hypoglycemia or after epinephrine stimulation.

Blood cells: Glucose enters erythrocytes (except porcine), leukocytes, and platelets through insulin-independent processes and is used in glycolysis and the hexose monophosphate shunt. Pig erythrocytes lack a functional glucose transporter and use inosine as the major energy substrate.

(Modified from Stockham SL, Scott MA: Fundamentals of veterinary clinical pathology, Ames 2002, Iowa State Press.

11 What is diabetes mellitus?

According to a 2000 report of the American Diabetes Association, “Diabetes mellitus is a group of metabolic diseases characterized by hyperglycemia resulting from defects in insulin secretion, insulin action, or both.” Type 1 diabetes mellitus (DM) is caused by beta cell destruction, which usually leads to absolute insulin deficiency (formerly called insulin-dependent DM, type I DM, or juvenile-onset DM). Type 2 DM is caused by insulin resistance with an inadequate compensatory insulin secretory response (formerly called non-insulin-dependent DM, type II DM, or adult-onset DM). The other forms of DM are referred to by the pathologic state or condition that created the carbohydrate metabolism defects (e.g., pancreatic, endocrine, drug-induced, infectious, and genetic DM).

12 What are the diagnostic criteria for diabetes mellitus?

A diagnosis of DM is appropriate when a hyperglycemia is associated with the clinical signs of polyuria (due to glucosuria), polydipsia, and weight loss. A hyperglycemia without the associated clinical signs is probably a transient hyperglycemia, and the animal probably does not have a disorder that is causing DM. However, the animal could be in a preclinical stage of DM. Diagnostic criteria in the preclinical stage include documentation of a carbohydrate (or glucose) intolerance.

13 What diseases and conditions cause hyperglycemia in domestic mammals?

Box 34-1 lists the diseases and conditions that may cause hyperglycemia.

Tags: Veterinary Clinical Pathology Secrets

Aug 26, 2016 | Posted by admin in INTERNAL MEDICINE | Comments Off

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