CHEMICAL EXAMINATION OF URINE

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Thursday, 10 August 2017 23:29
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The chemical examination is carried out for substances in urine are listed below:
 
  • Proteins
  • Glucose
  • Ketones
  • Bilirubin
  • Bile salts
  • Urobilinogen
  • Blood
  • Hemoglobin
  • Myoglobin
  • Nitrite or leukocyte esterase
 
PROTEINS

Normally, kidneys excrete scant amount of protein in urine (up to 150 mg/24 hours). These proteins include proteins from plasma (albumin) and proteins derived from urinary tract (Tamm-Horsfall protein, secretory IgA, and proteins from tubular epithelial cells, leucocytes, and other desquamated cells); this amount of proteinuria cannot be detected by routine tests. (Tamm-Horsfall protein is a normal mucoprotein secreted by ascending limb of the loop of Henle).
 
Proteinuria refers to protein excretion in urine greater than 150 mg/24 hours in adults.
 
Causes of Proteinuria
 

Box 826.1: Causes of proteinuria

• Glomerular proteinuria
• Tubular proteinuria
• Overflow proteinuria
• Hemodynamic (functional) proteinuria
• Post-renal proteinuria
Causes of proteinuria can be grouped as shown in Box 826.1.
 
  • Glomerular proteinuria: Proteinuria due to increased permeability of glomerular capillary wall is called as glomerular proteinuria.

    There are two types of glomerular proteinuria: selective and nonselective. In early stages of glomerular disease, there is increased excretion of lower molecular weight proteins like albumin and transferrin. When glomeruli can retain larger molecular weight proteins but allow passage of comparatively lower molecular weight proteins, the proteinuria is called as selective. With further glomerular damage, this selectivity is lost and larger molecular weight proteins (γ globulins) are also excreted along with albumin; this is called as nonselective proteinuria.

    Selective and nonselective proteinuria can be distinguished by urine protein electrophoresis. In selective proteinuria, albumin and transferrin bands are seen, while in nonselective type, the pattern resembles that of serum (Figure 826.1).

     
    • Massive proteinuria (>3.5 gm/24 hr)
    • Hypoalbuminemia (<3.0 gm/dl)
    • Generalised edema
    Hyperlipidemia (serum cholesterol >350 mg/dl)
    • Lipiduria
    Causes of glomerular proteinuria are glomerular diseases that cause increased permeability of glomerular basement membrane. The degree of glomerular proteinuria correlates with severity of disease and prognosis. Serial estimations of urinary protein are also helpful in monitoring response to treatment. Most severe degree of proteinuria occurs in nephrotic syndrome (Box 826.2).

  • Tubular proteinuria: Normally, glomerular membrane, although impermeable to high molecular weight proteins, allows ready passage to low molecular weight proteins like β2-microglobulin, retinol-binding protein, lysozyme, α1-microglobulin, and free immunoglobulin light chains. These low molecular weight proteins are actively reabsorbed by proximal renal tubules. In diseases involving mainly tubules, these proteins are excreted in urine while albumin excretion is minimal.

    Urine electrophoresis shows prominent α- and β-bands (where low molecular weight proteins migrate) and a faint albumin band (Figure 826.1).

    Tubular type of proteinuria is commonly seen in acute and chronic pyelonephritis, heavy metal poisoning, tuberculosis of kidney, interstitial nephritis, cystinosis, Fanconi syndrome and rejection of kidney transplant.

    Purely tubular proteinuria cannot be detected by reagent strip test (which is sensitive to albumin), but heat and acetic acid test and sulphosalicylic acid test are positive.

  • Overflow proteinuria: When concentration of a low molecular weight protein rises in plasma, it “overflows” from plasma into the urine. Such proteins are immunoglobulin light chains or Bence Jones proteins (plasma cell dyscrasias), hemoglobin (intravascular hemolysis), myoglobin (skeletal muscle trauma), and lysozyme (acute myeloid leukemia type M4 or M5).

  • Hemodynamic proteinuria: Alteration of blood flow through the glomeruli causes increased filtration of proteins. Protein excretion, however, is transient. It is seen in high fever, hypertension, heavy exercise, congestive cardiac failure, seizures, and exposure to cold.

    Postural (orthostatic) proteinuria occurs when the subject is standing or ambulatory, but is absent in recumbent position. It is common in adolescents (3-5%) and is probably due to lordotic posture that causes inferior venacaval compression between the liver and vertebral column. The condition disappears in adulthood. Amount of proteinuria is <1000 mg/day. First-morning urine after rising is negative for proteins, while another urine sample collected after patient performs normal activities is positive for proteins. In such patients, periodic testing for proteinuria should be done to rule out renal disease.

  • Post-renal proteinuria: This is caused by inflammatory or neoplastic conditions in renal pelvis, ureter, bladder, prostate, or urethra.
 
 
Figure 826.1 Glomerular and tubular proteinuria
Figure 826.1 Glomerular and tubular proteinuria. Upper figure shows normal serum protein electrophoresis pattern. Lower part shows comparison of serum and urine electrophoresis in (1) selective proteinuria, (2) non-selective proteinuria, and (3) tubular proteinuria
 
GLUCOSE
 
The main indication for testing for glucose in urine is detection of unsuspected diabetes mellitus or follow-up of known diabetic patients.
 
Box 826.3: Urine glucose
 
• Urine should be tested for glucose within 2 hours of collection (due to lowering of glucose by glycolysis and by contaminating bacteria which degrade glucose rapidly)
• Reagent strip test is a rapid, inexpensive, and semi-quantitative test
• In the past this test was used for home-monitoring of glucose; the test is replaced by glucometers.
• Urine glucose cannot be used to monitor control of diabetes since renal threshold is variable amongst individuals, no information about level of blood glucose below renal threshold is obtained, and urinary glucose value is affected by concentration of urine.
Practically all of the glucose filtered by the glomeruli is reabsorbed by the proximal renal tubules and returned to circulation. Normally a very small amount of glucose is excreted in urine (< 500 mg/24 hours or <15 mg/dl) that cannot be detected by the routine tests. Presence of detectable amounts of glucose in urine is called as glucosuria or glycosuria (Box 826.3). Glycosuria results if the filtered glucose load exceeds the capacity of renal tubular reabsorption. Most common cause is hyperglycemia from diabetes mellitus.
 
Causes of Glycosuria
 
1. Glycosuria with hyperglycemia:
 
  • Endocrine diseases: diabetes mellitus, acromegaly, Cushing’s syndrome, hyperthyroidism, pancreatic disease
  • Non-endocrine diseases: central nervous system diseases, liver disorders
  • Drugs: adrenocorticotrophic hormone, corticosteroids, thiazides
  • Alimentary glycosuria (Lag-storage glycosuria): After a meal, there is rapid intestinal absorption of glucose leading to transient elevation of blood glucose above renal threshold. This can occur in persons with gastrectomy or gastrojejunostomy and in hyperthyroidism. Glucose tolerance test reveals a peak at 1 hour above renal threshold (which causes glycosuria); the fasting and 2-hour glucose values are normal.
 
2. Glycosuria without hyperglycemia
 
  • Renal glycosuria: This accounts for 5% of cases of glycosuria in general population. Renal threshold is the highest glucose level in blood at which glucose appears in urine and which is detectable by routine laboratory tests. The normal renal threshold for glucose is 180 mg/dl. Threshold substances need a carrier to transport them from tubular lumen to blood. When the carrier is saturated, the threshold is reached and the substance is excreted. Up to this level glucose filtered by the glomeruli is efficiently reabsorbed by tubules. Renal glycosuria is a benign condition in which renal threshold is set below 180 mgs/dl but glucose tolerance is normal; the disorder is transmitted as autosomal dominant. Other conditions in which glycosuria can occur with blood glucose level remaining below 180 mgs/dl are renal tubular diseases in which there is decreased glucose reabsorption like Fanconi’s syndrome, and toxic renal tubular damage. During pregnancy, renal threshold for glucose is decreased. Therefore it is necessary to estimate blood glucose when glucose is first detected in urine.
 
 
KETONES
 
Excretion of ketone bodies (acetoacetic acid, β-hydroxybutyric acid, and acetone) in urine is called as ketonuria. Ketones are breakdown products of fatty acids and their presence in urine is indicative of excessive fatty acid metabolism to provide energy.
 
Causes of Ketonuria
 
Box 826.4: Urine ketones in diabetes
 
Indications for testing
 
• At diagnosis of diabetes mellitus
• At regular intervals in all known cases of diabetes, and in gestational diabetes
• In known diabetic patients during acute illness, persistent hyperglycemia (>300 mg/dl), pregnancy, clinical evidence of diabetic acidosis (nausea, vomiting, abdominal pain)
Normally ketone bodies are not detectable in the urine of healthy persons. If energy requirements cannot be met by metabolism of glucose (due to defective carbohydrate metabolism, low carbohydrate intake, or increased metabolic needs), then energy is derived from breakdown of fats. This leads to the formation of ketone bodies (Figure 826.2).
 
  1. Decreased utilization of carbohydrates:
    a. Uncontrolled diabetes mellitus with ketoacidosis: In diabetes, because of poor glucose utilization, there is compensatory increased lipolysis. This causes increase in the level of free fatty acids in plasma. Degradation of free fatty acids in the liver leads to the formation of acetoacetyl CoA which then forms ketone bodies. Ketone bodies are strong acids and produce H+ ions, which are neutralized by bicarbonate ions; fall in bicarbonate (i.e. alkali) level produces ketoacidosis. Ketone bodies also increase the plasma osmolality and cause cellular dehydration. Children and young adults with type 1 diabetes are especially prone to ketoacidosis during acute illness and stress. If glycosuria is present, then test for ketone bodies must be done. If both glucose and ketone bodies are present in urine, then it indicates presence of diabetes mellitus with ketoacidosis (Box 826.4).
    In some cases of diabetes, ketone bodies are increased in blood but do not appear in urine.
    Presence of ketone bodies in urine may be a warning of impending ketoacidotic coma.
    b. Glycogen storage disease (von Gierke’s disease)
  2. Decreased availability of carbohydrates in the diet:
    a. Starvation
    b. Persistent vomiting in children
    c. Weight reduction program (severe carbohydrate restriction with normal fat intake)
  3. Increased metabolic needs:
    a. Fever in children
    b. Severe thyrotoxicosis
    c. Pregnancy
    d. Protein calorie malnutrition
 
 
Figure 826.2 Formation of ketone bodies
Figure 826.2 Formation of ketone bodies. A small part of acetoacetate is spontaneously and irreversibly converted to acetone. Most is converted reversibly to β-hydroxybutyrate
 
BILE PIGMENT (BILIRUNIN)
 
Bilirubin (a breakdown product of hemoglobin) is undetectable in the urine of normal persons. Presence of bilirubin in urine is called as bilirubinuria.
 
There are two forms of bilirubin: conjugated and unconjugated. After its formation from hemoglobin in reticuloendothelial system, bilirubin circulates in blood bound to albumin. This is called as unconjugated bilirubin. Unconjugated bilirubin is not water-soluble, is bound to albumin, and cannot pass through the glomeruli; therefore it does not appear in urine. The liver takes up unconjugated bilirubin where it combines with glucuronic acid to form bilirubin diglucuronide (conjugated bilirubiun). Conjugated bilirubin is watersoluble, is filtered by the glomeruli, and therefore appears in urine.
 
Detection of bilirubin in urine (along with urobilinogen) is helpful in the differential diagnosis of jaundice (Table 826.1).
 
Table 826.1 Urine bilirubin and urobilinogen in jaundice
Urine test Hemolytic jaundice Hepatocellular jaundice Obstructive jaundice
1. Bilirubin Absent Present Present
2. Urobilinogen Increased Increased Absent
 
In acute viral hepatitis, bilirubin appears in urine even before jaundice is clinically apparent. In a fever of unknown origin bilirubinuria suggests hepatitis.
 
Presence of bilirubin in urine indicates conjugated hyperbilirubinemia (obstructive or hepatocellular jaundice). This is because only conjugated bilirubin is water-soluble. Bilirubin in urine is absent in hemolytic jaundice; this is because unconjugated bilirubin is water-insoluble.
 
 
BILE SALTS
 
Bile salts are salts of four different types of bile acids: cholic, deoxycholic, chenodeoxycholic, and lithocholic. These bile acids combine with glycine or taurine to form complex salts or acids. Bile salts enter the small intestine through the bile and act as detergents to emulsify fat and reduce the surface tension on fat droplets so that enzymes (lipases) can breakdown the fat. In the terminal ileum, bile salts are absorbed and enter in the blood stream from where they are taken up by the liver and re-excreted in bile (enterohepatic circulation).
 
 
UROBILINOGEN
 
Conjugated bilirubin excreted into the duodenum through bile is converted by bacterial action to urobilinogen in the intestine. Major part is eliminated in the feces. A portion of urobilinogen is absorbed in blood, which undergoes recycling (enterohepatic circulation); a small amount, which is not taken up by the liver, is excreted in urine. Urobilinogen is colorless; upon oxidation it is converted to urobilin, which is orange-yellow in color. Normally about 0.5-4 mg of urobilinogen is excreted in urine in 24 hours. Therefore, a small amount of urobilinogen is normally detectable in urine.
 
Urinary excretion of urobilinogen shows diurnal variation with highest levels in afternoon. Therefore, a 2-hour post-meal sample is preferred.
 
Causes of Increased Urobilinogen in Urine
 
  1. Hemolysis: Excessive destruction of red cells leads to hyperbilirubinemia and therefore increased formation of urobilinogen in the gut. Bilirubin, being of unconjugated type, does not appear in urine. Increased urobilinogen in urine without bilirubin is typical of hemolytic anemia. This also occurs in megaloblastic anemia due to premature destruction of erythroid precursors in bone marrow (ineffective erythropoiesis).
  2. Hemorrhage in tissues: There is increased formation of bilirubin from destruction of red cells.
 
Causes of Reduced Urobilinogen in Urine
 
  1. Obstructive jaundice: In biliary tract obstruction, delivery of bilirubin to the intestine is restricted and very little or no urobilinogen is formed. This causes stools to become clay-colored.
  2. Reduction of intestinal bacterial flora: This prevents conversion of bilirubin to urobilinogen in the intestine. It is observed in neonates and following antibiotic treatment.
 
Testing of urine for both bilirubin and urobilinogen can provide helpful information in a case of jaundice (Table 826.1).
 
 
BLOOD
 
The presence of abnormal number of intact red blood cells in urine is called as hematuria. It implies presence of a bleeding lesion in the urinary tract. Bleeding in urine may be noted macroscopically or with naked eye (gross hematuria). If bleeding is noted only by microscopic examination or by chemical tests, then it is called as occult, microscopic or hidden hematuria.
 
Causes of Hematuria
 
1. Diseases of urinary tract:
 
  • Glomerular diseases: Glomerulonephritis, Berger’s disease, lupus nephritis, Henoch-Schonlein purpura
  • Nonglomerular diseases: Calculus, tumor, infection, tuberculosis, pyelonephritis, hydronephrosis, polycystic kidney disease, trauma, after strenuous physical exercise, diseases of prostate (benign hyperplasia of prostate, carcinoma of prostate).
 
2. Hematological conditions:
 
Coagulation disorders, sickle cell disease Presence of red cell casts and proteinuria along with hematuria suggests glomerular cause of hematuria.
 
 
HEMOGLOBIN
 
Presence of free hemoglobin in urine is called as hemoglobinuria.
 
Causes of Hemoglobinuria
 
  1. Hematuria with subsequent lysis of red blood cells in urine of low specific gravity.
  2. Intravascular hemolysis: Hemoglobin will appear in urine when haptoglobin (to which hemoglobin binds in plasma) is completely saturated with hemoglobin. Intravascular hemolysis occurs in infections (severe falciparum malaria, clostridial infection, E. coli septicemia), trauma to red cells (march hemoglobinuria, extensive burns, prosthetic heart valves), glucose-6-phosphate dehydrogenase deficiency following exposure to oxidant drugs, immune hemolysis (mismatched blood transfusion, paroxysmal cold hemoglobinuria), paroxysmal nocturnal hemoglobinuria, hemolytic uremic syndrome, and disseminated intravascular coagulation.
 
Tests for Detection of Hemoglobinuria
 
Tests for detection of hemoglobinuria are benzidine test, orthotoluidine test, and reagent strip test.
 
HEMOSIDERIN
 
Hemosiderin in urine (hemosiderinuria) indicates presence of free hemoglobin in plasma. Hemosiderin appears as blue granules when urine sediment is stained with Prussian blue stain (Figure 826.3). Granules are located inside tubular epithelial cells or may be free if cells have disintegrated. Hemosiderinuria is seen in intravascular hemolysis.
 
Figure 826.3 Staining of urine sediment with Prussian blue stain
Figure 826.3 Staining of urine sediment with Prussian blue stain to demonstrate hemosiderin granules (blue)
 
MYOGLOBIN
 
Myoglobin is a protein present in striated muscle (skeletal and cardiac) which binds oxygen. Causes of myoglobinuria include injury to skeletal or cardiac muscle, e.g. crush injury, myocardial infarction, dermatomyositis, severe electric shock, and thermal burns.
 
Chemical tests used for detection of blood or hemoglobin also give positive reaction with myoglobin (as both hemoglobin and myoglobin have peroxidase activity). Ammonium sulfate solubility test is used as a screening test for myoglobinuria (Myoglobin is soluble in 80% saturated solution of ammonium sulfate, while hemoglobin is insoluble and is precipitated. A positive chemical test for blood done on supernatant indicates myoglobinuria).
 
Distinction between hematuria, hemoglobinuria, and myoglobinuria is shown in Table 826.2
 
Table 826.2 Differentiation between hematuria, hemoglobinuria, and myoglobinuria
Parameter Hematuria Hemoglobinuria Myoglobinuria
 1. Urine color  Normal, smoky, red, or brown  Pink, red, or brown  Red or brown
 2. Plasma color  Normal  Pink  Normal
 3. Urine test based on peroxidase activity  Positive  Positive  Positive
 4. Urine microscopy  Many red cells  Occasional red cell  Occasional red cell
 5. Serum haptoglobin  Normal  Low  Normal
 6. Serum creatine kinase  Normal  Normal  Markedly increased
.
Chemical Tests for Significant Bacteriuria (Indirect Tests for Urinary Tract Infection)
 
In addition to direct microscopic examination of urine sample, chemical tests are commercially available in a reagent strip format that can detect significant bacteriuria: nitrite test and leucocyte esterase test. These tests are helpful at places where urine microscopy is not available. If these tests are positive, urine culture is indicated.
 
1. Nitrite test: Nitrites are not present in normal urine; ingested nitrites are converted to nitrate and excreted in urine. If gram-negative bacteria (e.g. E.coli, Salmonella, Proteus, Klebsiella, etc.) are present in urine, they will reduce the nitrates to nitrites through the action of bacterial enzyme nitrate reductase. Nitrites are then detected in urine by reagent strip tests. As E. coli is the commonest organism causing urinary tract infection, this test is helpful as a screening test for urinary tract infection.
 
Some organisms like Staphylococci or Pseudomonas do not reduce nitrate to nitrite and therefore in such infections nitrite test is negative. Also, urine must be retained in the bladder for minimum of 4 hours for conversion of nitrate to nitrite to occur; therefore, fresh early morning specimen is preferred. Sufficient dietary intake of nitrate is necessary. Therefore a negative nitrite test does not necessarily indicate absence of urinary tract infection. The test detects about 70% cases of urinary tract infections.
 
2. Leucocyte esterase test: It detects esterase enzyme released in urine from granules of leucocytes. Thus the test is positive in pyuria. If this test is positive, urine culture should be done. The test is not sensitive to leucocytes < 5/HPF.
Last modified on Sunday, 20 August 2017 15:21
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    4. Pituitary TSH-secreting tumor.
     
    Causes of Decreased Total T4
     
    1. Primary hypothyroidism: The combination of decreased T4 and elevated TSH are indicative of primary hypothyroidism.
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    Free Thyroxine (FT4)
     
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    Total and Free Triiodothyronine (T3)
     
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    Thyrotropin Releasing Hormone (TRH) Stimulation Test
     
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    1. Confirmation of diagnosis of secondary hypothyroidism
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    Procedure
     
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    Interpretation
     
    1. Normal response: A rise of TSH > 2 mU/L at 20 minutes, and a small decline at 60 minutes.
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    Antithyroid Antibodies
     
    Box 864.1 Thyroid autoantibodies
     
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    Various autoantibodies (TSH receptor, antimicrosomal, and antithyroglobulin) are detected in thyroid disorders like Hashimoto’s thyroiditis and Graves’ disease. Antimicrosomal (also called as thyroid peroxidase) and anti-thyroglobulin antibodies are observed in almost all patients with Hashimoto’s disease. TSH receptor antibodies (TRAb) are mainly tested in Graves’ disease to predict the outcome after treatment (Box 864.1).
     
    Radioactive Iodine Uptake (RAIU) Test
     
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    • Hyperthyroidism due to Graves’ disease, toxic multinodular goiter, toxic adenoma, TSH-secreting tumor.
     
    Causes of Decreased Uptake
     
    • Hyperthyroidism due to administration of thyroid hormone, factitious hyperthyroidism, subacute thyroiditis.
     
    Uses
     
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    Thyroid Scintiscanning
     
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    Interpretation
     
    • Differential diagnosis of high RAIU thyrotoxicosis:
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    Table 864.1 Interpretation of thyroid function tests
    Test results Interpretations
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    2. Low TSH, Low FT4 Secondary hypothyroidism
    3. High TSH, Normal FT4 Subclinical hypothyroidism
    4. High TSH, Low FT4 Primary hypothyroidism
    5. Low TSH, Normal FT4, Normal FT3 Subclinical hyperthyroidism
    6. Low TSH, Normal FT4, High FT3 T3 toxicosis
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  • DISORDERS OF THYROID
    Box 863.1 Terminology in thyroid disorders
    • Primary hyper-/hypothyroidism: Increased or decreased function of thyroid gland due to disease of thyroid itself and not due to increased or decreased levels of TRH or TSH.
    • Secondary hyper-/hypothyroidism: Increased or decreased function of thyroid gland due to increased or decreased levels of TSH.
    • Tertiary hypothyroidism: Decreased function of thyroid gland due to decreased function of hypothalamus.
    • Subclinical thyroid disease: A condition with abnormality of thyroid hormone levels in blood but without specific clinical manifestations of thyroid disease and without any history of thyroid dysfunction or therapy.
    • Subclinical hyperthyroidism: A condition with normal thyroid hormone levels but with low or undetectable TSH level.
    • Subclinical hypothyroidism: A condition with normal thyroxine and triiodothyronine level along with mildly elevated TSH level.
    Among the endocrine disorders, disorders of thyroid are common and are only next in frequency to diabetes mellitus. They are more common in women than in men. Functional thyroid disorders can be divided into two types depending on activity of the thyroid gland: hypothyroidism (low thyroid hormones), and hyperthyroidism (excess thyroid hormones). Any enlargement of thyroid gland is called as a goiter. Terminology related to thyroid disorders is shown in Box 863.1.
     
    Hyperthyroidism
     
    Hyperthyroidism is a condition caused by excessive secretion of thyroid hormone. Causes of hyperthyroidism are listed in Table 863.1.
     
    Table 863.1 Causes of hyperthyroidism
    1. Graves’ disease (Diffuse toxic goiter)
    2. Toxicity in multinodular goiter
    3. Toxicity in adenoma
    4. Subacute thyroiditis
    5. TSH-secreting pituitary adenoma (secondary hyperthyroidism)
    6. Trophoblastic tumours that secrete TSH-like hormone: choriocarcinoma, hydatidiform mole
    7. Factitious hyperthyroidism
     
    Clinical Characteristics
     
    Clinical characteristics of hyperthyroidism are nervousness, anxiety, irritability, insomnia, fine tremors; weight loss despite normal or increased appetite; heat intolerance; increased sweating; dyspnea on exertion; amenorrhea and infertility; palpitations, tachycardia, cardiac arrhythmias, heart failure (especially in elderly); and muscle weakness, proximal myopathy, and osteoporosis (especially in elderly).
     
    The triad of Graves’ disease consists of hyperthyroidism, ophthalmopathy (exophthalmos, lid retraction, lid lag, corneal ulceration, impaired eye muscle function), and dermopathy (pretibial myxoedema).
     
    Box 863.2 Thyroid function tests in hyperthyroidism
    • Thyrotoxicosis:
      Serum TSH low or undetectable
      – Raised total T4 and free T4.
    • T3 toxicosis:
      – Serum TSH undetectable
      – Normal total T4 and free T4
      – Raised T3
    Laboratory Features
     
    In most patients, free serum T3 and T4 are elevated. In T3 thyrotoxicosis (5% cases of thyrotoxicosis), serum T4 levels are normal while T3 is elevated. Serum TSH is low or undetectable (< 0.1 mU/L) (Box 863.2).
     
    Undetectable or low serum TSH along with normal levels of T3 and T4 is called as subclinical hyperthyroidism; subtle signs and symptoms of thyrotoxicosis may or may not be present. Subclinical hyperthyroidism is associated with risk of atrial fibrillation, osteoporosis, and progression to overt thyroid disease.
     
    Features of primary and secondary hyperthyroidism are compared in Table 863.2.
     
    Table 863.2 Differences between primary and secondary hyperthyroidism
    Parameter Primary hyperthyroidism Secondary hyperthyroidism
    1. Serum TSH Low Normal or high
    2. Serum free thyroxine High High
    3. TSH receptor antibodies May be positive Negative
    4. Causes Graves’ disease, toxic multinodular goiter, toxic adenoma Pituitary adenoma
     
    Evaluation of hyperthyroidism is presented in Figure 863.1.
     
    Figure 863.1 Evaluation of hyperthyroidism
    Figure 863.1 Evaluation of hyperthyroidism. TSH: thyroid stimulating hormone; FT4: free T4; FT3: free T3; TRAb: TSH receptor antibody; TRH: Thyrotropin releasing hormone
     
    Hypothyroidism
     
    Hypothyroidism is a condition caused by deficiency of thyroid hormones. Causes of hypothyroidism are listed in Table 863.3. Primary hypothyroidism results from deficient thyroid hormone biosynthesis that is not due to disorders of hypothalamus or pituitary. Secondary hypothyroidism results from deficient secretion of TSH from pituitary. Deficient or loss of secretion of thyro-tropin releasing hormone from hypothalamus results in tertiary hypothyroidism. Secondary and tertiary hypothyroidism are much less common than primary. Plasma TSH is high in primary and low in secondary and tertiary hypothyroidism. Differences between primary and secondary hypothyroidism are shown in Table 863.4.
     
    Table 863.3 Causes of hypothyroidism 
    1. Primary hypothyroidism (Increased TSH)
      • Iodine deficiency
      • Hashimoto’s thyroiditis
      Exogenous goitrogens
      • Iatrogenic: surgery, drugs, radiation
    2. Secondary hypothyroidism (Low TSH): Diseases of pituitary
    3. Tertiary hypothyroidism (Low TSH, Low TRH): Diseases of hypothalamus
     
    Table 863.4 Differences between primary and secondary hypothyroidism
    Parameter Primary hypothyroidism Secondary hypothyroidism
    1. Cause Hashimoto’s thyroiditis Pituitary disease
    2. Serum TSH High Low
    3. Thyrotropin releasing hormone stimulation test Exaggerated response No response
    4. Antimicrosomal antibodies Present Absent
     
    Box 863.3 Thyroid function tests in hypothyroidism
    • Primary hypothyroidism
      – Serum TSH: Increased (proportional to degree of hypofunction)
      – Free T4: Decreased
      – TRH stimulation test: Exaggerated response
    • Secondary hypothyroidism
      – Serum TSH: Decreased
      – Free T4: Decreased
      – TRH stimulation test: Absent response
    • Tertiary hypothyroidism
      – Serum TSH: Decreased
      – FT4: Decreased
      – TRH stimulation test: Delayed response
    Clinical features of primary hypothyroidism are: lethargy, mild depression, disturbances in menstruation, weight gain, cold intolerance, dry skin, myopathy, constipation, and firm and lobulated thyroid gland (in Hashimoto’s thyroiditis).
     
    In severe cases, myxoedema coma (an advanced stage with stupor, hypoventilation, and hypothermia) can occur.
     
    Laboratory Features
     
    Laboratory features in hypothyroidism are shown in Box 863.3.
     
    Normal serum thyroxine (T4 and FT4) coupled with a moderately raised TSH (>10 mU/L) is referred to as subclinical hypothyroidism. It is associated with bad obstetrical outcome, poor cognitive development in children, and high risk of hypercholesterolemia and progression to overt hypothyroidism.
     
    Evaluation of hypothyroidism is presented in Figure 863.2
     
    Figure 863.2 Evaluation of hypothyroidism
    Figure 863.2 Evaluation of hypothyroidism. TSH: thyroid stimulating hormone; FT4: free T4; TRH: Thyrotropin releasing hormone
  • FEMALE INFERTILITY: CAUSES AND INVESTIGATIONS
    The ovaries are the sites of production of female gametes or ova by the process of oogenesis. The ova are released by the process of ovulation in a cyclical manner at regular intervals. Ovary contains numerous follicles that contain ova in various stages of development. During each menstrual cycle, up to 20 primordial follicles are activated for maturation; however, only one follicle becomes fully mature; this dominant follicle ruptures to release the secondary oocyte from the ovary. Maturation of the follicle is stimulated by follicle stimulating hormone (FSH) secreted by anterior pituitary (Figure 862.1). Maturing follicle secretes estrogen that causes proliferation of endometrium of the uterus (proliferative phase). Follicular cells also secrete inhibin which regulates release of FSH by the anterior pituitary. Fall in FSH level is followed by secretion of luteinizing hormone (LH) by the anterior pituitary (LH surge). This causes follicle to rupture and the ovum is expelled into the peritoneal cavity near the fimbrial end of the fallopian tube. The fallopian tubes conduct ova from the ovaries to the uterus. Fertilization of ovum by the sperm occurs in the fallopian tube.
     
    Figure 862.1 The hypothalamus pituitary ovarian axis
    Figure 862.1 The hypothalamus-pituitary-ovarian axis 
     
    The ovum consists of the secondary oocyte, zona pellucida and corona radiata. The ruptured follicle in the ovary collapses and fills with blood clot (corpus luteum). LH converts granulose cells in the follicle to lutein cells which begin to secrete progesterone. Progesterone stimulates secretion from the endometrial glands (secretory phase) that were earlier under the influence of estrogen. Rising progesterone levels inhibit LH production from the anterior pituitary. Without LH, the corpus luteum regresses and becomes functionless corpus albicans. After regression of corpus luteum, production of estrogen and progesterone stops and endometrium collapses, causing onset of menstruation. If the ovum is fertilized and implanted in the uterine wall, human chorionic gonadotropin (hCG) is secreted by the developing placenta into the maternal circulation. Human chorionic gonadotropin maintains the corpus luteum for secetion of estrogen and progesterone till 12th week of pregnancy. After 12th week, corpus luteum regresses to corpus albicans and the function of synthesis of estrogen and progesterone is taken over by placenta till parturition.
     
    The average duration of the normal menstrual cycle is 28 days. Ovulation occurs around 14th day of the cycle. The time interval between ovulation and menstruation is called as luteal phase and is fairly constant (14 days) (Figure 862.2).
     
    Figure 862.2 Normal menstrual cycle
    Figure 862.2 Normal menstrual cycle
     
    Causes of Female Infertility
     
    Causes of female infertility are shown in Table 862.1.
     
    Table 862.1 Causes of female infertility
    1. Hypothalamic-pituitary dysfunction:
    • Hypothalamic causes
      – Excessive exercise
      – Excess stress
      – Low weight
      – Kallman’s syndrome
      Idiopathic
    • Pituitary causes
      – Hyperprolactinemia
      Hypopituitarism (Sheehan’s syndrome, Simmond’s disease)
      – Craniopharyngioma
      – Cerebral irradiation
     2. Ovarian dysfunction:
    • Polycystic ovarian disease (Stein-Leventhal syndrome)
    • Luteinized unruptured follicle
    • Turner’s syndrome
    • Radiation or chemotherapy
    • Surgical removal of ovaries
    • Idiopathic
     3. Dysfunction in passages:
    • Fallopian tubes
      Infections: Tuberculosis, gonorrhea, Chlamydia
      – Previous surgery (e.g. laparotomy)
      – Tubectomy
      Congenital hypoplasia, non-canalization
      Endometriosis
    • Uterus
      – Uterine malformations
      – Asherman’s syndrome
      – Tuberculous endometritis
      Fibroid
    • Cervix: Sperm antibodies
    • Vagina: Septum
     4. Dysfunction of sexual act: Dyspareunia
     
    Investigations
     
    Evaluation of female infertility is shown in Figure 862.3.
     
    Figure 862.3 Evaluation of female infertility
    Figure 862.3 Evaluation of female infertility. FSH: Follicle stimulating hormone; LH: Luteinizing hormone; DHEA-S: Dihydroepiandrosterone; TSH: Thyroid stimulating hormone; ↑ : Increased; ↓ : Decreased
     
    Tests for Ovulation
     
    Most common cause of female infertility is anovulation.
     
    1. Regular cycles, mastalgia, and laparoscopic direct visualization of corpus luteum indicate ovulatory cycles. Anovulatory cycles are clinically characterized by amenorrhea, oligomenorrhea, or irregular menstruation. However, apparently regular cycles may be associated with anovulation.
    2. Endometrial biopsy: Endometrial biopsy is done during premenstrual period (21st-23rd day of the cycle). The secretory endometrium during the later half of the cycle is an evidence of ovulation.
    3. Ultrasonography (USG): Serial ultrasonography is done from 10th day of the cycle and the size of the dominant follicle is measured. Size >18 mm is indicative of imminent ovulation. Collapse of the follicle with presence of few ml of fluid in the pouch of Douglas is suggestive of ovulation. USG also is helpful for treatment (i.e. timing of coitus or of intrauterine insemination) and diagnosis of luteinized unruptured follicle (absence of collapse of dominant follicle). Transvaginal USG is more sensitive than abdominal USG.
    4. Basal body temperature (BBT): Patient takes her oral temperature at the same time every morning before arising. BBT falls by about 0.5°F at the time of ovulation. During the second (progestational) half of the cycle, temperature is slightly raised above the preovulatory level (rise of 0.5° to 1°F). This is due to the slight pyrogenic action of progesterone and is therefore presumptive evidence of functional corpus luteum.
    5. Cervical mucus study:
      Fern test: During estrogenic phase, a characteristic pattern of fern formation is seen when cervical mucus is spread on a glass slide (Figure 862.4). This ferning disappears after the 21st day of the cycle. If previously observed, its disappearance is presumptive evidence of corpus luteum activity.
      Spinnbarkeit test: Cervical mucus is elastic and withstands stretching upto a distance of over 10 cm. This phenomenon is called Spinnbarkeit or the thread test for the estrogen activity. During the secretory phase, viscosity of the cervical mucus increases and it gets fractured when stretched. This change in cervical mucus is evidence of ovulation.
    6. Vaginal cytology: Karyopyknotic index (KI) is high during estrogenic phase, while it becomes low in secretory phase. This refers to percentage of super-ficial squamous cells with pyknotic nuclei to all mature squamous cells in a lateral vaginal wall smear. Usually minimum of 300 cells are evaluated. The peak KI usually corresponds with time of ovulation and may reach upto 50 to 85.
    7. Estimation of progesterone in mid-luteal phase (day 21 or 7 days before expected menstruation): Progesterone level > 10 nmol/L is a reliable evidence of ovulation if cycles are regular (Figure 862.5). A mistimed sample is a common cause of abnormal result.
     
    Figure 862.4 Ferning of cervical mucosa
    Figure 862.4 Ferning of cervical mucosa
     
    Figure 862.5 Serum progesterone during normal menstrual cycle
    Figure 862.5 Serum progesterone during normal menstrual cycle
     
    Tests to Determine the Cause of Anovulation
     
    1. Measurement of LH, FSH, and estradiol during days 2 to 6: All values are low in hypogonadotropic hypogonadism (hypothalamic or pituitary failure).
    2. Measurement of TSH, prolactin, and testosterone if cycles are irregular or absent:
      Increased TSH: Hypothyroidism
      Increased prolactin: Pituitary adenoma
      Increased testosterone: Polycystic ovarian disease (PCOD), congenital adrenal hyperplasia (To differentiate PCOD from congenital adrenal hyperplasia, ultrasound and estimation of dihydroepiandrosterone or DHEA are done).
    3. Transvaginal ultrasonography: This is done for detection of PCOD.
     
    Investigations to Assess Tubal and Uterine Status
     
    1. Infectious disease: These tests include endometrial biopsy for tuberculosis and test for chlamydial IgG antibodies for tubal factor in infertility.
    2. Hysterosalpingography (HSG): HSG is a radiological contrast study for investigation of the shape of the uterine cavity and for blockage of fallopian tubes (Figure 862.6). A catheter is introduced into the cervical canal and a radiocontrast dye is injected into the uterine cavity. A real time X-ray imaging is carried out to observe the flow of the dye into the uterine cavity, tubes, and spillage into the uterine cavity.
    3. Hysterosalpingo-contrast sonography: A catheter is introduced into the cervical canal and an echocontrast fluid is introduced into the uterine cavity. Shape of the uterine cavity, filling of fallopian tubes, and spillage of contrast fluid are noted. In addition, ultrasound scan of the pelvis provides information about any fibroids or polycystic ovarian disease.
    4. Laparoscopy and dye hydrotubation test with hysteroscopy: In this test, a cannula is inserted into the cervix and methylene blue dye is introduced into the uterine cavity. If tubes are patent, spillage of the dye is observed from the ends of both tubes. This technique also allows visualization of pelvic organs, endometriosis, and pelvic adhesions. If required, endometriosis and tubal blockage can be treated during the procedure.
     
    Possible pregnancy and active pelvic or vaginal infection are contraindications to tubal patency tests.
     
    Figure 862.6 Hysterosalpingography
    Figure 862.6 Hysterosalpingography
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