PREGNANCY TESTS

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Wednesday, 16 August 2017 12:56
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Pregnancy tests detect human chorionic gonadotropin (hCG) in serum or urine. Although pregnancy is the most common reason for ordering the test for hCG, measurement of hCG is also indicated in other conditions as shown in Box 836.1.
 
Box 836.1 Indications for measurement of β human chorionic gonadotropin

• Early diagnosis of pregnancy
• Diagnosis and management of gestational trophoblastic disease
• As a part of maternal triple test screen
• Follow-up of malignant tumors that produce β human chorionic gonadotropin.
Human chorionic gonadotropin is a glycoprotein hormone produced by placenta that circulates in maternal blood and excreted intact by the kidneys. It consists of two polypeptide subunits: α (92 amino acids) and β (145 amino acids) which are non-covalently bound to each other. Structurally, hCG is closely related to three other glycoprotein hormones, namely, luteinizing hormone (LH), follicle-stimulating hormone (FSH), and thyroid-stimulating hormone (TSH). The α subunits of hCG, LH, FSH, and TSH are similar, while β subunits differ and confer specific biologic and immunologic properties. Immunological tests use antibodies directed against β-subunit of hCG to avoid cross-reactivity against LH, FSH, and TSH.
 
Syncytiotrophoblastic cells of conceptus and later of placenta synthesize hCG. Human chorionic gonadotropin supports the corpus luteum of ovary during early pregnancy. Progesterone, produced by corpus luteum, prevents ovulation and thus maintains pregnancy. After 7-10 weeks of gestation, sufficient amounts of progesterone are synthesized by placenta, and hCG is no longer needed and its level declines.
 
CLINICAL APPLICATIONS OF TESTS FOR HUMAN CHORIONIC GONADOTROPIN
 
  1. Early diagnosis of pregnancy: Qualitative serum hCG test becomes positive 3 weeks after last menstrual period (LMP), while urine hCG test becomes positive 5 weeks after LMP.
  2. Exclusion of pregnancy before prescribing certain medications (like oral contraceptives, steroids, some antibiotics), and before ordering radiological studies, radiotherapy, or chemotherapy. This is necessary to prevent any teratogenic effect on the fetus.
  3. Early diagnosis of ectopic pregnancy: Trans-vaginal ultrasonography (USG) and quantitative estimation of hCG are helpful in early diagnosis of ectopic pregnancy (before rupture).
  4. Evaluation of threatened abortion: Serial quantitative estimation of hCG is helpful in following the course of threatened abortion.
  5. Diagnosis and follow-up of gestational trophoblastic disease (GTD).
  6. Maternal triple test screen: This consists of measurement of hCG, α-fetoprotein, and unconjugated estriol in maternal serum at 14-19 weeks of gestation. The maternal triple screen identifies pregnant women with increased risk of Down syndrome and major congenital anomalies like neural tube defects.
  7. Follow-up of ovarian or testicular germ cell tumors, which produce hCG.
 
Normal Pregnancy
 
In women with normal menstrual cycle, conception (fertilization of ovum to form a zygote) occurs on day 14 in the fallopian tube. Zygote travels down the fallopian tube into the uterus. Division of zygote produces a morula. At 50-60-cell stage, morula develops a primitive yolk sac and is then called as a blastocyst. About 5 days after fertilization, implantation of blastocyst occurs in the uterine wall. Trophoblastic cells (on the outer surface of the blastocyst) penetrate the endometrium and develop into chorionic villi. There are two main forms of trophoblasts—syncytiotrophoblast and cytotrophoblast. Placental development occurs from chorionic villi. After formation of placenta, the conceptus is called as an embryo. When embryo develops most major organs, it is called as fetus (after 10 weeks of gestation).
 
Figure 836.1 Level of human chorionic gonadotropin during pregnancy
Figure 836.1 Level of human chorionic gonadotropin during pregnancy
 
Box 836.2 Diagnosis of early pregnancy

• Positive serum hCG test: 8 days after conception or 3 weeks after last menstrual period (LMP)
• Positive urine hCG test: 21 days after conception or 5 weeks after LMP
• Ultrasonography for visualization of gestational sac:
– Transvaginal: 21 days after conception or 5 weeks after LMP
– Transabdominal: 28 days after conception or 6 weeks after LMP
Human chorionic gonadotropin is synthesized by syncytiotrophoblasts (of placenta) and detectable amounts (~5 mIU/ml) appear in maternal serum about 8 days after conception (3 weeks after LMP). In the first trimester (first 12 weeks, calculated from day 1 of LMP) of pregnancy, hCG levels rapidly rise with a doubling time of about 2 days. Highest or peak level is reached at 8-10 weeks (about 100,000 mIU/ml). This is followed by a gradual fall, and from 15-16 weeks onwards, a steady level of 10,000-20,000 mIU/ml is maintained for the rest of the pregnancy (Figure 836.1). After delivery, hCG becomes non-detectable by about 2 weeks.
 
Box 836.2 shows minimum time required for the earliest diagnosis of pregnancy by hCG test and ultrasonography (USG).
 
Two types of pregnancy tests are available:
 
  • Qualitative tests: These are positive/negative result types that are done on urine sample.
  • Quantitative tests: These give numerical result and are done on serum or urine. They are also used for evaluation of ectopic pregnancy, failing pregnancy, and for follow-up of gestational trophoblastic disease.
 
Ectopic Pregnancy
 
Ectopic pregnancy refers to the implantation of blastocyst at a site other than the cavity of uterus. The most common of such sites (>95% cases) is fallopian tube. Early diagnosis and treatment of tubal ectopic pregnancy is essential since it can lead to maternal mortality (from rupture and hemorrhage) and future infertility. Ectopic pregnancy is a leading cause of maternal death during first trimester. Diagnosis of ectopic pregnancy can be readily made in most cases by ultrasonography and estimation of β-subunit of human chorionic gonadotropin.
 
Early diagnosis of unruptured tubal pregnancy can be made by quantitative estimation of serum hCG and ultrasonography. In normal intrauterine pregnancy, hCG titer doubles every 2 days until first 40 days of gestation. If hCG rise is abnormally slow, then an unviable pregnancy (either ectopic or abnormal intrauterine pregnancy) should be suspected.
 
Transabdominal USG can detect gestational sac in intrauterine pregnancy 6 weeks after LMP. The level of hCG in serum at this stage is >6500 mIU/ml. If gestational sac is not visualized at this level of hCG, then there is a possibility of ectopic pregnancy. Transvaginal ultrasonography can detect ectopic pregnancy average 1 week earlier than abdominal ultrasonography; it can detect gestational sac if β-hCG level is 1000-1500 mIU/ml. Therefore, if gestational sac is not visualized in the presence of >1500 mIU/ml of β-hCG level, an ectopic pregnancy can be suspected.
 
Early diagnosis of ectopic pregnancy provides the option of administration of intramuscular methotrexate (rather than surgery), which causes dissolution of conceptus. This improves the chances of patient’s future fertility. Serial measurements of hCG after surgical removal of ectopic pregnancy can help in detecting persistence of trophoblastic tissue.
 
Abortion
 
Termination of pregnancy before fetus becomes viable (i.e. before 20 weeks) is called as abortion.
 
In threatened abortion, vaginal bleeding is present but internal os is closed and process of abortion, though started, is still reversible. It is possible that pregnancy will continue.
 
Serial quantitative titers of hCG showing lack of expected doubling of hCG level and USG are helpful in diagnosis and management of abortion.
 
Gestational Trophoblastic Disease (GTD)
 
It is characterized by proliferation of pregnancyassociated trophoblastic tissue. The two main forms of GTD are hydatidiform (vesicular) mole (benign) and choriocarcinoma (malignant). Clinical features of GTD are as follows:
 
  • Short history of amenorrhea followed by vaginal bleeding.
  • Size of uterus larger than gestational age; uterus is soft and doughy on palpation with no fetal parts and no fetal heart sounds.
  • Excessive nausea and vomiting due to high hCG.
  • Characteristic snowstorm appearance on pelvic USG.
 
Quantitative estimation of hCG is helpful in diagnosis and management of GTD.
 
Trophoblastic cells of GTD produce more hCG as compared to the trophoblasts of normal pregnancy for the same gestational age. Concentration of hCG parallels tumor load. Also, hCG continues to rise beyond 10 weeks of gestation without reaching plateau (as expected at the end of first trimester).
 
After evacuation of uterus, weekly estimation of hCG is advised till subsequent three (weekly) results are negative; following evacuation of vesicular mole, hCG becomes undetectable (after 2-3 months) on follow-up in 80% of cases. Plateau or rising hCG indicates persistent GTD. In such cases, chemotherapy is indicated.
 
Negative results for hCG after therapy should be regularly followed up every 3 months for 1-2 years.
 
LABORATORY TESTS FOR HUMAN CHORIONIC GONADOTROPIN
 
These are classified into two main groups:
 
  • Biological assays or bioassays
  • Immunological assays
 
Bioassays
 
In bioassay, effect of hCG is tested on laboratory animals under standardized conditions. There are several limitations of bioassays like need for animal facilities, need for standardization of animals, long time required for the test results, low sensitivity, and high cost. Therefore, bioassays have been replaced by immunological assays.
 
In Ascheim-Zondek test, urine from pregnant woman is injected into immature female mice. Formation of hemorrhagic corpora lutea in ovaries (after 4 days) is a positive test. Friedman test is similar except that urine is injected into female rabbit. In rapid rat test, injection of urine containing hCG into female rats is followed by hyperaemia and hemorrhage in ovaries. Yet another test measures release of spermatozoa from male frog after injection of urine containing hCG.
 
Immunological Assays
 
These are rapid and sensitive tests for detection and quantitation of hCG. Variable results are obtained by different immunological tests with the same serum sample; this is due to differences in specificity of different immunoassays to complete hCG, β-subunit, and β-core fragment. A number of immunological tests are commercially available based on different principles like agglutination inhibition assay, enzyme immunoassay including enzyme linked immunosorbent assay or ELISA, radioimmunoassay (RIA), and immunoradiometric assay.
 
A commonly used qualitative urine test is agglutination inhibition assay. Early morning urine specimen is preferred because it contains the highest concentration of hCG. Causes of false-positive test include red cells, leukocytes, bacteria, some drugs, proteins, and excess luteinizing hormone (menopause, midcycle LH surge) in urine. Some patients have anti-mouse antibodies (that are used in the test), while others have hCG-like material in circulation, producing false-positive test. Anti-mouse antibodies also interfere with other antibody-based tests and are known as ‘heterophil’ antibodies. Fetal death, abortion, dilute urine, and low sensitivity of a particular test are causes of false-negative test. Renal failure leads to accumulation of interfering substances causing incorrect results.
 
Figure 836.2 Principle of agglutination inhibition test for diagnosis of pregnancy
Figure 836.2 Principle of agglutination inhibition test for diagnosis of pregnancy
 
In latex particle agglutination inhibition test (Figure 836.2), anti-hCG antibodies are incubated with patient’s urine. This is followed by addition of hCGcoated latex particles. If hCG is present in urine, anti-hCG serum is neutralized, and no agglutination of latex particles occurs (positive test). If there is no hCG in urine, there is agglutination of latex particles (negative test). This is commonly used as a slide test and requires only a few minutes.
 
Sensitivity of agglutination inhibition test is >200 units/liter of hCG.
 
Radioimmunoassay, enzyme immunoassay, and radioimmunometric assay are more sensitive and reliable than agglutination inhibition assay.
 
Quantitative tests are employed for detection of very early pregnancy, estimation of gestational age, diagnosis of ectopic pregnancy, evaluation of threatened abortion, and management of GTD.
 
REFERENCE RANGES
 
  • Serum human chorionic gonadotropin:
    – Non-pregnant females: <5.0 mIU/ml
    – Pregnancy: 4 weeks after LMP: 5-100 mIU/ml
    – 5 weeks after LMP: 200-3000 mIU/ml
    – 6 weeks after LMP: 10,000-80,000 mIU/ml
    – 7-14 weeks: 90,000-500,000 mIU/ml
    – 15-26 weeks: 5000-80000 mIU/ml
    – 27-40 weks: 3000-15000 mIU/ml
 
 Further Reading:
 
Last modified on Sunday, 20 August 2017 15:25
Dayyal Dg.

Clinical laboratory professional specialized to external quality assessment (proficiency testing) schemes for Laboratory medicine and clinical pathology. | Author/Writer/Blogger

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  • TOTAL THYROXINE (T4)
    Total serum thyroxine includes both free and protein-bound thyroxine and is usually measured by competitive immunoassay. Normal level in adults is 5.0-12.0 μg/dl.
     
    Test for total thyroxine or free thyroxine is usually combined with TSH measurement and together they give the best assessment of thyroid function.
     
    Causes of Increased Total T4
     
    1. Hyperthyroidism: Elevation of both T4 and T3 values along with decrease of TSH are indicative of primary hyperthyroidism.
    2. Increased thyroxine-binding globulin: If concentration of TBG increases, free hormone level falls, release of TSH from pituitary is stimulated, and free hormone concentration is restored to normal. Reverse occurs if concentration of binding proteins falls. In either case, level of free hormones remains normal, while concentration of total hormone is altered. Therefore, estimation of only total T4 concentration can cause misinterpretation of results in situations that alter concentration of TBG.
    3. Factitious hyperthyroidism
    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.
    2. Secondary or pituitary hypothyroidism
    3. Tertiary or hypothalamic hypothyroidism
    4. Hypoproteinaemia, e.g. nephrotic syndrome
    5. Drugs: oestrogen, danazol
    6. Severe non-thyroidal illness.
     
    Free Thyroxine (FT4)
     
    FT4 comprises of only a small fraction of total T4, is unbound to proteins, and is the metabolically active form of the hormone. It constitutes about 0.05% of total T4. Normal range is 0.7 to 1.9 ng/dl. Free hormone concentrations (FT4 and FT3) correlate better with metabolic state than total hormone levels (since they are not affected by changes in TBG concentrations).
     
    Measurement of FT4 is helpful in those situations in which total T4 level is likely to be altered due to alteration in TBG level (e.g. pregnancy, oral contraceptives, nephrotic syndrome).
     
    Total and Free Triiodothyronine (T3)
     
    Uses
     
    1. Diagnosis of T3 thyrotoxicosis: Hyperthyroidism with low TSH and elevated T3, and normal T4/FT4 is termed T3 thyrotoxicosis.
    2. Early diagnosis of hyperthyroidism: In early stage of hyperthyroidism, total T4 and free T4 levels are normal, but T3 is elevated.
     
    A low T3 level is not useful for diagnosis of hypothyroidism since it is observed in about 25% of normal individuals.
     
    For routine assessment of thyroid function, TSH and T4 are measured. T3 is not routinely estimated because normal plasma levels are very low.
     
    Normal T3 level is 80-180 ng/dl.
     
    Free T3: Measurement of free T3 gives true values in patients with altered serum protein levels (like pregnancy, intake of estrogens or oral contraceptives, and nephrotic syndrome). It represents 0.5% of total T3.
     
    Thyrotropin Releasing Hormone (TRH) Stimulation Test
     
    Uses
     
    1. Confirmation of diagnosis of secondary hypothyroidism
    2. Evaluation of suspected hypothalamic disease
    3. Suspected hyperthyroidism
     
    This test is not much used nowadays due to the availability of sensitive TSH assays.
     
    Procedure
     
    • A baseline blood sample is collected for estimation of basal serum TSH level.
    • TRH is injected intravenously (200 or 500 μg) followed by measurement of serum TSH at 20 and 60 minutes.
     
    Interpretation
     
    1. Normal response: A rise of TSH > 2 mU/L at 20 minutes, and a small decline at 60 minutes.
    2. Exaggerated response: A further significant rise in already elevated TSH level at 20 minutes followed by a slight decrease at 60 minutes; occurs in primary hypothyroidism.
    3. Flat response: There is no response; occurs in secondary (pituitary) hypothyroidism.
    4. Delayed response: TSH is higher at 60 minutes as compared to its level at 20 minutes; seen in tertiary (hypothalamic) hypothyroidism.
     
    Antithyroid Antibodies
     
    Box 864.1 Thyroid autoantibodies
     
    • Useful for diagnosis and monitoring of autoimmune thyroid diseases.
    • Antimicrosomal or antithyroid peroxidase antibodies: Hashimoto’s thyroiditis
    • Anti-TSH receptor antibodies: Graves’ disease
    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
     
    This is a direct test that assesses the trapping of iodide by thyroid gland (through the iodine symporters or pumps in follicular cells) for thyroid hormone synthesis. Patient is administered a tracer dose of radioactive iodine (131I or 123I) orally. This is followed by measurement of amount of radioactivity over the thyroid gland at 2 to 6 hours and again at 24 hours. RAIU correlates directly with the functional activity of the thyroid gland. Normal RAIU is about 10-30% of administered dose at 24 hours, but varies according to the geographic location due to differences in dietary intake.
     
    Causes of Increased Uptake
     
    • 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
     
    RAIU is most helpful in differential diagnosis of hyperthyroidism by separating causes into those due to increased uptake and due to decreased uptake.
     
    Thyroid Scintiscanning
     
    An isotope (99mTc-pertechnetate) is administered and a gamma counter assesses its distribution within the thyroid gland.
     
    Interpretation
     
    • Differential diagnosis of high RAIU thyrotoxicosis:
      – Graves’ disease: Uniform or diffuse increase in uptake
      – Toxic multinodular goiter: Multiple discrete areas of increased uptake
      – Adenoma: Single area of increased uptake
    • Evaluation of a solitary thyroid nodule:
      – ‘Hot’ nodule: Hyperfunctioning
      – ‘Cold’ nodule: Non-functioning; about 20% cases are malignant.
     
    Interpretation of thyroid function tests is shown in Table 164.1.
     
    Table 864.1 Interpretation of thyroid function tests
    Test results Interpretations
    1. TSH Normal, FT4 Normal Euthyroid
    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
    7. Low TSH, High FT4 Primary hyperthyroidism
     
    Neonatal Screening for Hypothyroidism
     
    Thyroid hormone deficiency during neonatal period can cause severe mental retardation (cretinism) that can be prevented by early detection and treatment. Estimation of TSH is done on dry blood spots on filter paper or cord serum between 3rd to 5th days of life. Elevated TSH is diagnostic of hypothyroidism. In infants with confirmed hypothyroidism, RAIU (123I) scan should be done to distinguish between thyroid agenesis and dyshormonogenesis.
  • 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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