Pediatrics clinical chemistry

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A seminar discussing the role of the Clinical Chemistry lab with pediatrics
Ola Elgaddar

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Pediatrics clinical chemistry

  1. 1. Pediatrics Clinical Chemistry
  2. 2. Pediatrics involves both intrauterine & extra uterine periods
  3. 3. Definitions Child: • Legal definition: A human being below the age of 18 years unless under the law applicable to the child, majority is attained earlier. • Biological definition: Anyone in the developmental stage of childhood, between infancy and adulthood.
  4. 4. - Neonate: newborn up to first 28 days of life - Infant: comprises neonatal period up to 12 months - Toddler: 1-3 years - Pre-school: 3-5 years - School-age: 6-10 years - Adolescent: 11-14 years
  5. 5. Some pediatrics clinical chemistry reference intervals
  6. 6. - Laboratory evaluation starts from the prenatal period. - The most important clinical conditions are: 1) Evaluation of maturity. 2) Diagnosis of congenital abnormalities
  7. 7. Intrauterine Growth Retardation (IUGR) AGA (Appropriate for Gestational Age): Birth weight is between 10th and 90th percentile for infant’s gestational age (GA). SGA (Small for Gestational Age): Birth weight <10th percentile for GA. IUGR (Intra Uterine Growth Retardation): Deviation and reduction in expected fetal growth pattern. Not all IUGR infants are SGA
  8. 8. ASYMMETRIC vs. SYMMETRIC GROWTH RETARDATION - Most growth retarded infants have asymmetric growth restriction. First there is restriction of weight and then length, with a relative “head sparing” effect. - This asymmetric growth is more commonly due to extrinsic influences that affect the fetus later in gestation, such as preeclampsia, chronic hypertension, and uterine anomalies.
  9. 9. - In the human brain, most neurons develop prior to the 18th week of gestation. Early gestational growth retardation would be expected to affect the fetus in a symmetric manner. - Examples of etiologies for symmetric growth retardation include genetic or chromosomal causes, early gestational intrauterine infections (TORCH) and maternal alcohol use.
  10. 10. CAUSES: A. Maternal • Low pre-pregnancy weight • Recent pregnancy and/or high parity • Chronic illness - such as malabsorption, diabetes, renal disease • Inadequate or poorly balanced dietary intake • Decreased O2 availability to fetus (e.g., high altitude, severe maternal anemia)
  11. 11. B. Uterine and placental factors: Inadequate placental growth, uterine malformations, decreased utero-placental blood flow (e.g., toxemias of pregnancy, diabetic vasculopathy) and multiple gestations C. Fetal causes: Include Chromosomal abnormalities and intrauterine infections (i.e., TORCH)
  12. 12. The most important organ we are after for maturity is the lung Assessment of Fetal Lung Maturity: -Fetal lung maturation is marked by production of a detergent-like material, surfactant, which forms a film on the alveolar surfaces.
  13. 13. -Prior to 35th week of gestation, the major component of surfactant is α-palmitic β-myristic lecithin. - After that time, dipalmitic lecithin predominates and phosphatidyl glycerol (PG) appears about a week later. - Minor phospholipid components of surfactant include phosphatidyl inositol, phosphatidyl ethanolamine, phosphatidyl serine, and sphingomyelins.
  14. 14. -Since sphingomyelin (S) concentration in AF is constant during the third trimester, it serves as a reference material against which surfactant lecithin (L) can be compared. - Measurement of L/S ratio avoids problems associated with variability in chemical extraction and inaccuracy in estimates of absolute concentration per AF volume
  15. 15. 1) Quantification of Pulmonary Surfactant: L/S Ratio (Lecithin / sphingomyelin): -It is the most valuable assay for the assessment of fetal pulmonary maturity. -At 32 weeks the L/S ratio reaches 1. Lecithin then rises rapidly, and an L/S ratio of 2.0 is observed at 35 weeks. - A ratio of 2.0 or greater has repeatedly been associated with pulmonary maturity.
  16. 16. -A mature L/S ratio predicted the absence of RDS in 98 percent of neonates. With a ratio of 1.5 to 1.9, approximately 50 percent of infants will develop RDS. Below 1.5, the risk of subsequent RDS increases to 73 percent. Other tests to evaluate surfactant: - Evaluation of Amniotic Fluid Turbidity visually - Shake Test - Foam Stability Index - Tap Test
  17. 17. 2) Test for PG: Amniostat-FLM: -A rapid immunologic semiquantitative agglutination test that can be used to determine the presence of Phosphatidylglycerol (PG) - Results are reported as: Negative Low- positive (PG 0.5-2ug/ml) High -positive (PG ≥2ug/ml) - RDS rarely develops if PG is ≥2ug/ml
  18. 18. 3) Fluorescent Polarization FLM Tests: - The TDx analyzer is an automated fluorescence polarimeter that determines surfactant albumin ratio. - The test requires 1 ml of uncentrifuged amniotic fluid and can be run in less than 1 hour. - The surfactant albumin ratio (SAR) is determined with amniotic fluid albumin used as an internal reference. -A ratio of 50 to 70 mg surfactant per gram of albumin is considered mature. - Correlates with L/S and has a better precision.
  19. 19. 4) Lamellar Body Counts: - Lamellar bodies are the storage form of surfactant. - They scatter light & can be counted directly using the platelet channel of most cell counters - A lamellar body count >30,000/μl uncentrifuged AF is highly predictive of pulmonary maturity, while a count <10,000/μl suggests a risk for RDS. - Neither meconium nor lysed blood has an effect on the lamellar body count.
  20. 20. Assessment of Premature Rupture of Membranes - Pre mature rupture of the membranes is the most common cause for pre maturity. - Diagnosing premature rupture of membranes is done by: 1) PH assessment of vaginal discharge: Unlike vaginal secretions whose pH is acidic, AF is alkaline. The vaginal pool aspirate of a gravida with watery discharge can be tested with nitrazine paper to estimate pH visually.
  21. 21. 2) An aliquot of the aspirated fluid can also be applied to a glass microscope slide, dried for 5 minutes and examined microscopically for a ‘fern' pattern, which indicates the presence of AF in the vaginal fluid pool.
  22. 22. 3) Fetal fibronectin in maternal plasma or AF: - Fetal fibronectin is a chorionic trophoblast protein. - If fetal fibronectin is increased in maternal plasma or AF or cervicovaginal secretions between 22-34 weeks gestation, it denotes loss of integrity of the fetal membranes. - A POC device is available at the obstetrician office for measuring fetal fibronectin. - It has a high predictive value for impending early delivery of the baby & RDS
  23. 23. Sampling in pediatrics
  24. 24. Sampling in pediatrics Pre analytical considerations: - Difficult sampling (Capillary; heal, thumb): needs an expert - Small patient’s size, number of times for blood to be drawn for repeat analysis. - Different sizes of tubes.
  25. 25. Recommended blood draw volumes for pediatric patients
  26. 26. Choice of the analyzer: - Dead volume? The smaller, the better - Clot detector - Sample cubs & primary tubes
  27. 27. Point of care analysis: • • • • • • Portable testing devices that are easy to use Small sample volume No sample preparation is required Provide rapid bed side results Can be connected to the hospital LIS Mainly used for: bilirubin, glucose, Hb, electrolytes & blood gases
  28. 28. Point of care analysis: • High cost, so its use should be rationalized for tests that needs short TAT • Neither their performance nor their dynamic ranges are as good as the tradition lab equipments. • Should be properly validated using appropriate quality assurance procedures • Very low or very high results should be checked in the main laboratory
  29. 29. The most important items that need evaluation in pediatrics are: 1. Regulation of blood gases & pH. 2. Kidney function with regulation of water & electrolytes 3. Liver function: physiologic jaundice & energy metabolism 4. Calcium and bone metabolism 5. Endocrine functions 6. Genetic diseases & neonatal screening
  30. 30. 1) REGULATION OF BLOOD GASES &PH -Requires fully mature lungs & kidneys after birth. - Immature lungs or surfactant may result in respiratory distress syndrome (RDS), where there is failure in excreting CO2 resulting in respiratory acidosis. (Plasma bicarbonate is not affected early, before renal compensation takes place) -
  31. 31. - The trauma & relative anoxia during delivery causes an increase in lactic acid production & accordingly, metabolic acidosis. (Reduced bicarbonate level, supplying bicarbonate reverses the condition) -Persistent acidosis that is not corrected by bicarbonate administration is an indication for possible inborn errors of metabolism.
  32. 32. -Alkalosis is uncommon in pediatric patients, most common causes: 1) Hyperammonemia: Secondary to diseases & inborn errors of metabolism 2) Pyloric stenosis & loss of gastric acids 3) Hypokalemia liver
  33. 33. 2) KIDNEY FUNCTION - From the 35th week of gestation, rapid kidney development takes place in preparation for extra uterine life GFR: - At birth, GFR is about 25 % of its value in older children.
  34. 34. - Tubular function shows a similar pattern, where the concentrating power of the kidney in the early months of life is only about 78 % of its value in the adult kidney. - This gradual process of renal development results in serum electrolytes level shift seen in the neonatal period.
  35. 35. - The kidney primarily maintains water homeostasis - Other causes of water loss in the neonatal period: 1) Insensible water loss through the skin 2) The use of radiant heaters (radiated heat) to maintain body temperature. 3) Insensible water loss from the lungs in RDS
  36. 36. Disorders affecting electrolytes and water balance
  37. 37. 3) Liver function Physiologic Jaundice: - The processing of many normal metabolic pathways and the metabolism of exogenous compounds proceed slower in neonates. - The most striking effect of an immature liver, even in a full term baby, is the failure to adequately metabolize bilirubin.
  38. 38. - Fetal blood is produced first by the embryonic yolk sac, then by the liver, and finally by the fetal bone marrow. - With the switch of erythropoiesis to the fetal liver, fetal hemoglobin production begins. - HbF consists of two α- and two γ-chains. - As the fetal bone marrow begins red cell production, HbA production increases. - At birth fetal blood contains 75% HbF and 25% HbA. HbF production rapidly diminishes during the first year of postnatal life. In normal adults, less than 1% of hemoglobin is HbF.
  39. 39. -HbF has a higher affinity for oxygen than does HbA. Thus in the placenta, oxygen is released from the maternal HbA, diffuses into the chorionic villi, and binds to the fetal HbF. -Bilirubin accumulates as fetal hemoglobin is rapidly destroyed and replaced by adult hemoglobin. - At birth, UDP - glucuronoyltransferase, the enzyme responsible for conjugating bilirubin, is immature. This results in the accumulation of unconjugated bilirubin and the development of physiologic jaundice.
  40. 40. -A normal baby may have a serum bilirubin up to 15 mg / dl, most of it unconjugated. This level should fall to the base line by the age of 10 days. -If severe and passes the immature BBB, it might lead to kernicterus (Bilirubin encephalopathy). - Complete absence of the bilirubin conjugating enzyme results in severe persistent jaundice (Crigler – Najjar syndrome)
  41. 41. Carbohydrate metabolism: -At birth, a full term baby has sufficient glycogen stores to provide glucose as an energy source. -If the delivery is stressful, these energy reserves may become depleted prematurely. - At that time, the normal physiologic rule of the gluconeogenesis pathway becomes essential where there is conversion of alanine into glucose.
  42. 42. -The later pathway is not always mature at birth which may result in what is termed “physiologic hypoglycemia”. - This condition usually corrects quickly as the enzyme system matures.
  43. 43. -Persistent and severe hypoglycemia should alert the physician towards a possible inborn error of metabolism, such a galactosemia. - Galactosemia is due to failure of conversion of galactose to glucose as a result of genetic deficiency in any of the following enzymes: galactose-1-phosphate uridyltransferase (GALT), galactokinase (GALK) or uridine diphosphate galactose-4-epimerase (GALE)
  44. 44. Nitrogen metabolism: -The liver is involved in the metabolic inter conversions of amino acids and in the synthesis of non-essential amino acids. - The liver synthesis most of the plasma proteins including albumin, transferrin, complements and coagulation factors.
  45. 45. -Important in nitrogen metabolism through the Urea cycle. -Owing to the immaturity of urea cycle enzymes early in life, the level of ammonia in the plasma of neonates is highly elevated than its level in a one year old child. - Persistently elevated ammonia level, should alert the investigator to possible liver damage.
  46. 46. 4) CALCIUM AND BONE METABOLISM - Normal bone growth requires integration of calcium, phosphate and magnesium metabolism with endocrine regulation from vitamin D, parathyroid hormone & calcitonin. -The active metabolite of vitamin D is 1,25 dihydroxy vitamin D. -Hydroxylation of Vitamin D from diet takes place in liver and kidneys and requires normal functioning of these organs.
  47. 47. - Absorption of vitamin D from the gastrointestinal tract, conversion to its active form in the kidney, and incorporation of calcium and phosphate into the growing bone requires normally active PTH. - Secretion of PTH is in turn modulated by serum calcium & magnesium levels, where low level of both divalent cations inhibits PTH secretion.
  48. 48. - Rapid bone growth occurring in infancy and puberty requires optimal coordination of mineral absorption, transport and endocrinecontrolled incorporation of the minerals into growing bone. - Approximately 98% of total body calcium is present in bone and less than 2 % is measurable in blood.
  49. 49. Hypocalcemia: - Hypocalcemia is defined as total serum calcium below 7.0 mg / dl or ionized calcium below 3.0 mg / dl. - In the newborn, particularly immature, these levels may be commonly encountered with few symptoms. However, hypocalcemia can result in irritability, twitching and seizers. Serum calcium is usually measured in children with seizers of unknown etiology. - Prolonged hypocalcemia can result in reduced bone growth and rickets.
  50. 50. Causes of hypocalcemia: • Prematurity • Metabolic acidosis • Vitamin D deficiency • Liver diseases • Kidney diseases • Hypoparathyroidism • Low calcium intake • High phosphorus intake • Hypomagnesemia (Inhibits PTH secretion)
  51. 51. Rickets: - Disease caused by a mineralization defect during bone formation resulting in increase in osteoid, the unmineralized organic matrix Of bone.
  52. 52. Causes: 1. Vitamin D deficiency: inadequate exposure to sun, poor vitamin D diet, malabsorption 2. End organ resistance to vitamin D 3. Phosphate depletion: e.g. Fanconi syndrome Biochemical findings: 1. Low serum calcium & phosphates 2. High serum ALP (Due to increased osteoblastic activity) 3. 25 (OH) D: to assess vitamin D status.
  53. 53. N.B: - Vitamin D-dependent rickets type I is an inherited defect in 25(OH) D-1α hydroxylase causing impaired formation of 1, 25(OH) vitamin D. - Vitamin D-dependent rickets type II is an inherited disorder characterized by very high serum concentration of 1, 25(OH) vitamin D. This syndrome is due to resistance to 1, 25(OH) vitamin D, secondary to defects in the 1,25(OH) vitamin D receptor.
  54. 54. Hypercalcemia: - Defined as total serum calcium > 11.0 mg / dl. - This is unusual in pediatrics, but has potentially severe clinical implications. - Patients with hypercalcemia have poor muscle tone, constipation, failure to thrive and may develop kidney stones leading to renal failure.
  55. 55. 5) ENDOCRINE FUNCTION Hypothalamic – pituitary – thyroid axis: Primary hypothyroidism (Congenital hypothyroidism): - Results from any defect that causes failure of the thyroid gland to synthesize and secrete thyroid hormones. - Incidence: 1/4000 births - Untreated patients with this condition have severe mental retardation with unusual facial features.
  56. 56. - Treatment by thyroid replacement therapy is usually successful when diagnosis is established. - The best diagnostic test is to measure serum TSH level which is high as a result of failure of the long feedback loop (between pituitary & thyroid gland). Thyroid hormone levels in untreated patients are very low. - The only neonatal screening program in Egypt.
  57. 57. Secondary hypothyroidism: - It is a result of pituitary failure to secrete TSH which results in lack of thyroid gland stimulation and subsequent production of thyroid hormones. - Diagnosed by low TSH. - It is important to study the other pituitary pathways to determine whether this is an isolated TSH defect or panhypopituitarism.
  58. 58. Neonatal Graves‘ disease: -The fetal thyroid-pituitary axis functions independently from the mother's axis in most cases. -However, if the mother has preexisting Graves' disease, her auto antibodies can cross the placenta and stimulate the fetal thyroid gland. Thus the fetus can develop hyperthyroidism. - Measurement of thyrotropin-binding inhibitory immunoglobulins is useful for assessing risk of fetal or neonatal Graves' disease.
  59. 59. Hypothalamic – pituitary – adrenal cortex axis: - The most important disorder is congenital adrenal hyperplasia (CAH) - CAH: Congenital absence of one or more of the synthetic enzymes that lead to cortisol and aldosterone biosynthesis. This leads to compensatory increase in ACTH leading to stimulation of steroids biosynthesis till the block point causing:
  60. 60. 1. Hyperplasia of the adrenal cortex 2. Accumulation of intermediate compounds proximal to the block 3. Shunting of the substrate towards the adrenal androgen pathway - The most common disorder is 21-Hydroxylase deficiency (1 / 5000 births)
  61. 61. Growth factors: - G.H deficiency results in poor growth and stunted growth. - When G.H acts on its receptors on the liver, the liver secretes IGF-1 & its binding protein IGF-BP3 - Due to the diurnal and pulsatile pattern of G.H secretion, a single measurement is not sufficient to diagnose its deficiency.
  62. 62. - Insulin induced hypoglycemia & clonidine induced hypotension are common stimulatory tests used to detect G.H deficiency. - Measuring IGF-1 & IGF-BP3 is considered an effective tool for assessing G.H deficiency because: 1. Their basal levels do not have the great variation that occurs for G.H 2. Infants with defects in IGF-1 & IGF-BP3 synthesis and secretion are unlikely to respond to G.H replacement.
  63. 63. Screening for diseases in pediatrics
  64. 64. Screening for diseases in pediatrics: I) Pre natal: - Maternal screening - Fetal screening: amniocentesis, chorionic villi sampling & pre implantation genetic diagnosis II) Post natal
  65. 65. MATERNAL SERUM SCREENING FOR FETAL DEFECTS: Multiple of the median (MoM): - The MoM is now universally used as a common currency for converting analyte values into an interpretative unit and is also the starting point for calculating risks for neural tube defects, Down syndrome, and trisomy 18.
  66. 66. - For each analyte, each lab should develop a set of median values for each week (or day) of gestation using the laboratory’s own assay values measured on the population to be screened. - Individual test results are then expressed as MoM by dividing each individual test result by the median for the relevant gestational week.
  67. 67. Screening for Down syndrome (trisomy 21) & trisomy 18: First trimester screening (Double test) – [11th- 14th gestational weeks] - Combining two serum markers (free beta HCG, PAPP-A), with ultrasound nuchal translucency (NT) - PAPP-A is lower in pregnancies complicated with Down syndrome than in normal ones. - HCG is higher & NT is thicker in Down syndrome pregnancies.
  68. 68. Second trimester screening (Triple test)[15th – 18th gestational weeks] A method combining measurements of 3 analytes, with maternal age into a single risk estimate. • AFP (alpha fetoprotein ) • uE3 (unconjugated estriol ) • HCG (Human chorionic gonadotrophin)
  69. 69. In Down syndrome: - AFP & uE3 are 25% lower than expected - HCG is two folds higher than expected - A 4th analyte, Dimeric Inhibin A (DIA), that is elevated in cases of Down syndrome is added and the test is called Quadruple test
  70. 70. Fetal trisomy 18: - AFP and uE3 concentrations are low - HCG concentrations are also very low - No role for DIA Women who test positive (or those who test negative but aged above 35) should be offered amniocentesis to obtain fetal cells for karyotyping or other DNA based techniques; the only way to confirm Down or trisomy 18 syndromes diagnosis.
  71. 71. Screening for Neural tube Defects (NTD): - Optimal screening is between 16 and 18 weeks of gestation. - The most commonly used AFP MoM cutoffs are between 2.0 and 2.5 MoM. - A second sample is needed for moderately elevated results (2.0 to 3.0 MoM) - If the result for the second AFP test is not elevated, the woman is considered to be screen-negative.
  72. 72. - If the result is still elevated: • 4-D U/S is used to verify gestational age • Identify other possible reasons for the increased AFP (ex: multiple pregnancy, abdominal hernias into the umbilical cord) - Patients still having an unexplained ↑↑ AFP test results amniocentesis for measurement of amniotic fluid AFP and acetylcholinesterase.
  73. 73. - Ultrasound diagnosis of open neural tube defects is now so reliable that it is often used for diagnosis in women with elevated maternal serum AFP without waiting for amniotic fluid measurements.
  74. 74. FETAL SCREENING: I) Amniocentesis: - Removal of amniotic fluid containing fetal cells, via a needle puncture from the uterus. - Any genetic analysis can be performed on these cells. - Performed 14-20th week of gestation.
  75. 75. FETAL SCREENING: I) Amniocentesis: - Removal of amniotic fluid containing fetal cells, via a needle puncture from the uterus. - Any genetic analysis can be performed on these cells. - Performed 14-20th week of gestation.
  76. 76. Amniotic fluid testing is done for: • Diagnosis of NTD (Confirming a screen positive mother) • Diagnosis of congenital diseases • Diagnosis of Isoimmunisation disease • Assessment of fetal lung maturity
  77. 77. 1) Diagnosis of NTD: AFP: - Patients with unexplained high maternal serum AFP levels and normal ultrasonography should be offered amniotic fluid testing - AFP values greater than or equal to 2.0 MoM are considered elevated. - A frequent interference is contamination of the fluid with fetal blood. Elevated amniotic fluid AFP should be tested for fetal hemoglobin, a sensitive marker of fetal blood contamination.
  78. 78. Acetylcholine Estrase (AChE): - AChE is a neural enzyme present in cerebrospinal fluid and fetal blood. - It is not present in maternal blood and is not normally detectable in amniotic fluid. - The abnormal presence of acetylcholinesterase in amniotic fluid is suggestive of an open fetal defect. - When AChE is detected, the ratio of AChE to pseudocholinesterase (PChE), a non-specific cholinesterase normally found in amniotic fluid, may help distinguish open neural tube defects from fetal blood contaminated fluid.
  79. 79. 2) Tests for isoimmunization disease: - Isoimmunization disease is a fetal haemolytic disorder caused by maternal antibodies directed against antigens on fetal erythrocytes. - The amount of bilirubin in the amniotic fluid is useful for determining the severity of the condition. - The most common cause of severe disease is sensitization of Rh-negative woman to the D antigen of the Rh system.
  80. 80. - An association exists between gestational age, severity of the disease, and bilirubin concentration. - The concentration of bilirubin is too low to be measured by standard photometric techniques (up to 0.03mgldL) but the determination can be done by absorption spectrophotometry. - The maximal absorbance of bilirubin is at 450 nm.
  81. 81. - In the absence of significant amounts of bilirubin, the absorbance spectrum for the amniotic fluid between 365 and 550 nm is nearly exponential.
  82. 82. -When plotted on a semi log scale (linear curve), the degree to which the curve deviates from a straight line at 450 nm is linearly proportional to the concentration of bilirubin (∆A 450)
  83. 83. - Results of (∆A 450) are established into 3 classification zones based on gestational age “ Liley's zones”
  84. 84. II) CVS: - Chorionic villi are precursors of the placenta and a good source of fetal tissue. - CVS can be performed safely by 10th week both transabdominally and transvaginally - Used for: •Cell culture & karyotyping •Enzyme assays •Direct gene analysis
  85. 85. III) Pre implantation genetic diagnosis (PGD): - For couples undergoing IVF. - Fertilized eggs are checked for the gene mutation and only the unaffected embryos are introduced into the uterus in the hope of a successful implantation. - Cells are isolated from the blastocyst (16 – 20 cells embryo), where DNA is extracted and tested for common or expected genetic mutations.
  86. 86. POST NATAL NEW BORN SCREENING: - NBS is a process of early identification of health conditions followed by their subsequent timely treatment before the onset of disease processes thereby minimizing the risk of long-term sequelae. - Key issues considered for NBS: • What are the effects of each genetic disorder? • What treatment is currently available for each genetic disorder, and at what age does treatment begin?
  87. 87. • Based on demographics, how many people are likely to be affected by each genetic disorder? • Should all newborn infants receive the same screening tests? Why or why not? • Can a test in question be performed on a large scale in laboratories? Why or why not? • What is the cost per test and the total for screening? • Will early identification of persons with the genetic disorder lead to cost savings in treatment or care? Why or why not?
  88. 88. Alpha-1-antitrypsin Deficiency: Clinical description: - Alpha-1-antitrypsin deficiency can lead to early onset of emphysema and/or liver failure. - These symptoms usually appear when a person is in their 30’s or 40’s. - Symptoms are more severe in smokers than in nonsmokers.
  89. 89. Genetics: - This disorder is caused by a mutation in the proteinase inhibitor (PI) gene on chromosome 14. - The normal protein coded for by this gene is involved in tissue repair. - Disorder symptoms depend on which type of mutation an individual has in the PI gene. - There are more than 70 different alleles of the PI gene. - The M allele is the wild variant. The mutant alleles S and Z are the most common disease causing variants.
  90. 90. Inheritance: Autosomal recessive Testing: - This disorder can be detected by testing the levels of alpha-1-antitrypsin in blood. If they are abnormally low, the next step is to identify the exact alpha-1-antitrypsin protein variants the person carries. - Abnormal forms of the alpha-1-antitrypsin protein can be detected using dried blood as a sample for gel electrophoresis.
  91. 91. Cystic Fibrosis: - Cystic fibrosis is caused by mutations in the cystic fibrosis transmembrane regulator (CFTR) gene on chromosome 7 which codes for the protein that controls ion transfer across cell membranes. - Disruption of salt transfer results in abnormal gland secretions and dehydration due to increased loss of salt and water during sweating. - CF affects almost all of the glands in the body that secrete fluid, resulting in a variety of symptoms.
  92. 92. - Secretions may be thick and cause blockage in the pancreas, intestines and lungs. - Mucus blockage also provides places for bacteria to multiply, increasing the probability of infection. - CF children show poor digestion, dehydration, coughing and vomiting. Molecular analysis has identified approximately 100 mutations in the CFTR gene. Different mutations determine the severity of symptoms seen in CF patients.
  93. 93. Testing: - Mutation in the CFTR gene results in an increase in an enzyme called trypsinogen. The initial newborn screen tests for this enzyme using a dried blood sample. - There are hundreds of mutations in the population, the most common is ∆F508. - Testing all mutations is very difficult unless a clinically accepted microchip technology is available.
  94. 94. - Measurement of chloride content in sweat collected after pilocarpine iontophoresis, is still the gold standard test for diagnosing CF Cystic Fibrosis Foundation Sweat Test2.flv
  95. 95. Huntington’s disease: - Huntington’s disease is characterized by the progressive death of certain neurons in the brain. - Symptoms generally appear between the ages of 3540 years and include depression, mood swings, amnesia, involuntary twitching and lack of coordination. - As the disease progresses, involuntary movements increase, memory declines, and walking, speaking and swallowing ability gradually diminish. - Death soon follows from choking, infections or heart failure.
  96. 96. Genetics: - Huntington’s is caused by excessive repeating of the DNA bases CAG (trinucleotide repeats) in the huntingtin gene on chromosome 4. - The normal number of repeats is 10 – 35, Huntington’s disease patients have 36 - 121 repeats. Inheritance Autosomal dominant Testing The huntingtin gene is analyzed in a blood sample to determine the number of CAG repeats.
  97. 97. Maple Syrup Urine Disease (MSUD) - Individuals with maple syrup urine disease (MSUD) are unable to properly metabolize three amino acids: leucine, isoleucine and valine. - The enzymes required to process these three amino acids are absent, inactive or only partially active. - Because these amino acids do not get broken down completely, high levels accumulate in the blood, urine and sweat.
  98. 98. -The by-product of isoleucine has a characteristic sweet smell which gives the disorder its name. - The three amino acids and their derivatives can be toxic at high levels and can lead to brain injury, mental retardation, seizures, vomiting, coma and even death. Genetics: The most common type is classic MSUD which is caused by a defect in the BCKDHA gene on chromosome 19.
  99. 99. Inheritance Autosomal recessive Testing - Newborn screening programs that test for MSUD use the same blood sample collected for PKU and galactosemia tests. - Generally, blood is analyzed for elevated levels of leucin
  100. 100. Phenylketonuria (PKU) - PKU is caused by the lack of phenylalanine hydroxylase , an enzyme that processes the amino acid phenylalanine. - Phenylalanine is not broken down and accumulates in the blood & it is toxic to the brain. - Untreated individuals with PKU show progressive developmental delay in the first year of life, mental retardation, seizures, autistic-like behavior and a peculiar body odor.
  101. 101. Genetics - In PKU individuals, the phenylalanine hydroxylase gene on chromosome 12 is disrupted. Inheritance Autosomal recessive
  102. 102. Testing - The blood phenylalanine level can be measured using a spot of dried blood. - The PKU test (the Guthrie test) was the first genetic screening test developed. - Automated tests (MS / MS) are now used in some screening programs. - The timing of the test is important; the test should be completed after the first day and before the seventh day of life. If done too soon, low levels in the newborn can be masked by the presence of maternal phenylalanine.
  103. 103. Sickle Cell Disease: - A group of inherited disorders of RBCs - If the gene encoding hemoglobin is mutated, it causes a change in the shape of the molecule. - When the mutated hemoglobin delivers oxygen to the tissues, the red blood cell collapses, resulting in a long, flat sickle-shaped cell. These cells clog blood flow, resulting in a variety of symptoms including pain, increased infections, lung blockage, kidney damage, delayed growth and anemia
  104. 104. Genetics - The gene encoding the beta chain of the hemoglobin molecule, located on chromosome 11, can be mutated in a variety of ways that result in different types of sickle cell disease. - Some mutations are more common than others. The three most common types of sickle cell disease are hemoglobin SS (Hb SS), hemoglobin SC (Hb SC), and hemoglobin sickle beta thalassemia (HbS beta-thalassemia).
  105. 105. Inheritance Autosomal recessive Testing: - Most screening programs utilize thin-layer isoelectric focusing (IEF) or high performance liquid chromatography (HPLC) techniques performed on capillary blood collected from a heel stick and absorbed onto filter paper.
  106. 106. Newborn Screening Card Rejection.flv
  107. 107. THANK YOU

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