HEMOGLOBINOPATHIES Flashcards
Disorders characterized by impaired synthesis of HEME
Porphyrias
Lead poisoning (plumbism/saturnism) causes
Lead inhibits enzymes, including ferrochelatase and pyrimidine-5’-nucleotidase
Causes basophilic stippling in reticulocytes (RNA remnants)
Lead inhibition of pyrimidine-5’-nucleotidase
Enzyme needed to completely remove RNA remnants from reticulocytes
Pyrimidine-5’-nucleotidase
Qualitative globin defects due to differences in amino acid arrangement in the polypeptide chain
Hemoglobinopathies
Major groups of hemoglobinopathies
Alpha-hemoglobinopathies, Beta-hemoglobinopathies, Gamma-hemoglobinopathies, Delta-hemoglobinopathies
Homozygous B-hemoglobinopathies (mutated B genes)
Hb A1 absent, abnormal hemoglobin predominant
Homozygous sickle cell disease genotype
Hb SS
Homozygous Hb C disease genotype
Hb CC
Heterozygous B-hemoglobinopathies (one normal, one mutated B gene)
Hb A1 predominant, abnormal Hb present
Heterozygous sickle cell trait genotype
Hb AS
Heterozygous Hb C trait genotype
Hb AC
Porphyrias affect
Heme (mitochondria, ferrous protophorphyrin IX)
Hemoglobinopathies affect
Globin (molecules and amino acids), hemoglobinopathies, thalassemias
Abnormal hemoglobins in β-hemoglobinopathies
Hb S, Hb C, Hb E
Hb S amino acid change
Glutamine to valine at position 6
Hb C amino acid change
Glutamine to lysine at position 6
Hb E amino acid change
Glutamine to lysine at position 26
M hemoglobins associated with methemoglobinemia, cyanosis, and chocolate brown color
Hb M-Saskatoon, Hb M-Milwaukee-1, Hb M-Milwaukee-2
Increased O2 affinity hemoglobins
Hb Hiroshima, Hb Rainier, Hb Bethesda
Decreased O2 affinity hemoglobins
Hb Agenogi, Hb Beth Israel, Hb Yoshizuka
Hemoglobin S mutation
Glutamic acid at position 6 replaced by valine
Sickle cell anemia vs. sickle cell trait
80-100% Hb S causes easy sickling, 20-40% Hb S requires lower O2 for sickling
Sickling mechanism
Conformational change and polymerization of Hb S upon oxygen release, forming tactoids or crystals
Effect of sickling on RBCs
Cells become rigid, obstruct blood flow, cause tissue death, organ infarction, and pain
Screening test for hemoglobin S
Sodium metabisulfite method, Sodium dithionite tube test (Solubility test), Hemoglobin Electrophoresis (Cellulose acetate)
Principle of sodium metabisulfite method
Reducing agent deoxygenates Hb, causing sickling in Hb S RBCs
Positive result in sodium metabisulfite method
Presence of sickle cells or ‘holly-leaf’ form (sickle cell trait)
Limitation of sodium metabisulfite method
Cannot differentiate sickle cell trait from sickle cell anemia
Confirmation test for hemoglobin S
Citrate agar
Negative result in sodium metabisulfite method
Normal or slightly crenated RBCs
Principle of sodium dithionite tube test
RBCs added to sodium dithionite and saponin; Hb S forms liquid crystals, producing a turbid appearance
Positive result in sodium dithionite tube test
Turbid solution, black lines on the reader scale not visible
Negative result in sodium dithionite tube test
Clear solution, black lines on the reader scale visible
Confirmatory test after positive sodium dithionite test
Hb electrophoresis
Test to confirm presence of Hb S
Positive sodium dithionite tube test (Solubility test)
Test for hemoglobin variants
Hemoglobin Electrophoresis (Cellulose Acetate)
Buffer used in hemoglobin electrophoresis
Alkaline buffer (pH 8.4 to 8.6)
Charge of hemoglobin in electrophoresis
Negatively charged molecule
Direction of hemoglobin migration in electrophoresis
Toward the anode (+) due to negative charge
How hemoglobin variants are detected
Difference in mobility based on net charge reveals variants
Primary screening procedure for abnormal hemoglobin
Hemoglobin Electrophoresis (Cellulose Acetate)
Abnormality in Hemoglobin C
Glutamic acid replaced by lysine at position 6 of beta chain
Crystal associated with Hb SC
Washington monument shape, protruding from RBC membrane
Crystal associated with Hb CC
Bar of Gold shape, within the RBC membrane
