week 5: morphology and Hbopathies

Question 1

• What are Heinz bodies?

Answer 1

o Formed by damage to hemoglobin through oxidative stress leading to irreversible hemichrome oxidation and precipitation.
o Purple-blue inclusions visible only after supravital stain
o Seen in: Alpha thalassemia, Congenital hemolytic anemia, G6PD deficiency

Question 2

• What is basophilic stippling?

Answer 2

o Indicates toxic injury to the bone marrow. May also appear with severe anemia, such as megaloblastic anemia
o Variable size of granules distributed throughout cell
o Seen with Lead poisoning, Myelofibrosis

Question 3

• What is the fate of heme after hemolysis?

Answer 3

o Excess Hb dimers in blood= hemoglobinemia
o Excess dimers in kidney -> hemoglobinuria
o Hb complex turned to bilirubin in M0 or liver -> biliary system

Question 4

• What is Erythrocyte Sedimentation rate (ESR)?

Answer 4

o Rate at which RBCs settle in an hour
o Commonly performed, simple and inexpensive lab test to assess overall inflammation.
o Used to track the progress of a disease or to monitor treatment. “sickness index”
o Not used diagnostically
o Results directly correlate with the amount of fibrinogen in anticoagulated blood.

Question 5

• What causes the ESR to increase?

Answer 5

o The normal negative cell surface charge of RBCs, “zeta potential”, makes RBCs repel each other
o Inflammatory proteins (acute phase reactants), mainly fibrinogen, neutralize the normal zeta
o RBCs no longer repel, but stack on each other= Rouleaux formation
o Larger mass of cell settles faster

Question 6

• What is ESR used for?

Answer 6

o Follow the course of a disease – in general, as the disease worsens the ESR increases and as the disease resolves the ESR decreases
o Can be used to monitor therapy – especially for inflammatory autoimmune diseases

Question 7

• What are the limitations to ESR?

Answer 7

o Nonspecific, nonsensitive
o May NOT be elevated in active disease
o Many factors can alter the test results

Question 8

• What factors can interfere with ESR?

Answer 8

o Low results if test not set up within 3 hours of sample collection
o Pregnancy (2nd & 3rd tri) may elevate
o Menstruation may elevate
o Sedimentation tube must be vertical
o Some anemia’s falsely increase – correction nomograms are available
o Polycythemia & sickle cell disease decreases
o Protein-producing malignancies falsely increase value
o Any bubble in column invalidates the test

Question 9

• What are some dzs that can increase ESR?

Answer 9

o Chronic renal failure
o Malignant diseases
o Bacterial infections
o Inflammatory diseases / Autoimmune Disease
o Necrotic diseases, MI
o Diseases assoc. with increased plasma proteins
o Assists in the diagnosis for: polymyalgia rheumatica and temporal arteritis (may be > 100 mm/hr)

Question 10

• What szs can falsely decrease the ESR?

Answer 10

o Sickle cell disease – cells severely distorted & cannot form rouleaux
o Hereditary spherocytosis – cells are too spherical, do not stack in rouleaux
o Hypofibrinogenemia – decreased protein inhibits aggregation & settling of RBCs
o Polycythemia vera – increased RBC counts inhibit settling due to simple concentration interference

Question 11

• What is the structure of hemoglobin?

Answer 11

o 4 polypeptides (globins), 2 alpha, 2 beta
o Each globin has a heme (porphyrin (red pigement) with Fe=metallo-porphyrin), which has O2 attracting properties
o Oxy-Hb or deoxyHb

Question 12

• What are the normal variants of Hb?

Answer 12

o Hb A: 97% of total cells Hb; 2a and 2B
o HbA2: <3%; 2a and 2d
o HbF: 1%; 2a and 2g; predominant during fetal development; 50-80% at birth, 8% at 6 months

Question 13

• What are hemoglobinopathies?

Answer 13

o Inherited defects in globin structure mostly involving a single amino acid substitution

o family of genetic disorders caused by:
o 1- Production of a structurally abnormal hemoglobin molecule (Qualitative hemoglobinopathies)
o Or: 2- Synthesis of insufficient quantities of normal hemoglobin (Quantitative hemoglobinopathies)
o Or: 3- both (rare).

Question 14

• What is Thalassemia?

Answer 14

o A genetic blood disorder where the body makes reduced amounts of globin chains.
o 2 main types: affects either alpha or beta chains (ie Alpha Thalassemia)
o In major or minor forms: major=homozygous, minor=heterozygous

Question 15

• What is beta thalassemia major?

Answer 15

o Children (cooley’s anemia): anemia in first year, facial bone deformities, fatigue, jaundice, growth failure, SOB
o HbA1 5-20%, HbA2 2-3%, HbF 65-100%
o Poikylocytic cells: elliptocytes, schistocytes, target cells, tear drop, spherocytes, hypochromic

Question 16

• What is thalassemia minor?

Answer 16

o Beta thalassemia minor is most common form (one beta chain is affected)
o Results in mild microcytic hypochromic anemia otherwise no clinical symptoms

Question 17

• What are the abnormal, clinically significant Hb variations?

Answer 17

o Hb S: Predominant in people with sickle cell disease. Disease exists on beta chain.
o Hb C: Disease is relatively benign with mild hemolytic anemia and splenomegaly
o Hb E: Mild hemolytic anemia and splenomegaly. Extremely common in SE Asia
o Hb Constant Spring: Alpha chain is abnormally long resulting in a thalassemic phenotype.
o Hb H: Composed of 4 beta chains. Happens in extreme limitation of alpha chains.
o Hb Barts: No alpha chains are produced. Most individuals die in utero

Question 18

• How can you see sickle cell Hb on electrophoresis?

Answer 18

o	HbA (normal) is more negatively charged than HbS, so moves farther
o	Val in sickle cell is neutral, but Glu is normal is negative

Question 19

• Why is beta thalassemia clinically more severe?

Answer 19

o A chains dissociate into monomers more readily than B chins, so B chains form hemichromes at a faster rate

Question 20

• What is sickle cell anemia? Hb? Incidence? Signs?

Answer 20

o Genetic trait causing hemoglobin irregularities
o Glutamic acid is substituted for valine; Allowing the polymerization of sickle hemoglobin when deoxygenated
o S 80-100%; A1 0%; A2 2-3%; F 2%
o Incidence is 6 months of age; Anemia is moderate to severe with slight jaundice; Significant number of patients die before age 40
o Sickle crisis may produce small infarcts in various organs, abdominal & bone pain most common

Question 21

• What is the difference between normal and sickle red cells?

Answer 21

o Normal: disc-shaped, deformable, life span 120 day

o Sickle: sickle-shaped, rigid, lives 20 days or less

Question 22

• What is the significance of hemolysis and vaso-occlusion in sickle cell anemia?

Answer 22

o Hemolysis: red cell destruction causes anemia, varying degrees between pts; bone marrow dramatically increases production of RBCs, but can’t keep up with destruction
o Vaso-occlusion: sickle cells block small vessels, can produce tissue hypoxia; result is pain or damage to organs

Question 23

• What is sickle cell trait?

Answer 23

o Heterozugous, SA
o Hb distribution: S 20-40%; A1 60-80%; A2 2-3%; F2%
o Incidence is about 8% of African Americans
o No anemia or clinical evidence of disease
o May develop splenic infarcts under hypoxic conditions
o Some persons develop hematuria

Question 24

• What is Hemoglobin C disease?

Answer 24

o Autosomal recessive disorder
o Similar to Hb S but glutamic acid to lysine substitution
o Considered a benign hemoglobinopathy
o Causes mild hemolytic anemia
o Unstable hemoglobin precipitates in RBCs and forms crystals
o Spleen removes defective RBCs from circulation
o Decreases the risk of malaria

Question 25

• Hb distribution in HbC dz? Incidence? Sxs?

Answer 25

o C 90-100%; A1 0%; A2 2-3%; F 2%
o Incidence is about 3% of African Americans from northern Africa
o Mild to moderate reduction in RBC lifespan
o Abdominal & bone pain occur, less severe than sickle disease
o Large number of target cells on blood smear (30-90%)

Question 26

• What is the malaria hypothesis?

Answer 26

o Malaria is found in areas where there are high carrier rates of hemoglobinopathies
o Being a carrier protects you from Malaria so you have an evolutionary “advantage” over a non-carrier—Don’t know why
o Hb S – reduces oxygen tension in cells retarding growth of the parasite
o A thalassemia – confers susceptibility to a milder form of malaria; once contracted, the patient has immunity to the more severe form and increased survival
o B-thalassemia - ???; possibly a role in decreasing cell adhesions in the deadly cerebral form of malaria

Question 27

• How is heme synthesized?

Answer 27

o 85% of total heme synthesis occurs in red blood cells (RBC); Ceases when RBC’s mature
o Heme stimulates protein synthesis in reticulocytes
o The liver is the main non-RBC source of heme synthesis; used mainly for the synthesis of the cytochrome P450 class of enzymes that are involved in detoxification
o 2 succinyl CoA + glycine –(ALA synthase in mito IMS)-> –(porphobilinogen synthase in outer membrane)->cytosol->uroporphyrinogen III -> coporphyrinogen III ->IMS->protoporphyrinogen IX –(ferrochelatase)->heme

Question 28

• What are porphyrias?

Answer 28

o A group of disorders caused by deficiencies of enzymes of the heme biosynthetic pathway.
o Two cardinal symptoms in patients with porphyria are photosensitivity and neurologic disturbances
o Acute: Affect the nervous system. Ssx come on quickly and last a short time. Sx: abdominal pain, vomiting, constipation, diarrhea. Sometimes seizures, anxiety and hallucinations can occur; may be life threatening.
o Cutaneous: affect the skin. Once exposed to sun, skin becomes fragile and blistered, leading to infection, changes in skin pigmentation and hirsutism

Question 29

• How are porphyrias defined by where the damaging compounds originate?

Answer 29

o Erythropoietic (bone marrow): Low number of RBCs , enlargement of the spleen
o Hepatic (liver): Abnormal liver function, increased risk of developing liver cancer
o Mixed: Both hepatic and erythropoitic
o Acquired: No genetic component, drug induced or due to lead poisoning

Question 30

• What is an example of an acquired porphyria?

Answer 30

o Lead poisoning: Pb inhibits bone marrow delta-aminolevulinic acid dehydratase (ALAD), a key enzyme in heme synthesis.
o Then the highly reactive oxyradical Delta-aminolevulinic acid (ALA) heme precursor accumulates, and is reported to cause liver cancer.
o Basophilic stippling in RBC’s

Question 31

• What lab testing is done for porphyrias?

Answer 31

o from blood, random or 24 hour urine, and/or a stool sample
o If acute or neurologic porphyria is suspected, a sample should be taken during an acute attack.
o Testing measures porphorins and their toxic precursors that have built up in tissue and fluids.
o Specialty labs may also measure for involved enzymes or genetic testing

Question 32

• What labs would you do for different porphyria sxs?

Answer 32

o Acute neuroviscerial: urine PBG and ALA; if increased, further test to find which P
o Sun-induced: erythrocyte protoporphyrin conc; increased is EPP
o Skin lesions: plasma or urine porphyrin profile; increased, further differentiate

Question 33

• What is the relationship between Hb, HCT, and RBC indices?

Answer 33

o All are interrelated RBC parameters
o In general, decreases in RBC leads to decreases in Hemoglobin & Hematocrit
o The relationship between these 3 parameters allows calculation of the RBC indices

Question 34

• How is Hb measured?

Answer 34

o Most common method used to measure total hemoglobin content is spectrophotometric analysis. Based on the premise that substances absorb light energy at different wavelengths.
o Spectrophotometry: RBCs lysed, release Hb; blue/green wavelengths shone through; light not absorbed is analyzed (Beer’s law) to determine Hb content

Question 35

• What are normal Hb levels? Interfering factors?

Answer 35

o Males: 14.0-17.4 g/dL
o Females: 12.0-16.0 g/dL
o Children: 11.0-16.0 g/dl
o Very high WBC counts; Severe lipidemia; RBC abnormalities; Increased turbidity; Increased bilirubin

Question 36

• What can cause decreased Hb levels?

Answer 36

o Decreased RBCs: bone marrow defects, renal disease, cell destruction, blood loss
o Hemoglobinopathies
o Lead poisoning
o Iron deficiency
o Increased blood volume: Pregnancy, over-hydration

Question 37

What can cause increased Hb levels?

Answer 37

o	Increased RBCs:  Polycythemia vera; renal tumors
o	COPD    
o	Pulmonary fibrosis
o	Heart disease
o	Decreased blood volume: Dehydration

Question 38

What is hypochromasia?

Answer 38

o Variation in Hb content
o Increase in RBCs central pallor, which occupies more than the normal third of cell diameter
o Found in: Fe deficiency, thalassemia

Question 39

• What is hematocrit? How is it measured?

Answer 39

o Volume of RBCs, as a % of volume of whole blood
o Indirect measure of RBC number and volume, since it is directly affected by both
o Approx. 3x Hb value when RBCs are normal
o Old: centrifuge capillary tube, =”packed cell volume”, %
o New: computers based on RBC count and size; HCT=RBCxMCV / 10

Question 40

• What are normal HCT values?

Answer 40

o Males: 42-52%
o Females: 36-46%
o Pregnant female: >33%
o Elderly: may be slightly decreased

Question 41

• What can cause increased HCT values?

Answer 41

o	Erythrocytosis
o	Congenital heart disease
o	Polycythemia vera (PCV)
o	Severe dehydration
o	Severe COPD

Question 42

• What can cause decreased HCT values?

Answer 42

o Anemia; Hemoglobinopathies; Cirrhosis; Hemolytic anemia; Hemorrhage; Nutrient deficiencies; Bone marrow failure; Prosthetic valves; Renal disease; Normal Pg: 2nd trim; Rheumatoid/collagen vascular diseases; Blood cancers and malignancies

Question 43

• What are some interfering factors with HCT?

Answer 43

o 1. RBC size has influence: larger RBCs associated with higher Hct because larger cells take up greater percentage of total volume
o 2. Extremely high WBC counts falsely decrease
o 3. Over and under hydration
o 4. Pregnancy decreases due to hemodilution in 2nd trim.
o 5. High altitudes increase due to hypoxia
o 6. Post-hemorrhage values not reliable for several hours
o 7. Drugs may decrease – chloramphenicol, penicillin

Question 44

• What are RBC indices?

Answer 44

o describe the size and Hb content of RBCs.
o The relationships between HCT, Hb level, and the RBC are converted to RBC indices through mathematical formulas.
o used to help in the differential diagnosis of anemia.
o worked out and first applied to the classification of anemia’s by Maxwell Wintrobe in 1934

Question 45

• what is mean corpuscular volume?

Answer 45

o Average volume of RBCs; Reported as fL, femtoliters.
o Derived from the RBC histogram by multiplying the # of RBCs by the size of the RBCs and multiplied by a calibration constant, or the mean of the red blood cell distribution histogram= MCV.
o Normal: 82-97 fL
o decreased MCV = microcytic RBCs (100 fL)
o Most useful of the indices: anemia is micro, macro, or normocytic based on MCV

Question 46

• What is an RBC histogram?

Answer 46

o Represents relationship b/w RBC size and number

o If macrocytes are present, the peak will skew to right (left if microcytic)

Question 47

• What factors can interfere with MCV calculation?

Answer 47

o Anything that affects RBC counts or RDW:
o Very high WBC count
o High concentration of very large platelets
o Agglutinated RBCs
o RBC fragments that fall below the 36 fL threshold

Question 48

• What is mean corpuscular hemoglobin?

Answer 48

o The average weight of Hgb in a RBC, reported in pg, picograms. A calculated value.
o Normal range: 26-34 pg of Hgb/RBC.
o MCH = Hbx10 / #RBCs in millions
o decreased MCH: small RBCs contain less Hgb
o Increased MCH: large RBCs contain more Hgb

Question 49

• What are interfering factors with MCH?

Answer 49

o Anything that interferes with hemoglobin readings

o Anything that interferes with RBC counts

Question 50

What is the mean corpuscular hemoglobin concentration?

Answer 50

o A “weight to volume” ratio of the average concentration of Hgb in a given volume of RBCs. A Calculated value. Reported as g/dL.
o Normal MCHC = 32-36 g/dL
o MCHC= Hbx100 / HCT
o Used to classify anemia type; normal MCHC=normochromic anemia; <30 = hypochromic; hyperchromic doesn’t exits

Question 51

• Why can’t RBCs be considered hyperchromic?

Answer 51

o MCHC values have a theoretical limit of 37; only 37 g/dl of hemoglobin can fit into an RBC!
o Machines calc HCT based on MCV and #RBC -> as #RBCs decrease, MCHC increases
o machines may indicate MCHC >37 in those with spherocytosis, hyperlipidemia, cold agglutinins, & rouleaux formation (some RBC problem that falsely lowers the machine’s RBC count)

Question 52

what are some normocytic, normochromic anemias?

Answer 52

o Chronic illness – the “anemia of chronic disease” (early stages)
o Renal disease- EPO
o Acute blood loss
o Aplastic anemia
o Acquired hemolytic anemia
o Early Fe def: make less RBC b/c less Fe, but cells still have adequate Hb just fewer cells

Question 53

• What are some microcytic, hypochromic anemias?

Answer 53

o Iron deficiency due to: Malabsorption, Malnutrition, Loss of blood (non-acute)
o Sideroblastic
o Lead poisoning
o Anemia of chronic disease (later stages)
o Thalassemia

Question 54

• What are some macrocytic, normochromic anemias?

Answer 54

o Vitamin B12 or folic acid deficiency
o Pernicious anemia
o Side effects of chemotherapy
o Myelodysplastic syndrome: BM abnormalities that results in decreased production of RBCs & WBCs often a precursor to development of Acute Myelogenous Leukemia (AML)

Question 55

• Pattern 1:

Answer 55

o	RBC elevated
o	HB elevated
o	HCT elevated
o	MCV normal
o	MCH normal	
o	MCHC normal
o	RDW normal
o	EPO decreased

Question 56

• Pattern 2:

Answer 56

o	RBC decreased
o	Hb decreased
o	HCT decreased
o	MCV elevated
o	MCH elevated
o	MCHC normal
o	RDW elevated

Question 57

• Pattern 3:

Answer 57

o	RBC decreased
o	Hb decreased
o	HCT decreased
o	MCV decreased
o	MCH decreased
o	MCHC decreased
o	RDW elevated/normal

Question 58

• Pattern 4:

Answer 58

o	RBC decreased
o	Hb decreased
o	HCT decreased
o	MCV normal	
o	MCH normal
o	MCHC normal
o	RDW normal	
o	BUN increased
o	Creatinine increased
o	Urinalysis +1 protein
o	EPO decreased