Heme 05: Disorders Of Hemoglobin

Front 1

What are the types of Hemoglobin disorders?

Back 1

- Structural variants of globins
- Thalassemias (underproduction of normal globin)

Front 2

What kind of anemia do Thalassemias cause?

Back 2

Microcytic / Hypochromic Anemias of varying severity

Front 3

What chromosomes are the α and β globin genes on?

Back 3

- 1 β gene on Chromosome 11
- 2 α genes on Chromosome 16

Front 4

How many different types of abnormal hemoglobins are there? How do they form?

Back 4

- >500 structural variants
- Most are single AA replacements in globin molecules
- Occasionally 2 aa replaced, deletions, insertions, chain elongations, or fusion genes
- Most are clinically silent

Front 5

What are the consequences of structural globin abnormalities dependent on?

Back 5

- What globin gene is affected (eg, δ gene mutations are inconsequential)
- Location of substitution in tertiary structure and/or quarternary structures of globin or Hb molecules

Front 6

What are the potential consequences of structural globin abnormalities?

Back 6

- Sickling
- Instability (degrades prematurely)
- Altered O2 affinity (↑ or ↓)
- Increased susceptibility to oxidation to methemoglobin (ferrous iron oxidized to ferric iron and unable to bind O2)
- Under-production
- Various combinations of above

Front 7

How do you diagnose structurally abnormal Hb?

Back 7

- Hemoglobin electrophoresis (gel or capillary)
- High performance liquid chromatography (HPLC)
- Other advanced techniques (isoelectric focusing, globin chain electrophoresis, gene mutation analysis)

Front 8

What is the procedure for a routine hemoglobin electrophoresis?

Back 8

- Typically performed in parallel w/ alkaline and acid buffers
- HbA has isoelectric point of 6.8 so in alkaline buffers will migrate to anode (+) but in acidic buffers will migrate to cathode (-)

Front 9

How does the buffer affect the migration of Hemoglobin on electrophoresis?

Back 9

Alkaline Buffer:
- HbA has negative charge → migrates towards anode (+)
- Migration of other Hb depends on net charge

Acidic Buffer:
- HbA has positive charge → migrates towards cathode (-)
- Migration of other Hb depends on net charge and interaction w/ components of media (slightly more complicated)

Front 10

What is the procedure for an HPLC electrophoresis?

Back 10

1) Fully automated cation exchange chromatography method

2) Whole blood method (whole blood hemosylate)
- Hb adsorbed onto resin particle in column
- Different species differentially eluted based on affinity for resin by gradually changing ionic strength of elution buffer
- Hb come off column at highly predictable retention times

Front 11

What determines the extent of HbS polymerization in Sickle Cell Disease (SS)?

Back 11

Time and concentration dependent

Front 12

What are the factors affecting the concentration of HbS (and thus influencing the polymerization of HbS)?

Back 12

- Percentage of HbS of total Hb
- Total Hb concentration in RBC (MCHC)

Front 13

What affects the percentage of HbS of total Hb?

Back 13

- Homozygous vs heterozygous Sickle Cell Disease
- Presence of other Hb species (eg, HbF)

Front 14

What affects the total Hb concentration in RBCs (MCHC)?

Back 14

- Increased in states of cellular dehydration
- Decreased when there is co-existant thalassemia

Front 15

What are some examples of how time affects the amount of sickling in HbS?

Back 15

- Sickling is enhanced in anatomic sites w/ sluggish flow (eg, spleen and BM)
- Blood flow through microvasculature retarded in certain pathologic states (eg, inflammation)

Front 16

What clinical settings predipose to sickling? Why?

Back 16

- Hypoxia
- Acidosis (shifts O2 dissociation curve to R causing increased deoxygenation of HbS)
- Dehydration (hypertonicity causes RBC dehydration)
- Cold temperatures (probably a result of peripheral vasconstriction w/ resultant sluggish flow)
- Infections (multiple mechanisms)

Front 17

At what PaO2 do SS (HbS) cells begin to sickle?

Back 17

~40 mmHg

Front 18

What happens to a RBC that sickles?

Back 18

- Initially reversible process
- After multiple sickling/unsickling cycles, membrane damage produces an irreversibly sickled cell
- RBC lifespan decreased to 20 days (from 120)

Front 19

What are the effects of RBC sickling?

Back 19

- Chronic hemolysis (correlates w/ number of irrev. sickled cells)
- Microvascular occlusion w/ resultant tissue hypoxia and infarction (does not correlate w/ irrev. sickled cells, rather is related to increased "stickiness" of SS RBCs b/c of membrane damage)

Front 20

How are newborns affected by Sickle Cell Disease?

Back 20

- Clinically fine because of high HbF levels
- Hematologic manifestations begin by 10-12 weeks of age

Front 21

What are the clinical manifestations of Sickle Cell Disease?

Back 21

- Severe anemia
- Acute pain crises
- Aplastic crises
- Auto-splenectomy or splenic sequestration crises
- Acute Chest Syndrome
- Strokes
- Megaloblastic anemia
- Growth retardation
- Bony abnormalities
- Leg ulcers
- Infections
- Cholelithiasis

Front 22

What causes acute pain crises in Sickle Cell Disease?

Back 22

Vaso-occlusion, particularly in BM

Front 23

What causes auto-splenectomy in Sickle Cell Disease? How common?

Back 23

- Repeated episodes of splenic infarction, results in shrunken, fibrotic, non-functional spleen
- Seen in essentially all adults w/ SS disease

Front 24

What causes Acute Chest Syndrome in Sickle Cell Disease?

Back 24

- Severe complication, major cause of death
- Results from pulmonary infections or fat emboli from infarcted BM
- Sluggish blood flow from inflammation causes sickling and vaso-occlusion, triggering vicious cycle

Front 25

What is the major cause of death in Sickle Cell Disease

Back 25

Acute Chest Syndrome - results from pulmonary infections or fat emboli from infarcted BM

Front 26

What causes Aplastic Crises in Sickle Cell Disease?

Back 26

Acute decrease in RBC production, typically from parvovirus B19 infection

Front 27

What causes Splenic Sequestration Crises in Sickle Cell Disease? What can it cause?

Back 27

- Acute pooling of blood in spleen
- Precipitous drop in hemoglobin
- Potential for hypovolemic shock
- Important cause of morbidity and mortality in children w/ SS disease

Front 28

What is an important cause of morbidity and mortality in children w/ SS disease?

Back 28

Splenic Sequestration Crises

Front 29

What causes Megaloblastic Anemia in Sickle Cell Disease? What can it cause?

Back 29

- Folate consumption because of chronic erythroid hyperproliferation
- Insufficient folate levels

Front 30

What are the laboratory findings in Sickle Cell Disease?

Back 30

- Chronic Anemia (Hb: 5-11 g/dL, normally ~7g/dL)
- ↑ Bilirubin
- Sickled cells, target cells, and polychromasia
- ↑ Reticulocytes
- Normal MCV
- Post-splenectomy changes in adults

Front 31

How common is "S-trait"? Clinical implications?

Back 31

- 8% of African Americans have S-Trait

- Clinically benign: no anemia, normal RBC survival, no crises or other complications in majority, normal PB smear
- May be mild, sub-clinical kidney damage: impairment of urine concentration, microhematuria

Front 32

What kind of Hemoglobin is found in patients w/ "S-trait"?

Back 32

- 60% HbA
- 40% HbS

Front 33

What is Hemoglobin SC disease?

Back 33

- Compound heterozygous state
- HbC results from Glu6 to Lys6 substitution on β-globin chain
- HbS results from Glu6 to Val6 substitution on β-globin chain

Front 34

How does Hemoglobin SC disease compare to SS disease?

Back 34

- Generally SC is milder than SS, but it is highly variable
- SC: Hb levels 10-12 g/dL
- SS: Hb levels 5-11 g/dL (avg ~7 g/dL)

Front 35

What is Hemoglobin S / β-thalassemia disease?

Back 35

Heterozygous HbS w/ trans β-thalassemia allele resulting in decreased or absent production of normal β-chains

Front 36

What are the symptoms of Hemoglobin S / β-thalassemia disease?

Back 36

Ranges from asymptomatic to a disorder nearly identical to SS disease, depending on output of normal β-chains from thalassemia allele

Front 37

What are the hemoglobin lab findings for Hemoglobin S / β-thalassemia disease?

Back 37

HbS > HbA

Front 38

What are thalassemias?

Back 38

Inherited disorders characterized by decreased production of structurally normal globin chains

Front 39

Where is β-thalassemia more common?

Back 39

Wide distribution in Mediterranean, Middle East, parts of India and Pakistan, and Southeast Asia

Front 40

Where is α-thalassemia more common?

Back 40

Occurs throughout Africa, Mediterranean, Middle East, and SE Asia

Front 41

What kind of anemia is caused by Thalassemias?

Back 41

Microcytic, hypochromic anemia d/t decreased hemoglobin production

Front 42

What does the severity of the hematologic manifestations of Thalassemia depend on?

Back 42

Degree of chain imbalance

Front 43

What happens to the normal globin in Thalassemias?

Back 43

Excess normally produced globin chains accumulate and cause intramedullary cell death and/or decreased RBC survival

Front 44

What kind of mutations occur in β-thalassemias? Causes?

Back 44

- Decreases β-globin chain production from affected alleles
- 250 mutations: mutations causing splicing errors, mutations in promotors causing decreased transcription, translation errors (frameshift or nonsense codons), but gene deletions are rare

Front 45

What are the types of β-thalassemias?

Back 45

- β-thalassemia major (Cooley's anemia)
- β-thalassemia intermedia
- β-thalassemia minor

Front 46

How much β-chain is present in β-thalassemia major? α-chain?

Back 46

- Absent or marked decrease in β-chain production on both β alleles
- Excess of normal α-chains, which are unable to form tetramers, and precipitate in normoblasts and erythrocytes

Front 47

What happens to RBCs in β-thalassemia major?

Back 47

- Intramedullary cell death and decreased RBC lifespan
- Ineffective erythropoiesis
- Bizarre RBC morphology (hypochromia, targeting, and erythroblastosis)

Front 48

How are infants affected by β-thalassemia major?

Back 48

Infants are well at birth, but anemia develops over first few months of life

Front 49

How severe of anemia is in β-thalassemia major? How much Hb?

Back 49

- Severe anemia
- Hb: 2-3 g/dL (virtually all HbF)

Front 50

How do you treat β-thalassemia major?

Back 50

- Dependent on transfusions
- Severity of clinical effects depends on adequacy of transfusion program and efficacy of iron chelation

Front 51

What are the large dots?

Back 51

Nucleated RBCs

Front 52

What are the effects of inadequately transfused β-thalassemia major?

Back 52

- Stunted growth
- Frontal bossing
- "Mongoloid" facies
- Increased skin pigmentation
- Characteristic bony abnormalities
- Fever
- Wasting
- Hyperuricemia
- Spontaneous fractures
- Hepatosplenomegaly
- Infections
- Folate deficiency
- Death in childhood

Front 53

What are the features of adequately transfused β-thalassemia major?

Back 53

- Normal early development
- Avoidance of classic complications
- Additional features depend on whether there is adequate iron chelation therapy

Front 54

What are the features of adequately transfused β-thalassemia major without adequate iron chelation therapy? Prognosis?

Back 54

- Absence of pubertal growth spurt and menarche
- Endocrine disturbances such as DM, adrenal insufficiency
- Death from cardiac disease by end of 3rd decade

Front 55

What are the features of adequately transfused β-thalassemia major with aggressive iron chelation therapy? Prognosis?

Back 55

- Less severe cardiac disease and endocrine disturbances
- Significantly improved life-span

Front 56

What is necessary to have a good life-span with β-thalassemia major?

Back 56

Adequately transfused w/ aggressive iron chelation therapy

Front 57

What are the features of β-thalassemia minor?

Back 57

- Heterozygous
- Asymptomatic
- Discovered incidentally

Front 58

In what populations is β-thalassemia minor more common?

Back 58

- Common in Mediterranean and Asian populations
- 1.5% of African Americans

Front 59

What are the lab features of β-thalassemia minor?

Back 59

- Mild or no anemia (Hb>~10g)
- Microcytosis (50-70 fL)
- Mild anisopoikilocytosis w/ scattered target cells
- Basophilic stippling
- Elevated HbA2: 3.5-7%

Front 60

What does this image show?

Back 60

β-thalassemia minor:
- Microcytosis (50-70 fL)
- Mild anisopoikilocytosis w/ scattered target cells
- Basophilic stippling

Front 61

What are the features of β-thalassemia intermedia?

Back 61

- Heterogenous group
- Intermediate severity between major and minor

Front 62

What causes α-thalassemia?

Back 62

Gene deletion of α-gene(s)

Front 63

What are the clinical subtypes of α-thalassemia?

Back 63

- Silent carrier (1 α gene deleted)
- α-thalassemia trait (2 α genes deleted)
- Hemoglobin H disease (3 α genes deleted)
- Hydrop Fetalis (4 α genes deleted)

Front 64

What is it called when 2 α-globin genes are deleted?Symptoms?

Back 64

α-Thalassemia Trait
- Mild microcytic anemia similar to β-thalassemia minor
- Discovered incidentally

Front 65

What is it called when 3 α-globin genes are deleted?Symptoms?

Back 65

Hemoglobin H Disease
- Mild to moderate, chronic hemolytic anemia
- HbH represents β-tetramers (excess form tetramers) - does not effectively transfer O2
- HbH soluble, so does not initially precipitate in normoblasts (no intramedullary cell death) - unstable over time, so precipitates in circulating RBCs, causing hemolysis

Front 66

What is it called when 4 α-globin genes are deleted?Symptoms?

Back 66

Hydrops Fetalis
- Infants are either stillborn or die within first few hours of life