Red Cell Membrane Disorders

Question 1

What are red cell membrane disorders?

Answer 1

• intrinsic defects of the red cell membrane => premature red cell destruction
• majority are hereditary rather than acquired

Question 2

RBCs must have deformability in order to squeeze through small spaces. This is based on 3 main cellular properties

Answer 2

• biconcave discoid geometry
• viscoelastic nature (elasticity) of the membrane
• cytoplasmic viscosity

Question 3

viscoelastic property of red cell membrane is affected by

Answer 3

mutations that alter proteins responsible for membrane structure

Question 4

cytoplasmic viscosity of red cell may be affected by

Answer 4

numerous mechanisms but membrane disorders are a rare cause and are due to membrane transport protein defects

Question 5

mutations that alter membrane structure

Answer 5

• hereditary spherocytosis
• hereditary elliptocytosis
• hereditary ovalocytosis

Question 6

mutations that alter membrane transport proteins

Answer 6

• dehydrated hereditary stomatocytosis

- overhydrated hereditary stomatocytosis

Question 7

T or F. hemolysis secondary to intrinsic RBS membrane defect is immune and DAT positive

Answer 7

F! non-immune and DAT negative

Question 8

Lab manifestation of hemolysis

Answer 8

• increased total bilirubin, decreased serum haptoglobin, increased LDH
• reticulocytosis (response by bone marrow)
• if BM can’t keep up = anemia

Question 9

group of hemolytic anemias caused by defects in proteins that disrupt the vertical interactions between transmembrane proteins and the underlying protein cytoskeleton

Answer 9

hereditary spherocytosis

- usually autosomal dominant inheritance

Question 10

hereditary spherocytosis mutations

Answer 10

• most common defects are combined ankyrin/spectrin mutations
• band 3 protein (30%)
• sole alpha spectrin, protein 4.2, Rh protein, etc, mutations are rare

Question 11

What do the defects in vertical membrane protein interactions result in?

Answer 11

separation of the spectrin-phospholipid bilayer

- causes RBCs to lose unsupported lipid membrane via vesicle formation w/out corresponding loss of volume

Question 12

end result of defects in vertical membrane protein interactions

Answer 12

decreased surface-to-volume ratio = shape becomes spherical = affects deformability

Question 13

typical lab findings for hereditary spherocytosis

Answer 13

• Hb decreased
• MCHC increased
• RDW increased
• retic count increase
• increase bilirubin, LDH
• decrease haptaglobin
• DAT neg
• spherocytes, polychromasia

Question 14

other causes of spherocytosis

Answer 14

• immune hemolytic anemia (DAT +)
• burns
• MAHA
• Clostridial sepsis
• hyposplenism (includes normal neonate)

Question 15

Diagnosis of hereditary spherocytosis: if uncertain, two other tests:

Answer 15

• EMA (eosin-5 maleimide)

- osomotic fragility test

Question 16

Describe EMA testing

Answer 16

• flow cytometry; incubation with a fluorescent dye (eosin 5-maleimide); binds to transmem proteins - mostly band 3
• even though Hs has various causes (ankyrin mutation), band 3 protein sites are decreased in all forms of HS
• decreased levels of EMA binding sites = decreased fluorescence compared to normal controls

Question 17

Osmotic fragility test principle

Answer 17

• RBCs put into series of increasingly hypotonic NaCl soln = water entering into RBCs to achieve eqm
• cause swelling; when internal vol gets too great = lysis
• bc spherocytes already have decreased surface area-vol ratio = they lyse in LESS hypotonic solns than normal RBCs

Question 18

T or F. osmotic fragility is more sensitive and specific than EMA testing

Answer 18

F! reverse

- several mods to soln may increase sensitivity and specificity (glycerol, acidified glycerol and pink test)

Question 19

Which test identifies membrane protein deficiencies by separating the various proteins via electrophoresis and quantifying via densitometry?

Answer 19

SDS-PAGE

Question 20

This measures RBC deformability at various osmotic gradients using a laser

Answer 20

Ektacytometry; most specific

Question 21

hypertonic cryohemolysis

Answer 21

based on HS cells being more sensitive to cooling at 0 degrees C in hypertonic soln than normal cells = this phenomenon is dependent on primary membrane protein defects rather than SA:vol

Question 22

Clinical manifestations of HS

Answer 22

• rarely severe anemia; majority have well compensated hemolysis w few symptoms
• at risk for:
  1. hemolytic crisis: increased rate of hemolysis due to infection or drugs
  2. aplastic crisis: BM ability to maintain high compensatory output impaired, most commonly by parvovirus infection or drugs
  3. megaloblastic crisis: high BM production outstrips folate supply (pregnancy)

Question 23

HS treatment

Answer 23

• transfusion as needed
• vitamin supplementation
• if significant chronic anemia or recurrent crises = splenectomy

Question 24

heterogenous group of membrane disorders; incidence unknown and most are asymptomatic

Answer 24

hereditary elliptocytosis

- more common in African/Med poplns (malarial geography)

Question 25

these gene mutations result in defective proteins which disrupt horizontal linkages of the protein cytoskeleton

Answer 25

HE!

• alpha spectrin
• beta spectrin
• protein 4.1

Question 26

when RBCs become ellipticle after repeated exposure to stress in peripheral circulation

Answer 26

• loss of deformability

- severe cases = membrane skeleton weakened to the point of lysis

Question 27

HE heterozygotes

Answer 27

• RBC lifespan usually normal = asymptomatic (picked up on PBS incidentally)
• may have mild anemia + lab evidence of hemolysis (can worsen with infections, pregnancy, etc.
• proportion of elliptocytes does NOT correlate with severity

Question 28

other causes of elliptical cells have to be ruled out prior to diagnosing HE

Answer 28

• iron deficiency
• thalassemia
• myelodysplastic syndrome
• primary myelofibrosis

Question 29

Hereditary Pyropoikilocytosis (HPP)

Answer 29

• HE homozygous or compound heterozygous
• rare
• can have moderate to severe hemolytic anemia
• may or may not have jaundice or splenomegaly

Question 30

HPP lab findings

Answer 30

• PBS shows extreme poikilocytosis with fragmentation, microspherocytes + elliptocytes
• MCV very low (fragments)
• RDW increased
• increased reticulocytosis
• marked RBC thermal sensitivity
• increased osmotic fragility
• decreased EMA fluorescence (more so than HS)
• may need gel electroph. or genetic testing

Question 31

AKA Southeast Asian Ovalocytosis

Answer 31

Hereditary Ovalocytosis

Question 32

HO

Answer 32

• single gene mutation in band 3 protein (tighter binding to ankyrin) = increased rigidity of membrane and decreased elasticity
• autosomal dominant; all cases heterozygous
• hemolysis is mild or absent
• no treatment required
• oval RBCs + 1-2 transverse bars

Question 33

mutations that alter membrane transport proteins

Answer 33

dehydrated her. stomatocytosis and overhydrated her. stomatocytosis

• affect cytoplasmic viscosity
• rarer than mutations that alter membrane structure
• asymptomatic + not often picked up

Question 34

this results in loss of ability to regulate cell vol by leakage of cations, resulting in either overhydration or dehydration

Answer 34

mutations that alter membrane transport proteins

Question 35

T or F. acquired causes of stomatocytosis are much more common

Answer 35

T!

includes: drying artifact on blood smear, alcoholism, cirrhosis, neoplasms, lead intoxication

Question 36

membrane transport protein mutation resulting in influx of Na+ that exceeds loss of K+ = more water enters cell = swelling

Answer 36

Overhydrated hereditary stomatocytosis

Question 37

increased permeability to K+ which leaks out but not balanced by increase in Na+ = water loss from cell

Answer 37

mild to moderate anemia and low level of stomatocytosis (dehydrated)