Complement Mediated Coagulation (PNH, aHUS)

Question 1

What part of the immune system

is complement a part of ?

Answer 1

• innate immune system
  
  • over 30 soluble and membrane bound proteins
  • provides defense against bacteria, fungi and viruses
  • many regulatory proteins that are either soluble or membrane bound
• mutations and autoantibodies frequently lead to disease
  
  • ex: PNH, aHUS

Question 2

What is the clinical manifestation

of coagulation cascade disorders ?

Answer 2

• often associated with thrombosis
  
  • highlights the close association between coagulation cascade and complement

Question 3

Which membrane complement regulators

have been linked to human disease ?

Answer 3

• CD55
  
  • limits CD5 convertase and ultimately prevents formation of the MAC complex
• CD59
  
  • major inhibitor of the terminal complement pathway
  • inhibits pore formation in MAC
• CD46
  
  • accelerates the decay of C3 convertase
  • it is expressed on all cells except erythrocytes
• Thrombomodulin
  
  • endothelial cell receptor
  • helps with activation of protein C
  • regulates Factor I mediated CDb inactivation

Question 4

What are the soluble complement regulators ?

Answer 4

• Factor H
  
  • regulates the alternative pathway
  • disrupts C3 convertase
• Factor I
  
  • synthesized in the liver
  • regulates all pathways in complement
  • inactivates C3b
• Factor B
  
  • cleaved by Factor D to generate C3 convertase of alternative pathway

Question 5

Which complement molecule seems to be

the connection between the cascade and coagulation ?

Answer 5

• C5a anaphylotoxin
• how it does it:
  
  • increase inflammatory cytokines
  • downregulates ADAMTS-13
  • generates tissue factor and PAI-1
  • decreases levels of Protein S, increase protein C resistance
  • activates thrombin

Question 6

What aspects of the coagulation cascade

can activate complement ?

Answer 6

• Factor XII interacts with C1 of the classical pathway
• Thrombin cleaves C3 and also generates C5a in the absence of C3
• Fibrinolytic factors (plasmin, kallikrein)
  
  • directly cleave C3

Question 7

How do complement-mediated hemolytic anemias

lead to thrombosis ?

Answer 7

• free hemoglobin results in platelet activation as well as:
  
  • inhibition of ADAMTS13
  • release of endothelial and red cell procoagulants
  • increased levels of tissue factor
  • oxygen reactive species
  • depletion of nitric oxide

Question 8

What is PNH ?

Answer 8

• acquired clonal hematopoietic stem cell disorder
• caused by mutations in the GPI-anchored proteins on surfaces of affected cells

Question 9

What gene is PIGA located on ?

Answer 9

• X chromosome
• gene product of one of 7 enzymes involved in GPI anchor biosynthesis
• IMP
  
  • virtually all PNH cases are associated with PIGA mutations

Question 10

What are other mutations that cause

PNH other than PIGA ?

Answer 10

• mutations on CD59 and CD55
• these are GPI-anchored proteins and are key in complement regulation
• due to their absence on erythrocytes
  
  • PNH manifest with chronic hemolyitic anemia
  • caused by alternative complement pathway activation on mutant erythroid membranes

Question 11

How long can the mutant RBC s in PNH circulate ?

Answer 11

• unlike foreign RBCs PNH erythrocytes circulate for 6-60 days
  
  • they are protected by factor H
  • so patients with decreased levels of factor H have worse hemolytic disease

Question 12

What other disorder is closely associated

with PNH ?

Answer 12

• acquired aplastic anemia
  
  • autimmune entity
  • characterized by depletion of hematopoietic stem cells

Question 13

What tests should be ordered in PNH ?

Answer 13

• Gold standard: flow cytometry
  
  • cocktail of antibodies against individual GPI-anchored proteins
  • FLAER
    
    • variant antibody that binds to the GPI anchor
    • helps detect loss of GPI anchored proteins

Question 14

What was the historical test for PNH and

what is the principle behind it ?

Answer 14

• Ham test
  
  • principle: PNH cells are more vulnerable to acidified serum which serves to activate the alternative complement pathway
  • thus incubation with acidified serum causes hemolysis that is not observed with normal erythrocytes
  • test lacked sensitivity and specificity

Question 15

What are the general manifestations of PNH ?

Answer 15

• most patients have chronic hemolysis with bursts of hemoglobinuria
• patients with bone marrow failure exhibit low levels of chronic hemolysis
• reticulocyte count is elevated
  
  • but not to the level you would expect for the degree of anemia
• morphologically the cells look normal
• clinical symptoms of anemia

Question 16

Clinical findings in PNH include what ?

Answer 16

• anemia (intravascular and extravascular)
• venous thrombosis
  
  • intraabdominal veins ex: mesenteric
  • cerberal veins
  • deep venous thrombosis
  • pulmonary embolism
  • dermal veins
• arterial thrombus
• smooth muscle distonia
• chronic kidney disease

Question 17

What factors are important in identifying the

degree of hemolysis in PNH ?

Answer 17

• size and type of PNH clone
• degree of complement activation

Question 18

What is the most common site of thrombosis

in PNH ?

Answer 18

• hepatic vein thrombsosi
  
  • Budd Chiari Syndrome

Question 19

What is the treatment of choice for PNH ?

Answer 19

• Eculizumab
  
  • used in patients with severe hemolytic anemia and thrombosis
  • must be infused every 2 weeks IV (limitation)
  • mechanism:
    
    • binds the terminal complement component C5 and blocks MAC formation

IMP

• due to absence of CD55 PNH erythrocytes are susceptible to opsonization by CD3b and thus extravascular hemolysis when they are on eculizumab
  
  • known to have chronic extravascular hemolysis

Question 20

What is the only cure for PNH ?

Answer 20

• hematopoietic stem cell transplantation

Question 21

What is atypical hemolytic

uremic syndrome ?

Answer 21

• it is a thrombotic microangiopathy
  
  • includes a heterogeneous group
  • presentation is typically MAHA, thrombocytopenia and organ damage
  • peripheral smear shows schistocytes, anemia and thrombocytopenia
• in order to diagnose atypical hemolytic uremic syndrome you must exclude other TMAs

Question 22

What is the pathophysiology of

atypical HUS ?

Answer 22

• disease of excessive activation of the APC
  
  • can be from inherited mutations in APC proteins or in autoantibodies
• requires a two hit model
  
  • mutations and autoantibodies are not enough to cause it
• what are common triggers:
  
  • pregnancy
  • inflammation
  • autoimmunity
  • surgery

Question 23

What tests should be ordered

for aHUS ?

Answer 23

• it is a clinical diagnosis of exclusion
• modified Ham test
  
  • use PNH erythrocytes (sensitive to complement)
  • mix them with aHUS patient serum (activated)
    
    • cells do not release dye due to complement induced death
  • has been initially shown to differentiate well between aHUS and TTP

Question 24

What is the differential diagnosis of

aHUS /

Answer 24

• TTP
• Shiga-toxin producing E.coli induced HUS
• Secondary causes of microangiopathy:
  
  • DIC
  • Drugs
  • Malignancy
  • Slceroderma-associated renal crisis
  • Malignant hypertension

Question 25

What is considered a severe ADAMTS13 deficiency ?

Answer 25

• usually level <10%
  
  • inherited : Upshaw-Schulman Syndrome
  • acquired: IgG autoantibodies against the molecule
    
    • more common than inherited

IMP: if ADAMTS13 is >10% and shiga-toxin is negative than aHUS should be considered

Question 26

What are the clinical findings in

aHUS ?

Answer 26

• aHUS patients usually manifest by age 18
• generally have anemia, thrombocytopenia and acute kidney injury
• other possible presentations
  
  • peripheral gangrene
  • arterial stenosis
  • dilated cardiomyopathy
  • cardiorespiratory arrest
  • neurologic, pulmonary and gastrointestinal complications

Question 27

What is the treatment for

aHUS ?

Answer 27

• severe, life-threatening emergency requiring immediate treatment
• plasma exchange can help in some cases but lasting complement mediated damage to the kidneys and CNS often persist and may lead to death
• Eculizumab is the treatment of choice

Question 28

How is failure to respond to PLEX diagnosed ?

Answer 28

• most frequently used criteria:
  
  • failure to achieve a hematologic response (improvement in platelet count, decrease in LDH) over the first 4-5 days
  • progressive end-organ damage while on PLEX
  • ADAMTS13 <10%