1. The Complement System Andrew E Thompson MD FRCPC
Fellow in Rheumatology
University of British Columbia

2. Overview
The complement system is part of the innate immune system (vs adaptive)
It is named “complement system” because it was first identified as a heat-labile component of serum that “complemented” antibodies in the killing of bacteria
It is now known that it consists of over 30 proteins and contributes 3 g/L to overall serum protein quantities

3. “Classical” Pathway
Begins with antibody binding to a cell surface and ends with the lysis of the cell
The proteins in this pathway are named C1-C9 (the order they were discovered and not the order of the reaction)
When complement is activated it is split into two parts
a – smaller of the two
B – larger part and usually the active part (except with factor 2)
Remember 3 Key Words
ACTIVATION
AMPLIFICATION
ATTACK

4. “Classical Pathway” ACTIVATION
C1q portion of C1 attaches to the Fc portion of an antibody
Only IgG and IgM can activate complement
Once activated C1s is eventually cleaved which activates C4 and C2
C4b & C2a come together to form the C4b2a which is the C3 convertase
C3 convertase activates C3 to C3a and C3b

5. “Classical Pathway” ACTIVATION
C3a binds to receptors on basophils and mast cells triggering them to release there vasoactive compounds (enhances vasodilation and vasopermeability)
C3a is called an anaphylatoxin
C3b serves as an opsonin which facilitates immune complex clearance

6. “Classical Pathway” AMPLIFICATION
Each C1s creates many C4b and C2b fragments
Each C4bC2a creates many C3b (activated C3)
Each C3b goes on to create many Membrane Attack Complexes
Example
1 C1S makes 100 C4bC2b
100 C4bC2b makes 10,000 C3b
10,000 C3b makes 1,000,000 MAC

7. “Classical Pathway” ATTACK Most C3b serves an opsonin function
Some C3b binds to C4bC2a to form the C5 convertase C4bC2aC3b
C5 convertase cleaves C5 leading to the formation of the Membrane attack Complex (C5-C6-C7-C8-C9)
The MAC “punches holes” in cell walls resulting in lysis

8. C3a binds to receptors on basophils and mast cells triggering them to release there vasoactive compounds (enhances vasodilation and vasopermeability) - ANAPHYLATOXIN
C5a is a:
Potent anaphylatoxin
Chemoattractant for neutrophils C3
C1q C2 C4
C3a C5
C3b 2a 4a 4b 2b
C5b
C5a
C5-Convertase
C3-convertase C7 C8 C9 C6
Classical Pathway

9. “Alternative Pathway”
Requires no specific recognition of antigen in order to cause activation

10. “Alternative Pathway”
ACTIVATION
Spontaneous conversion from C3 to C3b occurs in body
Normally, C3b is very short lived and quickly inactivated by proteins on the surface of the body’s own cell walls
However, bacteria or other foreign material may lack these surface proteins allowing C3b to bind and stay active

11. “Alternative Pathway”
AMPLIFICATION
Factor B binds to C3b
Factor B is then cleaved by factor D into Ba and Bb
C3bBb remains which acts as a C3 convertase (C3  C3a and C3b)
C3bBbC3b is formed which acts as a C5 convertase

12. “Alternative Pathway”
ATTACK
C5 is cleaved to C5a and C5b
C5b then starts the assembly of the Membrane Attack Complex

13. Alternative Pathway D Bb B Ba C3a C3 C3b C3 Anaphylatoxin C5
C3-Convertase
C5-Convertase
C3a
C3b
C5b
C5a D C7 B Bb Ba C8 C6 C9

14. Summary - Activation
Complement can be activated by the binding of antibody (Classical) or by the adherance of C3b to foreign material (Alternative)
The two pathways converge at the formation of the C5 convertase (C4b2a3b or C3bBbC3b)
The final common pathway is the formation of the membrane attack complex

15. Summary - Function Opsonization – C3b
Chemotaxis – C5a (attracts neutrophils)
Increases vasodilation & permeability of capillary beds via mast cell and basophil activation – C3a & C5a (Anaphylatoxins)
Cellular Lysis via the MAC

16. Laboratory Measures C3 – Quantitative measure (nephelometry)
CH50
Functional assay of entire Classical Pathway
Measures the ability of the patients serum to lyse 50% of a standard suspension of sheep erythrocytes coated with rabbit antibody
A low CH50 suggests either
CONSUMPTION OF COMPLEMENT COMPONENTS
DEFICIENCY OF COMPLEMENT COMPONENTS
AH50

17. The Role of Complement in the Rheumatic Diseases
The Double Edged Sword!
Needed for proper handling of immune complexes
Also mediates tissue damage, especially in the setting of autoantibodies and excessive immune complex formation
Just as the complement system can destroy a microbe, it can lyse and erythrocyte, phagocytose a platelet, or disrupt a basement membrane

18. Autoimmunity and Inherited Complement Deficiencies
Inherited deficiencies of complement components can result in autoimmunity, especially SLE
C1q deficiency – 90% have SLE
C4 deficiency – 75-80% have SLE
C2 deficiency – 10-40% have SLE
Early age of onset, prominent cutaneous manifestations, and presence of anti-Ro antibodies are features suggestive of a complement deficiency

19. Autoimmunity and Inherited Complement Deficiencies
QUIZ TIME
Q: Dendritic Cells present antigen with MHC Class I or MHC Class II?
A: MHC Class II – It is an APC (antigen presenting cell)
Q: MHC Class II molecules present antigen derived from intracellular or extracellular processes?
A: Extra-cellular (e.g. apoptotic cells)
Q: MHC Class II molecules stimulate CD4 or CD8 cells
A: CD4 (IIx4=8, Ix8=8)
Q: In this scenario with the APC being a dendritic cell – the CD4 T-lymphocyte it interacts with is a Th1 or Th2?
A: Th1 is chiefly involved in cellular mediated immunity and Th2 is chiefly involved in humoral mediated immunity – Th2
How does SLE form with complement deficiencies?
Failure to clear autoantigens (apoptotic cells)
Immature dendritic cells uptake the antigen in the presence of inflammatory cytokines causing them to mature into antigen presenting dendritic cells – Presents to T-Cell
Autoreactive B-Cells take up antigen from apoptotic cells and (with the help CD4+ Th2-Cells) transform into plasma cells that secrete autoantibody

20. Indirect Laboratory uses of Complement – Detection of Immune Complexes
C1q binding Assay
Normally C1q has a very weak affinity for monomeric IgG and IgM
When IgG or IgM are part of an immune complex the Fc portion undergoes a conformational change
This results in a much higher affinity for C1q
This test is an ELISA which looks for immune complexes in a patients serum capable of binding C1q

21. Indirect Laboratory uses of Complement – Detection of Immune Complexes
Raji Cell Preparation
Raji cells are a human lymphoblastoid cell derived from a patient with Burkitt’s lymphoma
They are unique because
They have surface receptors for C1q, C3b, C3bi, and C3d
Lack of surface immunoglobulin
Surface IgG receptors are low in number and avidity
Therefore, immune complexes containing complement can bind to surface receptors on Raji Cells!
This can then help to detect immune complexes capable of binding complement
Sensitive test, however, warm reactive anti-lymphocyte antibodies and anti-ds-DNA antibodies may cause false positive results