The Complement The Complement SystemSystem

Andrew E Thompson MD Andrew E Thompson MD FRCPCFRCPC

Fellow in RheumatologyFellow in Rheumatology

University of British ColumbiaUniversity of British Columbia

OverviewOverview

The complement system is part of the The complement system is part of the innate immune systeminnate immune system (vs adaptive) (vs adaptive)

It is named “complement system” It is named “complement system” because it was first identified as a heat-because it was first identified as a heat-labile component of serum that labile component of serum that “complemented” antibodies in the “complemented” antibodies in the killing of bacteriakilling of bacteria

It is now known that it consists of over It is now known that it consists of over 30 30 proteinsproteins and contributes and contributes 3 g/L3 g/L to overall to overall serum protein quantitiesserum protein quantities

““Classical” PathwayClassical” Pathway

Begins with antibody binding to a cell surface and ends Begins with antibody binding to a cell surface and ends with the lysis of the cellwith the lysis of the cell

The proteins in this pathway are named C1-C9 (the The proteins in this pathway are named C1-C9 (the order they were discovered and not the order of the order they were discovered and not the order of the reaction)reaction)

When complement is activated it is split into two partsWhen complement is activated it is split into two parts– a – smaller of the twoa – smaller of the two– B – larger part and usually the active part (except with factor B – larger part and usually the active part (except with factor

2)2) Remember 3 Key WordsRemember 3 Key Words

– ACTIVATIONACTIVATION– AMPLIFICATIONAMPLIFICATION– ATTACKATTACK

““Classical Pathway”Classical Pathway”

ACTIVATIONACTIVATION– C1qC1q portion of C1 attaches to the Fc portion portion of C1 attaches to the Fc portion

of an of an antibodyantibody– Only Only IgGIgG and and IgMIgM can activate complement can activate complement– Once activated C1s is eventually cleaved Once activated C1s is eventually cleaved

which activates C4 and C2which activates C4 and C2– C4b & C2a come together to form the C4b & C2a come together to form the C4b2aC4b2a

which is the which is the C3 convertaseC3 convertase – C3 convertase activates C3 to C3a and C3bC3 convertase activates C3 to C3a and C3b

““Classical Pathway”Classical Pathway”

ACTIVATIONACTIVATION– C3aC3a binds to receptors on binds to receptors on basophilsbasophils

and and mast cellsmast cells triggering them to triggering them to release there vasoactive compounds release there vasoactive compounds (enhances vasodilation and (enhances vasodilation and vasopermeability)vasopermeability)

– C3aC3a is called an is called an anaphylatoxinanaphylatoxin– C3b C3b serves as an opsonin which serves as an opsonin which

facilitates immune complex clearancefacilitates immune complex clearance

““Classical Pathway”Classical Pathway”

AMPLIFICATIONAMPLIFICATION– Each C1s creates many C4b and C2b Each C1s creates many C4b and C2b

fragmentsfragments– Each C4bC2a creates many C3b Each C4bC2a creates many C3b

(activated C3)(activated C3)– Each C3b goes on to create many Each C3b goes on to create many

Membrane Attack ComplexesMembrane Attack Complexes– ExampleExample

1 C1S makes 100 C4bC2b1 C1S makes 100 C4bC2b 100 C4bC2b makes 10,000 C3b100 C4bC2b makes 10,000 C3b 10,000 C3b makes 1,000,000 MAC10,000 C3b makes 1,000,000 MAC

““Classical Pathway”Classical Pathway”

ATTACKATTACK– Most C3b serves an opsonin functionMost C3b serves an opsonin function– Some C3b binds to C4bC2a to form Some C3b binds to C4bC2a to form

the the C5 convertase C4bC2aC3bC5 convertase C4bC2aC3b– C5 convertase cleaves C5 leading to C5 convertase cleaves C5 leading to

the formation of the Membrane the formation of the Membrane attack Complex (C5-C6-C7-C8-C9)attack Complex (C5-C6-C7-C8-C9)

– The MAC “punches holes” in cell The MAC “punches holes” in cell walls resulting in lysiswalls resulting in lysis

C1qC2C4

2a2b4b4a

C3-convertase

C3

C3a C3b

C5-Convertase

C3aC3a binds to receptors on binds to receptors on basophilsbasophils and and mast cellsmast cells triggering them to release triggering them to release there vasoactive there vasoactive compounds (enhances compounds (enhances vasodilation and vasodilation and vasopermeability) - vasopermeability) - ANAPHYLATOXINANAPHYLATOXIN

C5

C5a

C5b

C5aC5a is a: is a:

1.1. Potent Potent anaphylatoxinanaphylatoxin

2.2. Chemoattractant Chemoattractant for neutrophilsfor neutrophils

C6

C7C8

C9

Classical Classical PathwayPathway

““Alternative Pathway”Alternative Pathway”

Requires no specific recognition Requires no specific recognition of antigen in order to cause of antigen in order to cause activationactivation

““Alternative Pathway”Alternative Pathway”

ACTIVATIONACTIVATION– Spontaneous conversion from C3 to Spontaneous conversion from C3 to

C3b occurs in bodyC3b occurs in body– Normally, C3b is very short lived and Normally, C3b is very short lived and

quickly inactivated by proteins on the quickly inactivated by proteins on the surface of the body’s own cell wallssurface of the body’s own cell walls

– However, bacteria or other foreign However, bacteria or other foreign material may lack these surface material may lack these surface proteins allowing C3b to bind and proteins allowing C3b to bind and stay activestay active

““Alternative Pathway”Alternative Pathway”

AMPLIFICATIONAMPLIFICATION– Factor B binds to C3bFactor B binds to C3b– Factor B is then cleaved by factor D Factor B is then cleaved by factor D

into Ba and Bbinto Ba and Bb– C3bBb remains which acts as a C3 C3bBb remains which acts as a C3

convertase (C3 convertase (C3 C3a and C3b) C3a and C3b)– C3bBbC3b is formed which acts as a C3bBbC3b is formed which acts as a

C5 convertaseC5 convertase

““Alternative Pathway”Alternative Pathway”

ATTACKATTACK– C5 is cleaved to C5a and C5bC5 is cleaved to C5a and C5b– C5b then starts the assembly of the C5b then starts the assembly of the

Membrane Attack ComplexMembrane Attack Complex

C3C3bC3a

Anaphylatoxin

BD

BbBa

C3

C3a

C3bC5-Convertase

C3-Convertase

C5

C5a

C5b

AlternativAlternative Pathwaye Pathway

C6

C7C8

C9

Summary - ActivationSummary - Activation

Complement can be activated by the Complement can be activated by the binding of antibody (Classical) or by the binding of antibody (Classical) or by the adherance of C3b to foreign material adherance of C3b to foreign material (Alternative)(Alternative)

The two pathways converge at the The two pathways converge at the formation of the C5 convertase (C4b2a3b formation of the C5 convertase (C4b2a3b or C3bBbC3b)or C3bBbC3b)

The final common pathway is the The final common pathway is the formation of the membrane attack formation of the membrane attack complex complex

Summary - FunctionSummary - Function

OpsonizationOpsonization – C3b – C3b ChemotaxisChemotaxis – C5a (attracts – C5a (attracts

neutrophils)neutrophils) Increases vasodilation & Increases vasodilation &

permeabilitypermeability of capillary beds via of capillary beds via mast cell and basophil activation – mast cell and basophil activation – C3a & C5a (Anaphylatoxins)C3a & C5a (Anaphylatoxins)

Cellular LysisCellular Lysis via the MAC via the MAC

Laboratory MeasuresLaboratory Measures

C3 – Quantitative measure (nephelometry)C3 – Quantitative measure (nephelometry) C4 – Quantitative measure (nephelometry)C4 – Quantitative measure (nephelometry) CH50CH50

– Functional assay of entire Classical PathwayFunctional assay of entire Classical Pathway– Measures the ability of the patients serum to Measures the ability of the patients serum to

lyse 50% of a standard suspension of sheep lyse 50% of a standard suspension of sheep erythrocytes coated with rabbit antibodyerythrocytes coated with rabbit antibody

– A low CH50 suggests eitherA low CH50 suggests either CONSUMPTION OF COMPLEMENT COMPONENTSCONSUMPTION OF COMPLEMENT COMPONENTS DEFICIENCY OF COMPLEMENT COMPONENTSDEFICIENCY OF COMPLEMENT COMPONENTS

AH50AH50

The Role of Complement The Role of Complement in the Rheumatic in the Rheumatic DiseasesDiseases The Double Edged Sword!The Double Edged Sword! Needed for proper handling of immune Needed for proper handling of immune

complexescomplexes Also mediates tissue damage, Also mediates tissue damage,

especially in the setting of especially in the setting of autoantibodies and excessive immune autoantibodies and excessive immune complex formationcomplex formation

Just as the complement system can Just as the complement system can destroy a microbe, it can lyse and destroy a microbe, it can lyse and erythrocyte, phagocytose a platelet, or erythrocyte, phagocytose a platelet, or disrupt a basement membranedisrupt a basement membrane

Autoimmunity and Autoimmunity and InheritedInherited Complement Complement DeficienciesDeficiencies Inherited deficiencies of complement Inherited deficiencies of complement

components can result in autoimmunity, components can result in autoimmunity, especially SLEespecially SLE– C1q deficiency – 90% have SLEC1q deficiency – 90% have SLE– C4 deficiency – 75-80% have SLEC4 deficiency – 75-80% have SLE– C2 deficiency – 10-40% have SLEC2 deficiency – 10-40% have SLE

Early age of onset, prominent cutaneous Early age of onset, prominent cutaneous manifestations, and presence of anti-Ro manifestations, and presence of anti-Ro antibodies are features suggestive of a antibodies are features suggestive of a complement deficiencycomplement deficiency

Autoimmunity and Autoimmunity and Inherited Complement Inherited Complement DeficienciesDeficiencies How does SLE form with complement How does SLE form with complement

deficiencies?deficiencies?– Failure to clear autoantigens (apoptotic cells)Failure to clear autoantigens (apoptotic cells)– Immature dendritic cells uptake the antigen Immature dendritic cells uptake the antigen

in the presence of inflammatory cytokines in the presence of inflammatory cytokines causing them to mature into antigen causing them to mature into antigen presenting dendritic cells – Presents to T-Cellpresenting dendritic cells – Presents to T-Cell

– Autoreactive B-Cells take up antigen from Autoreactive B-Cells take up antigen from apoptotic cells and (with the help CD4+ Th2-apoptotic cells and (with the help CD4+ Th2-Cells) transform into plasma cells that Cells) transform into plasma cells that secrete autoantibodysecrete autoantibody

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

Indirect Laboratory uses of Indirect Laboratory uses of Complement – Detection of Complement – Detection of Immune ComplexesImmune Complexes C1q binding AssayC1q binding Assay

– Normally C1q has a very weak affinity for Normally C1q has a very weak affinity for monomeric IgG and IgMmonomeric IgG and IgM

– When IgG or IgM are part of an immune When IgG or IgM are part of an immune complex the Fc portion undergoes a complex the Fc portion undergoes a conformational changeconformational change

– This results in a much higher affinity for C1qThis results in a much higher affinity for C1q– This test is an ELISA which looks for This test is an ELISA which looks for

immune complexes in a patients serum immune complexes in a patients serum capable of binding C1qcapable of binding C1q

Indirect Laboratory uses of Indirect Laboratory uses of Complement – Detection of Complement – Detection of Immune ComplexesImmune Complexes Raji Cell PreparationRaji Cell Preparation

– Raji cells are a human lymphoblastoid cell derived Raji cells are a human lymphoblastoid cell derived from a patient with Burkitt’s lymphomafrom a patient with Burkitt’s lymphoma

– They are unique becauseThey are unique because They have surface receptors for C1q, C3b, C3bi, and C3dThey have surface receptors for C1q, C3b, C3bi, and C3d Lack of surface immunoglobulinLack of surface immunoglobulin Surface IgG receptors are low in number and aviditySurface IgG receptors are low in number and avidity

– Therefore, immune complexes containing Therefore, immune complexes containing complement can bind to surface receptors on Raji complement can bind to surface receptors on Raji Cells!Cells!

– This can then help to detect immune complexes This can then help to detect immune complexes capable of binding complementcapable of binding complement

– Sensitive test, however, warm reactive anti-Sensitive test, however, warm reactive anti-lymphocyte antibodies and anti-ds-DNA antibodies lymphocyte antibodies and anti-ds-DNA antibodies may cause false positive resultsmay cause false positive results