2941 Complement System

8/3/2019 2941 Complement System

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COMPLEMENT SYSTEMCOMPLEMENT SYSTEM

A.EVANGELINE CHRISTINAA.EVANGELINE CHRISTINA

11STST M.Sc Applied Microbiology,M.Sc Applied Microbiology,

The Presidency College.The Presidency College.


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DEFINITIONDEFINITION

It refers to a system of factors which occurIt refers to a system of factors which occur

in normal serum and are activatedin normal serum and are activated

characteristically by ANTIGENcharacteristically by ANTIGEN--ANTIBODYANTIBODYinteraction and subsequently mediate ainteraction and subsequently mediate a

number of biologically significantnumber of biologically significant

consequences.consequences.


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HistoryHistory

By the end of the 19By the end of the 19thth century,it was noted thatcentury,it was noted thatbactericidal, bacteriostatic and hemolytic action ofbactericidal, bacteriostatic and hemolytic action ofthe appropriate antibodies required thethe appropriate antibodies required the

participation of a heat labile component present inparticipation of a heat labile component present inthe normal serum of humans and animals.the normal serum of humans and animals.

Buchner 1Buchner 1stst observed the bactericidal effect ofobserved the bactericidal effect ofserum was destroyed by heating at 55 C for 1 hr.serum was destroyed by heating at 55 C for 1 hr.

Pfieffer observed that cholera vibrios were lysedPfieffer observed that cholera vibrios were lysedwhen injected intraperitonially into specificallywhen injected intraperitonially into specificallyimmunised guniea pigs(bacteriolysisimmunised guniea pigs(bacteriolysis--PfieffersPfieffersphenomenon).phenomenon).


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Bordet established that immunologicalBordet established that immunologicalbacteriolysis and hemolysis required 2 factorsbacteriolysis and hemolysis required 2 factors1.a heat stable antibody and1.a heat stable antibody and

2. a heat labile factor called Alexine.2. a heat labile factor called Alexine.

The name Alexine was now replaced asThe name Alexine was now replaced asCOMPLEMENT by Ehrlich as it complementedCOMPLEMENT by Ehrlich as it complementedthe action of the antibody.the action of the antibody.

Bordet and Gengou described the ComplementBordet and Gengou described the Complementfixation test, the serological test to detectfixation test, the serological test to detectcomplement.complement.


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The other immunological test to detectThe other immunological test to detect

complement is Immuno adherence test.complement is Immuno adherence test.

The system belongs to the group of BiologicalThe system belongs to the group of Biological

Effector Mechanism called as Triggered enzymeEffector Mechanism called as Triggered enzyme

cascade which also includes Coagulation,cascade which also includes Coagulation,

Fibrinolytic and Kinin system. They areFibrinolytic and Kinin system. They are

advantageous.advantageous. Ex;Each enzyme in the cascade is able to activateEx;Each enzyme in the cascade is able to activate

many molecules of the succeeding componentmany molecules of the succeeding component

providing for amplification of the response at eachproviding for amplification of the response at each

step.step.


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Every step has its own control mechanismsEvery step has its own control mechanisms

so that the cascade can be regulated withso that the cascade can be regulated with

precision.precision.


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PROPERTIESPROPERTIES

Present in the sera of all mammals and also in thatPresent in the sera of all mammals and also in thatof most lower animals including birds,amphibiansof most lower animals including birds,amphibiansand fishes.and fishes.

It is a nonIt is a non--specific serological reagent in thatspecific serological reagent in thatcomplement from 1 species can react withcomplement from 1 species can react withantibody from other species though, the efficiencyantibody from other species though, the efficiencyof the reaction is influenced by the taxonomicof the reaction is influenced by the taxonomic

distance between the species.distance between the species. Complement constitutes about 5% of normalComplement constitutes about 5% of normal

serum proteins and is not increased as a result ofserum proteins and is not increased as a result ofimmunisation.immunisation.


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Though some of its components are heat stable,Though some of its components are heat stable,

complement as a whole is heat labile.complement as a whole is heat labile.

It is denatured spontaneously slowly at RT andIt is denatured spontaneously slowly at RT and

being destroyed in 30 mins at serum thus deprivedbeing destroyed in 30 mins at serum thus deprived

of its complement activity is said to be inactivated.of its complement activity is said to be inactivated.

It ordinarily does not bind to free antigen orIt ordinarily does not bind to free antigen or

antibody but only to antibody which has combinedantibody but only to antibody which has combinedwith its antigen.with its antigen.

Though some of its components are heat stable,Though some of its components are heat stable,

complement as a whole is heat labile.complement as a whole is heat labile.

It is denatured spontaneously slowly at RT andIt is denatured spontaneously slowly at RT and

being destroyed in 30 mins at serum thus deprivedbeing destroyed in 30 mins at serum thus deprived

of its complement activity is said to be inactivated.of its complement activity is said to be inactivated.

It ordinarily does not bind to free antigen orIt ordinarily does not bind to free antigen or

antibody but only to antibody which has combinedantibody but only to antibody which has combinedwith its antigen.with its antigen.


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Fixation of the complement to the AgFixation of the complement to the Ag--AbAb

complex leads to the activation of classicalcomplex leads to the activation of classical

C pathway.C pathway. All classes of Ig do not fix C. Only IgM 3,1All classes of Ig do not fix C. Only IgM 3,1

and 2 fix C but not IgG4, IgD, IgA or IgE.and 2 fix C but not IgG4, IgD, IgA or IgE.

It binds at the Fc portion of the Ig molecule.It binds at the Fc portion of the Ig molecule. Fixation of C is not influenced by the natureFixation of C is not influenced by the nature

of Ag, but only by the class of the Ig.of Ag, but only by the class of the Ig.


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COMPONENTSCOMPONENTS

20 Chemically and Immunologically distinct serum20 Chemically and Immunologically distinct serumproteins comprising the complement components,proteins comprising the complement components,the Properdin system and the control proteins.the Properdin system and the control proteins.

It is a complex of 9 different fractions C1 to C9.It is a complex of 9 different fractions C1 to C9. C1 present in the serum as a calcium ionC1 present in the serum as a calcium ion

dependent complex, which on chelation withdependent complex, which on chelation withEDTA yields 3 protein subunits called as C1q, r s.EDTA yields 3 protein subunits called as C1q, r s.

So C is made up of 11 proteins totally.So C is made up of 11 proteins totally.

C fractions are named C1 to C9 in the sequenceC fractions are named C1 to C9 in the sequenceof the cascading reaction except that C4 comesof the cascading reaction except that C4 comesafter C1 and before C2.after C1 and before C2.


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Erythrocyte Antibody complex called as EA, theErythrocyte Antibody complex called as EA, theproduct is called as EAC, followed by theproduct is called as EAC, followed by thecomponents that have reacted ex. EAC14235 orcomponents that have reacted ex. EAC14235 or

EAC1EAC1--5.5. When C component acquires enzymatic or otherWhen C component acquires enzymatic or other

biological activity, it is indicated by a bar over thebiological activity, it is indicated by a bar over thecomponent number. ex: enzymatically activatedcomponent number. ex: enzymatically activated

C1 is shown as C1.C1 is shown as C1. When it is inactivated, it is indicated by a prefix iWhen it is inactivated, it is indicated by a prefix i

iC3b.iC3b.

Fragments cleaved indicated in small lettersFragments cleaved indicated in small letters

smaller fragment a, and a larger fragment b.smaller fragment a, and a larger fragment b.


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C ACTIVATIONC ACTIVATION

Activation of the C system initiated either byActivation of the C system initiated either by

AgAg--Ab complexes or by a variety of nonAb complexes or by a variety of non--

immunologic molecules.immunologic molecules.


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Sequential activation of C componentsSequential activation of C components

occur through 3 pathways namely:occur through 3 pathways namely:


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1.1. The Classical pathwayThe Classical pathway

2.2. The Alternative pathwayThe Alternative pathway

3.3. The Lectin pathwayThe Lectin pathway


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The classical pathwayThe classical pathway

It is named so because it was the 1It is named so because it was the 1stst to beto be

identifiedidentified

EA+C1 EAC1+C4 EAC14b+C2

C3+EAC14b2aEAC14b2a3bEAC14b2a3b5

b67

EAC14b2a3b5b6789 CYTOLYSIS

C1q,r,s

Ca+

C4a

Mg+ C2b

C3aC5,6,7

C8,9

c5aMAC


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Classical pathway consists of the followingClassical pathway consists of the following

steps;steps;


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Binding of C1 to AgBinding of C1 to Ag--Ab complex. The recognitionAb complex. The recognitionunit of C1 is C1q, which reacts with Fc piece ofunit of C1 is C1q, which reacts with Fc piece ofthe bound IgM or IgG. C1q binding in thethe bound IgM or IgG. C1q binding in the

presence of calcium ions leads to sequentialpresence of calcium ions leads to sequentialactivation of C1r, s.activation of C1r, s.

Activated C1s is an esterase. 1 molecule ofActivated C1s is an esterase. 1 molecule ofwhich can cleave several molecules of C4.C4 iswhich can cleave several molecules of C4.C4 is

split into C4a, which is an anaphylatoxin, andsplit into C4a, which is an anaphylatoxin, andC4b, which binds to cell membranes along withC4b, which binds to cell membranes along withC1.C1.


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C14b in the presence of Mg+ ions cleaves C2C14b in the presence of Mg+ ions cleaves C2into C2a, which remains bound to C4b and C2binto C2a, which remains bound to C4b and C2bis released. C4b2a has enzymatic activity and isis released. C4b2a has enzymatic activity and is

referred to as the CLASSICAL PATHWAY C3referred to as the CLASSICAL PATHWAY C3CONVERTASE.CONVERTASE.

C3 convertase splits C3 into 2 fragments: C3aC3 convertase splits C3 into 2 fragments: C3aan anaphylatoxin and C3b which remains cellan anaphylatoxin and C3b which remains cell

bound along with C4b2a and forms a tribound along with C4b2a and forms a tri--molecular complex C4b2a3b which hasmolecular complex C4b2a3b which hasenzymatic activity and is called C5enzymatic activity and is called C5CONVERTASE.CONVERTASE.


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THE MEMBRANE ATTACK phase of

complement activity begins at this stage

with C5 convertase cleaving C5 into C5a,an anaphylatoxin and C5b which

continues the cascade.

C6 and C7 then join to form C567 which

prepares the cell for lysis by C8 and C9

which join the reaction subsequently.


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Most of C567 escape and serve to amplify theMost of C567 escape and serve to amplify thereaction by adsorbing onto unsensitisedreaction by adsorbing onto unsensitisedbystander cells and rendering them susceptible tobystander cells and rendering them susceptible to

lysis by C8 and C9.lysis by C8 and C9. The unbound C567 has chemotactic activity.The unbound C567 has chemotactic activity.

The mechanism of complement mediatedThe mechanism of complement mediatedcytolysis is the production of HOLES on the cellcytolysis is the production of HOLES on the cell

membrane which disrupts the OSMOTICmembrane which disrupts the OSMOTICINTEGRITY of the membrane, leading to theINTEGRITY of the membrane, leading to therelease of the cell contents.release of the cell contents.


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The Alternative C PathwayThe Alternative C Pathway

It is a central process which involves theIt is a central process which involves the

activation of C3.activation of C3.

In the Classical Pathway, activation of C3 isIn the Classical Pathway, activation of C3 isachieved by C42.achieved by C42.

Activation of C3 without prior participation ofActivation of C3 without prior participation of

C142 is known as THE ALTERNATIVEC142 is known as THE ALTERNATIVEPATHWAY.PATHWAY.


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Example of the alternative pathway was theExample of the alternative pathway was thedemonstration by Pillemer of thedemonstration by Pillemer of thePROPERDIN SYSTEM as a group of serumPROPERDIN SYSTEM as a group of serum

proteins contributing to antiproteins contributing to anti--microbialmicrobialdefense without requiring specific antibody.defense without requiring specific antibody.

Activator ZYMOSAN, a polysaccharide fromActivator ZYMOSAN, a polysaccharide from

yeast cell wall. Other activators includeyeast cell wall. Other activators includeBactericidal endotoxin, IgA, IgD, cobraBactericidal endotoxin, IgA, IgD, cobravenom etc.venom etc.


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C3b activator complex is inactivated by 2C3b activator complex is inactivated by 2

protein factors H and I.protein factors H and I.

Bound C3b protected from such inactivationBound C3b protected from such inactivationinteracts with a magnesium dependentinteracts with a magnesium dependent

serum protein factor called Factor B which isserum protein factor called Factor B which is

called C3 Procalled C3 Pro--activator.activator.


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C3b+B C3bB+ Factor D (C3 proactivator

convertase)

P+C3bBb

Ba

Alternate

pathway C3convertase

stablize

Further steps in classical

pathway follows


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The Lectin PathwayThe Lectin Pathway

Lectins are proteins that recognize and bindLectins are proteins that recognize and bind

to specific carbohydrate targets (mannose).to specific carbohydrate targets (mannose).

Like Alternative pathway it does not dependLike Alternative pathway it does not dependon antibody for its activation, however theon antibody for its activation, however the

mechanism is more like that of classicalmechanism is more like that of classical

pathway, because after initiation it proceedspathway, because after initiation it proceeds

through the action of C4 and C2, to producethrough the action of C4 and C2, to produce

a C5 CONVRETASE.a C5 CONVRETASE.


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The Lectin pathway is activated by theThe Lectin pathway is activated by thebinding of mannose binding lectin (MBL) tobinding of mannose binding lectin (MBL) tomannose residues on glycomannose residues on glyco--proteins orproteins or

carbohydrates on the surface of microcarbohydrates on the surface of microorganisms.organisms.

MBL is an ACUTE PHASE PROTEINMBL is an ACUTE PHASE PROTEIN

produced in inflammatory response.produced in inflammatory response. Its function is similar to that of C1q which itIts function is similar to that of C1q which it

resembles in structure.resembles in structure.


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After MBL binds to the surface of a cell or aAfter MBL binds to the surface of a cell or a

pathogen, MBL assosiated serine proteases,pathogen, MBL assosiated serine proteases,

MASPMASP--1 and MASP1 and MASP--2, bind to MBL.2, bind to MBL.

The active complex formed by this associationThe active complex formed by this association

causes cleavage and activation of C4 and C2.causes cleavage and activation of C4 and C2.

The MASPThe MASP--1 and MASP1 and MASP--2 proteins have structural2 proteins have structural

similarity to C1r and C1s and mimic their activities.similarity to C1r and C1s and mimic their activities.


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This means of activating the C2This means of activating the C2--C4C4

components to form a C3 convertasecomponents to form a C3 convertase

without need for specific Ab bindingwithout need for specific Ab bindingrepresents an important innate defenserepresents an important innate defense

mechanism comparable to the alternativemechanism comparable to the alternative

pathway, but utilizing the elements of thepathway, but utilizing the elements of the

classical pathway except for the C1classical pathway except for the C1proteins.proteins.


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MBL binds

foreign surfaceMASP-1 AND 2 bind

MBL, generateactivated C1like

complex

Bind to activatedC1 & the cascade

continues


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REGULATIONREGULATION

Uncontrolled C activity can cause not onlyUncontrolled C activity can cause not only

exhaustion of the complement system butexhaustion of the complement system but

also serious damage to tissues.also serious damage to tissues. Several inbuilt control mechanisms regulateSeveral inbuilt control mechanisms regulate

the complement cascade at different steps.the complement cascade at different steps.

Regulation is by means of 2 mechanisms:Regulation is by means of 2 mechanisms:


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INHIBITORSINHIBITORS which halt the C activitywhich halt the C activity

INACTIVATORSINACTIVATORS which destroy C proteinswhich destroy C proteins


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InhibitorsInhibitors

Normal serum contains inhibitor of C1Normal serum contains inhibitor of C1esterase. This heat labile alphaesterase. This heat labile alphaneuraminoglycoprotein also inhibits manyneuraminoglycoprotein also inhibits many

other esterases found in blood such asother esterases found in blood such asPlasmin, Kininogen and the HagemanPlasmin, Kininogen and the Hagemanfactors.factors.

S protein in normal serum binds to C567S protein in normal serum binds to C567and modulates the cytolytic action of theand modulates the cytolytic action of themembrane attack complex.membrane attack complex.


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InactivatorsInactivators

A serum beta globulin called as factor I, providesA serum beta globulin called as factor I, provideshomeostatic control of C3 activation, particularlyhomeostatic control of C3 activation, particularlyby alternative pathway.by alternative pathway.

Factor H along with Factor I modulates C3Factor H along with Factor I modulates C3activation.activation.

Anaphylatoxin inactivator is an alpha globulin thatAnaphylatoxin inactivator is an alpha globulin thatenzymatically degrades C3a, C4a, and C5a whichenzymatically degrades C3a, C4a, and C5a whichare anaphylatoxins released during the Care anaphylatoxins released during the C

cascade.cascade. C4 binding protein controls the activity of cellC4 binding protein controls the activity of cell

bound C4b.bound C4b.


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Biological activityBiological activity

OPSONISATIONOPSONISATION Cells, AgCells, Ag--Ab complexesAb complexes

and other particles are phagocytosed muchand other particles are phagocytosed much

more efficiently in the presence of C3bmore efficiently in the presence of C3bbecause of the presence of C3b receptorsbecause of the presence of C3b receptors

on the surface of many phagocytes.on the surface of many phagocytes.

CHEMOTAXISCHEMOTAXIS C5a stimulates movementC5a stimulates movement

of neutrophils and monocytes towards sitesof neutrophils and monocytes towards sites

of Ag deposition.of Ag deposition.


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ANAPHYLATOXINSANAPHYLATOXINS C5a,C4a and C5aC5a,C4a and C5acan produce increased vascularcan produce increased vascularpermeability and smooth musclepermeability and smooth muscle

contraction. C3a and C5a also stimulatecontraction. C3a and C5a also stimulatemast cells to release Histamine.mast cells to release Histamine.

CYTOLYSISCYTOLYSIS Insertion of C56789 complexInsertion of C56789 complex

into the cell surface leads to killing or lysis ofinto the cell surface leads to killing or lysis ofmany types of cells, including Erythrocytes,many types of cells, including Erythrocytes,Bacteria and Tumor cells.Bacteria and Tumor cells.


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BIOSYNTHESIS OF CBIOSYNTHESIS OF C

Intestinal epitheliumIntestinal epithelium C1C1

MacrophagesMacrophages C2,C4C2,C4

SpleenSpleen C5,C8C5,C8LiverLiver C3,C6,C9C3,C6,C9

C4,C3,C5,C6C4,C3,C5,C6 some extent found in acutesome extent found in acute

phase of inflammationphase of inflammation


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DEFICIENCY OF C SYSTEMDEFICIENCY OF C SYSTEM

C1 inhibitor deficiencyC1 inhibitor deficiency heriditary angioneuroticheriditary angioneuroticedema (may be fatal)edema (may be fatal)

C2 deficiencyC2 deficiency serious pyogenic bacterial infectionserious pyogenic bacterial infection

Membrane attack complex deficiencyMembrane attack complex deficiency enhancesenhancessusceptibility to Neisserial infectionsusceptibility to Neisserial infection

Deficiency in components of the alternativeDeficiency in components of the alternativepathwaypathway ex: Properdin deficiency is associatedex: Properdin deficiency is associated

with greater susceptibility to Meningococcalwith greater susceptibility to Meningococcaldiseasedisease

C5 to C8C5 to C8 BacterimiaBacterimia


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Generalized deficiency of the complementGeneralized deficiency of the complement

components lead to the lack or impairmentcomponents lead to the lack or impairment

of nonof non--specific defense system of the body.specific defense system of the body.

2941 Complement System

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