Sherko A Omer MB ChB, MSc., PhD
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1 Immunology IMMUNOLOGY Sherko A Omer MB ChB, MSc., PhD
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THE COMPLEMENT SYSTEM Complement system includes more than 30 soluble and cell-bound proteins. The biological activities of this system affect both innate and acquired immunity. They are proteins or glycoproteins synthesized mainly by hepatocytes, although significant amounts are also produced by blood monocytes, tissue macrophages, and epithelial cells of the gastrointestinal and genitourinary tracts.
Transcript of Sherko A Omer MB ChB, MSc., PhD
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Immunology
IMMUNOLOGY
Sherko A Omer MB ChB, MSc., PhD
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Immunology
THE COMPLEMENT SYSTEM Complement system includes more than 30 soluble and cell-bound proteins.
The biological activities of this system affect both innate and acquired immunity.
They are proteins or glycoproteins synthesized mainly by hepatocytes, although significant amounts are also produced by blood monocytes, tissue macrophages, and epithelial cells of the gastrointestinal and genitourinary tracts.
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Immunology
THE COMPLEMENT SYSTEM These components constitute 5% (by weight) of the serum globulin fraction.
Most circulate in the serum in functionally inactive forms as proenzymes, or zymogens, which are inactive until proteolytic cleavage, which removes an inhibitory fragment and exposes the active site.
The complement-reaction sequence starts with an enzyme cascade.
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THE COMPLEMENT SYSTEM NomenclatureClassical pathway components are labelled with a C and a number (e.g., C1, C3).
Alternative pathway components are lettered (e.g., B, P, D). Some components are called factors (e.g., factor B, factor D).
Activated components or complexes have a bar over them to indicate activation (e.g., C4b2a).
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THE COMPLEMENT SYSTEM NomenclatureCleavage fragments are designated with a small letter after the component (e.g., C3a and C3b are fragments of C3).
Inactive C3b is designated iC3b.
Polypeptide cell membrane receptors for C3 are abbreviated CR1, CR2, CR3, and CR4.
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THE COMPLEMENT SYSTEM Complement activation can occur by three different mechanisms:
• The classical pathway• The alternative pathway• The lectin pathway
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THE COMPLEMENT SYSTEM
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THE CLASSICAL PATHWAY Classical pathway is initiated byFormation of soluble antigen-antibody complexes (immune complexes) or with the binding of antibody to antigen on a suitable target, such as a bacterial cell.
IgM and certain subclasses of IgG (human IgG1, IgG2, and IgG3) can activate the classical complement pathway.
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Structure of the C1 macromolecular complex. (a) Diagram of C1qr2s2 complex. A C1q molecule consists of 18 polypeptide chains arranged into six triplets, each of which contains one A, one B, and one C chain. Each C1r and C1s monomer contains a catalytic domain with enzymatic activity and an interaction domain that facilitates binding with C1q or with each other. (b) Electron micrograph of C1q molecule
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CLASSICAL PATHWAY
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CLASSICAL PATHWAY
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CLASSICAL PATHWAY
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CLASSICAL PATHWAY
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THE ALTERNATIVE PATHWAYThe alternative pathway is initiated by pathogens and particles of microbial origin (lipopolysaccharides from gram-negative bacteria, teichoic acid from gram-positive cell walls, fungal and yeast cell walls, some viruses and parasites)
Non pathogen materials (cobra venom factor, nephritic factor, heterologous erythrocytes and pure carbohydrates) and aggregated IgA.
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THE ALTERNATIVE PATHWAY
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THE ALTERNATIVE PATHWAY
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THE LECTIN PATHWAYThe lectin pathway is activated by the binding of mannose-binding lectin (MBL) to mannose residues on glycoproteins or carbohydrates on the surface of microorganisms including certain Salmonella, Listeria, and Neisseria strains, as well as Cryptococcus neoformans and Candida albicans.
MBL is an acute phase protein produced in inflammatory responses. Its function in the complement pathway is similar to that of C1q.
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THE LECTIN PATHWAYThe lectin pathway, like the alternative pathway, does not depend on antibody for its activation.
However, the mechanism is more like that of the classical pathway, because after initiation, it proceeds, through the action of C4 and C2, to produce a C5 convertase, this mechanism also called the MB Lectin pathway or mannan-binding lectin pathway.
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MEMBRANE ATTACK COMPLEX
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MEMBRANE ATTACK COMPLEX
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MEMBRANE ATTACK COMPLEX
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REGULATION OF COMPLEMENT SYSTEM
The classical pathway is regulated by C1 inhibitor (C1 Inh), a serine esterase inhibitor that causes C1r2s2 to dissociate from C1q preventing further activation of C4 or C2. C1Ihb absence leads to a condition called hereditary angioedema.
Factor J is a cationic glycoprotein that also inhibits C1 activity.
C4-binding protein (C4bBP) disassembles the C4b2a complex, allowing factor I to inactivate C4b.
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REGULATION OF COMPLEMENT SYSTEM
Factor H or decay- accelerating factor (DAF) compete with factor B for binding to C3b (e.g., to produce C3bH), decreasing the half-life of the C3bBb complex and causing dissociation of the complex into C3b and Bb.
Factor I acts on C3bH to degrade C3b.
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REGULATION OF COMPLEMENT SYSTEM
The MAC is regulated by S protein which binds soluble C5b67 and prevents its insertion into cell membrane.
Membrane bound factors such as homologous restriction factor (HRF) and membrane inhibitor of reactive lysis (MIRL) bind to C5b678 on autologous cells, blocking binding of C9.
Anaphylatoxin inactivator are soluble factors that inactivates anaphylatoxin activity of C3a, C4a, and C5a by carboxypeptidase N removal of C-terminal Arg.
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REGULATION OF COMPLEMENT SYSTEM
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REGULATION OF COMPLEMENT SYSTEM
MCP: membrane cofactor protein
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REGULATION OF COMPLEMENT SYSTEM
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BIOLOGIC CONSEQUENCES OF COMPLEMENT ACTIVATION
Cell lysis is achieved through MAC, The MAC formed by complement activation can lyse gram-negative bacteria, parasites, viruses, erythrocytes, and nucleated cells.
Cleavage products of complement components mediate inflammation. C3a, C4a and C5a have anaphylatoxin activity. Anaphylatoxins, bind to receptors on mast cells and blood basophils and induce degranulation, with release of histamine and other pharmacologically active mediators.
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BIOLOGIC CONSEQUENCES OF COMPLEMENT ACTIVATION
The anaphylatoxins also induce smooth muscle contraction and increased vascular permeability.
C3a, C5a, and C5b67 each can induce monocytes and neutrophils to adhere to vascular endothelial cells, extravasate through the endothelial lining of the capillary, and migrate toward the site of complement activation in the tissues.C5a is most potent in mediating these processes.
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BIOLOGIC CONSEQUENCES OF COMPLEMENT ACTIVATION
C3b is the major opsonin of the complement system, although C4b and iC3b also have opsonizing activity.
The amplification that occurs with C3 activation resultin a coating of C3b on immune complexes and particulate antigens.
Phagocytic cells, as well as some other cells, express complement receptors (CR1, CR3, and CR4) that bind C3b, C4b, or iC3b.
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CLEARING IMMUNE COMPLEXES FROM CIRCULATION
The coating of soluble immune complexes with C3b is thought to facilitate their binding to CR1 on erythrocytes.
Erythrocytes play an important role in binding C3b-coated immune complexes and carrying these complexes to the liver and spleen.
In these organs, immune complexes are stripped from the red blood cells and are phagocytosed, thereby preventing their deposition in tissues.
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BIOLOGIC CONSEQUENCES OF COMPLEMENT ACTIVATION
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BIOLOGIC CONSEQUENCES OF COMPLEMENT ACTIVATION
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COMPLEMENT-BINDING RECEPTORSReceptor Major ligand Activity Cellular Distribution
CR1 (CD35) C3b, C4b Blocks formation of C3, convertase; binds immune complexes to cells
Erythrocytes, neutrophils, monocytes, macrophages, eosinophils, follicular dendritic cells, B cells, some T cells
CR2 (CD21) C3d, C3dg, iC3b Part of B-cell co receptor; binds Epstein-Barr virus
B cells, follicular dendritic cells, Some T cells
CR3 (CD11b/18) CR4 (CD11c/18)
iC3b Bind cell-adhesion molecules on neutrophils, facilitating their extravasation; bind immune complexes, enhancing their phagocytosis
Monocytes, macrophages, neutrophils, natural killer cells , some T cells
C3a/C4a receptor C3a, C4a Induces degranulation of mast cell and basophils
Mast cells, basophils, granulocytes
C5a receptor C5a Induces degranulation of mast cells and basophils,
Mast cells, basophils, granulocytes, monocytes, macrophages, platelets, endothelial cells
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COMPLEMENT ASSAYS
Complement protein levels assayed by: NephelometryAgar gel diffusionRadial immunodiffusionELISA.
Functional assays include hemolytic assays to measure functional activity of specific components of either pathways.
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COMPLEMENT ASSAYS
The total hemolytic complement assay (CH50) measures the ability of the classical pathway and the MAC to lyse sheep RBC to which antibodies has been attached.
The alternative pathway CH50 measures the ability of the alternative pathway and the MAC to lyse rabbit RBC.
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ANTIGEN-ANTIBODY INTERACTIONA bimolecular association similar to an enzyme-substrate interaction.
It does not lead to an irreversible chemical alteration in either the antibody or the antigen.
The association between an antibody and an antigen involves various noncovalent interactions between the antigenic determinant (epitope), of the antigen and the paratope region of the antibody, (VH/VL) domain, particularly the hypervariable regions, or complementarity-determining regions (CDRs).
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ANTIGEN-ANTIBODY INTERACTION
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ANTIGEN-ANTIBODY INTERACTIONThe antigen antibody complex is not bounded firmly and may dissociate spontaneously
Binding is affected by environmental factors like pH in which binding is weaker in pH <4 or >10, increased salt concentration leads to weaker binding.
Temperatures of 50-55 C cause stronger binding.
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ANTIGEN-ANTIBODY INTERACTIONThe noncovalent binding is critically dependent on the distance (d) between the interacting groups.
As force is proportional to 1/d2 for electrostatic force and 1/d7 for Vander Waals force, so accordingly there must be a high degree of fitness between antigen and antibody (complementary binding) in order to these forces come to work.
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ANTIGEN-ANTIBODY INTERACTION
Affinity
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ANTIGEN-ANTIBODY INTERACTIONThe combined strength of the noncovalent interactions between a single antigen-binding site on an antibody and a single epitope is the affinity of the antibody for that epitope.
Low-affinity antibodies bind antigen weakly and tend to dissociate readily, whereas high-affinity antibodies bind antigen more tightly and remain bound longer.
In some biological reactions high affinity is superior to low affinity like in haemagglutination, haemolysis, complement fixation and enzyme inactivation.
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ANTIGEN-ANTIBODY INTERACTIONExperimentally antigen antibody complexes containing low affinity antibody persist longer in circulation and localized in glomerular basement membranes, this may lead to impairment of renal function.
In contrast high affinity antigen-antibody complexes are readily removed from circulation and tend to localize in mesangium of kidney and have little effect on kidney’s function.
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ANTIGEN-ANTIBODY INTERACTIONThe strength with which a multivalent antibody binds a multivalent antigen, avidity, is affected by affinity and valency.
Multivalent means that the molecule has more than one binding sites. A simple IgG molecules is multivalent as it has two antigen binding sites while an antigen may be monovalent (e.g. in hapten) or multivalent.
When an antigen binds an antibody with more than two binding sites the avidity become grater than the sum of individual binding sites (individual affinities).
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ANTIGEN-ANTIBODY INTERACTIONAlthough Ag-Ab reactions are highly specific, in some cases antibody elicited by one antigen can cross-react with an unrelated antigen.
Cross-reactivity occurs if two different antigens share an identical or very similar epitope.
In the latter case, the antibody’s affinity for the cross-reacting epitope is usually less than that for the original epitope.
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ANTIGEN-ANTIBODY INTERACTIONCross-reactivity usually characterised by less avidity than specific reaction which occur between antibody and the original antigen.
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ANTIGEN-ANTIBODY INTERACTION
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ANTIGEN-ANTIBODY INTERACTION
A number of viruses and bacteria have antigenic determinants identical or similar to normal host-cell components.
In some cases, these microbial antigens have been shown to elicit antibody that cross-reacts with the host-cell components, resulting in a tissue-damaging autoimmune reaction.
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ANTIGEN-ANTIBODY INTERACTIONThe bacterium Streptococcus pyogenes, expresses cell-wall proteins called M antigens. Antibodies produced to streptococcal M antigens have been shown to cross-react with several myocardial and skeletal muscle proteins and have been implicated in heart and kidney damage following streptococcal infections.
Some vaccines also exhibit cross-reactivity, vaccinia virus, which causes cowpox, expresses cross-reacting epitopes with variola virus, the causative agent of smallpox.
