Chapter 9: Complement
Complements are essential proteins. These are called complement because it complements the antibacterial activity of some of the antibodies. Compliments are given numerical names as C1-C9, which are 20 including subunits. Their concentration is 3-4 G/L in the blood.
Activation of complements leads to inflammation and localize the antigen or cause lysis. These are not increased by infection or other antigens.
But IL-1 and γ-interferon increase the synthesis.
Site of Action
The main site is cell surface or other biological membranes.
Changes by Activation of Complement
- There are ultrastructural changes in the cell membrane.
- There are changes in the electrical charges of the cell membrane.
- There is swelling of the membrane.
- Ultimately there are circular holes 8-12 nm in diameter.
Site of Formation
The complement may be synthesized in the intestinal epithelium, macrophagic cells, and spleen.
|Complement||Site of Synthesis|
|C1||Synthesized in Epithelial cells and fibroblasts.|
|C3 4 6 7 8 9||The liver is the major site of formation.|
|C1 2 3 4 5||Synthesized by macrophagic cells.|
|Factor B, D, H, P||Synthesized by macrophagic cells.|
Table X – Site of Complement Formation
Fetus: Starts forming complement by the second month of pregnancy.
After Activation Complement is cleaved into:
- Larger molecule labeled as “b”. It leads to further activation of the chain reaction.
- A smaller molecule labeled as “a” it promotes inflammation and produces pharmacological action.
- Further proteolysis gives inactive component labeled as “iC3 b”.
- Complement is catabolized in the body 1-3% per hour.
The following example shows the role of Antibodies and complement in the process of Bacteriolysis and Agglutination.
Complements are present in an inactive form in blood circulation, once activated they give chain reaction like blood coagulation factors.
- Classical pathway
- Alternative pathway
The classical pathway may be activated by:
- Immunological stimuli-like Ag & Ab complex, tissue injury or aggregated IgG.
- non-immunological stimuli like CRP, DNA, Trypsin, E. Coli, Salmonella, viruses, endotoxin and urate crystals.
Alternative Pathway may be activated by:
- Immunological stimuli like aggregated IgA, sometimes IgG.
- Non- immunological stimuli are lipopolysaccharides (endotoxin), Trypsin & trypsin-like enzyme, cobra venom. Parasite & teichoic acid of gram-positive bacteria.
The following diagram gives a summary of the Classical pathway and Alternative pathway.
Activation of Complement leads to:
- Lysis of Bacteria, viruses, and cells.
- Mediate acute inflammation.
- This leads to the release of histamine.
- It helps in opsonization and phagocytosis.
- It has a regulatory role and it regulates acute inflammation in the immune response.
Various Complement-Proteins lead to:
- Vasodilatation at the site of inflammation.
- Increase adherence of phagocytic cells to blood vessels endothelium.
- It directs movements of phagocytic cells to the area of inflammation.
- Ultimately clears of the infection.
The outcome of Complement Activation:
- Recruitment of acute inflammatory cells.
- Opsonization of pathogens.
- Ultimately killing of pathogens.
Suppose this reaction is taking place on the cell membrane of the RBC and sequence of the event may be as follows:
This can also be explained in the following diagram in a more simple way.
In the classic pathway antigen and antibody complex is recognized by the complement (C1q) and it starts chain reaction by stimulatingC1s and C1r which will stimulate C4 and C2. Then there is the stimulation of C3 C5 C6, 7, and ultimately C9.
This will give rise to MAC (membrane attacking complex) and leads to the osmotic death of the target cells.
Details of Classical Pathway Activation
Now the details of the classical pathway can be elaborated when suppose the reaction is occurring on the cell membrane where there is an antigenic determinant (Epitope).
- C1q recognize the Ag-Ab complex and then activate C1r.
- C1r– cleaves C1S (C1S–).
- C1S cleaves C4 to C4b and C4a.
- C1S also cleaves C2 to C2a and C2b.
- C4b and C2b act as C3-convertase and cleaves C3 to C3a & C3b.
- C4b C2b C3b acts as a C5 convertase and cleaves C5 into C5a & C5b.
- C5b binds to C6 and make a labile reactive site for C7.
- C5b 6, 7 is highly lipophilic and binds to the membrane where it acts high-affinity receptors for C8.
- C8 has three chains α, β, and γ where g
- γ-chains insert into the membrane.
- C5b678 polymerizes C9 forming the MAC (membrane attacking complex), which gives rise to hole formation in the cell-membrane Like drill machine.
- C5b678 makes a 3nm hole and cells become leaky.
- C5b6789 makes transmembrane holes, 15-16nm long and 8-12nm in diameter. This leads to the passage of electrolytes and water and causes osmotic lysis (death).
Functions of Various Components
- It is a weak anaphylatoxin and weak mediator of Inflammation.
- It increases vascular permeability.
- It causes contraction of smooth muscles.
- Some say that functions are unknown.
- It is a strong anaphylatoxin.
- It leads to an increase in vascular permeability and smooth muscle contraction.
- This leads to the release of vasoactive amines.
- Leads to release of lysosomal enzyme.
- It is chemotactic.
- Functions just like C3a.
- It increases neutrophil activity.
- More potent chemotactic factor than C3a.
These are C3a, C5a, and C4a. This name is because of their role in anaphylaxis, where these directly activate mast cells and basophils, through their receptor for C5a, and lesser extent C3a.
|Complement Protein||Complement Fraction|
|Binding to Ag-Ab complex (Recognition unit)||C1q|
|Activating enzymes||C1r, C1s, C2b, Bb & D|
|Membrane binding proteins and Opsonin||C3b & C4b|
|Mediate inflammation||C5a, C3a, C4a|
Table XI – Complement proteins
Check Mechanism or Inhibitors of the Classical Pathway
Nature has given a check mechanism, which controls unlimited damage caused by activated complement system even to the innocent bystander cells. These inhibitors are:
- C1-inhibitor: It is a glycoprotein and also known as C1 esterase. It destroys C1r, C1s.
- The factor I, It is proteolytic enzymes, it cleaves C3b into iC3b (inactive form). It also destroys C4b.
- Vitronectin (S-protein): Prevents binding of C5, 6, 7 to the membrane.
- Decay Accelerating Factor (DAF): Prevents activation of C4.
- C4 binding protein: Prevents activation of C4.
- Factor H: It binds C3b and accelerates the destructive action of factor I.
This is a primitive defense system. In this pathway, there is a bypass of C1, 4, 2 and there is direct stimulation of C3.
After the stimulation of C3 then there is a common pathway with the help of factor B and factor D., In the end, there is the formation of MAC (membrane attacking complex).
Binds to pathogen surface.
Binds factor B for cleavage by D.
|C3bBb||It is C3 convertase.|
|C3b2Bb||It is C5 convertase.|
|Ba||It is a small fragment and function is unknown|
|Bb||It is an active enzyme and C3 convertase.|
|C3bBb||It is C3 convertase.|
|C3b2Bb||It is C5 convertase|
|C3bBb||It is C5 convertase.|
|Factor D||Plasma serine protease cleaves factor B when it is bound To C3b to Ba and Bb.|
Table XII – Proteins of the alternative pathway
Positive Regulators for Alternative Pathways are certain substances, which enhance and give positive regulation of the alternative pathways. These are:
- Factor P (Properdin).
- Cobra Venom Factor (CoVF):- This complexes with Bb (CoVFBb) which acts as C3/C5 convertase. It is resistant to factor H & I.
Inhibitors of Alternative Pathways
These are natural check system or inhibitors.
- Factor H which binds to C3b.
- The factor I which inactivate and cleaves C3b.
Complement Activation is an Amplification Process.
- Roughly 1200C3b opsonin can deposit with a single molecule of IgM.
- Single IgM molecule cleaves one C1q and C1S and these will cleave 100 C4 molecules.
- 20 or 50 C4b cleaves about 100 C2 molecules.
- Many C5 initiates MAC (membrane-attacking complex).
The most important function of complement is to facilitate the uptake and destruction of the pathogen by phagocytic cells. This occurs by specific recognition of bound complement components by complement receptors (CR) present on phagocytic cells.
Similar complement receptors are present on RBC and RBC play an important role in the clearance of soluble Ag + Ab complexes from the circulation by carrying these (Ag + Ab) complexes to a reticuloendothelial system like the spleen.
|C3a & C5a Receptors||
Present on Mast cells and Basophils.
Receptors identified on Neutrophil, Monocytes, B and Null cells.
|C1q||are present on Mast cells and Basophils|
|C3b||Receptors identified on Neutrophil, Monocytes, B and Null cells.|
|C4b||Present on phagocytic cells, lymphocytes, and RBC.|
Table XIII – Receptors present on various cells
|Receptor||Binds to||Present on||Function|
|CR1||C3bC4b||RBC, Macrophages, Monocytes, Neutrophils & B-cells.||Promote C3b & C4b decay, stimulate phagocytes, RBC transport of immune complexes|
|B-cells||It is part of B-L co-receptor & EBV receptor|
|CR3||iC3b||Macrophages, Monocytes, Neutrophils||Stimulate phagocytes|
|CR4||iC3b||Macrophages, Monocytes, Neutrophils||Stimulate phagocytes|
|C1q||C1q||B-cells, Macrophages, Monocytes, platelets and endothelial cells||Binds immune complexes to Monocytes|
Table XIV – Various receptors
Biological Functions of Complements
|C1 and C4||Virus neutralization.|
|C3b||Immune adherence, phagocytosis & take part in Arthus reaction.|
|C3a, C5a, and C6 C7||Chemotaxis|
|C5 to C9||Lysis|
|C8 and C9||Cytotoxic action|
Human Diseases Associated with Complement Deficiency
Deficiency of any component of complement leads to various diseases e.g.
|C1q deficiency||SLE, Nephritis & hypogammaglobulinemia|
|C1r deficiency||Renal diseases, SLE, recurrent infection and Rheumatoid arthritis|
|C3 deficiency||Recurrent infections (pyogenic)|
|C5 deficiency||Recurrent infections + Gonococcal infections and SLE|
|C6 deficiency||Recurrent infections + Meningococcal infection|
|C7 deficiency||Recurrent infections + Glomerulonephritis|
|C8 deficiency||Recurrent infections + Gonococcal & Meningococcal infections|
Complement Fixation Test
The complement may be used as a diagnostic tool. The basic principle is its activity which leads to hemolysis of RBC. In this test, we need:
- Serum of the patients.
- Indicator system which consists of antibody-coated RBC.
- Known antigen.
This test is used to find Unknown Antibody or Antigen. The following diagram shows the basic mechanism.
This test may be manipulated by making dilution of serum or complement or one can have known antibody and unknown antigen or vice versa. Even the titer of antibody or concentration of complement may be estimated by serial dilution.
Use of Complement fixation test
This test can be used to diagnose various diseases e.g Pneumococcal pneumonia, syphilis and etc. with the help of this test one can quantitate antigen or antibody. This test was very famous for the diagnosis of syphilis.