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Complement cascade

The complement cascade (aka complement system or just complement) has seven functions -- shown in the table below -- all of which are essential to the immune system.

Complements consists of three different simultaneous pathways of activating the membrane attack complex: the classical pathway, part of adaptive immunity; and alternative and lectin pathways, part of innate immunity. Three different pathways are involved in complement activation: classical pathway, which is stimulate by antigen-antibody complexes; alternative pathway, which spontaneously activates on contact with pathogenic cell surfaces; and mannose-binding lectin pathway, which recognizes mannose sugars which are usually present only on pathogenic cell surfaces.

FunctionOverview
Lysis of CellsThe first complement cascade function discovered, cell lysis begins with pores forming in the cell membrane, and ends with fluid rushing into and bursting the cell (an event called hypotonic cell death). This complement function is not effective against organisms with cell walls, including Gram-positive microbes and fungi.
OpsonizationOpsonization (preparation of cells for phagocytosis) is performed by macrophages and PMNs. A cleave product called C3b is formed by binding of macrophage and PMN F(c) receptors to antibodies. C3b is the primary opsonin, coating antigens to form an antigen-C3b complex. This complex then stimulates phagocytosis by binding phagocytices' complement receptors.
InflammationAnaphylatoxins C3a, C4a and C5a are small peptides produced during the complement cascade, and induce inflammation. Anaphylatoxins bind mast-cell and basophil receptors, induce degranulation to cause smooth muscle contraction and vascular permeability. Also, anaphylatoxins amplify the inflammatory response by inducing synthesis of pro-inflammatory cytokines. C5a is the most potent anaphylatoxin. Along with C3a and C5b67, C5a also is a chemoattractant an activator of white blood cells -- it induces monocytes and neutrophils to bind capillaries' vascular endothelium, extravasate out of the capillaries and migrate to tissue where complement has been activated.
Immune ClearanceImmune clearance is transfer of immune complexes from blood to the spleen and liver. C3b facilitates binding of immune complexes to CR1 on red blood cells. As these red blood cells pass through the spleen and liver, the immune complexes are removed and phagocytosed. In addition, complement helps make immune complexes more soluble.
Enhanced ImmunityC3 is required for optimal expansion of CD8 T cells during systemic viral infections.
Virus NeutralizationComplement helps eliminate virions by inducing them to aggregate, and by coating the capsid.

Classical pathway

In the classical pathway of complement, the F(c) of an antigen-antibody complex binds to the C1q component of C1. The F(c) must be immobilized as part of an antigen-antibody complex -- an antibody enough is not alone to bind C1q and activate C1q's two protease components (C1r and C1s). C1q has low affinity for F(c), so multiple F(c)s must be bound to C1q before C1r and C1s are activated. Once these proteases are activated, they cleave C4 and then C2. Two of the cleavage products -- C4b and C2a -- form a new protease (C3 convertase, aka C4b2a) which cleaves C3 into C3a and C3b. C3 convertase binds to C3b to form a new protease (C5 convertase, aka C4b2a3b) that cleaves C5 into C5a and C5b. C5b is the first protein of the membrane attack complex (MAC).

Because IgM has multiple F(c)s, it is more effective than other antibodies at activating complement. However, IgM must first change its conformation to expose its C1q binding sites. Also, please note that C3a and C5a just float away.

Lectin pathway

The lectin pathway of complement is identical to the classical pathway, except the lectin pathway begins with mannose-binding protein instead of C1. Mannose-binding protein (aka MBP, mannose-binding lectin or MBL) is a primitive and non-clonal form of humoral immunity (immunity mediated by antibodies). Mannose-binding protein binds carbohydrate antigens frequently found on pathogen cell surfaces, but infrequently expressed by host cells. MBP is structurally similar to C1, with a binding region and two proteases, mannan-binding lectin associated proteases (or MASPs for short). Once MBP binds its carbohydrate antigen, the MASPs are activated to cleave C4 and then C2, with subsequent events identical to the classical pathway.

Alternative pathway

Complement Cascade Alternative Fluid Phase

There are two concepts behind the alternative pathway of complement: what occurs when a non-self cell is absent; and what occurs when a non-self cell is present. When a non-self cell is absent (meaning the tissue is healthy) then there is fluid-phase activation. Fluid-phase activation occurs continuously, spontaneously and very slowly. In fluid-phase activation, C3 spontaneously activates via hydrolysis to form C3H2O -- since it is unstable, C3H2O usually reverts to C3. However, if C3H2O encounters Factor B, then the two molecules bind to form a more stable C3H2OB molecule. Factor D then cleaves C3H2OB molecule to yield the enzyme C3H2OBb (aka fluid-phase C3 convertase). C3H2OBb has an active site on Bb; to culminate fluid-phase activation, this active site cleaves C3 into C3a and C3b. Fluid-phase activation is depicted in the figure to the left.

When a non-self cell is present, then a much faster process occurs. C3b binds to the surface of the non-self cell, then Factor B binds to the C3b. This cell-surface C3b-B complex is cleaved by Factor D to form C3b-Bb, a potent C3 convertase which begins cleaving C3 at an enormous rate to yield lots of C3a and C3b. The C3b eventually smothers the cell-surface, marking it for phagocytosis by macrophages, and granulocytes with C3b receptors. As the concentration of Factor B is depleted, a C3b-Bb begins to bind C3b to form the enzyme C3b-Bb-C3b (C5 convertase). This C5 convertase cleaves C5 into C5a and C5b: as in the classical pathway of complement, C3a and C5a just float away; C5b, already stuck to the cell surface, binds other proteins to form the membrane attack complex.

Membrane Attack Complex

The membrane attack complex (MAC) is an extremely potent agent of cell lysis. After cleavage of C5 into C5a and C5b, C5b binds one molecule each of C6, C7 and C8. After this, C5b binds an additional 6-10 molecules of C9 to form the complete membrane attack complex. The large multi-subunit membrane attack complex inserts into the lipid bilayer, allowing fluid to rush into the cell and kill the cell. Just one membrane attack complex per cell is enough to destroy the cell.

Complement Cascade Peptides

PeptideOverview
C3C3 is an abundant serum protein with a labile (changeable) thioester bond. When C3 is cleaved, the thioester bond breaks and an active group is exposed. This C3 active group attaches to other proteins (either free or membrane-bound) through NH2 or OH groups. Further cleavage products of C3 have important activity and bind with cellular receptors.
C3bC3b is an extremely important opsonin, an agent which stimulates phagocytosis.
C5aIn addition to being an anaphylatoxin, C5 potently attracts and activates white blood cells.
C3a
C4a
C5a
C3a, C4a and C5a are anaphylatoxins. Anaphylatoxins are small peptides which stimulate inflammation via: smooth muscle contractions (increasing lymph flow); vascular permeability; induction of inflammatory cytokines; and mast cell and basophil degranulation. Receptors for C3a, C4a and C5a have been found on circulating leukocytes, mast cells, macrophages, hepatocytes, lung epithelial cells, endothelial cells, astrocytes and brain microglia. Anaphylatoxins are responsible for the symptoms of many diseases, with symptoms ranging from mild to deadly.

Discovery of Complement

Cell lysis -- a function of complement -- was identified in the early 1900s as critical to the immune system yet independent of antibodies.

An experiment was devised to identify the agents of complement.

StepOverview
ImmunizationRabbits were injected (immunized) with sheep red blood cells (SRBCs).
Observation 1SRBC are lysed by immunized-rabbit serum.
Observation 2SRBC are not lysed by immunized-rabbit serum that has been cooked at 56°C for 30 minutes.
Observation 3SRBC are not lysed by non-immunized-rabbit serum.
Observation 4SRBC are lysed by non-immunized-rabbit serum that is mixed with cooked immunized-rabbit serum
ConclusionThere are two factors initiating lysis: a heat-stable antibody which is dependent upon cells of the immune system; and a heat-labile agent that does not necessarily bind antigens.
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