The Complement System
Biological Activities of the Complement
1. Opsonization - Macrophages and neutrophils express a cell-surface receptor, called CRI, which binds to C3b. Microorganisms coated with C3b will therefore be brought into contact with these phagocytic cells where they will be readily phagocytosed. 2. Induction of Local Inflammation - During complement activation the C3a, C4a and C5a fragments produce inflammation at the site of activation. This occurs by two mechanisms: • These fragments act as chemo attractants which direct the migration of phagocytic cells towards the site of complement activation. • These fragments may cause the degranulation of mast cells and basophils which release inflammatory mediators such as histamine and other vasoactive substances that increase capillary permeability. For this reason these fragments are often referred to as anaphylatoxins. 3. Lysis of Pathogens - The end result of the membrane attack pathway is the appearance of holes in the surface membrane of the original inciting microorganism. This is particularly important in destroying Neisseria organisms (the bacteria responsible for meningitis and the sexually transmitted disease gonorrhea) and any defect in the membrane attack complex may predispose the individual to infection with these organisms. 4. Viral Neutralization - Mannan binding lectin binds to a variety of viruses and leads to opsonization and lysis of the organism. It may also interfere with the ability of the virus to interact with its membrane receptor and thus blocks its entrance into the host cell. 5. Clearance of Immune Complexes - Soluble immune complexes coated with C3b become bound to complement receptors (CRI) present on phagocytic cells but especially on circulating erythrocytes. On the surface of the red cells they are carried through the circulation to the liver and spleen where they are removed by macrophages.
Functions of the Complement System
A series of proteins working in a cascading manner, to generate important host defense mechanisms. The complement system, comprising approximately 30 circulating and membrane-expressed proteins, is an important effector arm of both the innate and antibody-mediated acquired immune responses. Named from some of the earliest observations of its activity - a heat-sensitive material in serum that "complemented" the ability of antibody to kill bacteria - we now know that complement plays a major role in defense against many infectious organisms. Three different pathways of complement activation: 1) Classical 2) Alternative 3) Mannan binding ligand (MBL)
Hereditary Angiodema
Rare autosomal dominant disorder resulting in the deficiency of the regulatory protein C1-INH, the only control protein for the classical pathway components C1r and C1s, results in uncontrolled cleavage of C2 and C4. Genetic deficiency of C1-INH results in hereditary angioedema (HAE). The condition is characterized by localized edema in the skin and mucosa resulting from dilatation and increased permeability of the capillaries. The symptoms are recurrent attacks of swollen tissues, such as the face and limbs, pain in the abdomen, and swelling of the larynx, which can compromise breathing. The condition is due to deficiency of C1-INH which normally inhibits the activity of enzymes in serum cascades other than the complement cascade. One of these serum pathways forms kinins, including bradykinin, which is a potent vasodilator and inducer of increased vascular permeability and smooth muscle contraction. This results in swelling or edema, which is free of cellular components. These swellings do not itch in contrast to the swellings of acute allergic reactions that are intensely itchy due to histamine release from mast cells and basophils. It is worth noting that not all patients with HAE have decreased levels of C1-INH. About 15% of these patients have normal levels, but the protein is abnormal and fails to inhibit C1 activity. **Other parts of the complement system are not directly affected and can still function through other means, such as the alternative pathway. Therefore, MAC complex formation, which would cause pore formation and cell lysis of targets, splitting of C3 into C3a &C3b, and formation of the anaphylatoxins C3a and C5a would still be intact.
The Alternative Pathway
The alternative pathway of complement activation is triggered by a variety of bacterial and viral products. The most widely studied include lipopolysaccharide (LPS) also referred to as endotoxin, from the cell walls of gram-negative bacteria) and the cell walls of some yeasts. Activation of the alternative pathway occurs in the absence of specific antibody. Thus, the alternative pathway of complement activation is an effector arm of the innate immune defenses. Some of the components of the alternative pathway are unique - the serum factors B and D, and properdin (also known as factor P) - while others, C3, C3b, C5, C6, C7, C8, and C9, are common to those used in the classical pathway. Initiation of the pathway occurs when C3b normally present in the circulation due to spontaneous hydrolysis of C3, binds to the foreign microbial products. A series of reactions then occurs culminating in the formation of a C3 convertase called C3BbBP by C3b, Factor B, Factor D, and Properdin/Factor P. C3bBbP/C3 convertase then splits C3 into C3a and C3b and continues the complement cascade. Because small amounts of C3b are always present in the circulation, the alternative pathway is always primed and ready to go. Activation therefore must be tightly controlled and two inhibitory proteins Factor H and Factor I constantly inactivate C3b thereby inhibiting or controlling the alternative complement pathway activity.
Classical Complement Pathway
The classical pathway was so named because it was the first complement pathway to be worked out. The component proteins are designated C1, C2, etc up to C9. (The numbers designate the order in which the components were discovered, rather than their position in the activation sequence.) Antigen-antibody complexes are the predominant activators of the classical pathway; thus, this pathway is a major effector pathway of the humoral adaptive immune response. 1. The first step involves the binding of C1 to the Fc region of an antibody attached to its specific antigen. This leads to the generation of an enzyme C1 esterase following a complex interaction between C1q, C1r and C1s, which together is known as Activated C1 (acts as the esterase). This activation is controlled by a protein called C1-inhibitor (C1-INH). 2. C1 esterase/Activated C1 acts on C4 to yield a small fragment C4a which is released and a larger fragment C4b. The latter binds covalently to Activated C1, forming a new Activated C14b complex. This new complex binds C2 causing the C2 molecule to become susceptible to the action of C1 esterase which cleaves it to form C2a and C2b. 3. C2b remains attached to C14b on the surface membrane of the antigen and this complex (C14b2b) functions as a C3 convertase. 4. The C14b2b/C3 convertase cleaves C3 to give a small C3a fragment and a larger C3b fragment. C3a (together with C5a) stimulates inflammation by attracting neutrophil polymorphs (its chemotactic effect) and by causing degranulation of basophils and mast cells. C14b2b3b triggers the membrane attack complex by splitting C5 into C5a and C5b, and C5b then binds the remaining components of the attack complex in C6, C7, C8, and poly-C9. The MAC forms a pore on the antigenic surface to cause the influx of water and efflux of important molecules, leading to its death. **Important Functions of the Complement System: 1) The production of opsonins, molecules that enhance the ability of macrophages and neutrophils to phagocytose material, aided by C3b 2) Chemotaxis via the recruitment of neutrophils, aided by C3a and C5a 3) the production of anaphylatoxins, peptides that induce local and systemic inflammatory responses by causing degranulation of basophils and mast cells, which contain potent inflammatory chemicals such as histamine, done by C3a and C5a 4) Lysis of organisms coated with specific antibody via MAC, done by C8 and C9
The Membrane Attack Pathway
The membrane attack pathway is the sequence of events between C3 and C9 culminating in the "punching of holes" in the membrane of the initiating antigen. 1. C5 is cleaved to form C5a and C5b. 2. C5b binds to cell membranes and binds C6. The C5b6 complex then combines spontaneously with C7. 3. C8 binds to the complex C5b67 and the entire new complex inserts itself into cell-membranes. 4. Incorporation of many molecules of C9 causes further penetration of the lipid bilayer and results in the formation of a transmembrane channel that disturbs the cell's osmotic equilibrium: ions pass through the channel and water enters the cell. The cell swells and the membrane becomes permeable to macromolecules, which then escape from the cell. The result is cell lysis.
Activation of the Complement
There are three different pathways whereby the complement system recognizes and is activated by microorganisms. Each involves a series of proteins converging at C3 and each will result in the attachment of complement proteins to microbial surfaces: 1. The classical pathway is triggered by antibodies bound to antigens on the microbial surface. 2. The alternative pathway is triggered directly by bacterial cell surfaces. 3. The lectin pathway is initiated when mannan-binding lectin binds to carbohydrates on the surface of microbes.
The Lectin Binding Pathway
This pathway is activated by mannose residues found on the surface of many bacteria. The unique proteins of this system include mannose-binding lectin (MBL) and two associated proteases, known as mannose-associated serine protease (MASP) 1 and 2. Activation through MBL results in the cleavage of the classical complement pathway components C4 and C2 to form C4b2b, the classical pathway C3 convertase, on the surface of the bacterium. Thus, the lectin pathway converges with the classical pathway at the activation of C3. The lectin pathway is activated by terminal mannose residues of proteins and polysaccharides found on the surface of bacteria. These terminal mannose residues are not found on the surface of mammalian cells, and so the lectin pathway of complement activation may be thought of as discriminating between self and non-self. Because this pathway is activated in the absence of antibody, it is part of the innate immune defenses. More important in children because as you age, you develop more mechanisms for fighting off infections
Regulation of the Complement
Uncontrolled complement activation can rapidly deplete complement components, leaving the host unable to remove infectious agents. In addition, as we shall describe later in the chapter, complement activation generates activated fragments (especially the cleavage products of C3, C4, and C5) that induce potent inflammatory responses to remove infectious agents. However, the very potency of these responses may damage the host. Normally, inappropriate activation of complement does not occur, because many steps in the complement pathways are negatively regulated by specific inhibitors. Many of the molecules that regulate complement activation are expressed on the surface of mammalian cells, but not microbial cells. Consequently, the regulators of complement activity limit damage to the individual, while allowing activated complement components to focus on removing microbial pathogens.An important inhibitory protein is C1 esterase inhibitor (C1 INH) that inhibits the first step in the activation of the classical complement pathway. C1 INH binds to C1r and C1s, causing them to dissociate from C1q, and preventing complement activation. As we shall describe below, C1INH also inhibits the function of enzymes in other serum cascades, particularly those involved in clotting and in the formation of kinins, potent mediators of vascular effects. Absence of this protein results in the disease Hereditary angioedema
Complement Deficiencies
the complement system plays an important role in defending the host against microorganisms. It is particularly important in defending against what are known as pyogenic (pus-forming) bacteria, which include Neisseria species, Streptococcus pneumoniae, Haemophilus influenza, and Staphylococcus aureus. The major pathways of defense against these organisms appear to be the production of IgG antibody that binds to the bacteria accompanied by opsonization, complement activation, phagocytosis, and intracellular killing. Thus, situations in which any one of these activities are diminished, resulting from either genetic deficiencies or acquired conditions, renders a person particularly susceptible to these organisms. In addition, complement is important in removing immune complexes from the circulation and deficiencies of certain complement components can result in immune complexes depositing in tissues, leading to autoimmune conditions. Individuals genetically deficient in specific complement components are relatively rare (approximately 1 in 10,000 people), and deficiencies are not always associated with disease. Deficiencies include: 1. C1q, C1r, C2, C4 = SLE 2. C3, C5, Factor I, Factor D, Factor H, Factor B = each deficiency leads to recurrent infection 3. C6, C7, C8 = overwhelming Neisserial Infection 4. C1 Inhibitor = Hereditary Angiodema (HAE) 5. higher C5-C9 = no disease Deficiencies in any of the classical pathway components C1, C4, or C2 are associated with increased susceptibility to autoimmune diseases such as systemic lupus erythematosus. This predisposition for SLE in these individuals appears to be the result of an impaired ability to process and clear immune complexes. C3 deficiency is rare, but can be severe and even life-threatening because C3 is central to all the complement pathways. C3-deficient individuals are susceptible to recurrent pyogenic infections, and may also develop inflammatory disorders associated with circulating immune complexes. Deficiency of the alternative pathway component properdin, or of factors B or D, is associated with pyogenic infections. Deficiency of mannose-binding lectin can be a major problem in early life manifesting as severe recurrent infections. Individuals lacking components of the membrane attack complex C5b-C9 are prone to recurrent Neisserial infections.