REGULATION OF COMPLEMENT
Complement is non-specific in its acitivity. Able to attack host cells as well as microorganisms. One way that activity is regulated is by the fact that the complement components are very labile when activated. They will undergo spontaneous inactivation unless bound by the next complement component. i.e. C3b undergoes spontaneous hydrolysis by the time it has diffused 40nm In addition there are a series of regulatory proteins which regulate complement activity: * C1 inhibitor [C1 INH] forms a complex with C1r2s2 causing it to dissociate from C1q Circulating RBCs are exposed to 1000s of C3b molecules every day. Damage is prevented by the activity of: RCA proteins regulators of complement activation whole family of proteins - all of which are C3 convertase regulatory proteins Share a consensus seqence termed an SCR (short consensus repeat) of ~60residues All encoded at a single chromosome location Called the RCA gene cluster A number of the RCA proteins prevent the assembly of the C3 convertase C4b-binding protein (C4bBP) all bind to C4b preventing C2b CR1 from binding Membrane cofactor protein (MCP) Once they are bound to C4b - Factor I cleaves C4b----------C4d and C4c Similarly, in the alternative pathway: CR1 MCP all bind to C3b and prevent its association with Factor H FactorB Once they are bound, Factor I cleaves C3b RCA proteins may also cause assembled C3 convertase to dissociate C4b binding protein CR1 Factor H DAF all accelerate the inactivation (decay) of the convertase Regulatory Proteins at the level of the MAC S protein - binds to C5b67 and prevents its insertion into host cells Homologous restriction factor (HRF) Membrane inhibitor of reactive lysis (MIRL) bind to C8 and prevent assembly of poly C-9 *Both must be from the same species. This is why complement activation is more efficient when you use complement from another source. BIOLOGICAL CONSEQUENCES OF COMPLEMENT ACTIVTION 1) Lysis of microorganisms (gram - bacteria, fungi, protozoa, enveloped viruses, etc.) Gram + bacteria are somewhat impervious to MAC due to thick peptidoglycan layer. Many pathogens have mechanisms for avoiding complement lysis. 2) inflammation 3) opsonization 4) viral neutralization 5) clearance of immune complexes C3a, C4a, and C5a are anaphylatoxins They bind to complement receptors on mast cells and basophils and induce degranulation histamine release, etc. leads to: a) increased smooth muscle contraction and b) increased vascular permeability------leads to c) influx of phagocytic cells and antibody C5a and C3a can also cause eosinophil degranulation. C3a and especially *C5a and C5b67 attract monocytes and neutrophils to the site of complement activation and lead to increased rates of extravasation [With C5a, Picomolar levels are effective] These fragments also up regulate the expression of complement receptors leading to enhanced phagocytic activity by macrophages and neutrophils. C3b, C4b and C3bi are all opsonins C3b MAJOR OPSININ! All phagocytic cells have C3b receptors Viral Neutralization Antibody + complement form a thick layer around viruses leading to inability to bind to their host cells. Antibody + complement can also cause viral agglutination Also direct lysis of viral envelope Solubilization of Immune Complexes SLE - Systemic Lupus Erythematosus Type III Hypersensitivity Reaction (Immune Complex Hypersensitivity) Deficiencies in C1, C2, C4 and CR1 predispose an individual to SLE. In fact, 90% of C4 deficient patients develop SLE. These factors of course lead to deposition of C3b and C3b seems to be required for Clearance. When immune complexes are coated with C3b the complexes can bind to RBCs and are carried to the liver and spleen where the immune complexes are stripped and phagocytosed. CR1 on phagocytes binding to C3b enhances phagocytosis. SLE patients tend to have lower levels of CR1 on their RBC.
LEUCOCYTE HOMING AND EXTRAVASATION Lymphocytes are capable of a remarkable level of recirculation, continuously moving through the blood and lymph to various lymphoid organs. ~45% of the lymphocytes go to the spleen, ~42% go to various peripheral lymph nodes and the remainder go to various mucosal-associated lymphoid tissues (MALT) or SALT (skin associated lymphoid tissues). In order for recirculating lymphocytes to enter various lymphoid organs or inflammatory-tissue spaces, the lymphocytes must adhere to an pass between the endothelial cells lining the walls of blood vessels by extravasation. Vascular endothelial cells possess cell adhesion molecules on their surface. Some of these are expressed constitutively and others are inducible (expressed primarily in response to certain cytokines produced during an inflammatory response). Most CAMs belong to one of four families of proteins: Selectins Mucins Integrins Immunoglobulins Selectins- membrane glycoproteins with a distal lectin-like domain which enables the protein to bind to specific carbohydrate moities [L-selectin on all leucocytes} E and P selectin are expressed on vascular endothelial cells. Mucins-serine and threonine-rich proteins that are heavily glycosylated. They present sialyated carbohydrate ligands to selectins. For instance, L selectin on leucocytes recognizes sialylated carbohydrates on two mucin-like molecules (CD34 and GlyCAM-1) expressed on certain endothelial cells of lymph nodes. PSGL-1 on neutrophils interacts with E and P-selectin on inflamed endothelium. Integrins- a b heterodimers , integral membrane proteins expressed by leucocytes which facilitate adherence to the vascular endothelium. Different integrins [ex: LFA-1] are expressed by different populations of leukocytes, allowing these cells to bind to different CAMs belonging to the Ig family expressed along the vascular endothelium. [ LAD (leucocyte adhesion deficiency) autosomal recessive disease characterized by recurrent bacterial infections and impaired wound healing. Abnormal synthesis of the B chain. Leucocytes cannot extravasate.] Ig superfamily - proteins which contain at least one Ig-domain ICAM-1,2,3, VCAM on vascular endothelial cells, MAdCAM-1 - has both Ig and mucine-like domains expressed by mucosal endothelial cells. It directs lymphocytes into the mucosa. It binds to integrins via its immunoglobulin-like domain and to selectins via its mucin-like domain. NEUTROPHIL EXTRAVASATION During an Inflammatory response cytokines and other chemicals act on local blood vessels (dilation and increased permeability) to show an increased expression of CAMs [Cell Adhesion Molecules] When this happens the vascular endothelium is said to be inflamed or activated Neutrophils are among the first cells to bind and extravasate (process similar for monocytes but better studied for neutrophils): rolling chemoattractant activating stimulus arrest and adhesion transendothelial migration rolling [due to tether--blood flow pressure to dislodge----tether] initially low affinity selectin-carbohydrate interaction during inflammation, E and P selectins expressed in higher levels on vascular endothelial cells and these bind to mucin-like cell adhesion molecules on the neutrophil membrane . chemoattractant as the neutrophil rolls, it is activated by various chemoattractants that are localized on the endothelial cell surface or secreted locally by cells involved in the inflammatory process. chemokines - chemoattractive cytokines (IL-8, and macrophage inflammatory protein (MIP), PAF, C5a) adhesion binding to chemokines triggers G protein activated signal transduction pathway which induces a conformational change in integrins increasing their affinity for Ig superfamily adhesion molecules (CAMs) on the endothelium. extravasation directed migration of the neutrophil through interendothelial junctions. This phase has not been well characterized.
LYMPHOCYTE HOMING AND EXTRAVASATION
Lymphocytes are capable of a remarkable level of recirculation, continuously moving through the blood and lymph to various lymphoid organs. In order for recirculating lymphocytes to enter various lymphoid organs or inflammatory-tissue spaces, the lymphocytes must adhere to an pass between the endothelial cells lining the walls of blood vessels by extravasation. Antigen specific T cells are known to disappear from the circulation in less than 48 hours. Occurs especially at locations called High Endothelial Venules (HEVs) [plump, cuboidal cells] present in all secondary lymphoid sites except the spleen. Appearance or expression of HEVs dependent upon antigen stimulation. These HEVs express on their surface special molecules called cell-adhesion molecules (CAMs). Antigen stimulation results in an increased expression of these CAMs thereby facilitating extravasation. [germ-free animals do not have HEVs] Re-circulating lymphocytes, monocytes, and granulocytes bear adhesion-molecule receptors for: CAMs of selectin family (E and P-selectin) mucin family (GlyCAM-1 and CD34) Ig superfamily (ICAM-1-3, VCAM-1, and MAdCAM-1) When distributed in a tissue-specific way these molecules are termed: vascular adressins. The corresponding receptors on WBCs have come to be called HOMING RECEPTORS. Homing different subsets of lymphocytes migrate differentially into different tissues trafficking or homing Due to differential expression of homing receptors Also due to chemokine secretion patterns and differential responses: MIP (macrophage inhibitory protein( preferentially attracts naive T cells Monocyte chemoattractant protein (MCP) and RANTES preferentially attract memory T cells Naive Lymphocytes initial attachment to HEVs generally mediated by binding of homing receptor L-selectin to vascular adressins such as GlycCAM-1 and CD34 on HEVs If they encounter antigen within the node, they become activated and enlarge into lymphoblasts and are retained for at least 48 hours. Rapid proliferation and differentiation to effector and memory cells which then leave the node. Effector and Memory Lymphocytes Effectors tend to home to regions of infection Memory lymphocytes are attracted to the type of tissue where they originally encoutnered the antigen. These cells express high levels of certain homing receptors, this allows tissue-specific homing behavior. Actualy process of Extravasation is a multi-step process which involves a cascade of adhesion molecule interactions. First step is selectin-carbohydrate interaction Naive lymphocytes bind to HEV by L-selectin Rolling less pronounced because of slow blood flow in postcapillary venules Then Integrin activating stimulus is mediated by chemokines that are either localized on the endothelial surface or secreted locally. HEVs secrete lymphocyte-specific chemokines )neutrophils dont extravasate at HEVs even though they express L-selection Chemokine binding to G-protein coupled recptors leads to increased affinity of integrins ( LFA-1 ) which then interact with high affinity to Ig superfamily CAMs (ICAM-1) so that there is a firm binding to endothelium. Entire process is regulated by the immune system.
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Lecture 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29