Complement determinations in human disease - …

1 CME review articleThis feature is supported by an unrestricted educational grant from AstraZeneca LPComplement determinations in human diseaseM. Michael Glovsky, MD*; Peter A. Ward, MD ; and Kent J. Johnson, MD Objective:To define techniques used for Complement measurements and examine the clinical relevance of alterations ofcomplement determinations in Sources:Data have been assembled from the authors research, original articles, and reviews, as well as chapters andcomplete books on Selection:Studies were chosen for inclusion by the opinions of the authors, relevant Complement reviews, publications,and : Complement has been shown to possess approximately 31 proteins, some of which are enzymes (C1r, C1s, C2, factorB, factor D), some cofactors, some inhibitors or inactivators, and others composed of membrane-integrated proteins. All of thecomplement proteins have been purified, and many of the respective genes have been identified.

2 The Complement cascade is adual-edged sword, causing protection against bacterial and viral invasion by promoting phagocytosis and , Complement can cause substantial damage to blood vessels (vasculitis), kidney basement membrane and attachedendothelial and epithelial cells (nephritis), joint synovium (arthritis), and erythrocytes (hemolysis) if it is not :Definitive evidence is available that Complement -mediated tissue destruction occurs after immune complexinjury in the kidney and lung and may be important in lupus erythematosus and adult respiratory distress syndrome. Future studieson Complement receptor structure and function may provide clues to treat effectively lupus, hemolytic anemias, and addition, gene therapy and antibody therapy need further refinement to treat immunodeficiency Allergy Asthma ;93:513 disclosure:Drs. Glovsky, Ward, and Johnson have indicated that this article does not include the discussion of unapproved/investigative use ofa commercial disclosure:Drs.

3 Glovsky, Ward, and Johnson have indicated that in the last 12 months they have not had any financial relationship, affiliation,or arrangement with any corporate sponsors or commercial entities that provide financial support, eduction grants, honoraria, or research supportorinvolvement as a consultant, speaker s bureau member, or major stock shareholder whose products are prominently featured either in this article or with thegroups who provide general financial support for this CME for CME credit1. Read the CME review article in this issue carefully and complete the activity by answering the self-assessment examination questions on the form To receive CME credit, complete the entire form and submit it to the ACAAI office within 1 year after receipt of this issue of the discovery of Complement by Jules Bordet more than100 years ago, the importance of the Complement system inproducing lysis of bacteria and protecting humans and exper-imental animals against infectious microorganisms has 9 Within the past 30 years, Complement hasbeen shown to possess approximately 31 proteins, some ofwhich are enzymes (C1r, C1s, C2, factor B, factor D), somecofactors, some inhibitors or inactivators, and others com-posed of membrane-integrated proteins.

4 All of the comple-ment proteins have been purified, and many of the respectivegenes have been proteins of the comple-ment system include the receptors for C1q, C3a/C4a, C5a,CR1, CR2, and CR3 (C3b, C3bi, and C3d receptors). Inaddition, membrane components (decay-accelerating factor,CD55 and CD59, and membrane inhibitor of C8 and C9insertion) are important regulating proteins. The complementcascade is a dual-edged sword, causing protection againstbacterial and viral invasion by promoting phagocytosis andinflammation. Pathologically, Complement can cause sub-stantial damage to blood vessels (vasculitis), kidney base-ment membrane and attached endothelial and epithelial cells* Huntington Medical Research Institute, Pasadena, California. Department of Pathology, University of Michigan Medical School, AnnArbor, for publication March 1, for publication in revised form May 19, 93, DECEMBER, 2004513(nephritis), joint synovium (arthritis), and erythrocytes (he-molysis) if it is not adequately controlled.

5 This review definestechniques used for Complement measurements and examinesthe clinical relevance of alterations of Complement determi-nations in disease . Data have been assembled from the au-thors research, original articles, and reviews, as well aschapters and complete books on ACTIVATION MECHANISMST here are 3 currently known Complement activation mecha-nisms (Fig 1): (1) the classical pathway, (2) a recently de-scribed pathway (the mannose-binding lectin pathway), and(3) the alternative complexes, apoptotic cells, or C1q bound to itsligand can activate the classical pathway. On activation, C1rand C1s are converted from proenzymes to activated enzymesand cleave their natural substrates C4 and C2 to C4a and C4band C2a and C2b, respectively. C4b and C2a form C4b2a (C3convertase of the classical pathway), which splits C3 to C3aand C3b. C3b combines with C4b2a to form C4b2a3b (C5convertase), which cleaves C5 to C5b and C5a. This leads toformation of the C5b, C6, C7, C8, and C9 membrane attackcomplex (MAC).

6 Mannose-binding lectin, when bound to mannose residueson microbial surfaces, can interact with 2 serine proteases:MASP-2 and MASP-1. MASP-2 cleaves and inactivates C4and C2 to form C4b2a. MASP-1 may cleave C3 directly,promoting the formation of C4b2a3b, the C5 convertase ofthe classical alternative pathway can be activated in the absence ofantibody by insoluble polysaccharides, yeast cell walls, andaggregated IgA and IgE at high concentrations. Factor D, aserine protease, cleaves factor B to Ba and Bb. The largerfragment of factor B combines with the large fragment of C3,Figure 1. Complement activation OF ALLERGY, ASTHMA & IMMUNOLOGYC3b, to form the alternative pathway C3 convertase, stabilizes the C3 convertase by binding to thecomplex, PC3bBb. C3bBb then can further cleave C3 to formadditional C3b molecules, some of which combine withC3bBb to form C3bBbC3b, the C5 convertase of the alter-native pathway, which cleaves C5 to C5a and C5b.

7 Thus, theclassical pathway and the mannose-binding lectin pathway,as well as the alternative pathway, converge on C3 to initiatethe formation of C3b, the Complement factor, when bound tomicroorganisms, which initiates phagocytosis and destructionof the invading microbe. C3a, a proinflammatory anaphyla-toxin, is also MAC complex (C5b-C9) and C5a (the most potentanaphylotoxin) are generated by enzymatic cleavage of C5 toC5b and C5a. Table 1 lists alterations in the complementactivation pathways and some of the diseases associated withactivation of the classical Complement pathway such asseen in immune complex disease associated with DNA anti-DNA antibodies is often associated with low CH50, low C4,and low C3. Alternative pathway activation of factor B andC3 is seen in endotoxin shock syndromes, as well as in seraof patients with membranoproliferative , activation of the classical pathway occurs in the relativeabsence of the C1 inhibitor as seen in hereditary angioedema(HAE).

8 Fluid-phase activation of the alternative pathwayoccurs with factor I deficiency, when C3 convertase activityis not controlled. With chronic inflammation that occurs inReiter syndrome, acute-phase reactants, such as C4, factor B,and haptoglobin, may be increased. Young children with lowmannan-binding lectin levels frequently are infected withbacterial and yeast pathogens. The decreased levels of CH50and C4 reflect classical pathway activation, whereas de-creased C3 and factor B levels with normal C4 levels areindicative of alternative pathway activation. Deficiencies ofthe regulator proteins C1 inhibitor and factor I are reflectedby increased classical and alternative pathway activation, MECHANISMS OF THE COMPLEMENTSYSTEMTo control activation of the classical pathway, C1 inhibitorbinds to activated C1r and C1s and inactivates these binding is in a 1:1 ratio. The C4b2a complex (C3convertase) is controlled by C4 binding protein, which bindsto the complex.

9 C4b is then cleaved by factor I. Underphysiologic conditions, the classical pathway control stopsthe further breakdown of classical pathway components. Thealternative pathway C3 convertase (C3bBb) is regulated byfactor H, which competes with Bb, the split product of factorB, for the binding site on C3b, forming (C3bH).9 The C3b isfurther degraded by factor I to smaller breakdown the similar control mechanisms of the classical andalternative classical and alternative pathwayconvertases are broken up by C4b binding protein and factorH, respectively. C4b and C3b are then cleaved by factor I toinactive MEASUREMENTC omplement components can be measured either as proteinsor based on their functional ,4,10 19 Because the pro-teins that comprise the classical and alternative complementcascades have been isolated in chemically pure form, anti-bodies have been produced to these proteins in several animalspecies. These antibodies may be used to quantitate comple-ment components through one of several radial immunodiffusion, antibody to a single comple-ment component is incorporated into an agar gel.

10 Test serumis placed in wells cut into the agar. During a subsequentincubation period that lasted 1 to 3 days, the specific com-plement protein diffuses into the agar, forming a precipitinring in its interaction with antibody. The diameter of the ringreflects the concentration of Complement proteins present inthe test serum. The precise amount is determined by compar-ing the diameter of the precipitin ring formed with standardsolutions of Complement 1. Complement Profiles in DiseaseActivation pathwayCH50C4C3 Factor BRelated diseasesClassical pathway22 2 2 Systemic lupus erythematosus, some immune complex diseases,some cases of urticaria and angioedemaNorAlternative pathway2N22 Endotoxin shock, membrano-proliferative glomerulonephritisFluid-phase activation of classical22pathwayN orNN Hereditary angioedemaFluid-phase activation of alternative222pathwayN or NC3b inactivator deficiencyAcute-phase reactions11 1 1 Chronic nonimmune complex diseases such as seen in Table 5 Activation pathwayMBLMASP-1, -2 Related diseasesMannose-binding lectin pathway2?