Complement Flashcards

1
Q

Complement

Overview

A
  • Named for its ability to complement the killing action of antibody on bacteria.
  • Significant role in:
    • Promoting inflammation
    • Defense against certain forms of extracellular bacteria
    • Tissue damage
      • Immune complex diseases like systemic lupus erythematous
  • Can provide some protection within minutes/hours of an infection ⇒ innate immunity
  • Works more efficiently with antigen specific antibodies produced by adaptive immunity
  • > 30 proteins found in plasma and tissue fluids
    • Concentrations at steady state levels unless system activated
    • Made by liver
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2
Q

Complement

Cascade

A
  • Successive proteins are converted from inactive to active forms
    • Many are pro-enzymes that require proteolytic cleavage to become active
    • Other are activated by a conformation change due to protein binding
  • Product of one reaction initiates another
  • There are 3 pathways of activation
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3
Q

Complement

Pathways

A
  1. Classical
    • Prefix “C” followed by a number 1-9
  2. Lectin
    • Descriptive names
  3. Alternative
    • ​Letters

Classical pathway requires generation of antibodies and requires 1 week or more for activation.

Alternative & Lectin pathways independent of adaptive immunity and are readily available for defense.

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4
Q

Classical Pathway

A

1) Activation

(Requires Ca2+)

  • IgG or IgM binds antigen causing conformational change exposing specific FC region sequences
  • C1q of the C1qrs complex binds to the FC region of a single IgM or at least 2 IgG molecules
  • When at least 2/6 knobs of C1q are bound at the same time a conformation change is induced
  • Brings active site on C1r into contact with C1s which splits and activates C1s (an esterase)

2) C4 and C2 Cleavage by C1qrs

  • C1qrs cleaves C4 into C4a and C4b
    • Hundreds of C4 can be cleaved by a single C1qrs ⇒ amplification
  • C4a
    • Released into local environment
    • Very short half-life
    • Weak biological activity
  • C4b
    • Cleavage revealed a thioester bond
      • Can be covalently linked to pathogen surface
      • Can be rapidly spontaneously hydrolyzed and inactivated by water
    • Also revealed a region that binds C2b
  • C1qrs then cleaves C2 into C2a and C2b
  • C2a diffuses away
  • C2b binds to C4b on membrane surface
    • Forms C4b2b = classical C3 convertase
      • Half-life of a few minutes

3) C3 cleavage by C3 convertase

  • Classical C3 convertase cleaves C3 into C3a and C3b
    • C3 is the most prevalent complement protein in plasma
    • A single C3 convertase can cleave thousands of C3’s ⇒ significant amplification
  • C3a
    • Released into local environment
    • Significant biological effects
  • C3b
    • Most C3b covalently binds to surface membrane and functions as opsonin
    • Eventually one C3b fragment binds to the C4b2b complex forming C4b2b3b = classical C5 convertase

4) Terminal Pathway

Common to classical, alternative and lectin pathways.

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5
Q

Acute Phase Proteins

A

Acute inflammation causes macrophages to produce IL-6.

IL-6 induces liver to produce acute phase proteins including:

Mannose-binding protein (MBP)

C-reactive protein (CRP)

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6
Q

Mannose Binding Protein

(MBP)

aka

Mannan-Binding Lectin

(MBL)

A
  • Similar structure to C1q
  • Can bind terminal mannose groups on bacterial and yeast carbohydrates but not human cells
  • Associated with 2 serum proteases (MBP-associated serine proteases)
    • MASP-1 and MASP-2
    • Function similar to C1r and C1s
  • When bound to a surface, MBP/MASP-1/MASP-2 complex able to cleave C4 and C2 to generate classical C3 convertase
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7
Q

C-Reactive Protein

(CRP)

A
  • Binds to the phosphorylcholine components on bacterial and fungi cell
  • Does not bind to phosphorylcholine present in the phospholipids of humans
  • Functions like an antibody
  • Once bound to a surface, CRP is able to bind C1qrs and activate the classical pathway
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8
Q

Lectin Pathway

A
  • Initiated by binding of MBP or CRP to conserved motifs on the surface of certain bacteria and fungi
  • Binding activates the rest of the classical cascade
    • MBP functions like C1qrs
    • CRP functions like an antibody
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9
Q

Alternative Pathway

Inducers

A

Promotes the alternative pathway by sheltering C3b from proteolytic cleavage.

  • gram-negative bacteria (specifically LPS)
  • cell wall components of certain gram-positive bacteria and yeast (zymosan)
  • certain viruses and parasites
  • cobra venom factor
  • depositions of IgA immune complexes
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10
Q

Tickover

A

C3 is spontaneously hydrolyzed by H2O into C3a and activated C3b(H2O).

Normally C3b(H2O) is rapidly degraded but protection by an “activator surface” results in a rapid increase of C3b(H2O) concentration

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11
Q

Alternative Pathway

A
  • C3b (from C3 convertase) or C3b(H2O) (from tickover) binds factor B.
  • Binding facilitates cleavage of factor B by plasma protease factor D into Ba and Bb.
    • Ba diffuses into local environment.
    • Bb forms C3bBb or C3b(H2O)Bb
    • Both forms can act as the alternative C3 convertase.
    • C3bBb bound and stabilized by factor P (Properdin)
  • Alternative C3 convertase cleaves C3 into C3a and C3b.
    • C3a enters local environment & has significant metabolic effects
    • C3b complexes with more factor B and becomes activated by factor D ⇒ amplification
  • When a C3b binds to form C3bBb3b it becomes the alternative C5 convertase.
  • Alternative C5 convertase cleaves C5 into C5a and C5b.
    • C5a with significant metabolic effects.
    • C5b enters the terminal pathway.
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12
Q

Terminal Pathway

A

Common to the classical, alternative, and lectin pathways of activation.

  1. Activated classical C5 convertase (C4b2b3b) or alternative C5 convertase (C3bBb3b) splits C5 into C5a and C5b.
    • C5a released into the local environment
      • Proinflammatory effects
    • C5b covalently binds to the membrane
      • Initiates assembly of the terminal complement components to form the membrane attack complex (MAC)
  2. Remainder of MAC complex is assembled non-enzymatically.
    • C5b binds C6, C7, and C8.
    • C8 inserts into the membrane.
  3. C9 polymerization
    • 6-16 molecules of C9 bind to the complex, polymerize, and generate a transmembrane pore in the membrane called the membrane attack complex (MAC).
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13
Q

Membrane Attack Complex

(MAC)

A
  • Transmembrane pore formed of C9
  • Allows flow of solute & electrolytes across the cell membrane
  • Degree of damage depends on cell type
    • Typically gram negative more suseptible
    • Can lyse RBC’s
    • Compromises nucleated cells until lysis
  • Some bacteria have developed strategies to inhibit or block complement cascade
    • Capsule to prevent MAC formation
    • Enzymes in cell wall that inactivate components of cascade
  • Even if MAC fails to eliminate the cell, C3b remains intact on membrane & promotes phagocytosis
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14
Q

Complement Cascade

Regulation

A
  1. Short half-life of products of complement cleavage
    • eg C5a, C3b, C4b2b3b
  2. Inhibitors
    • C1 inhibitor (C1INH)
      • Inhibits proteolytic cleavage activity of C1qrs by binding to C1r and C1s causing dissociation from C1q
    • Factors H and I
      • Found in serum or bodily fluid
      • Factor H binds C3b and facilitates binding of factor I forming inactivated iC3b
  3. Host cell protection mechanisms
    • Complement receptor 1 (CR1)
      • Found on RBC, B cells, macrophages, and dendritic cells
      • Functions like Factor H binding to C3b making it susceptible to factor I
    • Decay accelerating factor (DAF)
      • Found on RBC’s
      • Promotes dissociation of classical and alternative C3 convertase
    • CD59 (Protectin)
      • Prevents MAC formation on host cells
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15
Q

Complement

Effector Activites

A
  1. Phagocytic recruitment
    • anaphylatoxin: C5a > C3a >>>> C4a
    • activation of the vascular endothelium: C5a > C3a
    • chemoattractant: C5a most potent
  2. Phagocytic killing
    • activates phagocytes: C5a >>> C3a
    • opsonization: C3b (binds CR1 on phagocytes)
  3. Direct damage/killing
    • cellular damage and lysis by MAC (C5b789n)
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16
Q

Anaphylatoxin

A

C5a > C3a >>> C4a

  • Causes mast cell degranulation
    • Increase histamine release
  • Causes:
    • smooth muscle contraction
    • increased vascular permeability
    • increased plasma protein concentration in tissues (e.g. complement and antibodies)
    • facilitates neutrophil migration into tissues
17
Q

Activation of Vascular Endothelium

A

C5a > C3a

  • C5a/C3a induces expression of adhesion molecules on vascular endothelium
  • Promotes WBC binding
  • Facilitates diapedesis
18
Q

Chemoattractant

A

C5a >>>> C3a

  • C5a (and C3a) promotes changes in neutrophils and monocytes that promote their firm adherence to vascular endothelium and diapedesis
  • Also creates a chemotaxic gradient in the tissue for cells to follow to site of inflammation
19
Q

Phagocyte Activation

A

C5a >>> C3a

  • C5a induces the respiratory burst
  • Also induces expression of C1 receptors to increase phagocytosis
    • C1 receptors enhance opsonization by degrading C3b into iC3b which binds to CR3.
    • CR3 induces phagocytosis better than CR1.
20
Q

Opsonization

A

C3b primarily

Some C4b

  • Phagocytes (monocytes and neutrophils) express complement receptors:
    • CR1 binds C3b
    • CR3 binds iC3b
21
Q

Deficiencies in

Early C components

A

C1q, C1r, C1s, C4, and C2

  • increases in immune complex diseases
    • glomerulonephritis
    • vasculitis
  • increases in extracellular bacterial infections
    • primarily staph and strep
22
Q

Deficiency of C3

A
  • increases in all bacterial infections
    • abnormalities in MAC formation, opsonization, inflammation, and neutrophil recruitment
  • increases incidence of immune complex diseases
    • due to decreased clearance by reticuloendothelial system
23
Q

Deficiencies in the

Late Components

A

C5, C6, C7, C8

  • Increased susceptibility and recurrent bacterial infections with Neisseria
    • causes gonorrhea and some forms of meningitis
24
Q

Hereditary Angioedema

A
  • Deficiency in the C1 inhibitor
    • Autosomal dominant
    • ​Can be acquired later in life due to autoantibodies for C1INH
  • Results in excessive activation of C4 and C2 by C1
    • C2 converted to C2 kinin
    • Results in excessive swelling
  • C1INH also involved in clearance of clotting, kinin, and plasmin systems.
25
Q

Factor I Deficiency

A
  • Results in uninhibited activation of the alternative pathway
  • Excessive complement consumption
    • Decreased serum C3 levels
  • Causes abnormal opsonization leading to increased susceptiblity to infections
  • Hives seen due to excessive C’ activation and release of anaphylatoxins
26
Q

CH50 Assay

A
  • The dilution of serum that causes lysis of 50% of opsonized RBC’s
    • Reported as the reciprocal of the greatest dilution where 50% lysis achieved
  • Measures the total activity of the classical lytic pathway
    • CH50 decreases if at least one specific complement component is decreased
  • Individual components need to be measued seperately.
  • Interpretation:
    • no lysis - congenital (null) deficiencies of C1 - C8
    • 1/2 decrease in CH50 = C9 deficiency
    • decreased lysis = systemic activation of complement cascade via infection, immune-complex diseases, or auto-immune processes due to consumption
27
Q

C3 and C4

Assays

A

Measures the intact C3 and C4 levels in the serum and not the biologically active productions generated during complement activation.

  • Low C4 concentration = intrinsic abnormality or consumption by activation of the classical pathway
  • Low C3 concentration = intrinsic abnormality or consumption by activation of either the classical or alternative pathways