Complement System and Additional Topics - Part 2 of Innate Immunity
COMPLEMENT SYSTEM
What Is the Complement System?
The complement system is a cascade of approximately 30 proteins found in blood plasma.
Key functions:
Kill pathogens directly by forming holes (membrane perforation).
Opsonize pathogens to tag them for phagocytosis (enhanced ingestion by immune cells).
Recruit immune cells, promoting inflammation.
Clear immune complexes (aggregates of antibodies and antigens).
Metaphor: Think of it like a set of dominoes, where the activation of one protein triggers the next, amplifying the response.
THE THREE PATHWAYS - Overview
Introduction
All three complement pathways initiate differently but converge at the complement component C3, after which they follow the same route to form the Membrane Attack Complex (MAC).
PATHWAY 1: CLASSICAL PATHWAY
Trigger
Activation occurs when antibodies (specifically IgM or IgG) bind to the surface of a pathogen.
Notable:
Considered the "classical" pathway due to its initial discovery.
Steps
C1 Activation
The C1 complex consists of three components: C1q, C1r, C1s.
C1q binds to the Fc region of antibodies (at least two IgG molecules or one IgM, which has five binding sites).
Binding of C1q activates the serine protease C1r, which in turn activates C1s.
C1s becomes a protease (enzyme that cleaves proteins).
C4 and C2 Cleavage
Activated C1s cleaves C4 into C4a (a small fragment) and C4b (a large fragment).
C4b attaches to the pathogen's surface.
C1s also cleaves C2 into C2a (large) and C2b (small).
C4b and C2a combine to form the C3 convertase (C4b2a).
Key Point: The C3 convertase for the classical pathway is C4b2a.
PATHWAY 2: LECTIN PATHWAY
Trigger
Initiated by Mannose-Binding Lectin (MBL), which recognizes mannose residues on the surfaces of pathogens like bacteria, fungi, and viruses (absent from human cells).
This pathway does not require antibodies.
Steps
MBL Activation
MBL binds to mannose on pathogens.
Structurally, MBL resembles C1q.
Activation of MBL leads to activating MBL-associated serine proteases (MASP-1, MASP-2), analogous to C1r and C1s.
C4 and C2 Cleavage
MASP-2 cleaves C4 into C4a and C4b, and C2 into C2a and C2b, similar to the classical pathway.
C4b and C2a come together to form the C3 convertase (C4b2a).
Key Point: The lectin pathway also uses the C3 convertase C4b2a, the same as the classical pathway.
PATHWAY 3: ALTERNATIVE PATHWAY
Trigger
Triggered by spontaneous “tickover,” involving factors that amplify the response on pathogen surfaces.
This pathway does not require antibodies or lectins; it is continuously active at low levels.
Steps
Spontaneous C3 Hydrolysis (“Tickover”)
C3 undergoes spontaneous hydrolysis in the circulation to form C3(H2O).
C3(H2O) binds to Factor B.
Factor D cleaves Factor B into Ba (small) and Bb (large).
C3(H2O)Bb forms a fluid-phase C3 convertase (inefficient).
Amplification on Pathogen Surface
C3(H2O)Bb cleaves additional C3 into C3a and C3b.
C3b binds to pathogen surfaces.
Human cells possess regulatory proteins (DAF, Factor H, Factor I) that quickly inactivate C3b.
On pathogen surfaces, C3b binds to Factor B, leading to its cleavage by Factor D to form Bb, thus creating the C3bBb convertase (alternative pathway).
This form is more stable than the fluid-phase convertase.
Stabilization
Properdin (Factor P) stabilizes C3bBb, leading to increased C3b production and an amplification loop.
Key Point: The C3 convertase of the alternative pathway is C3bBb, stabilized by properdin.
CONVERGENCE POINT - C3 ACTIVATION
All three pathways lead to active C3 convertases that cleave C3 into two fragments:
C3 convertase for classical pathway: C4b2a
C3 convertase for lectin pathway: C4b2a
C3 convertase for alternative pathway: C3bBb
C3 cleavage results in:
C3a (small fragment)
C3b (large fragment)
Central step for complement activity.
C3 FUNCTIONS - Importance of C3
C3a
Anaphylatoxin:
C3a binds to C3a receptors on mast cells and basophils, leading to:
Degranulation and histamine release
Causes inflammation characterized by vasodilation, increased vascular permeability, and smooth muscle contraction.
C3a aids in recruiting immune cells.
C3b - The Workhorse
Function 1: Opsonization (Most Important)
C3b coats the pathogen’s surface enhancing phagocytosis.
Phagocyte receptors include:
CR1 (CD35): Binds C3b
CR3 (Mac-1, CD11b/CD18): Binds iC3b (inactive C3b)
C3b serves as the primary opsonin.
Function 2: Forms C5 Convertase
C3b combines with C4b2a to form the C5 convertase in classical/lectin pathways.
C3b combines with C3bBb for the C5 convertase in the alternative pathway.
Function 3: Amplification
C3b can join with the alternative pathway components to enhance C3 convertase production.
Positive feedback loop amplifying the complement response.
Regulation of C3b
Factor I is a protease that cleaves C3b into iC3b (inactive) and C3dg.
Factor H and DAF (Decay Accelerating Factor) assist Factor I in this regulation.
Human cells have these regulators, which protect them from complement attack, whereas bacterial cells do not.
THE TERMINAL PATHWAY - MAKING THE MAC
C5 Activation
C5 convertase cleaves C5 resulting in:
C5a (small)
C5b (large)
C5a
The Most Powerful Anaphylatoxin:
Functions include:
Being a potent chemotactic factor, effectively recruiting neutrophils.
Activates neutrophils and macrophages.
Causes increased vascular permeability and stimulation of respiratory burst in phagocytes.
Major clinical mediator of septic shock.
C5b Initiates MAC Formation
C5b remains attached to the pathogen surface and starts the assembly of the Membrane Attack Complex (MAC).
Formation of the Membrane Attack Complex (MAC)
Steps
C5b binds to C6 to form C5b6.
C5b6 binds to C7, forming C5b67 (which inserts into the membrane).
C5b67 binds to C8, forming C5b678 (creating a small pore).
C5b678 recruits multiple C9 molecules (approximately 10-16), which polymerize to form a larger transmembrane pore.
Result: MAC = C5b6789 (with multiple C9s) creates large pores in the bacterial membrane.
Water and ions enter the bacteria, leading to cell swelling and lysis (bursting).
What Gets Killed by MAC?
MAC is most effective against gram-negative bacteria (due to their thin cell wall).
Particularly susceptible: Neisseria species (e.g., meningococcus, gonococcus).
Also targets some enveloped viruses and certain parasites.
What Is Resistant?
Gram-positive bacteria are typically resistant to MAC due to their thick peptidoglycan cell wall, relying instead on opsonization (via C3b) for clearance.
Protection of Human Cells
CD59 (Protectin): Prevents C9 polymerization on human cells.
DAF (CD55): Accelerates decay of C3 convertases, protecting host cells.
SUMMARY OF COMPLEMENT FRAGMENTS
Mnemonic
C3a: Anaphylatoxin → inflammation ("a" = anaphylatoxin)
C3b: Opsonization, C5 convertase ("b" = binds bacteria)
iC3b: Opsonization (binds CR3) ("i" = inactive)
C5a: Anaphylatoxin, Chemotaxis ("a" = attracts neutrophils)
C5b: Starts MAC assembly ("b" = begins MAC)
C5b-9 (MAC): Lyses pathogen (Membrane Attack Complex)
Memory Trick: "Small fragments (a) = Anaphylatoxins" (C3a, C4a, C5a)
"Big fragments (b) = Bind to stuff" (C3b binds bacteria, C5b binds C6)
COMPLEMENT REGULATION
Why We Don’t Attack Ourselves:
Fluid Phase Regulators (in blood)
C1 Inhibitor (C1-INH): Inactivates C1r and C1s, halting classical pathway.
Factor H: Binds C3b, aiding Factor I in cleaving it.
Factor I: Protease that cleaves C3b into iC3b.
C4-binding protein (C4BP): Accelerates decay of C3 convertase (C4b2a).
Cell Surface Regulators (on human cells)
DAF (CD55): Dissociates C3 convertase.
MCP (CD46): Helps Factor I cleave C3b.
CR1 (CD35): Binds C3b/C4b, assisting inactivation.
CD59 (Protectin): Prevents C9 polymerization, inhibiting MAC formation.
Importance: Human cells contain these regulatory proteins, making them immune to complement attack while non-human cells lack in this protection.
COMPLEMENT DEFICIENCIES - CLINICAL CORRELATIONS
EARLY CLASSICAL PATHWAY DEFECTS (C1, C2, C4)
Disease Risks:
C1q, C1r, C1s: Patients at risk for Systemic Lupus Erythematosus (SLE) due to inability to clear immune complexes, leading to deposition in tissues.
C2 Deficiency: SLE, recurrent infections are common; C2 deficiency is the most prevalent complement deficiency.
C4 Deficiency: Associated with SLE due to impaired immune complex clearance.
Key Concept: The classical pathway is crucial in clearing immune complexes; defects can lead to tissue-specific immune complex deposits causing diseases like SLE.
C3 DEFICIENCY
Disease Risks:
Severe recurrent pyogenic infections (caused by Staphylococcus, Streptococcus, and Pseudomonas) due to lack of opsonization (C3b).
Membranoproliferative glomerulonephritis (MPGN) due to dysregulation of the alternative pathway.
Key Concept: C3 is the central component for the complement system; absence leads to a lack of opsonization and severe vulnerability to infections.
ALTERNATIVE PATHWAY DEFECTS
Disease Risks:
Factor D Deficiency: Leads to recurrent Neisseria infections, as it is impossible to activate the alternative pathway.
Properdin Deficiency: X-linked condition resulting in recurrent Neisseria infections due to instability of the alternative pathway.
TERMINAL PATHWAY DEFECTS (C5-C9, MAC)
Disease Risks:
Deficiencies in C5, C6, C7, C8, or C9 lead to recurrent Neisseria infections, as the MAC cannot form effectively.
Bacterial infections can lead to meningococcal and gonococcal diseases due to these specific vulnerabilities.
Key Concept: The MAC is especially critical for lysing Neisseria species.
CLINICAL PEARL
Patient presenting with recurrent meningitis should raise suspicion of complement deficiencies, particularly involving C5-C9 components.
REGULATORY PROTEIN DEFECTS
Disease Risks:
C1 Inhibitor Deficiency (C1-INH): Leads to Hereditary Angioedema (HAE), characterized by uncontrolled classical pathway activation resulting in bradykinin-induced angioedema.
Factor H Deficiency: Associated with Hemolytic Uremic Syndrome (HUS) and MPGN, resulting from uncontrolled activation of the alternative pathway causing damage to kidneys and RBCs.
Factor I Deficiency: Related to HUS and MPGN, leading to recurrent infections due to inability to inactivate C3b.
CD55 (DAF) Deficiency: Seen in Paroxysmal Nocturnal Hemoglobinuria (PNH), leading to complement-mediated lysis of RBCs.
CD59 Deficiency: Also results in PNH, failing to inhibit MAC formation, thus causing intravascular hemolysis.
Important Details: Hereditary Angioedema (HAE)
Condition arises from deficiency or dysfunction of the C1 inhibitor, leading to unchecked C1 activation that influences the kallikrein-kinin pathway.
Bradykinin production increases vascular permeability, leading to recurrent severe swelling episodes (e.g., face, lips, airway).
Triggered by stress, trauma, or infections; can be life-threatening if airway swelling occurs.
Treatment involves C1-INH replacement and bradykinin receptor antagonists.
Important Details: Paroxysmal Nocturnal Hemoglobinuria (PNH)
PNH emerges from acquired mutation in hematopoietic stem cells leading to GPI anchor deficiency, inhibiting the attachment of CD55 and CD59.
Clinical manifestation includes complement attack on RBCs.
Symptoms include dark urine in the morning (hemoglobinuria), fatigue, risk of thrombosis, and pancytopenia.
Diagnosis through flow cytometry to detect absence of CD55/CD59 and treated with eculizumab, an anti-C5 antibody.
ANAPHYLATOXINS (C3a, C4a, C5a)
What Are Anaphylatoxins?
Small complement fragments that induce inflammation through activation of mast cells and other immune cells.
The Three Anaphylatoxins
Potency and Functions:
C4a: Weakest; causes mild inflammation.
C3a: Intermediate potency; drives inflammation, with some chemotactic abilities.
C5a: Strongest; potent chemotactic factor for neutrophil recruitment and activation.
What Do Anaphylatoxins Do?
Mast Cell Activation:
Bind to receptors on mast cells and basophils, triggering degranulation and release of histamine and other mediators.
Effects of Histamine Release:
Causes vasodilation, increasing blood vessel diameter and causing redness and heat.
Increases vascular permeability leading to edema (fluid leakage).
Smooth muscle contraction in bronchioles (bronchoconstriction, leading to wheezing) and in the gastrointestinal tract (causing abdominal pain).
Chemotaxis (especially C5a):
C5a is a potent neutrophil chemotactic factor, creating a gradient that guides neutrophils toward the infection site.
Neutrophil Activation (C5a):
Stimulates increased expression of adhesion molecules enhances phagocytosis and respiratory burst in neutrophils, promoting reactive oxidative species (ROS) production and degranulation.
Clinical Relevance of Anaphylatoxins
Overproduction of C5a can occur in diseases like septic shock, leading to systemic inflammation and hypotension.
High C5a levels are observed in ARDS, recruiting neutrophils that can cause lung damage.
Complement activation can also contribute to ischemia-reperfusion injuries.
Why Called “Anaphylatoxins”?
These fragments can induce symptoms similar to anaphylaxis, characterized by vasodilation, bronchoconstriction, and shock-like features, although they are not true anaphylactic reactions (the latter is IgE mediated).
CYTOKINES
What Are Cytokines?
Cytokines are small proteins secreted by cells that act as signals for communication among cells, often referred to as "cellular messengers".
They regulate various processes including immune responses, inflammation, and hematopoiesis.
Categories of Cytokines
Interleukins (IL): Cytokines involved in communication between leukocytes.
Interferons (IFN): Proteins that inhibit viral replication.
Tumor Necrosis Factors (TNF): Originally identified for their tumorigenic effects.
Colony-Stimulating Factors (CSF): Promote the production of blood cells.
Chemokines: A subcategory that attracts specific cells (chemotactic).
KEY CYTOKINES - Details Table
IL-1:
Source: Macrophages, epithelial cells.
Targets: Hypothalamus, endothelium, T cells.
Functions: Induces fever, activates T cells, promotes acute phase response (endogenous pyrogen).
IL-2:
Source: Activated T cells.
Targets: T cells, NK cells.
Functions: Promotes T cell proliferation and NK cell activation (therapeutically used as Aldesleukin).
IL-3:
Source: Activated T cells.
Target: Hematopoietic stem cells.
Functions: Supports growth of all hematopoietic cells.
IL-4:
Source: TH2 cells, mast cells.
Targets: B cells, T cells.
Functions: Promotes IgE class switching and TH2 differentiation; pivotal in allergies.
IL-5:
Source: TH2 cells, mast cells.
Targets: Eosinophils, B cells.
Functions: Activates and recruits eosinophils and promotes IgA class switching; elevated in allergic and parasitic infections.
IL-6:
Source: Macrophages, TH2 cells.
Targets: Liver, B cells.
Functions: Induces acute phase protein synthesis and fever; used as inflammation biomarker.
IL-7:
Source: Bone marrow stroma.
Targets: Lymphoid progenitors.
Functions: Critical for lymphocyte development and lineage commitment.
IL-8 (CXCL8):
Source: Macrophages, endothelium.
Target: Neutrophils.
Functions: The main chemotactic signal for neutrophils (targeted by many anti-inflammatory drugs).
IL-10:
Source: TH2 cells, Tregs, macrophages.
Target: T cells, macrophages.
Functions: Anti-inflammatory properties, inhibits TH1 responses, suppresses macrophage activation.
IL-12:
Source: Macrophages, dendritic cells.
Targets: T cells, NK cells.
Functions: Induces TH1 differentiation, activates NK cells, stimulates IFN-γ production, essential for cell-mediated immunity.
TNF-α:
Source: Macrophages, T cells.
Functions: Induces fever, promotes acute phase response, and apoptosis; a critical target for therapeutic biologics (anti-TNF drugs).
IFN-α/β (Type I):
Source: Infected cells, plasmacytoid dendritic cells.
Functions: Induces an antiviral state by upregulating MHC I and activating NK cells; used therapeutically for Hepatitis infections.
IFN-γ (Type II):
Source: TH1 cells, NK cells.
Functions: Activates macrophages, enhancing their abilities (M1), upregulates MHC II; critical for immune response against mycobacteria.
CYTOKINE-BLOCKING DRUGS (BIOLOGICS)
Monoclonal Antibodies - Detailed:
Adalimumab (Humira): Targets TNF-α; used for rheumatoid arthritis, Crohn's, and psoriasis; notable side effects include increased infection risk and heart failure.
Infliximab (Remicade): Chimeric anti-TNF; utilized in similar conditions; associated with risks such as TB reactivation and infusion reactions.
Etanercept (Enbrel): Soluble receptor for TNF; indications are the same but distinct side effects include demyelinating disease.
Tocilizumab (Actemra): Blocks IL-6 receptors; indicated for rheumatoid arthritis and cytokine storm; watch for liver enzyme elevation.
Anakinra (Kineret): IL-1 receptor antagonist; targets autoinflammatory diseases but can cause injection site reactions.
Canakinumab (Ilaris): Neutralizes IL-1β; similarly indicating for autoinflammatory conditions.
Ustekinumab (Stelara): Targets IL-12/IL-23 for psoriasis and Crohn's; risks include infectious complications.
Secukinumab (Cosentyx): Targets IL-17A, useful for psoriasis but be aware of Candida infections.
Natalizumab (Tysabri): Blocks leukocyte migration, indicated for MS; linked to progressive multifocal leukoencephalopathy (PML).
Omalizumab (Xolair): Anti-IgE; is useful in severe allergic asthma but can occasionally trigger anaphylaxis.
Rituximab (Rituxan): Targets CD20 on B cells for various conditions, leading to B cell depletion.
PAIN MEDIATORS
Sources and Functions
Bradykinin: Produced from the kinin system, induces vasodilation, increases vascular permeability, and causes pain.
Prostaglandin E2 (PGE2): Derived from the cyclooxygenase pathway, causes fever, pain, and inflammation.
Clinical Note: Non-steroidal anti-inflammatory drugs (NSAIDs) like ibuprofen block COX, thereby decreasing PGE2 and its related effects.
ACUTE PHASE REACTANTS
What Are Acute Phase Proteins?
Acute phase proteins are proteins synthesized by the liver that increase or decrease in concentration during acute inflammation.
Triggers: Produced in response to cytokines IL-1, IL-6, and TNF-α from macrophages that signal liver hepatocytes.
POSITIVE ACUTE PHASE PROTEINS
Function and Clinical Use:
C-Reactive Protein (CRP): Acts as an opsonin, binding to bacteria; serves as a biomarker for inflammation (elevated in infections, myocardial infarctions, autoimmune diseases).
Serum Amyloid A (SAA): Recruits immune cells; can lead to amyloid deposit formation if inflammation is chronic.
Fibrinogen: Involved in clotting and raises the erythrocyte sedimentation rate (ESR) during inflammation, reflecting increased ESR response.
Hepcidin: Regulates iron transport and is used for monitoring diseases associated with chronic inflammation.
Complement proteins (e.g., C3, C4): Enhance immune response.
α1-Antitrypsin: Protects lungs by inhibiting neutrophil elastase; deficiency associated with emphysema.
Other acute phase reactants include haptoglobin (which binds free hemoglobin) and ceruloplasmin (copper transport).
NEGATIVE ACUTE PHASE PROTEINS
Normal Functions and Clinical Relevance:
Albumin: Maintains oncotic pressure and serves as a transport protein; decreases in concentration as liver shifts production to acute phase reactants during inflammation.
Transferrin: A transport protein that reduces during inflammation as the body attempts to hide iron from bacteria.
ESR (Erythrocyte Sedimentation Rate):
Measures the speed of RBCs falling in a tube; elevated during inflammation due to increased fibrinogen levels causing RBC clumping.
Useful in monitoring conditions like infections and autoimmune diseases.
ANEMIA OF CHRONIC DISEASE
Caused by hepcidin-induced iron sequestration leading to low serum iron levels despite normal iron stores in the body, contrasting with traditional iron deficiency anemia.