Lecture 11: Effector Response Against Infection (1)

Immunity to a Pathogen

• Early innate immune response – Provides rapid protection (within hours or days) after the pathogen enters the body, helping to limit damage and keep us alive.

• Adaptive immune response – Involves B-cells and T-cells; develops over days, clears the infection, and creates long-lasting immunity through memory cells.

Innate Immunity

• 1st line of defence - aims to deal with the pathogen quickly → rapidly drives immune response

• Involves cells from the myeloid lineage (derived from a common myeloid progenitor cell precursor in the bone marrow)

• Cells mediating this response are activated by pattern recognition receptors e.g. TLRs → recognise LPS from bacteria rapidly to generate a response follow danger recognition

Myeloid Cells

• Cells differ in size and shape

• They contain granules packed with enzymes and noxious chemicals that allow them to kill pathogens

• They are composed of 2 main families

  • Phagocytes e.g. macrophages, DCs and neutrophils → engulf bacteria

  • Granulocytes, e.g. basophils and eosinophils → full of granules (+ unusual shaped nuclei

Phagocytes

• Macrophages & Neutrophils:

  • Perform phagocytosis: engulf pathogens and kill them using internal compartments.

  • Macrophages can later act as antigen-presenting cells (APCs).

• Dendritic Cells:

  • Phagocytose pathogens and present antigenic peptides to T-cells.

  • Crucial for activating the adaptive immune response.

Macrophages

• Most abundant immune cells, found in all tissues from embryonic development.

• Act as professional phagocytes – engulf pathogens and dead/dying cells.

• Origins:

  • From monocytes (bone marrow-derived, circulate in blood, differentiate in tissues).

  • From embryonic cells (seed tissues early on, long-lived, tissue-resident, don’t reproduce).

• Use special receptors on the surface to detect and phagocytose pathogens e.g. TLR2, TLR4, FcγRIIA (CD32), FcyRIIIA

• Name means “big eaters” (Macro = big, phage = eater)

Neutrophils

• Make up 90% of circulating granulocytes

• Very readily activated – circulating and are rapidly activated in response to a pathogen

• Move into sites of inflammation from the circulation to provide rapid innate immunity

• Short lifespan (dead neutrophils = pus)

  • Pus is a build-up of white cellular mass from neutrophils that have entered an area and ‘killed everything’ and then died

• Highly phagocytic

• Can sense cues from and move towards pathogens = chemotaxis (as can macrophages)

Process of Phagocytosis

• Recognition: receptors on the phagocyte’s surface chemically bind to pathogens (e.g. LPS, phosphoproteins).

• Internalisation: Pathogen is engulfed into a phagosome (vacuole containing the pathogen),

• Destruction in Phagolysosome:

  • Phagosome fuses with a lysosome → forms a phagolysosome.

  • Killing mechanisms:

    • Acidification – low pH is lethal.

    • Toxic contents – noxious chemicals e.g. enzymes & reactive oxygen species damage pathogen walls.

    • In neutrophils, granules with enzymes (MPO)/antimicrobial peptides (a-defensins) also fuse with the phagosome to enhance killing.

Granulocytes

• Contain pre-stored, pre-synthesised granules: allow for rapid pathogen killing without needing new protein synthesis.

  • Granule contents = toxic molecules & chemical mediators.

• Types:

  • Neutrophils: effective phagocytes, full of granules.

  • Eosinophils & Basophils: poor at phagocytosis, act via granule release.

• Become highly abundant in disease states (e.g. asthma).

  • When inappropriately activated, it can contribute to disease.

Eosinophils

• Rare BCs: 2-5% blood white blood cells

• Toxic Granules: Eosinophil cationic protein and Major Basic Protein –

  • Highly enzymatic and noxious molecules that help to kill off pathogens

• Activated to degranulate by a number of mechanisms:

  • e.g. complement, antibody, cytokines

Basophils

• Rare Blood cells: 0.2% of white blood cells

• Toxic Granules: Histamine, Prostaglandins, Heparin and Leukotrienes

  • Histamine linked to allergies – immune system drives the release of histamine molecules = symptoms of an allergic response

• Activated to degranulate by number of mechanisms:

  • e.g. complement, antibody, cytokines

Mast Cells

• Basophil-like, but found in the tissue, not the blood

  • Foundin barrier tissues e.g skin the lung and the intestine - barriers to the outside world and areas we are more likely to encounter an infection – the immune system is highly active

• Bind and detect IgE antibody, often produced in a type 2 setting, on cell surface with high affinity at rest

• Mast cell are activated when IgE binds an antigen (multivalent) = release granules

Natural Killer (NK) Cells

• Innate immune cells that act rapidly and non-specifically.

• Kill altered cells (e.g. virus-infected or tumor cells) by releasing cytotoxic molecules e.g. TNF-a, perforin, granzymes A, B, H,K M

  • FasL (CD95L) binds to Fas to trigger apoptosis

• Require direct physical contact with target cells.

• Provide a backup defence when pathogens evade other immune responses.

• Evolved multiple killing mechanisms to adapt to pathogen escape strategies.

NK Response

• Cells recognise self-antigen peptides presented by MHC I, which normally act as anti-apoptotic signals.

• Use two types of receptors:

  • Inhibitory receptors bind to MHC I → sends “off” signal, prevents killing of healthy cells.

  • Activating receptors turn NK cells on when abnormal signals are detected.

• If a cell is normal (MHC I present) → inhibitory signal prevents killing and attack.

• If a cell is abnormal (e.g. virus-infected or tumour cell causing modification and MHC I is missing) → inhibitory signal lost, NK cell is activated.

• NK cell physically interacts with the target and releases cytotoxic molecules to kill it.

Bidirectional Communication of Innate and Adaptive Immune Response

• Innate immune cells produce cytokines that help drive specific T and B cell responses (adaptive immunity).

• Adaptive immune cells produce cytokines and antibodies that enhance and regulate the innate immune response.

  • This bidirectional communication ensures a coordinated and effective immune defence.

Complement System

• A major component of the innate immune system → preformed soluble molecules present in the circulation that help to mediate immunity

• Collection of 30+ soluble proteins, produced in the liver as inactive precursors.

  • Circulates in blood and body fluids (inactive)

• Enhances antibody-mediated killing of bacteria (discovered in the 1890s).

• Also provides early, antibody-independent killing of pathogens.

Complement Proteins

• Proteins produced in the liver have enzymatic function,

• They are produced as inactive precursors, which once cleaved (broken down via an enzymatic reaction into 2 components) become active.

• This cleaved component can often have roles in inflammation, but the remaining fragment has further enzymatic sites.

• Complement protein cleavage and activation is a cascade, with activation of one component activating further steps to provide a powerful form of immunity through this soluble enzymatic process.

  • May have an active enzymatic site which may help to cleave bacteria and a binding site for the next protein in the cascade.

3 Pathways on Complement Activation

• Classical Pathway: Complement binds to antibody bound to pathogen surface

• Alternative Pathway: Complement binds to directly to pathogen surfaces (via recognition of conversed sequences)

• Lectin Pathway: Complement binds to Mannose binding protein bound to pathogen surface

  • All converge to produce a combination of molecules → C3 convertase

C3 Convertase

• An enzyme that cleaves the C3 complement to form C3b which then kills the pathogen

Classical pathway

• C1 is the first component of the pathway and has a pentameric structure with 3 subunits: C1q, C1r, C1s.

• C1q binds to antibodies (usually IgM or IgG) that are already bound to pathogens.

• Binding of C1q activates C1r, which activates C1s, starting the complement cascade.

Classical Pathway: Activation of C1s

• Oten involves an antibody.

• It targets pathogens that have already been bound by an antibody.

• Innate antibodies like IgM are produced by B-cells and are pre-optimised to bind pathogens.

  • IgM has a pentameric structure (5 Ab molecules together) and binds to pathogens.

  • C1q (with its pentameric structure) binds to IgM bound to pathogens for molecular recognition.

• IgG antibodies, also pre-optimised, can also be recognised by C1q, even though they don’t have the same exact molecular pattern.

• Binding of C1q activates the C1r component, leading to cleavage and activation of C1s.

Classical Pathway: Activation of C3 Convertase

• C1 cleaves C4 into C4a and C4b

• C4b sticks to the bacteria and has an enzymatic function and cleaves C2 to form C2a and C2b

• C2a then fuses to C4b to form C4B2A

• Cascade of cleavage to form a complex of complement cascade molecules on the surface of the bacteria e.g. with C1, C4 C2, which acts as a ‘Master Regulator’ of C3 cleavage -C3 Convertase, which then cleaves C3 into C3a and C3b

• C3b then kills the pathogen

Alternative Pathway

• Does NOT require antibody

• Involves other complement factors B and D – same result of C3 3b generation

• Complement factors B and D are able to activate C3 on their own resulting in C3 convertase that kills the pathogen via C3b generation

  • C3 spotaneously forms C3 H2O

  • Factor B binds to C3(H₂O), creating a complex

  • Factor D (a serine protease) cleaves the bound Factor B into Ba and Bb fragments

  • The resulting C3(H₂O)Bb complex serves as the initial C3 convertase

Lectin Pathway

• Mannose-binding lectin (MBL) is present in serum and binds to mannose, a conserved glycoprotein molecule found on the surface of pathogens (e.g., bacteria, fungi).

• When MBL binds to pathogens, MBL-associated proteases (MASPs) are recruited, activating enzymatic activity.

• The MBL-MASP complex cleaves C4 and C2, just like the classical pathway.

• Instead of antibodies, MBL recognises conserved patterns on the pathogen’s surface, and recruits the complement cascade to activate C3

• The cleavage of C4b follows the same sequence as the classical pathway, leading to C3 activation.

Modes of Pathogen Removal Following C3 Cleavage

• 1. Induction of cell lysis – forms holes in the pathogen

• 2. Opsonisation of pathogen (“prepare the table”) --> signals the macrophage to the presence of the pathogen

• 3. Induction of chemotaxis and inflammation – chemical/soluble cues to recruit other immune cells to the site of damage/ pathogen

• 4. Immune complex clearance (small antibody-soluble antigen complexes – immune complexes, not enough antibody to mediate phagocytosis)

First Cellular Defence Against Pathogens

• Mediated by phagocytes

  • Macrophages

  • Neutrophils

  • Kill pathogen inside the cell

    • Phagosome/phagolysosome

• Other important cellular arms of innate immunity:

  • Granulocytes (neutrophils, eosinophils, basophils)

  • Mast cells

  • NK cells

Removal of Pathogens

• Mediated by the complement system

  • most proteins are inactive initially

  • system involves cascade of complement component activation 

• 3 pathways that can trigger the complement cascade

  • Classical (antibody-mediated)

  • Alternative (doesn’t require antibody)

  • Lectin pathway (via mannose-binding lectin)