Innate Immune Function
Lecture 4
Dr. Khalid Mehmood
Overview….
• To respond quickly, components of the innate immune system
are genetically programmed to recognize molecules from broad
classes of pathogens.
• Innate immune responses include
• the rapid destruction of an infectious organism,
• activation of phagocytic cells, and
• the localized protective response known as inflammation
• In inflammation, innate (and sometimes adaptive) cells and
molecules are stimulated to isolate and destroy infectious
agents and trigger tissue repair.
Recognition
• First important function - to recognize the immunogen
• Microbes have conserved molecular signature patterns
pathogen associated molecular patterns (PAMPs)
conserved, structural features expressed by microbes but not by the host.
• The innate immune system uses their pattern
recognition receptors (PRRs) to recognize PAMPs
• Because the host does not produce PAMPs, innate
immune system is able to discriminate between self
and non-self.
A. Pathogen Associated Molecular Patterns (PAMPs)
• PAMPs are products of normal microbial physiology; Sugars,
Proteins, Lipids, Nucleic acids, or combinations of these.
• Well known examples of PAMPs are lipopolysaccharide (LPS) of
Gram -ive and peptidoglycan of Gram +ive bacteria
• Bacterial lipopolysaccharide (LPS) is a major constituent of the
outer cell membrane of Gram -ive bacteria.
• Cell-surface molecules on phagocytes (monocytes, macrophages,
dendritic cells, mast cells) have toll-like receptor 4 (TLR4) & other
cell-surface molecules that bind LPS.
• Peptidoglycans are major components of cell wall of Gram +ive
bacteria and are recognized by TLR2 receptors on phagocytic cells.
Binding of PAMPs
on microbial
surfaces by PRRs
on the surfaces of
phagocytes
activates the
phagocytes to
ingest and degrade
the microbes,
cytokine
production and
inflammation.
B. Pattern Recognition Receptors
(PRRs)
• Innate immune cells (phagocytes) recognize PAMPs by Cell surface PRRs, or Soluble
molecules (e.g., complement)
• PRRs are present as extracellular or membrane-bound proteins on phagocytic cells.
1. Toll-like receptors (TLRs): TLRs recognize diverse pathogens.
• After binding to PAMPs, TLR signals nucleus to increased activation of genes for
cytokines and antimicrobial molecules.
• This results in synthesis & secretion of pro inflammatory cytokines and recruitment of
leukocytes at site of infection.
2. Scavenger receptors:
• These are involved in binding of small lipoproteins, polysaccharides, and nucleic acids.
• Cause internalization of bacteria and phagocytosis of host cells undergoing apoptosis.
3. Opsonins: Opsonins bind to microbial surfaces.
• Make microbes more attractive to phagocytic cells, thus facilitating their destruction.
• Receptors for opsonins are present on phagocytic cells.
• Subsequent increased phagocytic destruction of microbes is called opsonization.
C. Markers of abnormal self
Microorganisms may change the host cells.
• MHC class I molecules on all nucleated host
cells are “markers of self”.
• Some viruses cause an infected host cell to
reduce expression of MHC 1 on their surface.
• Similar changes also occur in cancerous cells.
• Such abnormal host cells can alert immune system by expressing stress
signal molecules on their surfaces.
• In humans, these include some heat shock proteins and two molecules
known as MICA & MICB.
• These stress signals are detected by various receptors like TLRs (TLR2 &
TLR4) and the killer activation receptors (KARs) of natural killer (NK) cells
Soluble Defense Mechanisms
Innate immune system also uses soluble molecules to
protect from viral infection, for destruction of
microbes, or increasing phagocytosis.
A. Type I interferons:
• Produced by dendritic cells (IFN-α), fibroblasts (IFN-
β), and other cells in response to viral infection.
• IFNs induce activation of several antiviral defenses
(in both virally infected & non-infected cells) like
RNA-dependent protein kinase (PKR) & apoptosis.
• Also influence activities of phagocytes
B. Microcidal molecules:
Cells in skin & mucous membranes like epithelial cells, neutrophils & macrophages
secrete microcidal peptides called defensins. These peptides form channels in the
cell membranes of bacteria, causing influx of certain ions & lysis.
Other microcidal molecules are cathelicidin, lysozyme, DNases & Rnases
C. Complement:
• Complement is a collective term for a system of enzymes and proteins
that function in both the innate and adaptive immune system.
• In innate immune system, complement can be activated in two ways; via
– alternative pathway (antigen is recognized by its surface characteristics)
– mannan-binding lectin (MBL) pathway
• in the adaptive immune system, complement is activated via the classical
pathway that begins with antigen-antibody complex.
Functions of Complement:
• Lysis of bacteria, cells, & viruses;
• Promotion of phagocytosis (opsonization);
• Triggering of inflammation & secretion of
immunoregulatory molecules; and
• Clearance of immune complexes from
circulation
Complement: Collective term for a system of enzymes & proteins
In innate immunity, complement can be activated in two ways; via
– Alternative pathway (antigen is recognized by its surface characteristics)
– Mannan-binding lectin (MBL) pathway
Complement nomenclature
• Components C1-C9, B, D, & P are native complement (protein)
components.
• Fragments of native complement components (C4a, C5b, Bb etc)
are indicted by lowercase letter .
• Smaller cleavage fragments are assigned the letter “a,” and
• Larger fragments are assigned the letter “b.”
• A horizontal bar above a component or complex indicates
enzymatic activity, e.g., C4͞bC2b.
1. The alternative pathway
• This pathway is initiated by cell-surface constituents that are
recognized as foreign to the host, such as LPS
• A variety of enzymes (e.g., kallikrein, plasmin, elastase)
cleave C3, the most abundant serum complement
component, into several smaller fragments.
• One of these, C3b fragment (unstable) is the major opsonin
of the complement system and readily attaches to receptors
on cell surfaces and starts following cascade…
1. C3b binds Factor B.
2. Factor B in the complex is cleaved by Factor D to produce
C3bBb, an unstable C3 convertase.
3. Two proteins, C3b inactivator (Factor I)
& β1H-globulin (Factor H) act as
negative regulator by inactivating C3b
to avoid overfunctioning.
4. Alternatively, C3bBb binds properdin
(Factor P) to produce stabilized C3
convertase, C3bBbP.
5. Additional C3b fragments join the
complex to make C3bBbP3b, also known
as C5 convertase. It cleaves C5 into C5a
& C5b.
6. C5b inserts into the cell membrane and
is the necessary step leading to
formation of the membrane attack
complex (MAC) and cell lysis.
2. Terminal or lytic pathway
• Result of alternative, MBL or classical
pathway of complement activation.
• The attachment of C5b leads to the
addition of components C6, C7, & C8.
• C8 provides a strong anchor into the
membrane and facilitates the
subsequent addition of multiple C9
molecules to form a pore in the
membrane.
• Loss of membrane integrity results in
the unregulated flow of electrolytes
and causes the lytic death of the cell.
3. Mannan-binding lectin pathway
• Lectins are proteins that bind to specific
carbohydrates.
• This pathway is activated by binding of
mannan-binding lectin (MBL) to mannose-
containing residues of glycoproteins on
microbial surface (Listeria & Salmonella spp).
• MBL is an acute phase protein (serum
proteins whose levels rise rapidly in response
to infection, inflammation or stress).
• MBL, once bound to appropriate mannose-
containing residues, can interact with MBL-
activated serine protease (MASP).
• Activation of MASP leads to subsequent
activation of components C2, C4, and C3,
resulting in formation of C 5 convertase that
initiates cell lysis
4. Anaphylotoxins
• The small fragments (C3a, C4a, C5a) generated by the
cleavage of C3 and C5 in alternative pathway and of C3,
C4, and C5 in the MBL pathway act as anaphylotoxins
• Anaphylotoxins attract and activate different types of
leukocytes
• They draw additional cells to the site of infection to
help eliminate the microbes
• C5a has the most potent effect, followed by C3a & C4a
• Cytokines are secreted by D. Cytokines and chemokines
leukocytes & other cells
• Important in innate,
adaptive immunity and
inflammation
• Cytokines are involved in
many activities e.g,
– chemotaxis,
– activation of specific
cells,
– induction of broad
physiologic changes
• Chemokines is subgroup
of low molecular weight
cytokines
• These are involved in the
chemotaxis (chemical-
induced migration) of
leukocytes.
Cellular Defense Mechanisms
In addition to soluble means of defense, Innate immune system employs
cellular mechanisms to combat infection:
Receptors that recognize pathogens trigger inflammation, and
destruction of microbes by phagocytosis
Also, NK cells detect & destroy infected or transformed host cells
A. Phagocytosis:
Phagocytosis is engulfment & degradation of microbes and other particulate
matter by innate immune cells such as macrophages, dendritic cells,
neutrophils, and even B lymphocytes (prior to their activation).
These cells defend the body by ingesting microbes, & remove cellular debris
that arise from normal physiologic functions.
Phagocytosis involves cell-surface receptors associated with specialized
regions of the plasma membrane called clathrin coated pits.
Dendritic cells use an additional mechanism of macropinocytosis. This process
does not involve clathrin. Instead, plasma membrane “ruffles” or projections
fold back upon the membrane to engulf extracellular fluids.
1. Recognition & attachment of microbes by phagocytes
Phagocytosis initiates when a phagocyte binds a
cell or molecule that has penetrated the body's
barriers.
The binding occurs at following receptors on the
phagocyte surface:
• PRRs (including TLRs) that recognize PAMPs,
• Complement receptors (CR) that recognize
certain fragments of complement (especially
C3b) that adhere to microbial surfaces,
• Fc receptors that recognize immunoglobulins
that have bound to microbial surfaces or
other particles,
• Scavenger receptors, and
• Others.
2. Ingestion of microbes and other material
After attachment,
Microbe is engulfed by pseudopodia
(extensions of cytoplasm & cell membrane)
Internalization or endocytosis
Dendritic cells can extend projections &
encircle extracellular fluids to form
cytoplasmic vesicles without attachment.
Once internalized, bacteria are trapped
within phagocytic vacuoles (phagosomes) or
cytoplasmic vesicles in the cytoplasm.
This trigger changes within the phagocyte i.e,
increases in size, more aggressive &
increased production of certain molecules
Some of these molecules contribute to
destruction of ingested microbes; others act
as chemotactic agents and activators for
other leukocytes (Cytokines & Chemokines)
Phagosomes, the membrane-bound organelles 3. Destruction of
containing ingested microbes/materials, fuse ingested microbes &
with lysosomes to form phagolysosomes.
other materials
Lysosomes employ multiple mechanisms for
killing and degrading ingested matter. These
include:
• Lysosomal acid hydrolases, including
proteases and nucleases.
• Oxygen radicals, including superoxide radicals
(O2–), hypochlorite (HOCl–), hydrogen peroxide
(H2O2), and hydroxyl radicals, that are highly
toxic to microbes. The combined action of
these molecules causes oxidative burst
• Nitrous oxide (NO)
• Decreased pH
• Other microcidal molecules
4. Secretion of cytokines and chemokines
Once activated, phagocytes secrete cytokines and
chemokines that attract and activate other cells involved
in innate immune responses.
Cytokines or chemical messengers such as interleukin-1
(IL-1) and interleukin-6 (IL-6) induce the production of
proteins that lead to elevation of body temperature.
Other cytokines, such as tumor necrosis factor-α (TNF-α),
increase the permeability of local vascular epithelia to
increase movement of cells and soluble molecules from
the vasculature into the tissues.
Still others, such as interleukin-8 (IL-8) and interleukin-12
(IL-12), attract and activate leukocytes such as
neutrophils and NK cells.
Cellular Defenses… B. Natural Killer Cell
NK cells detect & target abnormal host cells for
destruction.
NK cells possess killer activation receptors (KAR)
that recognize stress-associated molecules,
including MICA and MICB on the surface of
infected and transformed host cells.
Binding of KAR to MICA and MICB generates a kill
signal.
Killer inhibition receptor (KIR) regulates such
killing
Inflammation
Components of both the innate and adaptive immune systems may respond
to certain antigens to initiate a process known as inflammation.
The cardinal signs of inflammation are pain (dolor), heat (calor), redness
(rubor), swelling (tumor), and loss of function (functio laesa).
Enlarged capillaries that result from vasodilation cause redness (erythema)
and an increase in tissue temperature.
Increased capillary permeability allows for an influx of fluid and cells, causes
swelling (edema).
Phagocytic cells release lytic enzymes at the site, damaging healthy cells.
An accumulation of dead cells and fluid forms pus, while mediators released
by phagocytic cells stimulate nerves and cause pain.
The innate immune system contributes to inflammation by
– activating the alternative and lectin-binding complement pathways,
– attracting & activating phagocytic cells to secrete cytokines & chemokines,
– activating NK cells,
– altering vascular permeability and increasing body temperature
Physiological Events of the Acute Inflammatory Response.
(a) At the site of injury (splinter), chemical messengers are
released from the damaged tissue, mast cells, and the
blood plasma. These inflammatory chemicals stimulate
neutrophil migration, diapedesis, chemotaxis, and
phagocytosis.
(b) Neutrophil integrins interact with endothelial
selectins (1) to facilitate margination (2) and
diapedesis (3).
