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Innate Immunity
Jason Ryan, MD, MPH
Barriers to Infection

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Connexions/Wikipedia

Skin/ Innate Adaptive

Mucous Immune Immune
Membranes System System
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Immune Systems
Innate Adaptive
• Fast-acting system • Slow-acting (days)
• Non-specific reaction • Highly specific
• Same cells, same reaction • Unique cells activated to
to many invaders respond to a single
• No memory invader
• 2nd infection same • Memory
response as 1st infection • 2nd infection: faster
response
Antigen Presentation
• Innate system can be activated by “free” antigen
• Pathogenic molecules detected freely in blood, tissue
• Adaptive system requires “antigen presentation”
• Pathogens must be engulfed by cells, broken down
• Pieces of protein transported to surface
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• Antigen “presented” to T-cells for activation
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Cytokines
• Cell signaling proteins
• Often released by immune cells
• Stimulate inflammatory response
• Various subsets
• Chemokine: Attracts immune cells (chemotaxis)
• Interleukins: IL-1, IL,2, etc
• Tumor necrosis factor (TNF): Can cause tumor death
• Transforming growth factor (TGF)
• Interferons: Named for interfering with viral replication
Cluster of Differentiation (CD)
• Cellular surface molecules
• CD3, CD4, CD8
• Found on many immune cells (T-cells, B-cells)
• Used to identify cell types
• Some used as receptor/cell binding
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Innate Immune System

• Phagocytes
• Macrophages (hallmark cell)
• Neutrophils
• Complement
• Natural Killer Cells
• Eosinophils
• Mast cells and Basophils
Innate Immunity
General Principles

• Recognize molecules that are “foreign”

• “Pathogen-associated molecular patterns” (PAMPs)
• Present on many microbes
• Not present on human cells
• Pattern recognition receptors
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• Key receptor class: “Toll-like receptors” (TLRs)

• Macrophages, dendritic cells, mast cells
• Recognize PAMPs → secrete cytokines
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Innate Immunity
Pattern Recognition

• Endotoxin (LPS)
• LPS binds LPS-binding protein (found in plasma)
• Binds CD14 on Macrophages
• Triggers TLR4
• Cytokine production: IL-1, IL-6, IL-8, TNF
• Peptidoglycan cell wall
• NOD receptors (intracellular)
• Nucleotide-binding oligomerization domain
• Cytokine expression
Innate Immunity
Pattern Recognition

• Mannose (polysaccharide on bacteria/yeast)

• Mannose-binding lectin (MBL) from liver
• Activates lectin pathway of complement activation
• Lipoteichoic acid on Gram-positive bacteria
• Double stranded RNAAfraTafreeh.com
• Unmethylated DNA
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Monocytes and Macrophages

• Macrophages: guardians of innate immunity
• Produced in bone marrow as monocytes
• Circulate in blood ~3 days
• Enter tissues → macrophages
• Kupffer cells (liver)
• Microglia (CNS)
• Osteoclasts (bone)

Dr Graham Beards/Wikipedia
Monocytes and Macrophages
• Three key functions:
• Phagocytosis
• Cytokine production
• Antigen presentation
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Dr Graham Beards/Wikipedia
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Phagocytosis
• Macrophages engulf pathogens into phagosome
• Phagosome merges with lysosome
• Lysosomes contain deadly enzymes
• Death of bacteria, viruses

Graham Colm/Wikipedia
Phagocytosis
• Reactive oxygen species (superoxides)
• Produced by NADPH Oxidase (respiratory burst)
• Generate hydrogen peroxide H2O2 and O2-
• Reactive nitrogen intermediates
• NO (nitric oxide) + O2− AfraTafreeh.com
(superoxide) → ONOO− (peroxynitrite)
• Enzymes:
• Proteases
• Nucleases
• Lysozymes (hydrolyze peptidoglycans)
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Lysosome Enzyme Secretion

Lung Abscess

Yale Rosen/Wikipedia
Phagocytosis
• Some pathogens block this process
• Tuberculosis modifies phagosome
• Unable to fuse with lysosome
• Proliferation inside macrophages
• Protection from antibodies
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• Chediak-Higashi Syndrome
• Immune deficiency syndrome
• Failure of lysosomes to fuse with phagosomes
• Recurrent bacterial infections

CDC/Public Domain/Wikipedia
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Macrophages
• Macrophages can exist in several “states”
• Resting: Debris removal
• Activated (“primed”): more effective
• Major activators (via surface TLRs):
• LPS from bacteria
• Peptidoglycan
• Bacterial DNA (no methylation)
• Also, IFN-γ from T-cells, NKC
• Attracted by C5a (complement)
Macrophages
Key Surface Receptors

Surface Molecule Signal

CD14 LPS (gram negatives)
Fc receptor Fc (antibodies)
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C3b receptor C3b (complement)
T-Cell Molecules
MHCII CD4
B7 CD28
CD40 CD40L
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Macrophages
Cytokines

• Key cytokines are IL-1 and TNF-α

• Others: IL-6, IL-8, IL-12
IL-1 and TNF-α
• Both ↑synthesis endothelial adhesion molecules
• Allows neutrophils to enter inflamed tissue
• IL-1
• “Endogenous pyrogen” (causes fever)
• Acts on hypothalamus AfraTafreeh.com
• TNF-α
• Can cause vascular leak, septic shock
• “Cachectin:” Inhibits lipoprotein lipase in fat tissue
• Reduces utilization of fatty acids → cachexia
• Kills tumors in animals (“tumor necrosis factor”)
• Can cause intravascular coagulation → DIC
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IL-6, IL-8, IL-12

• IL-6
• Fever
• Stimulates acute phase protein production in liver (CRP)
• IL-8
• Attracts neutrophils
• IL-12
• Promotes Th1 development (cell-mediated response)
Neutrophil
• Derived from bone marrow
• Granules stain pink with Wright stain
• Eosinophils=red, Basophils=blue
• Circulate ~5 days and die unless activated
• Drawn from blood stream to sites of inflammation
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• Enter tissues: Phagocytosis

• Granules are lysosomes (bactericidal enzymes)
• Provide extra support to macrophages

Dr Graham Beards/Wikipedia
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Neutrophil
Blood stream exit

• Rolling
• Selectin ligand neutrophils (Sialyl-Lewis X)
• Binds E-selectin or P-selectin endothelial cells
• Crawling (tight binding)
• Neutrophils express integrin
• Bind ICAM on endothelial cells
• Transmigration
• Neutrophils bind PECAM-1 between endothelial cells
• Migration to site of inflammation
• Chemokines: C5a, IL-8
Neutrophil
Blood stream exit

PMN
SL AfraTafreeh.com

Selectin

Step 1:
IL-1 and TNF stimulate expression selectin
PMNs bind selectin via selectin ligand
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Neutrophil
Blood stream exit

PMN
INT

ICAM

Step 2:
LPS or C5a stimulates integrin on PMNs
Integrin binds ICAM on endothelium
Neutrophils
• Small granules (specific or secondary)
• Alkaline phosphatase, collagenase, lysozyme, lactoferrin
• Fuse with phagosomes → kill pathogens
• Also can be released in extracellular space
• Larger (azurophilic orAfraTafreeh.com
primary)
• Acid phosphatase, myeloperoxidase
• Fuse with phagosomes only
• Band forms
• Immature neutrophils
• Seen in bacterial infections
• “Left shift”

A. Rad/Wikipedia
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Neutrophil
• Do not present antigen
• Phagocytosis only
• Contrast with macrophages: APCs and phagocytes
• Chemotaxins (attracters of neutrophils)
• IL-8 (from macrophages)
• C5a
• Opsonin: IgG (only antibody that binds neutrophils)
Complement
• Complement proteins produced by liver
• Most abundant is C3
• Frequent, spontaneous conversion C3 → C3b
• C3b binds amino and hydroxyl groups
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• Commonly found on surface of pathogens
• Failure of C3b to bind leads to rapid destruction
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Complement
• C3b → MAC formation
• Membrane attack complex
• Forms pores in bacteria
leading to cell death

Wikipedia/Public Domain
Natural Killer Cells
• Two key roles:
• Kill human cells infected by viruses
• Produce IFN-γ to activates macrophages

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Natural Killer Cells

• MHC Class I
• Surface molecule of most human cells
• Presents antigen to CD8 T-cells
• Activates adaptive immunity against intracellular pathogens
• Some viruses block MHC class I
• NKC destroy human cells with reduced MHC I
Natural Killer Cells
• CD16 on surface
• Binds Fc of IgG → enhanced activity
• Antibody-dependent cell-mediated cytotoxicity
• CD56
• Also called NCAM (Neural Cell Adhesion Molecule)
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• Expressed on surface of NK cells (useful marker)
• Also found in brain and neuromuscular junctions
• Aids in binding to other cells
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ADCC
Antibody-dependent cellular cytotoxicity

• Antibodies coat pathogen or cell

• Pathogen destroyed by immune cells
• Non-phagocytic process
• Classic examples: NK cells and Eosinophils

Satchmo2000/Wikipedia
ADCC
Antibody-dependent cellular cytotoxicity

• Natural Killer Cells

• IgG binds to pathogen-infected cells
• CD16 on NK binds Fc of IgG
• NKC kills cell
• Eosinophils AfraTafreeh.com
• IgE binds to pathogens, especially large parasites
• Eosinophils bind Fc of IgE
• Release of toxic enzymes onto parasite
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Natural Killer Cells

• Lymphocytes (same lineage as T-cells and B-cells)
• Do not mature in thymus
• No memory
• Do not require antigen presentation by MHC
Eosinophils, Mast Cells, Basophils
• All contain granules with destructive enzymes
• All can be activated/triggered by IgE antibodies
• Important for defense against parasites (helminths)
• Too large for phagocytosis
• Release of toxic substances kills parasite
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• Main medical relevance is in allergic disease

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Eosinophil
• Granules appear red with Wright stain
• Major basic protein in eosinophils: (+) charge
• Eosin dye: (-) charge
• Discharge contents (cytotoxic enzymes) onto parasites
• Major basic protein (MBP)
• Eosinophilic cationic protein (ECP)
• Eosinophil peroxidase (EPO)
• Eosinophil-derived neurotoxin (EDN)

Bobjgalindo/Wikipedia
Eosinophil
• Activated by IgE
• Antibody-dependent cellular cytotoxicity
• Stimulated by IL-5 from Th2 cells
• ↑eosinophil count characteristic of helminth infection
• Normal % eosinophils <5% or <500 eosinophils/microL
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• Also seen in many allergic diseases

Bobjgalindo/Wikipedia
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Mast cells and Basophils

• Granules appear blue with Wright stain
• Basophils: blood stream
• Mast cells: Tissue
• Bind Fc portion of IgE antibodies
• IgE molecules crosslink → degranulation
• Histamine (vasodilation)
• Enzymes (peroxidases, hydrolases)

Wikipedia/Public Domain
Innate Immune System
• Phagocytes
• Macrophages (hallmark cell)
• Neutrophils
• Complement
• Natural Killer Cells AfraTafreeh.com
• Eosinophils
• Mast cells and Basophils
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Adaptive Immune System

• T-cells
• CD4: Cytokine production
• CD8: Destruction infected human cells
• B-cells
• Antibody production
• Inter-related with innate immunity
• Cytokines
• Antigen presentation
Dendritic Cells
Langerhans Cells

• Skin and mucosal membranes

• Antigen presenters
• Migrate to lymph nodes
• Activate T-cells
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Wikipedia/Judith Behnsen
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Immune Cell Terminology

Eosinophil Basophil Neutrophil
Mast Cell

Granulocytes

Agranulocytes

Lymphocytes Monocytes

Natural Killer T-cells B-cells Macrophage

Cells
Immune Cell Lineage

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A. Rad/Wikipedia