ABBAS 2

Question 1

Characteristics of innate immunity

Answer 1

 Defence mechanisms always present
 Recognise and respond to microbes
 Do not react against nonmicrobial substances
 Can be triggered by host cells that are damaged by microbes
 Differences from adaptive immunity
o Adaptive must be stimulated by and adapts to encounters with microbes before it can be effective
o Adaptive may be directed against microbial and nonmicrobial Ags
 Specifically targets microbes

Question 2

Recognition of microbes

Answer 2

1. Innate recognises structures shared by various classes of microbes and not present on host cells
2. Recognise structure of microbes that are essential for survival and infectivity of microbes
3. Recognise molecules released from stressed or necrotic cells : Damage associated molecular patterns
4. Receptors of innate encoded in germline, not produced by somatic recombination
5. Does not react against host
6. Responds in the same way to repeat encounters with the same microbes
7. 2 principle of reactions

Question 3

1. Innate recognises structures shared by various classes of microbes and not present on host cells

Answer 3

o Pathogen-Associated Molecular Patterns (PAMP)
 Bacterial lipopolysaccharide (LPS) / endotoxin – bacterial cell wall
 Terminal mannose residues – bacterial
 Double-stranded RNA – viruses
 Unmethylated CpG
oligonucleotides – microbial DNA
o Pattern Recognition Receptors (PRR) – receptors that recognise PAMP
o Innate immunity can bind to host cells but do not react against host cells due to lack of activation mediated by regulatory molecules present on host cells but not on microbes

Question 4

2. Recognises structures of microbes that are essential for survival and infectivity of microbes

Answer 4

o Thus microbe cannot evade innate immunity by mutating or not expressing targets of innate immune recognition
o Microbes w/o functional forms of recognition structures lose ability to infect and colonise host
o But microbes often evade adaptive immunity by mutating Ags recognised by lymphocytes as these Ags are not required for microbial life

Question 5

3. Recognises molecules released from stressed or necrotic cells

Answer 5

Damage associated molecular pattern (DAMPs)

Question 6

4. Receptor of innate immunity encoded in germline, not produced by somatic recombination

Answer 6

o Predetermined specificity for microbes
o Less diverse specificity than adaptive immunity
o Receptors are nonclonally distributed unlike adaptive
 Identical receptors expressed on all cells of the same type
 Many cells of innate recognise and respond to the same microbe

Question 7

5. Does not react against host

Answer 7

o Inherent specificity for microbial structures
o Host cells express regulatory molecules that prevent innate rxns
o Adaptive also diff bt self and non-self: lymphocytes that recognise self-Ag are killed or inactivated

Question 8

6. Responds in the same way to repeat encounters with microbes

Answer 8

(unlike adaptive: more efficient with each successive encounter with microbe)
o No immunologic memory (unlike adaptive)

Question 9

7. 2 Principle types of reactions

Answer 9

o Inflammation: recruit and activate leukocytes

o Anti-viral defence: mediated by NK cells, cytokines and interferons

Question 10

Cellular receptors for microbes

Answer 10

	Receptors are expressed on
   o	Phagocytes
   o	DCs
   o	Lymphocytes
   o	Epithelial and endothelial cells

 Expressed on
o Cell surface
o Endoplasmic reticulum - rapidly recruited to vesicles (endosomes) into which microbial products are ingested
o Cytoplasm - sensors of cytoplasmic microbes

 Molecular patterns: diff classes of receptors specific for diff types of microbial proteins

 Toll-like receptors (TLRs)

 Other receptors in innate immunity

 Receptors associate with inflammasome (multi-protein complex tt transmits signals tt activate enzyme to cleave a precursor of cytokine interleukin-1 (IL-1) into bioactive form)

Question 11

Toll-like receptors (TLRs)

Answer 11

o Cell surface receptors on phagocytes and other cell types

o Act as pattern recognition receptors in innate immune response to LPS and microbial products

o Similar to type 1 interleukin-1 receptor

o Specific for diff components of microbes
 TLR 2: bacterial lipoglycans
 TLR 3, 7, 8: viral nucleic acids (ds RNA)
 TLR 4: bacterial LPS
 TLR 5: bacterial flagellin
 TLR 9: unmethylated CG rich (CpG) oligonucleotides (more in bacterial cells than mammalian cells)

o Present on
 Cell surface: recognise products of extracellular microbes
 In endosomes into which microbes are ingested

o Engagement of TLRs
 Activate transcription factors: stimulate exp of cytokines, enzymes and proteins involved in antimicrobial fxns of activated phagocytes and DCs

o Activate transcription factors
 NF-κB (Nuclear Factor-κB): promotes exp of cytokines and endothelial adhesion molecules
 IRF-3 (Interferon Response Factor-3): promote production of type I interferons, cytokines tt block viral replication

Question 12

Other receptors in innate immunity

Answer 12

o Cell surface receptor: peptide that begin with N-formyl methionine (bacterial protein)
o Receptor for terminal mannose residues: phagocytosis of bacteria
o Cytoplasmic receptors
 Viral nucleic acids
 Bacterial peptides
 Microbes
 Components of dead cells eg. Uric acid, DNA

Question 13

Interleukin 1 (IL-1)

Answer 13

 Powerful inducer of inflammatory reaction to microbes and damaged tissues
 Gain-of-fxn mutations cause autoinflammatory syndromes: excessive IL-1; treatment: IL-1 antagonists

Question 14

Common sites of entry from epithelial cells

Answer 14

o	Skin (physical contact)
o	GIT (ingestion)
o	Respiratory tract (breathing)

Question 15

How does epithelial prevent entry of microbes

Answer 15

 Protected by continuous epithelia tt provide physical and chemical barriers against infection
 Epithelial cells produce peptide antibiotics to kill bacteria
 Intraepithelial lymphocytes (T cell lineage but express Ag receptors of limited diversity)
o Recognise microbial lipids
o Express receptors composed of γ and β chains unlike αβ TCR in most T cells

Question 16

Neutrophils (polymorphonuclear leukocytes - PMNs)

Answer 16

• Most abundant leukocyte
• Production stimulated by cytokines (colony-stimulating factors)
• Cytokines act on bone marrow stem cells to stimulate proliferation and maturation of neutrophil precursors
• 1st cell to respond to most infections
• Ingest microbes in circulation
• Rapidly enter extravascular tissues at sites of infection
• Short life

Question 17

Monocytes / Macrophages

Answer 17

o Less than neutrophils
o Ingest microbes in blood and tissues
o Survive for long period in extravascular tissue unlike neutrophils
o Differentiate into macrophages in extravascular tissue – Mononuclear phagocyte system
o Resident macrophages found in connective tissues and every organ in body

Question 18

Action of phagocytes

Answer 18

o Migrate to extravascular sites of infection
o Use receptors to recognise microbes in blood and extravascular tissues
o Phagocytose (ingest) microbes and destroy ingested microbes in intracellular vesicles
o Macrophages respond to products of T cells and fxn as effector cells of cell-mediated immunity

Question 19

How does leukocytes migrate to extravascular sites of infection?

Answer 19

o Binding to endothelial cells -> adhesion -> transmigration thru epithelium
o In response to chemoattractants produced on encounter with microbes

	Upon infection
o	Selectin-mediated rolling
o	Integrin-mediated firm adhesion
o	Chemokine-mediated motility
o	Migration of blood leukocytes to extravascular site of infection

Question 20

Which cytokines are produced by in response to infectious microbes that breach epithelium?

Answer 20

o Tumour Necrosis Factor (TNF)

o Interleukin-1 (IL-1)

Question 21

How does the cytokines produced act?

Answer 21

o Act on endothelium of small vessels at site of infection
o Stimulate endothelial cells to rapidly express 2 adhesion molecules (E-selectin and P-selectin)
o Selectin: carb-binding property of molecules
o Circulating neutrophils and monocytes express surface carbs tt bind weakly to selectins

Question 22

How does selectin produced recruit leukocytes?

Answer 22

 “Rolling” of leukocytes on endothelial surface
o Neutrophils bind to selectins on endothelial surface – become tethered to endothelium
o Flowing blood disrupts binding
o Bonds re-form downstream

Question 23

Integrins

Answer 23

o Adhesion molecules expressed by leukocytes
o “integrate” extrinsic signals into cytoskeletal alterations
o Present in low-affinity state on unactivated leukocytes

Question 24

What are chemokines?

Answer 24

o Chemoattractant cytokines
o Produced by tissue macrophages that encounter the microbe and endothelial cells responding to macrophage-derived TNF and IL-1
o Bind to glycoproteins on luminal surface of endothelial cells
o Displayed at high conc to leukocytes rolling on endothelium
o Stimulate rapid increase in affinity of leukocyte integrins for their ligands on endothelium
o TNF and IL-1 act on endothelium to stimulate expression of ligands for integrins
o Firm binding of integrins to their ligands arrests rolling leukocytes on endothelium
o Cytoskeleton of leukocytes reorganised -> cells spread out on endothelial surface
o Chemokines stimulate motility of leukocytes which then migrate bt endothelial cells -> thru vessel wall (along chemokine conc gradient from low to high to site of infection: chemotaxis)

Question 25

Inflammation

Answer 25

o Accumulation of leukocytes at sites of infection o Vascular dilation o Increased leakage of fluid and proteins

Question 26

Leukocyte adhesion deficiency

Answer 26

o Defective leukocyte recruitment to sites of infection o Due to inherited deficiencies in integrins and selectin ligands o Increase susceptibility to infections

Question 27

Action of receptors in neutrophils and macrophages

Answer 27

1. Activate phagocytes – TLRs, receptors for formyl methionine peptide, receptors for cytokines (IFN-γ, chemokines) 2. Phagocytosis of microbe - mannose receptors, scavenger receptors 3. Ingestion of microbes and activating phagocytes – receptors for products of complement activation and antibodies (receptors avidly bind microbes coated with complement proteins or Abs)

Question 28

Phagocytosis

Answer 28

 Membrane receptors bind to microbe  Extension of phagocyte plasma mb around microbe  Mb closes up and pinches off  Microbe internalised in a membrane-bound vesicle – phagosome  Phagosome fuses with lysosome – phagolysosome

Question 29

How does neutrophils and macrophages destroy microbes?

Answer 29

1. Phagocytosis 2. Receptors of phagocytes signals activation of enzymes in phagolysosomes 3. Microbicidal substances produced in lysosomes and phagolysosomes – act on ingested microbes but do not damage phagocytes o Inflammation can cause tissue injury as microbicidal enzymes and ROS can sometimes be liberated into extracellular space

Question 30

How does phagolysosomes destroy microbes?

Answer 30

 Phagocyte oxidase: convert molecular O2 into superoxide anion and free radicals – Reactive Oxygen Species (ROS) – toxic to ingested microbes  Inducible nitric oxide synthase: arginine -> nitric oxide (microbicidal)  Lysosomal proteases: break down microbial proteins

Question 31

Chronic granulomatous disease

Answer 31

 Inherited deficiency of phagocyte oxidase enzyme  Immunodeficiency disease  Phagocytes cannot eradicate intracellular microbes  Host tries to contain infection by recruiting more macrophages and lymphocytes -> cause granulomas (collection of cells around microbes)

Question 32

Action of macrophages

Answer 32

o Kill phagocytosed microbes o Produce cytokines tt recruit and activate leukocytes o Secrete growth factors and enzymes – repair injured tissue and replace with connective tissue o Stimulate T lymphocytes – enhance adaptive immunity o Respond to products of T cells o Effector cells of cell-mediated immunity

Question 33

Dendritic cells

Answer 33

 Present antigen  Respond to microbes by producing cytokines  Cytokines recruit leukocytes and initiate adaptive immune responses  Bridge bt innate and adaptive immunity

Question 34

NK cells

Answer 34

 Lymphocytes that recognise infected and stressed cells – kill them  Secrete macrophage-activating cytokine (IFN-γ) – macrophages become more effective at killing phagocytosed microbes  Make up 10% of lymphocytes in blood and peripheral lymphoid organs  Contain abundant cytoplasmic granules  Express characteristic surface markers  Do not express immunoglobulins and TCR  Activation triggers discharge of proteins in cytoplasmic granules toward infected cells – induce apoptosis of infected host cells

Question 35

Cooperative function between macrophage and NK cells

Answer 35

o Macrophages ingest microbes and produce IL-2 o IL-2 activates NK cells to secrete IFN-γ o IFN-γ activates macrophages to kill ingested microbes

Question 36

Activation of NK cells

Answer 36

o Determined by balance bt activating and inhibitory receptors ``` o Activating receptors recognise cell surface molecules commonly expressed on stressed cells o Types of stress  Virus  Intracellular bacteria  DNA damage  Malignant transformation ``` o NK cells eliminate irreparably injured and tumour cells o Activating receptors  NKG2D – recognise molecules that resemble class I MHC (expressed during stress)  Receptor specific for IgG Abs bound to cells – Antibody-Dependent Cellular Cytotoxicity (ADCC) – recognition of Ab-coated cells and subsequent killing of these cells o Activating receptors contain immunoreceptor tyrosine-based activation motifs (ITAMs) in cytoplasmic tails  ITAMs also present in subunits of lymphocyte antigen receptors  ITAMs are phosphorylated on tyrosine residues when receptors bind their ligands  Phosphorylated ITAMs then bind and promote activation of cytoplasmic protein tyrosine kinases -> signal transduction pathway -> cytotoxic granule exocytosis and production of IFN-γ

Question 37

Inhibition of NK cells

Answer 37

o Inhibitory receptors specific for self class I MHC expressed on ALL healthy nucleated cells (ie. All cells except RBC) o Block signalling by activating receptors o Inhibitory receptors  Killer cell immunoglobulin-like receptors (KIRs)  Receptors consisting of protein CD94 and lectin subunit NKG2 o Inhibitory receptors contains immunoreceptor tyrosine-based inhibitory motifs (ITIMs)  ITIMs are phosphorylated on tyrosine residues when receptors bind class I MHC  ITIMs then bind and promote activation of cytoplasmic protein tyrosine phosphatases -> remove phosphate groups from tyrosine residues of signalling molecules -> blocking activating receptors of NK cells o When inhibitory receptors of NK cells encounter self MHC, NK cells are shut off o Viruses block expression of class I MHC in infected cells  Evade killing by virus-specific CD8+ cytotoxic T cells  NK cell inhibitory receptors not engaged  Virus induces exp of activating ligands  NK cells are thus activated and eliminate virus-infected cells o Ability of NK cells to protect against infections enhanced by  Cytokines from macrophages  Dendritic cells that encountered microbes o NK-activating cytokines  Interleukin-15 (IL-15): devp and maturation of NK cells  Type I interferons (IFNs): enhance killing fxn of NK  IL-12: enhance killing of NK o Host use CTLs to recognise MHC-displayed viral Ags -> viruses shut off MHC exp -> NK cells evolve to respond to absence of MHC

Question 38

Other classes of lymphocytes that form part of the innate immunity

Answer 38

 Innate immunity  Express somatically rearranged Ag receptors (like B and T cells) but have limited diversity  γβ T cells o In epithelia  NK-T cells o Express surface molecules typically found on NK cells o In epithelia and lymphoid organs o Recognise microbial lipids bound to a class I MHC-related molecule CD1  B-1 cells o B cells found in peritoneal cavity and mucosal tissue o Produce Abs in response to microbes and microbial toxins tt pass thru intestinal walls o Produce most of circulating IgM Abs in blood (natural antibodies) – specific for carbs in bacterial cell wall  Marginal zone B cells o Present at edges of lymphoid follicles in spleen and other organs o Involved in rapid Ab responses to blood-borne polysaccharide-rich microbes  Respond like adaptive immunity (Ab production) but have features of innate immunity (rapid response, limited diversity of Ag recognition)

Question 39

Complement system

Answer 39

 Collection of circulating and membrane-associated proteins  Complement proteins: proteolytic enzymes  Complement activation: sequential activation of enzymes (enzymatic cascade)  Activation of complement system via 3 pathways o Alternative pathway (innate immunity) o Classical pathway (adaptive immunity) o Lectin pathway (innate immunity)  Activated complement proteins o Fxn as proteolytic enzymes to cleave other complement proteins – enzymatic cascade – can be rapidly amplified o Central plasma protein: C3 (cleaved by enzymes generated in early steps)  3 pathways differ in initiation but share late steps and same effector fxns

Question 40

Complement Classical pathway (adaptive immunity)

Answer 40

 Triggered when Ab bind to microbes/Ags

Question 41

Complement Alternative pathway (innate immunity)

Answer 41

 Triggered when complement proteins activated on microbial surfaces  Cannot be controlled since complement regulatory proteins absent on microbes (but present on host cells)

Question 42

Complement Lectin pathway (innate immunity)

Answer 42

 Activated when mannose-binding lectin (plasma protein) binds to terminal mannose residues on surface glycoproteins of microbes  Lectin activates proteins of classical pathway

Question 43

What is the major fragment of C3?

Answer 43

 Major proteolytic fragment of C3: C3b |  C3b covalently attaches to microbes, able to activate downstream complement proteins on microbial surface

Question 44

3 functions of complement system

Answer 44

1. Opsonisation of microbes with complement proteins 2. Promote leukocyte recruitment (inflammation) at site of complement activation 3. Induce osmotic lysis or apoptotic death of microbes

Question 45

Complement - 1. Opsonisation of microbes with complement proteinis

Answer 45

 C3b coats microbes  Promotes binding of microbes to phagocytes (which have receptors for C3b)  Opsonised microbes are quickly ingested and destroyed by phagocytes

Question 46

Complement - 2. Promote leukocyte recruitment at site of complement activation

Answer 46

 Proteolytic fragments of complement proteins esp C3a, C5a are chemoattractants for phagocytes

Question 47

Complement - 3. Induce osmotic lysis or apoptotic death of microbes

Answer 47

 Complement activation leads to formation of a polymeric protein complex  Complex inserts into microbial cell mb  Disrupt permeability barrier  Cause death of microbe

Question 48

TNF

Answer 48

 Promote thrombus formation on endothelium  Reduce BP by a combi of reduced myocardial contractility and vascular dilatation and leakiness  High levels (produced in response to bacteria) cause septic shock (low BP, disseminated intravascular coagulation, metabolic disturbances)

Question 49

IL-12

Answer 49

 Produced by DC and macrophages  In response to LPS and microbial molecules  Activate NK cells -> macrophage activation

Question 50

IFN-GAMMA

Answer 50

 Produced by NK cells  Activate macrophage  Cytokine of both innate and adaptive since T cell produces it too

Question 51

Type-1 interferons

Answer 51

 Produced by DC, macrophages and infected cells in viral infections  Inhibit viral replication  Prevent spread of infection to uninfected cells  Eg. IFN-α:used to treat chronic viral hepatitis

Question 52

Plasma proteins of innate immunity

Answer 52

- Plasma mannose-binding lectin - C-reactive protein ``` - Acute phase response (to infection) o Rapid increase of circulating levels of plasma proteins (acute phase reactants) after infection o Part of innate immune response o Proteins  CRP  Fibrinogen  Serum amyloid A protein o Acute phase reactants synthesised by liver in response to inflammatory cytokines esp IL-6, TNF ```

Question 53

Evasion of innate immunity by microbes

Answer 53

1. Intracellular bacteria resist destruction inside phagocytes o Listeria monocytogenes  Produce protein tt enables it to escape from phagocytic vesicles and enter cytoplasm of infected cells  No longer susceptible to ROS and NO (produced in phagolysosomes) 2. Contain lipid that inhibits fusion of vesicles containing ingested bacteria with lysosomes o Mycobacteria 3. Cell walls resistant to actions of complement proteins

Question 54

How innate immunity stimulates adaptive immunity?

Answer 54

 Innate immune responses generate molecules tt fxn as “second signals” (SS) which activate T and B cells tog with Ags ```  2 signals needed for full activation of Ag-specific lymphocytes o Signal 1: antigen o Signal 2 can be provided by 1. Microbe 2. Innate immune response to microbe 3. Host cell damaged by microbe ```  Need for microbe-dependent SS ensures lymphocytes respond to infectious agents and not to harmless non-infectious substances

Question 55

Adjuvant

Answer 55

o Substance distinct from Ag o Enhance T cell activation by  Promoting accumulation of APC at site of Ag exposure  Enhancing exp of costimulators and cytokines by APC

Question 56

Second Signals

Answer 56

 Costimulators: surface molecules expressed by DC and macrophages which bind to receptors on naive T cells, fxn tog with Ag recognition to activate T cells  Cytokine IL-12: produced by DC and macrophages – stimulate differentiation of naive T cells into effector cells of cell-mediated adaptive immunity o DC and macrophages stimulated by microbes to produce SS to activate T cells

Question 57

How are blood-borne microbes removed by innate and adaptive together

Answer 57

o Microbes activate complement system by alternative pathway o C3d produced is covalently attached to microbe o B cells recognise microbial Ags and C3d -> initiate process of B cell diff to secrete Ab o Complement product serves as SS for humoral immune response