Immune response to extracellular bacteria

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Immune response to extracellular bacteria
The task of the immune system is to eliminate the
bacteria and toxins.
The immune functions that are important for
eliminating extracellular bacteria
Complement
Monocyte/Macrophage
Neutrophil
B cell/antibodies
T cell/APCs
2
Complement
Origin produced primarily by hepatocytes in
liver. Activated macrophage can also produce it.
Location blood and tissue fluid.
Function
Lyse bacteria
Opsonize bacteria to facilitate phagocytosis
Induce inflammation
Clearance of antigen-antibody complex
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Lysis of bacteria
Complements can be activated by three pathways.
degradation
1. Classical pathway
IgM gt IgG
Specificity
Bacteria-antibody(Fc)-C1
C3b
Classical C3 convertase
Bb
C4, C2
C4b2a
RCA
B,D
Alternative C3 convertase
C3bBb
C3
C3b
C3b
C3 amplification loop
C3b2Bb (alternative C5 convertase)
C4b2a3b (Classical C5 convertase)
Require activation of B cells.
C6,7,8,9
C5b
C5b6789
C5
Natural antibodies by B1 cells and Cross-reacting
antibodies may initiate the reaction during
primary immune response.
MAC
HRF, CD59
Lysis of bacteria
C-reactive protein (acute phase response)
also activate complement through C1.
C5b678, no C9
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Lysis of bacteria
2. Lectin pathway
degradation
Specificity
Bacteria-MBL-MASP
C3b
Classical C3 convertase
Bb
C4, C2
C4b2a
RCA
B,D
Alternative C3 convertase
C3bBb
C3
C3b
C3b
C3 amplification loop
C3b2Bb (alternative C5 convertase)
C4b2a3b (Classical C5 convertase)
Early response. Upregulated during acute phase
response.
C6,7,8,9
C5b6789
C5b
C5
MAC
HRF, CD59
Lysis of bacteria
C5b678, no C9
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Lysis of bacteria
3. Alternative pathway
Degradation
C3b
Bb
RCA specificity
Slow spontaneous hydrolysis
B,D
Alternative C3 convertase
C3bBb
C3
C3b
C3b
C3 amplification loop
C3b2Bb (alternative C5 convertase)
Early response.
C6,7,8,9
C5b
C5b6789
C5
MAC
HRF, CD59 specificity
Lysis of bacteria
C5b678, no C9
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Lysis of Bacteria
MAC is effective against Gram negative bacteria.
The thick peptidoglycan in the cell wall of
Gram Positive bacteria prevents the insertion of
MAC.
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Opsonization
C3b is covalently attached to bacteria during
complement activation.
Phagocytes (macrophages, neutrophils) have
complement receptors that can recognize C3b.
C3d facilitate the clustering of B cell
coreceptor (CD21/CD19/TAPA1) with BCR and
stimulate B cell activation.
Inflammation
histamine
vasodilaton
(Increase blood flow)
Mast cells
Prostaglandin leukotrine
C5a, C3a C4a
inflammation
vascular permeability
(leakage of plasma)
Leukocyte extravasation
(Recruitment of neutrophil and monocyte)
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Clearing of soluble antigen-antibody complex
Small antigen-antibody complexes (e.g.
Toxin-antibody complex) are not phagocytosed.
Complement activation by the classical pathway
C3b covalently attached to the antigen
C3b-antigen-antibody complex is taken up by
erythrocytes through CR1
Erythrocytes transport the complex to spleen and
liver
The complex is transferred to Macrophages and
internalized for degradation
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Macrophage
Bone marrow
Hematopoietic stem cell (HSC)
Bone marrow
Primary lymphoid organ
thymus
Lymphocyte maturation
Lymphoid progenitor
Myeloid pregenitor
thymus
T cell progenitor
B cell progenitor
Granulocyte-monocyte progenitor
Blood
monocyte
T cell
Neutrophil
B cell
1-6
50-70
Lymphocyte (20-40)
Lymphocyte circulation
Tissue
Secondary lymphoid organ
Lymphocyte activation
macrophage
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Function of Macrophage
1. Phagoctyosis
2. Produce inflammatory cytokines
3. Antigen presentation to T cells
4. Scavenger to eliminate apoptotic cells and
immune complexes.
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Phagocytosis
Scavenger receptor
Mannose receptor
Phagocytic receptors
?-glucan receptor
bacteria
Bacteria (PAMPs)
Fc receptor (antibody)
phagosome
Complement receptor (C3b)
Opsonin receptors
Collectin receptor (MBL)
Pentraxin receptor (C-reactive protein)
exocytosis
Reactive oxygen intermediates
Oxygen-dependent
Degraded bacteria
Reactive nitrogen intermediates
Bacterial killing
bacteria
Antigen- presentation
Oxygen-independent
phagolysosome
Class II MHC
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Secretion of inflammatory mediators and antigen
presentation
IL-1
Local inflammation
MBL
TNF-?
Inflammatory cytokines
Acute phase response (liver)
IL-6
C-reactive protein
IL-8
Neutrophil activation And chemotaxis
Bacteria (PAMPs)
TLRs
M? activation
Prostaglandin, leukotriene
inflammation
Class II MHC
Antigen presentation T cell activation
B7
(Secondary lymphoid tissues)
Phagocytosis
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Neutrophil
Bone marrow
Hematopoietic stem cell (HSC)
Bone marrow
Primary lymphoid organ
thymus
Lymphocyte maturation
Lymphoid progenitor
Myeloid pregenitor
thymus
T cell progenitor
B cell progenitor
Granulocyte-monocyte progenitor
Blood
monocyte
T cell
Neutrophil
B cell
1-6
50-70
Lymphocyte (20-40)
Lymphocyte circulation
Tissue
Secondary lymphoid organ
Lymphocyte activation
macrophage
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Functions of Neutrophils
Neutrophils (blood circulation)
Induced by inflammatory mediators
Rolling (mucin like CAM - E,P-selectin)
Activation by chemoattractants (IL8(CXCL8), C5a,
N-formyl-peptides)
Inflamed endothelial cells
Activation of integrin (LFA-1, CR3, CR4, common ?
subnit CD18)
Stable interaction between LFA-1 with ICAM
Chemotaxis by IL8, C5a, N-formyl-peptides, etc
Phagocytosis
Degranulation
Kill bacteria
Eliminated by macrophages
apoptosis
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B cells
Bone marrow
Hematopoietic stem cell (HSC)
Bone marrow
Primary lymphoid organ
thymus
Lymphocyte maturation
Lymphoid progenitor
Myeloid pregenitor
thymus
T cell progenitor
B cell progenitor
Granulocyte-monocyte progenitor
Blood
monocyte
T cell
Neutrophil
B cell
1-6
50-70
Lymphocyte (20-40)
Lymphocyte circulation
Tissue
Secondary lymphoid organ
Lymphocyte activation
macrophage
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B cell maturation
Surface marker (flow cytometry)
V(D)J recombination
2nd lymphoid tissues
Bone marrow
GL- ProB
DJ- ProB
Small PreB
sIgM sIgD
Large PreB
HSC
CLP
sIgM
ImmatureB
Mature B
C-kit
IL7R
B220 (CD45R)
B cell marker
CD19
Ig?/?
SLC (?5/VpreB)
RAG-1,2
TdT
IgH (?)
IgL (?)
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B cell developmental stages are characterized by
surface markers and V(D)J recombination.
Cell surface marker is detected by Flow cytometry
Fluorescence-activated cell sorter (FACS)
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V(D)J recombination
1. Organization of the IgH and IgL loci and the
order of recombination.
IgH
D-J joining
V-DJ joining
IgL (? and ?)
V-J joining
Which parts correspond to the immunoglobulin V
and C regions.
2. Basic recombination mechanism.
spacer
heptamer
nanomer
Recombination occurs between 23-bp and 12-bp RSS
(12-23 rule).
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7
9
7
12
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RSS
V
D
J
RAG-1,2 cleaves between RSS and the gene segments
D
V
J
DNA repair to join the DNA breaks
Imprecise coding junction (P nucleotide, N
nucleotide)
Coding joint

Signal joint
D
V
J
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Immunoglobulin diversity
1. Combinatorial diversity
Many combinations of V,D,J gene segments
Many combinations of heavy and light chains.
2. Junction imprecision
CDR3
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Pre-BCR
Btk XLA
Proliferation
Activate transcription of k light chain
Shut off Rag1/2, TdT
Shut off SLC
Prevent V(D)J recombination on the second allele
Expand pre-B popultion
Progression to the next stage of maturation
VJ recombination of ? locus.
5-6 rounds x 30-70
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IgL rearrangement
1. Ordered rearrangement ?-?
2. Allelic exclusion.
3. Repeated rearrangement.
Negative selection
Multivalent self-antigen
Clonal deletion
Bone marrow and periphery
Anergy
Soluble self-antigen
Receptor editing (light chain) may rescue some
self-reactive B cells in Bone marrow.
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Alternative splicing vs polyA determines
membrane vs secreted Ig.
IgM is expressed first. IgD is expressed
through alternative splicing.
Ig diversity in chicken gene conversion.
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Lymphocyte Recirculation
The connection between lymphatic system and blood
circulation.
The major lymphoid tissues.
Efferent lymphatic vessele
Other 2nd lympoid tissues
B cell follicles
HEV (extravasation)
M cell (MALT)
T cell area
B,T cells
The circulation facilitates the encounter of
lymphocytes with antigens.
Blood circulation
Antigen
B cell follicles
White pulp
spleen
T cell area
Blood circulation
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T cells
Bone marrow
Hematopoietic stem cell (HSC)
Bone marrow
Primary lymphoid organ
thymus
Lymphocyte maturation
Lymphoid progenitor
Myeloid pregenitor
thymus
T cell progenitor
B cell progenitor
Granulocyte-monocyte progenitor
Blood
monocyte
T cell
Neutrophil
B cell
1-6
50-70
Lymphocyte (20-40)
Lymphocyte circulation
Tissue
Secondary lymphoid organ
Lymphocyte activation
macrophage
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T cell maturation
IL7
IL7R
RAG-1/2
TdT
SCF
c-kit
Double negative CD4-CD8-
Double positive CD4CD8
Single positive CD4 or CD8
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TCR loci
The structure and organization of TCR ??????????.
The order of rearrangement
??
?
???????
????
Diversity
Combinatorial diversity (many combinations of
gene segment and the two subunits)
Imprecise coding joint CDR3
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Commitment to ? or ?? lineage
Pre-TCR signals the progression for further
maturation along the ? lineage.
?-selection
Proliferation
Transcriptional activation of TCR? locus
Degradation of RAG-2 and repression of RAG-1/2
expression
Activation of CD4 and CD8 Expression (DP stage)
Expand ? precursors
(100x)
Rearrangement of TCR?
Prevent rearrangement Of the other ? allele
(allelic exclusion) Prevent rearrangement Of
the ? and ? loci (?? commitment)
(repeated rearrangement no allelic exclusion)
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Cortex
TCRCD4CD8
Positive selection CD4, CD8 lineage commitment
cortex epithelial cells
Apoptosis
Elimination by macrophage
TCRCD4
TCRCD8
medulla
Dendritic cells and macrophages
Negative selection
Medulla epithelial cells (AIRE)
Apoptosis
Elimination by macrophage
None self-reactive Mature T cells
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Lymphocyte Recirculation
The connection between lymphatic system and blood
circulation.
The major lymphoid tissues.
Efferent lymphatic vessele
Other 2nd lympoid tissues
B cell follicles
HEV (extravasation)
M cell (MALT)
T cell area
B,T cells
The circulation facilitates the encounter of
lymphocytes with antigens.
Blood circulation
Antigen
B cell follicles
White pulp
spleen
T cell area
Blood circulation
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MHC and antigen presentation
Gene structure and diversity
Class I MHC HLA-A,B,C (human) H-2D, L, K
(mouse).
Class II MHC HLA-DP, DQ, DR (human) H-2IA, IE
(mouse).
Polymorphism and polygene allows the presentation
of diverse peptides to T cells.
MHC interaction with TCR, CD4, CD8
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TCR binds to Class I MHC-peptide complex.
T cell
TCR
CDRs
peptide
MHC
CDR3 is the most variable part in TCR, and forms
the major contact with the peptide.
CDR3 coincides with imprecise junctions
Target cell
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CD4-Class II MHC, CD8-Class I MHC interaction
stabilize TCR-MHCpeptide interaction and
transmits activation signals.
Class II
Class I
Activation signal
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Antigen processing and presentation
Dendritic cells
virus
APCs
Macrophages
All nucleated cells
B cells
Thymus epithelial cells
8-10 amino acid Peptide Hydrophobic
C-terminus Anchor residue
13-18 amino acid peptide
Bacteria, toxin
Endocytosis Phagocytosis macropinocytosis
endosome
TCR antigen may derive from internal peptides
while Ab eptitope is generally on molecular
surface.
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T cell activation
Antigen presenting cells
Dendritic cells, macrophage, B cells
Antigen capture (type and location, migration)
Co-stimulatory molecule
Distinction from non-antigen presenting cells
T cell anergy
T cell activation
T cell activation
TCR
Phosphorylation cascade
Proliferation and differentiation Into effector
cells
clustering
CD4 or CD8
Lck, ZAP-70
ITAM
Co-stimuation (B7-CD28) stimulates IL-2
production.
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B cell activation
BCR
Ag
Phosphorylation cascade
clustering
B7
Co-receptor (CD19,CD21,TAPA1)
C3d
Lyn, Fyn, Blk Syk
TH cell activation
Ag
Class II MHC-peptide
endocytosis
BCR
Secret immunoglobulins
CD40
Plasma cells
CD40L
Activated B cells
Activated TH cells
Memory cells
Cytokines (IL4, IL5)
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Antibodies
Antibody structure and function domains
Antigen binding site (CDR1,2,3)
VH
VL
CH1
CL
Fab
CH2
Hinge region
Fc
CH3
IgH constant region determines antibody class.
Effector functions
Complement activation
Bind to Fc receptor on phagocytes
Opsonin-mediated phagocytosis
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The functions of antibodies
1. Neutralization of pathogen and toxin.
IgA is in external secretions (mucus, milk,
tears, and saliva). IgA prevents the attachment
of pathogen to mucous surface.
IgM also contributes to immunity in external
secretions.
High affinity IgG is important for neutralizing
toxins.
2. Oposonin for phagocytes. This is
important for eliminating encapsulated bacteria.
IgG is the most effective.
3. Complement activation.
IgM gt IgG
4. Maternal protection
IgG is transported across the placenta.
IgA in milk.
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Innate immunity, Adaptive immunity
(immediate)
Complement Mast cell Resident macrophage
Present before infection
Limited and fixed specificity
Innate immunity
Does not improve
(within hours)
Neutrophils Monocytes
No memory
Macrophage
infection
day
1
0
2
3
4
5
6
7
(after 5-7 days)
B and T cells
Memory B and T cells
Primary immune response
Secondary immune response
Adaptive immunity
Develop in response to infection
Diverse and specific
Improve in specificity
Immunological memory