Download - 14 February, 2006
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
Complement
Origin: produced primarily by hepatocytes in liver. Activated macrophage can also produce it.
Location: blood and tissue fluid.
Function:
Lyse bacteriaOpsonize bacteria to facilitate phagocytosis
Induce inflammation
Clearance of antigen-antibody complex
Lysis of bacteriaComplements can be activated by three pathways.
1. Classical pathway:
Bacteria-antibody(Fc)-C1
C4, C2 C4b2aClassical C3convertase
C3 C3bB,D
C3bBb
Specificity
Alternative C3 convertase
C3 amplification loopC3b
C3b2Bb (alternative C5 convertase)
C4b2a3b (Classical C5 convertase)
C5 C5b C5b6789MAC
C6,7,8,9
Lysis of bacteria
IgM > IgG
Require activation of B cells.
Natural antibodies by B1 cells andCross-reacting antibodies may initiatethe reaction during primary immune response.
C-reactive protein (acute phase response) alsoactivate complement through C1.
BbC3b
degradation
RCA
C5b678, no C9
HRF, CD59
Lysis of bacteria
2. Lectin pathway:
Bacteria-MBL-MASP
C4, C2 C4b2aClassical C3convertase
C3 C3bB,D
C3bBb
Specificity
Alternative C3 convertase
C3 amplification loopC3b
C3b2Bb (alternative C5 convertase)
C4b2a3b (Classical C5 convertase)
C5 C5b C5b6789MAC
C6,7,8,9
Lysis of bacteria
Early response.
Upregulated during acute phase response.
C3b
degradation
RCABb
C5b678, no C9
HRF, CD59
Lysis of bacteria
3. Alternative pathway:
C3 C3bB,D
C3bBbAlternative C3 convertase
C3 amplification loopC3b
C3b2Bb (alternative C5 convertase)
C5 C5b C5b6789MAC
C6,7,8,9
Lysis of bacteria
Slow spontaneoushydrolysis
C3bBb
Degradation
RCAspecificity
HRF, CD59specificity
C5b678, no C9
Early response.
MAC is effective against Gram negative bacteria.
The thick peptidoglycan in the cell wall of GramPositive bacteria prevents the insertion of MAC.
Lysis of Bacteria
Opsonization
C3b is covalently attached to bacteria during complement activation.
Phagocytes (macrophages, neutrophils) have complement receptors that can recognize C3b.
Inflammation
C5a, C3aC4a
Mast cells
histamine
Prostaglandinleukotrine
vasodilaton(Increase blood flow)
inflammationvascular permeability
Leukocyte extravasation
(leakage of plasma)
(Recruitment of neutrophil and monocyte)
C3d facilitate the clustering of B cell coreceptor (CD21/CD19/TAPA1) with BCR and stimulate B cell activation.
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
Macrophage
Hematopoietic stem cell(HSC)
Myeloid pregenitor
Granulocyte-monocyte progenitor
monocyte
Bone marrow
Neutrophil1-6% 50-70%
Blood
Lymphoid progenitor
B cell progenitor
B cell
T cell progenitor
T cell
thymus
Lymphocyte (20-40%)
Primary lymphoid organ
Bone marrow
thymusLymphocyte maturation
Secondary lymphoid organ
Lymphocyte activation
Lymphocyte circulation
macrophage
Tissue
Function of Macrophage
1. Phagoctyosis
2. Produce inflammatory cytokines
3. Antigen presentation to T cells
4. Scavenger to eliminate apoptotic cells and immune complexes.
Phagocytosis
Bacteria (PAMPs)
Phagocytic receptors
Scavenger receptor
Mannose receptor
-glucan receptor
Opsoninreceptors
Fc receptor (antibody)
Complement receptor (C3b)
Collectin receptor (MBL)
Pentraxin receptor (C-reactive protein)
bacteria
phagosome
bacteria
phagolysosome
Bacterial killing
Oxygen-dependent
Oxygen-independent
Reactive oxygenintermediates
Reactive nitrogenintermediates
Degradedbacteria
exocytosis
Class II MHC
Antigen-presentation
Secretion of inflammatory mediatorsand antigen presentation
Bacteria (PAMPs) TLRs M activation
Inflammatory cytokines
IL-1
TNF-
IL-8
IL-6
Neutrophil activationAnd chemotaxis
Class II MHC
B7Antigen presentationT cell activation
Acute phase response(liver)
Local inflammationMBL
C-reactiveprotein
Phagocytosis
Prostaglandin, leukotriene inflammation
(Secondary lymphoid tissues)
Neutrophil
Hematopoietic stem cell(HSC)
Myeloid pregenitor
Granulocyte-monocyte progenitor
monocyte
Bone marrow
Neutrophil1-6% 50-70%
Blood
Lymphoid progenitor
B cell progenitor
B cell
T cell progenitor
T cell
thymus
Lymphocyte (20-40%)
Primary lymphoid organ
Bone marrow
thymusLymphocyte maturation
Secondary lymphoid organ
Lymphocyte activation
Lymphocyte circulation
macrophage
Tissue
Functions of NeutrophilsNeutrophils (blood circulation)
Rolling (mucin like CAM - E,P-selectin)
Activation by chemoattractants (IL8(CXCL8), C5a, N-formyl-peptides)
Activation of integrin (LFA-1, CR3, CR4, common subnit: CD18)
Stable interaction between LFA-1 with ICAM
Inflamed endothelial cells
Induced by inflammatorymediators
Chemotaxis by IL8, C5a, N-formyl-peptides, etc
Phagocytosis DegranulationKill bacteria
apoptosis Eliminated by macrophages
B cells
Hematopoietic stem cell(HSC)
Myeloid pregenitor
Granulocyte-monocyte progenitor
monocyte
Bone marrow
Neutrophil1-6% 50-70%
Blood
Lymphoid progenitor
B cell progenitor
B cell
T cell progenitor
T cell
thymus
Lymphocyte (20-40%)
Primary lymphoid organ
Bone marrow
thymusLymphocyte maturation
Secondary lymphoid organ
Lymphocyte activation
Lymphocyte circulation
macrophage
Tissue
B cell maturation
HSC CLPGL-ProB
DJ-ProB
LargePreB
SmallPreB
ImmatureB
sIgM+sIgM+
sIgD+
Mature B
B220 (CD45R)
CD19
C-kit
IL7R
SLC (5/VpreB)
RAG-1,2
TdT
IgH ()
IgL ()
Ig/
B cell marker
Surface marker (flow cytometry) V(D)J recombination
Bone marrow
2nd lymphoidtissues
B cell developmental stages are characterizedby surface markers and V(D)J recombination.
Cell surface marker is detected by Flow cytometry
Fluorescence-activated cell sorter (FACS)
V(D)J recombination
IgH : V-DJ joining
IgL ( and ): V-J joining
D-J joining
1. Organization of the IgH and IgL loci and the order of recombination.
2. Basic recombination mechanism.
JD
RAG-1,2 cleaves between RSS and the gene segments
JD
DNA repair to join the DNA breaks
D J +
Coding joint
Signal joint
12 97
heptamerspacernanomer
Recombination occurs between23-bp and 12-bp RSS (12-23 rule).RSS
Which parts correspond to the immunoglobulin V and C regions.
Imprecise coding junction (P nucleotide, N nucleotide)
23 79
V
V
V
Immunoglobulin diversity
1. Combinatorial diversity
Many combinations of V,D,J gene segments
Many combinations of heavy and light chains.
2. Junction imprecision
CDR3
Pre-BCR
Proliferation
Expand pre-Bpopultion
Shut off Rag1/2, TdT
Prevent V(D)J recombinationon the second allele
Activate transcriptionof k light chain
VJ recombination of locus.
Shut off SLC
Progression tothe next stageof maturation
5-6 roundsx 30-70
Btk: XLA
IgL rearrangement
1. Ordered rearrangement: -
3. Repeated rearrangement.
2. Allelic exclusion.
Negative selection
Multivalent self-antigen Clonal deletion
Soluble self-antigen AnergyBone marrow and periphery
Receptor editing (light chain) may rescue some self-reactive B cells in Bone marrow.
Alternative splicing vs polyA determines membranevs secreted Ig.
IgM is expressed first. IgD is expressed throughalternative splicing.
Ig diversity in chicken: gene conversion.
Lymphocyte RecirculationThe connection between lymphatic system and blood circulation.
B,T cellsBlood circulation
White pulpB cell follicles
T cell area
HEV (extravasation)
B cell follicles
T cell area
Blood circulation
Efferent lymphatic vessele
The circulation facilitatesthe encounter of lymphocyteswith antigens.
The major lymphoid tissues.
spleen
Other 2nd lympoid tissues
Antigen
M cell(MALT)
T cells
Hematopoietic stem cell(HSC)
Myeloid pregenitor
Granulocyte-monocyte progenitor
monocyte
Bone marrow
Neutrophil1-6% 50-70%
Blood
Lymphoid progenitor
B cell progenitor
B cell
T cell progenitor
T cell
thymus
Lymphocyte (20-40%)
Primary lymphoid organ
Bone marrow
thymusLymphocyte maturation
Secondary lymphoid organ
Lymphocyte activation
Lymphocyte circulation
macrophage
Tissue
T cell maturation
Double negative CD4-CD8-
Double positive CD4+CD8+
Single positive CD4+ or CD8+
RAG-1/2TdT
IL7
SCF
IL7R
c-kit
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
Commitment to or lineage
Pre-TCR signals the progression for further maturation along the lineage.
Proliferation
Expand + precursors
Degradation of RAG-2 and repression of RAG-1/2 expression
Prevent rearrangementOf the other allele (allelic exclusion)
Prevent rearrangement Of the and loci ( commitment)
Activation of CD4 and CD8 Expression(DP stage)
Transcriptional activationof TCR locus
Rearrangement of TCR
(100x)
-selection
(repeated rearrangementno allelic exclusion)
Positive selectionCD4, CD8 lineage commitment
CortexTCR+CD4+CD8+
cortex epithelial cells
Apoptosis Elimination by macrophage
TCR+CD4+ TCR+CD8+
Negative selection
None self-reactive Mature T cells
Apoptosis Elimination by macrophage
medullaDendritic cells and macrophages
Medulla epithelial cells (AIRE)
Lymphocyte RecirculationThe connection between lymphatic system and blood circulation.
B,T cellsBlood circulation
White pulpB cell follicles
T cell area
HEV (extravasation)
B cell follicles
T cell area
Blood circulation
Efferent lymphatic vessele
The circulation facilitatesthe encounter of lymphocyteswith antigens.
The major lymphoid tissues.
spleen
Other 2nd lympoid tissues
Antigen
M cell(MALT)
MHC and antigen presentation
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).
Gene structure and diversity
Polymorphism and polygene allows the presentation of diverse peptides to T cells.
MHC interaction with TCR, CD4, CD8
TCR binds to Class I MHC-peptide complex.
TCR
T cell
Target cell
MHC
peptide
CDRs
CDR3 is the most variable part in TCR, and forms the major contact with the peptide.
CDR3 coincides with imprecisejunctions
CD4-Class II MHC, CD8-Class I MHC interaction stabilize TCR-MHC:peptide interaction and transmits activation signals.
Class II Class I
Activation signal
Antigen processing and presentation
EndocytosisPhagocytosismacropinocytosis
endosome
Dendritic cells
Macrophages
B cells
Thymus epithelial cells
APCs
All nucleated cells
8-10 amino acidPeptideHydrophobic C-terminusAnchor residue
13-18 amino acid peptide
TCR antigen may derive from internalpeptides while Ab eptitope is generallyon molecular surface.
Bacteria, toxin
virus
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 activation T cell anergy
T cell activation:
TCR
CD4 or CD8clustering Phosphorylation
cascade
Lck, ZAP-70ITAM
Proliferation and differentiationInto effector cells
Co-stimuation (B7-CD28) stimulates IL-2 production.
B cell activation
BCR
Co-receptor(CD19,CD21,TAPA1)
clusteringPhosphorylationcascade
Lyn, Fyn, BlkSyk
BCRAg
Ag
C3d
endocytosis Class II MHC-peptide
B7
TH cell activation
Activated TH cells
CD40L
Cytokines(IL4, IL5)
CD40
Activated B cells
Plasma cells
Memory cells
Secret immunoglobulins
AntibodiesAntibody structure and function domains
CH1
CH2
CH3
VH VL
CL
Hinge region
Antigen binding site (CDR1,2,3)
Fab
Fc
Complementactivation Bind to Fc receptor
on phagocytes
Opsonin-mediated phagocytosis
Effector functions
IgH constant region determines antibody class.
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 > IgG
4. Maternal protection
IgG is transported across the placenta.
IgA in milk.
infection
10 2 3 4 5 6 7day
ComplementMast cellResident macrophage
NeutrophilsMonocytes Macrophage
Innate immunity
B and T cells
Adaptive immunity
Primary immune response
(immediate)
(after 5-7 days)
Secondary immune response
Innate immunity, Adaptive immunity
(within hours)
Present before infection
Limited and fixed specificity
Does not improve
Memory B and T cells
Develop in response to infection
Diverse and specific
Improve in specificity
Immunological memory
No memory