HIS12 Humoral Immune Response Flashcards
Life history of B lymphocyte
HSC
—> Naive / Mature B cells (with BCR: IgM + IgD) (smaller, fewer cytoplasm)
—> Circulation
—> Secondary Lymphoid organs
—> Plasma cells (larger, more cytoplasm, actively producing Ab) + Memory cells (~ to Naive B cell)
—> Circulation / Reside in BM
B cells in Secondary Lymphoid organs:
1. ***Antigen recognition
—> Survey / Interact with antigens
—> induced to Effector B cells (Plasma cells) + Memory B cells
- ***Interaction with other immune cells
—> Expansion + Differentiation
—> Ab Production
Functional outcomes of B cell response:
- Different Ab subclass and functions
- Primary vs Secondary response: Quantitative and Qualitative changes
Antibody isotypes - variation in CH domains
F(ab) region:
- Antigen binding
—> Define antigen specificity
—> Huge variation in sequences
F(c) region:
- Limited variation
—> IgG, IgM, IgD, IgA, IgE
Different forms of Ab —> allow specialisation in different functions:
- Monomeric form (secretory form)
- Membrane-bound (IgM, IgD on naive B cell)
- IgA dimer
- IgM pentamer
Characteristics of humoral response
Primary response:
- **Long lag phase (~7-10 days)
—> **Clonal selection + Differentiation (of antigen-specific naive B cells)
—> Plasma cells + Memory cells
- Antigen-specific Ab levels: ↑ —> Plateau —> ↓
- IgM first —> IgG
- Persist for a few days to weeks depending on Nature of antigen
Secondary response / response under repeated exposure (e.g. Tertiary response):
- **Short lag phase
—> antigen-specific Memory B cells activated
—> quickly expand
—> Plasma cells + Memory cells
- **Rapid + Exponential production of Ab —> extended plateau + **last longer
- **IgG predominantly (+ other isotypes e.g. IgA, IgE) but little IgM
- Ab affinity for antigen much higher than primary response
***Primary vs Secondary humoral response
- Time course
- Primary: Longer induction time (~7-10 days)
- Secondary: Shorter induction time - Antibody titre
- Primary: Smaller
- Secondary: Higher (10-100 fold) - Antibody class
- Primary: IgM (first to be produced) —> IgG (~10 days later) (Both specific for initial antigens)
- Secondary: IgG (exponential ↑ in titre), IgM (~ magnitude to primary response) - Antibody affinity
- IgG»_space; IgM for same antigen (difference in Fab region)
Induction of B cell response - Secondary lymphoid organs
Primary lymphoid follicle (mostly B cell)
- Cortex of LN
Activation of B cells:
- Primary lymphoid follicles
—> Secondary lymphoid follicles with Germinal centre (Dark cap + Light zone (Expanded B cells + Differentiated plasma cells))
(Inactive B cell: Dark staining
Active B cell: Lighter staining)
Antigen-induced B cell activation
BCR:
- Ig Heavy + Light chain
- ***Ig α/β chain —> for intracellular signal transduction (∵ Heavy chain does not have long cytoplasmic tail —> incapable of initiating intracellular signaling)
Ig α/β chain:
- ***Immunoreceptor tyrosine-based activation motif (ITAM)
—> Tyrosine kinase activation
—> phosphorylation —> intracellular signaling
- **Key BCR signaling events involve:
1. Tyrosine kinases: LYN, SYK, BTK (Burton’s tyrosine kinase)
2. PLCγ (Phospholipase Cγ)
3. PI3K (Phosphatidyl inositol-3-kinases)
Binding of antigen to Fab region:
- Antigen cross-link Ig (BCR) on B cell surface
—> Recruitment + Phosphorylation of LYN, SYK to ITAM
—> Activation of BTK, PLCγ (↑ intracellular [Ca] as 2nd messenger), PI3K
—> Intracellular signaling cascades
—> Activation of transcription factors
—> Gene transcription
—> Protein synthesis:
- Cell ***proliferation (e.g. c-fos, c-myc) —> B cells at resting stage enter cell cycle —> mitotic division
- ***Receptors for cytokines (from T cell) (e.g. IL-4R, IL-6R, TGFβR, IFNγR)
- Surface molecules (e.g. ***MHC class II (antigen presenting molecules) for interaction with CD4+ T cells)
- Specific enzymes: ***Activation induced deaminase (AID) for somatic hypermutation in Ig
- Proteins required for cell differentiation
AND
**Antigen processing + presentation:
Endocytosis of antigens and processing
—> Antigen presentation on **MHC class II for T cell interaction
—> B cell act as APC
***Thymus-dependent (TD) antigen —> Effects of antigens on B cells
Thymus-dependent (TD) Antigen (***Protein Ag)
- mostly ***Protein Ag!!!
- require 2 signals (1 from Ag, 1 from T cell)
- require Contact help from T helper cell for optimal Ab response (i.e. need B cell to present Ag to T cell)
- e.g. via **CD40 (on B cells) + **CD40L (on T cells) molecular interaction
Modulation of B cell response by CD40-CD40L:
Antigen bind on BCR (**1st signaling)
—> Peptides from Ag processed + presented on MHC class II molecules (unregulated, more MHC class II on surface) (+ IgM pentamer release)
—> Interaction with T cell with specific TCR
—> Activation of T cell
—> Express CD40L on T cell surface
—> Interaction with CD40 on B cell surface
—> **2nd signaling / co-stimulatory signaling
—> B cell undergo further differentiation
—> ***Class switching
—> Produce IgG instead of IgM
CD40-CD40L: critical for Ig Class switching to IgA, IgG, IgE
CD40L deficiency (X-linked) in patients: - X-linked ***hyper IgM syndrome - low IgG / other isotypes despite pathogenic stimulation - also defect in macrophage function (∵ also interact with CD40L on T cell) —> immunodeficiency, infection susceptibility
***Thymus-independent (TI) antigen —> Effects of antigens on B cells
TI antigens: Elicit Ab response without need for Th cell contact
2 subtypes (different activation mechanisms):
- TI-1 antigens (**Mitogens)
- Mitogens e.g. LPS, microbial associated nucleic acids
- NOT bind to BCR
- Activation through **non-BCR: bind to additional receptors (e.g. **Toll-like receptors) on B cell
- **Polyclonal B cells activation (regardless of specificity) (咩B cell品種都activate) - TI-2 antigens (**Polysaccharide antigens)
- **Polysaccharide antigens with **repeating subunits (e.g. **ABO blood group antigens, Pneumococcal polysaccharide, Salmonella polymerised flagellin)
—> Cannot be presented on MHC-II
- Little class-switching (difference in Ab come from Fab region)
- Weaker and mainly **IgM Ab response
- **No / Little memory cell formation (∵ no T cell help)
- **Activate B cell by **extensively cross link Ig on B cell surface
- NOT presented to CD4 T cell —> NOT initiate T cell help
—> Plasma cells produce IgM without T cell help
***簡單而言:
TI1: Mitogens, Non-BCR, Polyclonal activation
TI2: Polysaccharide Repeating subunits, BCR cross-linking, IgM
***Germinal centre - Key B cell activation and differentiation events
Germinal centres:
- derived from Primary lymphoid follicles
- induced upon B cell activation by Ag
Dark zone:
- High cell density + High rate of proliferation
- B cell enter from circulation via afferent lymphatic vessel —> encounter Ag within lymphoid organ
1. **Clonal selection + Proliferation —> **Centroblasts
2. **Somatic hypermutation in Ig gene —> change **affinity to bind Ag during B cell replication —> subtle ***differences in Fab region among B cells
Light zone: - Centroblasts —> ***Centrocytes 1. ***Ig Affinity maturation (SHM) 2. ***Ig Class switching (淨係喺Light zone) 3. Differentiation of Plasma cells + Memory cells —> exit circulation, some reside in BM
***Somatic hypermutation in Ig gene
- Involves Activation-induced (cytidine) Deaminase (AID) (***B cell only)
- Function: Produce Fab region with better / reduced “fit” to Ag
AID (expressed in **activated B cell)
—> **Cytosine —> Uracil in DNA during cell replication
—> Uracil **recognised as error in DNA sequence
—> **removed and replaced by random nucleotide by error-prone DNA polymerase
—> **introduce mutations in Ig gene (“Hotspots” of SHM)
—> **Complementarity determining region (CDRs) / ***Hypervariable region of V domains of H + L chain (3 CDRs in Fab region)
—> Hypervariable loops from each domain brought together by folding
—> single Hypervariable surface
—> Antigen binding site at tip of each arm
—> SHM change affinity to bind Ag during B cell replication
Affinity maturation of Ab
Result of SHM
Enhance of humoral response via affinity maturation in repeated antigen exposure (越黎越high affinity)
- IgM —> IgG (primary) —> IgG (secondary) —> IgG (tertiary)
- Number of mutations induced in Fab region ↑ over time
- Kd (dissociation constant) between Ab and Ag ↓ —> Affinity ↑ (>100 fold)
B cell proliferate in response to Ag
—> High rate of random mutation in re-arranged V segment in Ig without changing overall specificity to Ag (大同小異)
—> several clones of B cells
—> slight difference in Fab region
—> compete for same Ag
—> B cells with high affinity Ig for Ag
—> ***selected for survival, others negatively selected
—> Ab affinity ↑ >100 fold in secondary response
***B cell differentiation and survival
2 cell types important for B cell development and survival:
- Follicular Th cell
- **Cell-cell contact via CD40-CD40L
- **Cytokines e.g. IL-4/IL-5/IL-13
- Critical for B cell ***Isotype class switching and survival (via Cytokine: survival factor) - Follicular Dendritic cells (FDC)
- Process and present antigens
- Compete with B cells for Ag (同B cell鬥搶, B cell搶贏就可以生存)
—> Only B cells with high affinity Ig compete effectively for Ag
—> induced by FDC to express cell ***survival protein bcl-2 (in B cell)
—> rescue them from cell death
—> allowed to become Plasma cells + Memory cells
***Antibody isotype class switching
Naive B cells:
Heavy chain:
- VDJ segment (V domain) rearranged first
- ALL Ig C region gene segments present (μ: IgM, γ: IgG, α: IgA, δ: IgD, ε: IgE)
- DNA contain ***switch regions in front of each Ig C region gene segment (contain specific motif for site specific recombination)
B cell activation:
- DNA rearrangement in C domain
—> DNA recombination involves enzymes: AID, UNG, APE
—> removal of certain Ig C region gene segments at ***DNA level (NOT RNA splicing)
—> produce specific Ig transcript
—> “class-switched” B cells
IgM / IgD on B cell surface:
Primary IgM / IgD transcript by **alternative RNA splicing (transcribed before switching —> **NOT involve DNA rearrangement of C gene segments)
IgG / IgA / IgE:
Synthesised in activated B cells and Plasma cells (***Involve DNA rearrangement —> require AID, UNG, APE)
***Class switch recombination (CSR) enzymes: AID, UNG, APE
- AID (Activation-induced Deaminase)
- change Cytosine —> Uracil - UNG (Uracil DNA glycosylase)
- removal of Uracil
—> AID, UNG present in ***switch regions
- APE (Apyrimidic/Apurinic-endonucleases)
- create a **nick in the phosphodiester backbone in DNA (in switch regions)
—> free ends of DNA rejoined by **non-homologous end joining (NHEJ)
—> 鞋帶兩邊loop join —> 中間唔要
Isotype class switching
- IgM / IgD are transcribed before switching
- IgM will be switched to other classes (e.g. IgA, IgG, IgE) upon appropriate stimulation
- ***NOT affect Ig affinity / specificity
- ***BUT biological effector activities varies
T cell help through
1. **CD40-CD40L interaction
2. **Cytokines from Th cells
—> switching involves transcription of DNA in rearranged C region which is ***cytokine-dependent
—> T cell cytokines are critical in determining which isotype to switch to
—> e.g. IL-4 promote IgE, TGFβ promote IgA, IFNγ promote IgG1, IgG3
