B cells & Antibodies Flashcards
1
Q
Molecules of the immune system
A
- antibodies
- TCRs
- MHC
2
Q
Types of epitopes
A
- Linear - formed by several (6) linear/adjacent AA residues - can be accessible to the ab as they are on external surface - if buried in protein (can only be accessible via denaturation)
- Conformational - formed by aa residues not in the linear sequence - become spatially juxtaposed in folder protein
3
Q
Antibody Class-Switching
A
Switch from primary (IgM) to secondary (IgG, IgA, IgE)
*IgG present on ab in circulation
4
Q
Immunological Memory
A
- can respond to subsequent infection w/ the same pathogen with a response
- much faster
- greater magnitude ….. than initial exposure
- can be very long lived (2009 Influenza A) or short lived (Covid-19)
5
Q
Effective vs Sterilising immunity
A
Effective
- if you encounter virus, you may get infected remain asymptomatic and transmit on the virus
- infection but subsequent successful eradication of virus
- vaccine induced immune response - unable to stop every virus from replicating
Sterilising
- someone who’s vaccinated or naturally infected will not catch the virus or transmit it further
- these abs neutralise and block virus entry into cells and prevent virus replication
6
Q
Subclasses of ab
A
IgM, IgG, IgA, IgD
7
Q
Clonal selection theory
A
- B cells specific for different Ag develop before exposure to these Ags (Ag-independent)
- Ag selects the appropriate B cell/Ab from pre-existing B cell pool
- All surface ab on any one naive B cell is the same - clonal
- Following Ag exposure - B cells expand, differentiate, proliferate and secrete ab (Ag-dependent)
8
Q
IgM
A
- monomer on surface of B lymphocyte
- pentameric form is secreted
- held together by J chains
- found mainly in blood - does not penetrate tissues well
- 1st ab made in response to a specific infection
- efficient at complement activation
- efficient at neutralisation of bacteria and viruses
- not involved in opsonisation or ADCC
9
Q
IgG
A
- most abundant ab in serum
- 4 subtypes - differ in hinge region
- longer half life than IgM ( 23 vs 5 days)
- All but IgG2 cross placenta - protection to fetus
- efficient at complement activation
- efficient at neutralisation of bacteria and viruses
- promotes opsonisation or ADCC
10
Q
IgA
A
- most abundant ab in the body in terms of quantity (70%)
- exists as monomeric but when secreted onto mucous surfaces - dimeric
- synthesised in the gut
- 2 types
- IgA1 - longer hinge region - found in serum
- IgA2 - found in secretions
- has J chain
- SIgA - secreted out of cell onto mucous surface
- prevents pathogen adherence to mucous surfaces via neutralisation
- promotes ADCC by interacting with FcR of NK cell
- poor at complement fixation and opsonisation
11
Q
IgE
A
- present in trace amounts in serum
- normal role - controlling worm infections
- greatly increased in hypersensitivity (asthma, hay fever, allergies)
12
Q
Ab structure
A
- Light chain
- Heavy chain
- held together by sulphide bridges between cysteine
- Fc region - receptors of phagocytes
- highly variable regions: CDR1,2,3 on L and H chain
- how these regions come together - dictates type of antigen will bind
13
Q
Mechanisms generating ab diversity
A
- Multiple germ line V genes
- V-j and V-D-J recombinations
- N-nucleotide addition
- Recombinational inaccuracies
- Somatic hypermutation (SHM)
- Assorted H and L chains
14
Q
V-D-J Junctional Diversity
A
- NTs are randomly deleted and inserted at junctional joining sites
- Mechanism to increase junctional diversity - increased complexity of abs
- TDT enzyme
15
Q
3 termination codons
A
- UGA
- UAA
- UAG
16
Q
Allelic Exclusion
A
- maintains law of clonal selection - each B cell is clonal and can only produce 1 ab
- each B lymphocyte has 2 copies of each H and L chain - maternal and paternal
- allelic exclusion prevents co-expression of both alleles
17
Q
Ab Repertoire
A
- junctional flexibility - N regions additions, deletions and somatic mutation - increases all possible combinations of B cells (~ 109 naive mature B cells circulating at one time)
- Many B cells don’t reach circulation
- naive B cells migrate to secondary lymphoid organs (LN and spleen)
18
Q
How do naive B cells undergo differentiation into ab-secreting plasma cells?
A
- binding of antigen to membrane anchored ab receptor leads to internalisation of the ab
19
Q
Ab class switching
A
- 1st ab made in response to particular infection is IgM (hl 4 days)
- Ab class switch from primary to secondary (IgG, IgA, IgE)
- Fab region - antigen binding region stays the same
- Fc region is exchanged
20
Q
Ab class switching: Switch Recombination
A
- rearranged VDJ combines with downstream C gene with removal by deletion of the intervening DNA
- T helper cell dependant/driven
21
Q
Cytokines that play a roles in switch recombination
A
- IL4: promotes switch to IgG4 and IgE
- IFNᵧ: promotes switch to IgG2 - from Th lymphocytes
22
Q
Mode of Action of Abs
A
Direct effector:
- Neutralisation - prevention of virus/bacterial toxin from interacting with its receptor - prevent infection
Secondary effector:
- Opsonisation - promotes phagocytosis by macrophages/NFs - Fc receptor required for uptake
- ADCC - promoted by opsonisation by CTLs and NKs - activated by ab attaching to surface of pathogen cell
Activation of complement - membrane attack complex - lysis of bacteria in some viruses
23
Q
Ab producing B cells
A
- plasmablasts
- SLPCs
- LLPCs
- Memory B cells
24
Q
Plasmablasts
A
- secrete low affinity abs but still receive sufficient stimulation signals to survive
- supply earliest ab to pathogens - short-lived
- most immature ab-producing B cell
- affinity for antigen is 1/100 of that of germinal centre B cell
25
Short-lived plasma cells (SLPCs)
* following antigen encounter naive B cells proliferate and terminally differentiate into SLPCs
* produces **large amounts** of antigen-specific ab of IgM and IgG - ab factory
* optimum response seen after 2 weeks
* following control of infection - undergo cell apoptosis
* don't play a role in long term memory
26
Long-lived Plasma Cells (LLPCs)
* B cells that have been successfully stimulated with antigen do not become SLPC - migrate further into the follicle and enter GC
* B cells can initiate a new GC or enter existing GC
* can live and produce ab for months-yrs
* relocate into the bone marrow or GALT
27
Memory B Cells
* GC-derived where they are class switched and hypermutated
* basis of long-term immunity
* B cells not receiving sufficient signals - Fas mediated apoptosis
28
T follicular helper (Tfh) cells
* Contain surface markers: CD4+, Bcl-6, CXCR5
* Interact with memory B cells at B-T cell junction
* crucial for long-term survival of MBCs
* Secrete cytokines: IL-4, IL-21, IFNᵧ
29
Role of memory B cells in response to future re-exposure to antigen
* MBCs are quiescent and require re-stimulation to stimulate memory response
* re-exposure to antigen - rapid activation of MBCs in spleen and LNs - produce ab-secreting SLPCs
* Some reactivated MBCs give rise to new LLPCs and others stay in GC
30
GC features in severe COVID
* **Bcl-6+ GC B cells and Tfh cells are diminished**
* **Low levels of somatic hypermutation in GC B cells**
* Plasmablasts accumulate in blood
* Formation of dysfunctional B cells and plasmablasts
* TNF-𝛼, IL-6, IL-8 production - damage of GC - lymphopenia
* Loss of GCs compromises production of long-lived B cell memory and high-affinity abs
31
Somatic Hypermutation of abs
* somatic mutations in ab genes with selective survival of B cells - produce abs with highest affinities
* takes place in GC in LN and spleen
* Follicular DCs - sequester B cell antigen and stimulate B cell
* crucial fro LLPC and memory B cell long term survival
* turn on anti-apoptosis pathways
32
Mechanism of Somatic Hypermutation
* somatic point mutation in variable gene of H and L chain genes
* more mutation seen after secondary and tertiary infection
* causes deamination of cytosine to uracil in DNA by activation-indeuced cytidine deaminase **(AID)**
* mismatch occurs (G:C becomes G:U)
33
How is a somatic mismatch hypermutation resolved?
* uracil not found in DNA but RNA
* U bases removed via repair enzyme - **uracil-DNA glycosylase**
34
Types of Immunity
* Natural Active Immunity
* Natural Passive Immunity
* Artificial Vaccination
* Artificial Passive
35
Natural Immunity
* Active Immunity - get disease, immune response (abs and T-cells) - develop lifelong or partial immunity (memory)
* Passive Immunity - abs from mother across placenta to foetus
36
Artificial Immunity
* Vaccination - develop lifelong or partial immunity
* Passive - direct injection of monoclonal abs - short lived, ab eventually degraded
37
Types of Vaccines
* mimic natural infection in the immune system - develops B and T cells immunity to vaccine
1st gen:
* killed - inactivated
* live - nonvirulent (attenuated)
38
Limitations of vaccines
* not all infections agents can be cultured
* extensive safety precautions required
* batch variation
* reversion to virulence
* limited shelf-life
39
Subunit vaccines
* derived from outer surface of pathogen
* viral or bacterial surface protein
Advantages
* no live components - no risk of inducing disease
* chemically defined
* safe and more stable than LAV (live)
Disadvantages
* expensive
* less immunogenic than LAV
E.g.
* Hep B sag vaccine - recombivax HB
* COVID-19 spike protein - Novavax
40
Nucleic Acid Vaccines
* mRNA viruses approved for COVID
* moderna
* Pfizer
41
Virus Vector Vaccines
* use virus vector to carry vaccinating agent
* Adenoviruses - persist in cells as extra-chromosomal episomes
e.g
* vaccinia virus - small pox
* AstraZeneca - replication-deficient monkey adenoviral vector vaccine encoding SARS-CoV-2 spike gene
41
Virus Vector Vaccines
* use virus vector to carry vaccinating agent
* Adenoviruses - persist in cells as extra-chromosomal episomes
e.g
* vaccinia virus - small pox
* AstraZeneca - replication-deficient monkey adenoviral vector vaccine encoding SARS-CoV-2 spike gene
42
mAbs production
* HAMA effect - person'e immune system sees murine mAb as a foreign protein - develops its own abs against the mAb
* CDR grafting - replace as much of mouse gene regions with human gene regions
e.g. Trastuzumab (Herceptin) - HER2 Metastatic breast cancer
43
Example of mAb application
Rheumatoid Arthritis
* mAbs to inflamm cytokines (TNF-𝛼, IL-6, IL-1)
* cell involved in RA - endothelial, synovial cell, leukocytes
44
Ab-drug conjugate (ADC)
**Kadcyla**
* consists of mAb trastuzumab (Herceptin) linked to cytotoxic agent mertansine (DM1) - Tubulin inhibitor
