9. Specialised Cells: B cells

Learning Outcomes: L18

The mechanism of B cell activation Importance of coreceptors for signalling and costimulatory molecules for amplification The functions of FcϒRIIB and shutting down of the B cell response The basic organisation of germinal centres and the roles of the various cells Follicular dendritic cells Follicular T helper cells; IL-21, Bcl-6 Somatic hypermutation and AID Antibody class switching and AID The different types of antibodies/immunoglobulins in our bodies and their role in the immune response Primary versus secondary responses Opsonisation, neutralisation, complement fixation Link to allergy, asthma and how microbes can promote such types of responses

Activation of B Cells

B cells recognise antigen via BCR  Lock and Key approach The part of antigen that BCR/Ab recognise= epitope of antigen  Continuous: Ab recognises sequence on antigen that is linear  Discontinuous: Forms the shape that Ab recognises when molecule folds Important for experiments: ​​​​​​​If Ab recognises linear epitope: ELISA is fine as whole protein in plate, western blot fine (protein denatured on gel) If Ab recognises discontinous epitope: western blot not fine, -ve result may not mean antigen not their but Ab may no longer recognise

Activation of B Cells

Like TCR, BCR signals through co-receptor, signal potentiated via co-stimulatory molecules- receptor alone has no capacity to signal: Co-receptors Igα and Igβ have tyrosine kinase activity Antigen binds BCR, conformational change in intracellular part, Igα and Igβ phosphorylated (activated) + phosphorylate costimulatory molecules CD19/CD81 form a co-receptor complex that provides costimulatory amplification of signals through the BCR C3d complement bound to pathogen also helps target pathogen to BCR, B cell has receptors for complement (CR2/CD21) CD4+ T cell help activation + differentiation of B cells into plasma cells  

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B Cell Response No Longer Needed

When B response not need, B cell shut down by signals through the negative regulator- FcϒRIIB (Fc receptor): Positive signals: ​​​​​Kinase activation proliferation of B cells CD19, CD81 act as costimulatory molecules for B cell activation Negative signals: Fc gamma receptor looks for excess IgG Dephosphorylation of key molecules for B cell activation (like CTLA4) Mutations in FcϒRIIB linked to autoimmune production of Ab's to host cell- cant shut down B cell response efficiently

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B Cell Response No Longer Needed

Activation In Secondary Lymphoid Tissues

Highly organised structures where cells are compartmentalised: Inside B cell follicle is germinal centre B cells in outer follicle, upon antigen encounter move to germinal centre: activation, proliferation and differentiation Germinal centre= less dense, only contains B cells with BCR triggered by antigen

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Germinal Centre

Critical for development of B cell responses: Within germinal centre have interactions between activated B cell, follicular DC and Th cell  B cell proliferation  Somatic hypermutation of B cell  Class switch Once these are achieved B cell has different fates: Memory cell, Plasma cell, Apoptosis (check point at germinal centre for autoreactivity- dies unlike in BM get receptor editing) 

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GC: CD4+ Follicular Th cell (Tfh)

Central to formation and propagation of germinal centre, resides in germinal centre tells B cell to do the 3 functions TF for differentiation into Tfh cell is Bcl-6 and it makes cytokine IL-21  

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GC: Function

IL-21 is criticial mediator in germinal centre function: Tfh express chemokine receptor CXCR5 , responsive to CXCL13 produced by GC to recruit and keep B cell and Tfh cell there  IL-21: Helps stromal cells of GC to maintain themselves Signals back to Tfh to keep alive Signal into B cell for proliferation) Tfh continually expresses antigens on its surface, that B cell responds to, to ensure B cell performs the 3 function

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GC: Function

Infection occurs in outer B cell follicle, interaction with conventional Th cell, activated and moves into GC In dark zone= clonal expansion occurs  Moves into light zone= interacts with follicular DC and Th cell and receives signals to transform into: Memory B cell  Short-lived plasma cell that secretes Abs  Moves into BM and remains as long-lived plasma cell, secreting small amounts of Ab to that pathogen (protective immunity)

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Somatic Hypermutation (SHM) by AID

Advanced Induced Deaminase (AID) is active in GC to facilitate SHM First encounter of R with antigen= not good fit but still triggers initial activation Mutations in Hv region enable a tighter fit (stronger interaction= better signal)- secondary response= memory cells Each subsequent encounter 'hones' the antigen binding site further  B cells that have undergone SHM will have their BCRs re-screened to determine if the mutations lead to autoreactive BCR (B cell dies if so)  

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Class Switching by AID

Need specific Ab for specific response- remove Fc region of IgM and replace with different Ab Fc region Upstream every Ab Fc region is a switch region (except igD so igM expressing cells also express igD) AID causes switch regions to form haripin loop that brings together Fc constant regions to change Cleave out intermediate regions Brings new Fc region to the VDJ so specifcity of Ab stays same

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Maturation of Humoral Response

Maturation of humoral response characterised by Ig with increased affinity for its cognate antigen and switching of Ig class: Primary response- takes few days to start making Ab's- need B cell activation, CD4 T help, functioning in GC to class switch  Initially get IgM- replaced by IgG after 15 days  Short lived, low-grade Ab production  Secondary infection- immune system stimulates IgG B cell to secrete Ab because antigen binding site better fit 

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IgM

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Forms pentameric str held together by a protein called J chain  5 weak binding sites, 2 per arm= 10 antigen binding sites  Ensures pathogens dont escape primary response Low affinity Key role=  protection during immune response but 10 binding sites compromise low affinity First Ig produced during initial immune response

IgD

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Found in serum- unknown function Usually found co-expressed with IgM on surface of unactivated B cells Acts as receptor

IgG

High affinity Key role= memory immune response Class switched in primary and secondary response  Occurs as disulphide linkage of 2 heavy chains and 2 light  Most common Ig in serum  Generated following initial infection due to class switching of IgM  Can cross placenta (receptors for Fc regions of IgG on placenta) Exceptional at activating C' and ADCC

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IgA

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Exists as dimer: two monomeric Ig units held together by J chain Present in serum and secretory fluids, most abundant in secretory fluids (tears, colostrum, saliva, intestinal tract Monomer in serum  Dimer in mucosal tissue  Secretory component (yellow chain) makes IgA well suited to reside in mucosa (wraps around dimer, protects J chain, stops dimer doesn't falling apart) Neutralises toxins (in blood stream)

IgE

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Least common Ig in serum  Additional constant region domain  Binds to specialised Fc epsilon receptors on mast cells, basophils and eosinophils  Strongly involved in allergic reactions and clearance of parasitic infections

IgE Roles: Deal with Parasitic Infections

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Enriched in mucosal tissues and associated with mast cells (innate) so are well placed to deal with parasites Parasitic antigen picked up by B cell, CD4 T cell help class switch to IgE Mast cells have receptors to bind IgE Fc region: parasitic antigen bound by IgE, bound by mast cell receptor  Triggers mast cell to secrete eosinophil chemoattractant factor  Eosinophils have toxic granules

Allergic Reactions

Susceptibility to allergic reactions influences by your genes: Allergy= immune response to something harmless Individuals predisposed to developing allergies= atopic individuals that have higher circulating levels of IgE than non-atopic individuals Allergies are inherited (not always) and strong link to maternal side of family

Allergic Reactions: Asthma

Narrower airways- can't take in as much air  IgE secreting B cell activates mast cell to release inflammatory mediators IgE binds mast cell causing secretion of histamines, leuktorienes, cytokines (inflammatory mediators) from mast cell  Secretory molecules affect airways by: Early response- bronchospasms, swelling and airflow obstruction Late response= airway inflammatiion, obstruction and hyperresponsiveness

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Allergic Reactions: Asthma

Microbes linked to asthma development: Up until toddler, immune system predominantly Th2 biased, becomes more Th1 with age  Children at risk of asthma/allergies retain high Th2 environment  Theories Th1 vs Th2 biased immunity: ​​​​​​​Hygiene hypothesis: Children with siblings and pets have lower risk of allergies/asthma  Siblings pick up infections as they grow, when a baby is exposed to wide range of infections, need Th1 response, shapes immune system to Th1 Infection-asthma hypothesis: ​​​​​​​Have own microbiome (gut bacteria) Infection when young, with beneficial microbes= shapes immune system to be less responsive to allergens Infection when young, with specific microbe combinations= more likely develop allergies and asthma

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Allergic Reactions: Asthma

Summary Ab Classes

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