L6 - Lymphocyte activation: B cells

Question 1

How do B cells act differently to T cells in response to antigens?

Answer 1

B cells don’t deal with antigens themselves

They release antibodies into the blood which recognise the pathogens

Question 2

What are memory B cells?

Answer 2

When some B cells keep the BCR on their surface – memory B cells

Question 3

What are antibodies used for?

Answer 3

Neutralisation
Opsonisation
Complement activation

Question 4

B cell antibody production

Answer 4

B cells also need several signals to become activated

Naïve B cells express membrane (m) Ig/BCR (IgM or IgD) & encounter non-self Ag in secondary lymphoid tissue

Binding of Ag to BCR provides signal 1 to the B cell

Question 5

Basis of Ag/BCR signal 1

Answer 5

BCR-associated polypeptides involved in signalling – Ig-alpha & Ig-beta allow BCR to go to the cell surface

Crosslinking BCR activates intracellular kinases

Has the same signalling motifs in its signalling molecules – ITAMs (yellow bits)

ITAM (immunoreceptor tyrosine-based activation motif) – series of AA that have 2 tyrosine residues that get phosphorylated

Question 6

How can signal 1 be increased/enhanced?

Answer 6

• If antigen has activated complement cascade
• Lots of C3b
• Complement receptor 2 (CR2) on B cell surface (CD21)
• CR2/CD19/CD81 form the BCR co-receptor complex
• Augments the signal

Question 7

What happens if an Ag that binds BCR is coated with complement?

Answer 7

it can also bind CR2 on B cells to give an increased signal 1

Other molecules in the complex: CD19 & CD81

Antigen binds to both the BCR & CR2 to induce a stronger signal

Question 8

How do B cells receive signal 2?

Answer 8

Bind signal 2 differently depending on the type of Ag they bind

Question 9

What are the types of Ag that B cells can bind to?

Answer 9

T (thymus)-independent Ag (TI)

T (thymus)-dependent Ag (TD)

Question 10

T-independent Ag (TI)

SIGNAL 2

Answer 10

Signal 2 is provided by either the:

1) Antigen itself
2) Extensive cross-linking of BCR

DOESN’T INVOLVE T CELLS
Induce antibodies in absence of T cells

Antibodies made this way cannot class switch so isn’t a great response

Question 11

T-dependent Ag (TD)

SIGNAL 2

Answer 11

Signal 2 is provided by CD4+ T cells

INVOLVES T CELLS – most antigens
• Drives a very good B cell response

B cells don’t go to the thymus – if you take out the thymus you get no T cells – no T cells means B cells don’t respond properly

MHC II presents peptide derived from antigen on the B cell

Signal 2 comes from the T cell in TD not the antigen itself like in TI

Question 12

What are the 2 types of TI antigens?

Answer 12

TI-1 Ag

TI-2 Ag

Question 13

TI-1 Ag

Answer 13

As well as binding to BCR, TI-1 Ag also bind to other receptors on all B cells providing signal 2

Eg. lipopolysaccharide from gram -ive bacteria binds to TLR4 expressed by B cells

In high concentrations can act as polyclonal activators (mitogens) for B cells

The 2 signals (1 from BCR, 2 from TLR) lead to B cell activation, proliferation & antibody secretion

Question 14

TI-2 Ag

Answer 14

Contain repeated epitopes
• Often polysaccharides – same sugar repeated lots
• Important in some bacterial infections – coated

Will therefore cross-link many BCR on the same B cell

Take longer to induce B cell activation

Develops after age 5 – need conjugative vaccines

Question 15

TD Ag response compared to TI Ag

Answer 15

Antibody responses seen to TD Ag are much better than those to TI Ag – the classical acquired response

Require T cells

Question 16

Process of TD Ag antibody response

Answer 16

1) B cell recognises coat protein of the virus, takes it up, processes it & presents it to the T cell
2) Doesn’t necessarily present bits of the coat protein
3) T recognises the internal parts of the virus – peptide shown by the MHC
4) When the T cell activates the B cell, B cell makes BCR to bind coat proteins
5) T cell epitope is inside the virus whereas B cell is outside the virus
6) Both parts end up being presented

B cells act as APC for TD Ag

Question 17

Why do you make conjugate vaccines?

Answer 17

Converting a TI Ag to a TD Ag is a way of improving the efficiency of a vaccine

Question 18

How do you make conjugate vaccines?

Answer 18

A sugar cannot be processed & presented –

1) Can take the sugar, add it to a protein (chemically or genetically)
2) Now have a conjugate protein – when it gets taken up by the B cell it creates peptide fragments
3) Can then present these fragments to the T cell
4) T helper cell recognises it & delivers signal 2 to the B cell
5) B cell now makes antibodies against the sugar that are T-dependent (TD)
6) Makes a better response – can now be classed switched

Couldn’t naturally do this as a response to a sugar

Question 19

Example of a conjugate vaccine

Answer 19

Haemophilus influenzae type b

Protective response requires antibodies to capsular polysaccharide (which is a TI Ag)

Coupling this to a protein such as tetanus toxoid converts it to a TD Ag

This allows young children to be immunized and protected (TI antibodies not produced until about 5 years old)

Question 20

What are B/CD4+ T cell interactions important for?

Answer 20

Good antibody responses

B cells enter lymph node from blood to find antigens & T cell to help

If a B cell comes into contact with its specific Ag it can then be activated

If Ag is TD, B cell presents peptide from Ag to CD4+ TH cells at the boundary of the T/B areas within the lymph node forming B/T cell conjugates

Question 21

Activated B cells

Answer 21

Conjugates of B lymphoblasts & T cells move to primary follicles (B cell areas)

Form germinal centres in follicle – B cells proliferating in the lymph node as a result of getting help from T cells

B cells divide rapidly to become centroblasts & undergo:
• Somatic hypermutation of Ig genes
• Isotype switching

Differentiate into non-dividing centrocytes (smaller)

Question 22

B/CD4+ T cell interactions

Answer 22

B cell binds Ag via BCR & presents peptide on MHC II from Ag to activated CD4+ TH cell
• As a result T cell then expresses CD40 ligand (CD40L)
• Secretes cytokines

B cell receives signal 2 from T cell via CD40/CD40L binding & via cytokine from T cells binding receptors –> B cell proliferation

CD40 signal also induces activation induced deaminase (AID) which is required for class switching & somatic hypermutatiom

Question 23

What is AID?

Answer 23

Activation induced deaminase (AID)

Required for class switching & somatic hypermutatiom

Question 24

Follicular T helper cells (TFH)

Answer 24

Found predominantly in the B cell follicles of the lymph node

Specialised to help B cells

Secrete either TH1 or TH2 type cytokines

Can be identified with specific markers that differ from other sub-sets of CD4 TH cells

Question 25

What can germinal centre (GC) B cells do?

Answer 25

Can either:

1. Differentiate into plasma cells
   • Secrete various isotypes
   • High affinity antibody, somatically mutated
2. Form long lived memory cells – recirculate
3. Die within lymphoid tissue
   • If BCR no longer binds antigen (as a result of unsuccessful V region created or somatic mutation)

Question 26

Somatic hypermutation

Answer 26

Introduces point mutations into V regions of the Ig

Approx. 1 mutation/V region/cell division – 106 x normal DNA mutation rate

Enzymes primarily involved include:
• AID & DNA repair genes

Question 27

Where are follicular dendritic cells (FDC) found?

Answer 27

In germinal centres

Question 28

What do follicular dendritic cells (FDC) do?

Answer 28

Not bone marrow derived dendritic APC

Only live in GC – not the same as DC

Cells in primary follicle that capture intact Ag for B cells to bind BCR

FDC capture Ag via FcR & CR as immune complexes

Oversee B cell affinity maturation

Question 29

How do FDC oversee B cell affinity maturation?

Answer 29

Centrocytes undergoing somatic hypermutation express mutated BCR on surface

Centrocytes thus compete with each other for Ag on FDC & for signals from T-FH cell

If mutated BCR binds Ag on FDC better than un-mutated BCR, will present more efficiently & receive CD40 signal from TFH cell

Failure = apoptosis

Those centrocytes that have generated higher affinity BCR survive to differentiate into plasma cells

Question 30

Role of CD40 expressed by B cells

Answer 30

Protects centrocytes from apoptosis

Allows isotype switching

CD40L deficiency/mutations = hyper-IgM syndrome – cannot switch isotypes

Question 31

Isotype (class) switching

Answer 31

IgM produced initially – co-expressed with IgD by differential mRNA splicing

IgG, IgA & IgE produced later – AID recombines DNA to change constant region of antibody without affecting existing VDJ region

Deficiency of AID also leads to hyper-IgM syndrome

Question 32

B cell activation summary

Answer 32

• B cell comes out of the bone marrow and hopefully isn’t auto-reactive
• While expanding its mutating its BCR to get a better BCR
• Affinity maturation process in the germinal centers
• If the BCR is really good & gets lots of T cell help it becomes a plasma cell
• May make IgM or told to switch depending on the cytokine
• DC tells T cell to what T cell to become
• T cell then tells B cell what antibody to make
• Some B cells wont release BCR and will keep them on the surface as memory B cells