Lecture 10 - B Cell Immunity

Question 1

3 kinds of B cell Subsets exist…

Which are the main B cells?

Which are considered innate like?

Which are T independent and which are T dependent?

Which cells mediate the adaptive immune response?

Answer 1

1. B-2 cells
2. B-1 and Marginal Zone B cells
3. Conventional B-2 cells are T DEPENDENT

( B1 and MZ are T independent)

4. B-2 = ADAPTIVE IMMUNITY

Question 2

Which of the following do not undergo somatic hypermutation? What is required for somatic hypermutation and what are these requirements similar to?

1. B-1
2. B-2
3. Marginal Zone B cells

Answer 2

B-1 and MZ B cells

• somatic hypermutation requires AID and Switch region (like isotope switching)

Question 3

Where are B-1 cells made? Where are conventional B-2 cells made?

State the following for B-1 Cells:

1. Primary Location
2. Where first produced
3. Repertoir (large or smalle) & why?
4. Long or short lived
5. Innate or Adaptive immunity
6. Memory? Somatic Hypermutation?

Answer 3

1. FETAL LIVER
2. Bone Marrow

B-1 Cells:

1. Body cavities –> peritoneal, pleural
2. FETAL LIVER
3. SMALL repertoire since no somatic hypermutation or isotype switch
4. LONG lived –> self-renewing
5. INNATE
6. NO MEMORY OR SOMATIC HYPERMUTATION

** self-renewing, unlike B-2 cells which are replaced from bone marrow

RESPOND TO CARB ANTIGEN (unlike B-2 which responds to peptide)

Question 4

State the following for MZ cells:

1. When made?
2. V-region repertoir
3. Primary location**
4. Respond to Carb Antigen?
5. Somatic hypermutation?
6. Memory?
7. Innate or adaptive?

Answer 4

1. After birth
2. Restricted V region (not diverse)
3. SPLEEN
4. YES respond to carb antigen (microorganisms)
5. NO somatic hypermutation
6. NO MEMORY
7. INNATE!!!!

** long lived!!

Question 5

Where are Marginal Zone B cells found?

Answer 5

SPLENIC marginal zone at the interface of circulation & lymphoid tissue

** innate like –> first line of defense**

Question 6

Protein antigens usually require adjuvent, what is the purpose of adjuvent?

What is the most common adjuvent used with vaccines?

Answer 6

Adjuvent enhances IMMUNOGENICITY

2. Use Alum –> Aluminum Hydroxide
   - allows for greater affinity of antigen to antibody
   - readily invested by APC’s
   - - can get an insoluble form which will be rapidly taken up by macrophages

** too little immunogen results in NO immune response**

Question 7

What are the main factors that influence Immunogenicity?

Answer 7

1. Size
   - large size
   - at least 10,000 daltons
   (ex: insulin injected from bovines was only 6,000D and did not elicit an immune response which is why it could be used for diabetic patients back in the day)
2. Subcutaneous
3. Need PARTICULATE form
4. Adjuvants
   - can get an insoluble form which will be rapidly taken up by macrophages

Question 8

The following describes what type of B cells:

1. T INDEPENDENT
2. No somatic Hypermutation
3. No memory

Answer 8

B-1 and MZ B cells

Question 9

____ of Immunogen affects the immune response.

Ex; Snake Venome

Answer 9

DOSE

• need at least 10,000 daltons

main point :
NEED a certain amount of antigen before the primary immune response is initiated to produce IgM, soon after IgG produced for secondary response

Snake Venom will not generate an immune response since it does not induce a large enough immunogenic antibody production

Question 10

State how T - Dependent B activation occurs.

Answer 10

1. BCR interacts with antigen
2. ANtigen is internalized with the BCR and is degraded
3. Peptides associate with MHC - II molecules and go to the surface of the B cell
4. TCR of the T cell recognizes the peptide in context of MHC-II and is stimulated to produce cytokines
5. Cytokines activate the B cell to proliferate and differentiate into antibody producing cells

Question 11

What are the 2 signals required for T- Dependent B cell activation?

Answer 11

1. Interaction of BCR with antigen
2. Interaction of TCR with peptide/MHC complex and interaction between COSTIMULATORY MOLECULES CD40 (B cell) and CD40L (T cell)

Question 12

How does T INDEPENDENT Activation occur?

Answer 12

1. Usually Carbohydrates (like B-1 and MZ B cells)
2. SECOND SIGNAL is provided by the ANTIGEN
   - can give a signal to the B cell to become activated via a TLR

Question 13

Activation of Th cells by TCR interaction with peptide/MHCII triggers T cells to secrete cytokines to activate and initiate B cell proliferation/differentiation/

What are these cytokines?
What second signal is required before the cytokines are released?

Answer 13

1. IL4, IL5, IL6
   IL-4
   - (involved in Th2 response, promotes growth of B cells, enhances class switching to IgE and IgG)

IL- 5

• promotes differentiation of B cells
• enhances switching to IgA
• stimulates growth of eosonophils

IL-6
- causes fever and stimulates production of acute phase proteins

2. CO-STIMULATORY signal by B cell CD40 binding to T cell CD40L

Question 14

What 2 things are required to push B cells into proliferation?

Answer 14

Cytokines (IL4,5,6)

and Co-stimulatory Signal (CD40 & CD40L)

Question 15

Once the B cells proliferate, what are formed?

Answer 15

1. GERMINAL CENTERS
2. Plasma Cells
3. memory B cells

Question 16

Because B and T cells must recognize the same antigen, how does one make Anti-Polysaccharide Antigen that functions as a T -dependent response (but is actually T-independent)?

State how this relates to the example of Hymophealus Influenza Vaccines.

Answer 16

B cells recognize 3D conformation of antigens
T cells recognize peptide in context of MHC

ex: hymopholus Influenza  cannot make an anti-carbohydrate antibody (which are T independent responses, but babies do not make T independent, ONLY T DEPENDENT)

how do we get a baby to make T independent antigen (anti carbohydrate)???

1. Have a B cell bind bacterial polysaccharide epitope linked to tetanus toxoid protein
2. The B cell recognizes and binds the polysaccharide, internalizes and degrades the whole conugate and then displays toxoid derived peptides on surface MHC Class II molecules.
3. Helper T cell generated in response to earlier vaccination against the toxoid recognize the complex on the B cell surface and activate the B cells to produce anti-polysaccharide antibody.
4. This antibody can then protect against infection with H. Infleunza type B.

MAIN = take a T independent antigen and make it a T dependent antigen
(germinal center, memory, somatic hypermutation)

Question 17

In regards to the primary and secondary response:

1. Which has better affinity? Why?
2. What antibodies are produced in both?

Answer 17

1. SECONDARY RESPONSE
   - due to somatic hypermutation at the end of primary response!!!!
   - antibody titer is higher in secondary response
2. IgM = primary, SLOW
   IgG = secondary, QUICK

Question 18

B and T cells must recognize the same _____ although they don’t recognize the same _____.

Answer 18

1. Antigen
2. Epitope

• H. Influenza Type B:
• babies don’t make T -independent responses to polysaccharides
• link the polysaccharide to protein (tetanus toxoid- t dependent)

B cells with specificity for the polysaccharide will bind the H. Influenza polysaccharide linked to tetanus toxoid and will present peptides of the tetanus toxoid to T cells
- T cells can interact and stimulate H. Influenza polysaccharide specific B cells

Question 19

How do we get IgG in the secondary response from IgM?

What is required for this? (2)

Where does this occur?

Answer 19

1. Isotope Switching
2. . AID & Switch region
3. Occurs in the PERIPHERY
   - in the germinal center

Question 20

What regulates Isotope Switching?

What is rearranged?

Can this be reversed?

Answer 20

1. Occurs during T dependent responses and is regulated by T Cell CYTOKINES
2. C H region
   - heavy chain of the constant region
   - antigen specificity remains the same! (same VDK associated with IgA that was with previously made IgM)
3. Cannot be reversed since all the intervening genes were spliced out!

Question 21

What 2 things from previous lectures rely on ALTERNATIVE SPLICING?

Answer 21

1. Membrane vs. Secreted IgM

2. IgM vs. IgD

Question 22

State the following for Somatic Rearrangements:

1. Mediated by what 2 things
2. Where does it occur
3. Does antigen specificity change?

Answer 22

1. RSS and RAG1/2
2. BONE MARROW & THYMUS (tcell maturation)
3. YES!!
   - Variable region VDJ changes

Question 23

State the following for Isotope Switching:

1. Mediated by what 2 things?
2. Where does it occur ?
3. Does antigen specificity change?

Answer 23

1. AID & Switch Region
2. PERIPHERY –> germinal center
3. NO –> epitope remains the same
   - only the constant region heavy chain changes (isotype determinant)

Question 24

What are the sites of B cell proliferation and differentiation?

What occurs here? (4 important events)

Answer 24

GERMINAL CENTERS
- form during immune response in SECONDARY lymphoid organs (spleen, lymph nodes, etc)

1. Isotope Switch
2. . Somatic Hypermutation
3. Development of Memory B cells
4. Development of Plasma cells

Question 25

What type of Bcells are killed by apoptosis once the B cells are activated? What type of B cells are POSITIVELY SELECTED?

Answer 25

1. Anti- self 2. No- affinity/Low-affinity Antibody HIGHER Affinity antibody - therefore little antigen will be required to allow for successful binding to antibody Produce 1. Memory B cell 2. Plasma B cell via T cell help and B cell receptor cross-linking to sustain B cell proliferation and differentiation

Question 26

As the Immune response changes from primary to secondary (and eventually tertiary), what increases? This occurs due to what mechanism?

Answer 26

1. HIGHER AFFINITY of Ig Genes | 2. Due to SOMATIC HYPERMUTATION

Question 27

What is found in the Germinal Centers in the following zones: 1. Dark Zone 2. Light Zone 3. Mantle Zone

Answer 27

1. Dark Zone - B cells! 2. Light Zone - Follicular Dendritic Cells 3. Mantle Zone - T cells!

Question 28

How do we get a B cell to select a high affinity antibody? Where is the antigen for this high affinity reaction?

Answer 28

1. B cell must present antigen (take it up) and express it in context of MHC II so that T cell can see it and react 2. the antigen is on FOLLICULAR DENDRITIC CELLS!!! - From the germinal centers

Question 29

What is the function of follicular dendritic cells?

Answer 29

FOLLICULAR DENDRITIC CELLS with ANTIGEN ON THE SURFACE (does not take up antigen, therefore not APC) - DISPLAY HIGH AFFINITY ANTIGEN 1. B cell sees the FOLLICULAR dendritic cell with antigen 2. if sees antigen will take it up, process, and present in context of MHC II to T cells 3. T cells interact and initiate co-stimulatory signal and cytokines necessary for B cell activation 4. Activation of B cells stimulate the B cell to become a memory B cell or a plasma B cell*** everything else dies by apoptosis!!

Question 30

What is Somatic Hypermutation? What enzyme is required?

Answer 30

1. Changes in the antigen binding specificity as a result of somatic mutations in V genes which can generate additional diversity 2. AID - AID deficient patients only have IgM ! - the Ig genes are not somatically diversified

Question 31

After somatic hypermutation, cells with high affinity receptors are rescued from cell death by antigen in the form of antigen-antibody complexes bound to _____ followed by B cell presentation of the antigen to Th cells.

Answer 31

FOLLICULAR DENDRITIC CELLS

Question 32

Primary B cell blasts enter the germinal center and leave as ____ or ____

Answer 32

1. Memory B cells | 2. Antibody Forming cells (plasma blasts)

Question 33

How do T cells enhance antibody production? 1) How are Th cells activated? 2) How are Plasma Cells & Memory cells produced?

Answer 33

1. Precursor T helper + APC = Activated Th cells 1. The APC takes-up antigen, processes it and expresses peptides in context of MHC II 2. The TCR recognizes peptide/MHCII complex and becomes activated Activated Th cell + Antigen Specific B cell --> activation of B cells = PLASMA cell + MEMORY B cell 1. The TCR recognizes peptide/MHCII complex on B cell and the TH cells secrete cytokines which stimulate the B cell to proliferate and differentiate

Question 34

What 4 things are missing from T- independent (TIAntigen Polysaccharide) RESPONSES?

Answer 34

1. No germinal center formation 2. No somatic Hypermutation 3. No memory 4. No isotype switch * * LOW AFFINITY TO ANTIBODY - No secondary signal of CD40 and CD40L - -> instead antigen is the secondary signal!!! - vaccines thus require boosters or adjuvants for antigens lacking a peptide component (lipopolysaccharides from gram negative bacteria) ex: pneumococcal polysaccharide vaccine

Question 35

What 4 events occur in T DEPENDENT responses?

Answer 35

1. Germinal center formed 2. Somatic Hypermutation 3. Memory 4. Isotype Switching ** secondary signal occurs: CD40 with CD40L**

Question 36

What antibodies are usually formed by T independent responses? What are some antigens for TI responses?

Answer 36

1. IgM - no somatic hypermutation - antibodies have lower affinity than those of T-dependent 1. Lipopolysaccharide, polymerized flagellin, pneumococal polysaccharide, natural polysaccharide, dextran, levans, and hyaluronic acid - rapid response to bacterial antigens - B cells may be triggered by these antigens by CROSS-LINKING the BCR * * usually no memory**