B-Lymphocyte Development and Antibody Production

Question 1

B-cell

Developmental Stages

Answer 1

1. Pluripotent hematopoietic stem cell becomes committed to the B-cell lineage in the bone marrow.
   
   • Ag independent.
   • Requires microenvironment and interactions with bone marrow stromal cells.
2. Pro-B cells undergo heavy chain rearragement.
3. Pre-B cell characterized by presence of cytoplasmic μ heavy chain.
   
   • 2 heavy chains combined with 2 surrogate light chains to form pre-B cell receptor.
   • Successful binding of the pre-B cell receptor signals via tyrosine kinase receptor Btk that rearrangement has resulted in a functional HC.
   • This halts HC gene rearrangement & drives multiple rounds of cell division.
   • LC rearragement occurs, kappa first then lambda.
   • Stage ends with the formation of a functional LC (75% kappa and 25% lamba).
4. Upon successful assembly of light-chain gene, LC synthesized and joined with μ-heavy chains to form membrane-bound IgM ⇒ immature B cell.
   
   • Signaling via IgM triggers cessation of LC gene rearrangement.
   • Tolerance occurs here.
     
     • Ag binding results in apoptosis or anergy.
5. Immature B cells leave bone marrow and circulate throughout blood and lymphatics sampling antigens.
6. Immature B cells enter secondary lymphoid organs.
   
   • Receive signals important for survival and maturation.
   • Start to express IgD on surface to become mature naive B cell.
7. Naive B cells that encouter Ag are stimulated by Ag-mediated BCR cross-linking becoming activated B-cells.
8. Activated B cells divide, differentiate, and begin producing secreted IgM.
   
   • Migrate towards T_H cell compartment
9. T_H cells cause activated B-cells to differentiate into plasma cells or memory cells.

Question 2

B-cell Checkpoints

Answer 2

Every step is checked to ensure lymphocyte is functional.

If heavy chain fails then cell tries with other copy.

Both alleles at 𝛋-locus attempted first prior to λ-locus.

2/3 will expression kappa and 1/3 express lambda.

Question 3

Tolerance

Answer 3

Removal of potentially autoreactive immature B-cells via deletion or anergy.

1. BCR activation on immature B cell (IgM⁺, IgD^-) results in the loss of the cell via:
   
   • clonal deletion - apoptosis
   • anergy - functionally inactivated
2. Immature B cells that do not recognize Ag mature as they move through the 2° lymphoid organs and becomes mature B cells (IgM⁺, IgD^{<span>+</span>})
3. BCR engagement of a mature naive B cell results in activation, proliferation, and Ab production.

Assumptions:

• “self” molecules are constantly present and induce tolerance during development
• “non-self” components are transiently present, therefore, mature ells react to eliminate them when they appear.
  
  • Immature B cells are likely to be tolerized to those foreign Ag
  • The longer the Ag is present the less robust the immune response

The small # of autoreactive Ab which escaped tolerance kept in check by T-cells.

Question 4

Clonal Selection Paradigm

Answer 4

For an effective B-cell response, the immune system must:

1. establish and maintain a diverse array of Ag-reactive B cell clones that are deviod of pathogenic autoreactivity.
2. Selectively activate, expand, and modify these clones based on antigenic experience.

Question 5

Soures of Antigen

Answer 5

Activation of humoral immunity occurs in defined anatomical sites.

When an antigen enters the body it becomes concentrated in various paripheral lymphoid organs.

Blood borne Ag filbered by the spleen.

Ag from tissue spaces and inflammatory processes drained by lymphatic system to regional lymph node or lymph nodules.

Question 6

B Cell Complex

Answer 6

• Membrane-bound Ig non-covalently linked to Igα and Igβ
  
  • BCR binds the Ag
    
    • Igα and Igβ transmits the signal into the cell causing activation

Question 7

B-cell Activation

Answer 7

1. Binding of a multivalent antigen induces a conformational change in BCR.
2. Crosslinking of surface bound Ig.
3. Clustering and aggregation of multiple Ig’s and other cell surface receptors (e.g. CD45).
4. Transphosphorylation and activation of various protein tyrosine kinases.
   
   • FYI only: Btk, Blk, Lck, Fyn
5. Activated tyrosine kinases phosphorylate ITAMs (immunoreceptor tyrosine-based activation motifs) found on the Igα/Igβ chains
6. Phosphotyrosines become docking sites for adapter proteins which are subsequently phosphorylated.
7. Phosphorylated adaptors recruit various signaling molecules.
8. Results in activation of transcription factors that initiate specific gene expression leading to cell proliferation and differentiation.
   
   • B-cells do require a second signal for proliferation which usually comes from helper T cells or components of innate immunity.

Question 8

Activated B-cell

Functions

Answer 8

Antigen-mediated B-cell activation with appropriate secondary signal results in:

• production of soluble IgM
  
  • able to make IgM without T-cell help
• B cell proliferation and differentiation
  
  • clonal expansion of cells
• B-cell mediated activation of T helper cells
  
  • Leads to T-cell dependent B-cell activation
    
    • increased B-cell proliferation
    • increase Ab production
    • affinity maturation
    • isotype switching
  • Occurs in the germinal center of lymph node

Question 9

Primary vs Secondary

Immune Responses

Answer 9

Question 10

sIgA

Characteristics

Answer 10

Mucosal Immunity:

• sIgA binds inhaled and ingested microbes.
• Prevents adhesion
• Promotes destruction and clearance
• Thus inhibiting microbial colonization

Synthesis:

• Most of sIgA produced in lymphoid tissue of lamina propria
• Transported across the epithelium into the lumen

Question 11

sIgA Transport

Answer 11

1. Dimeric IgA (2 IgA monomers joined by J-chain) binds to the poly-Ig receptor on the surface of endothelial cells.
2. IgA endocytosed in vesicle and transported to luminal surface.
3. In the lumen, poly-Ig receptor cleaved by protease and sIgA released.
4. sIgA remains bound by a portion of the poly-Ig receptor which becomes the secretory component.
   
   • Helps protect sIgA from degradation.

If IgA deficient, the poly-Ig receptor is able to transport IgM into secretions so there is some mucosal protection.

Receptor has higher affinity for IgA.

Question 12

Neonatal Immunity

Answer 12

Newborns with under-developed immune systems.

Receives passive immunity from the mother via two routes:

Placental Transfer

• Maternal IgG binds to neonatal Fc receptor (FcRn) expressed in the placenta.
• IgG actively transported into fetal circulation.
• Only occurs during 3rd trimester.
  
  • Do not consider reactive diseases such as Rh incompatibility until 3rd trimester.
  • Increased risk of infection in premies.
• Half-life of maternal IgG is 21 days.
• Lowest [IgG] occurs 3-6 months after birth when IgG switches from being maternally derived to infant produced.

Breast Milk

• Colostrum (1st milk secreted after pregnancy) and breast milk is a source of maternal sIgA.
• Provides passive immunity against GI and respiratory infections.
  
  • Protection against pathogens in maternal environment and those that the mother has been vaccinated against.
  • Infant likely to encouter these too.
• sIgA does not cross into the baby’s blood.

Question 13

Active Immunity

Answer 13

• Stimulated by immunization
  
  • Infection
  • Vaccination
• Your immune system generates the response
• Takes more time (days to weeks)
• Results in memory

Question 14

Passive Immunity

Answer 14

• Generated by the transfer of Ab
• Recipient becomes transiently resistent or immune without exposure to the pathogen
• Rapid
• Lasts as long as Ab does
• No memory
• Examples:
  
  • placental transfer of IgG to fetus
  • sIgA in breast milk
• Treatments:
  
  • Anti-pathogen Ig (anti-hepatitis after exposure)
  • Anti-toxin Ig (anti-tetanus toxin Ig)
  • Anti-venoms (anti-rattlesnake venom Ig - uses horse serum)