HIS09 Lymphocyte Development

Question 1

***B cell development process in Bone marrow

Answer 1

2 aspects:

1. Intracellular change —> **Ig gene rearrangement + **Expression of poIypeptide Ig H+L chains (Status of Ig genes)
2. Extracellular change —> ***Expression of molecules on cell surface

Process:
HSC / Lymphoid progenitor in bone marrow (mature in bone marrow as well)

—> Progenitor-B (Pro-B, Early / Late) cells: Intranuclear
- undergo Ig μ ***heavy chain gene rearrangement

—> Precursor-B (Pre-B) cells: Intracellular

• express μ ***heavy chain in cytoplasm
• ***light chain genes rearranged (κ first, if unsuccessful then λ later) —> expressed in cytoplasm (packed together, move to surface)
• **Pre-B cell receptor: small amounts of μ Heavy chain + Surrogate Light chain (not involved in antigen recognition, only interaction with **growth factor, cytokines (e.g. from stromal cells) in bone marrow micro-environment for delivering signal for ***survival, proliferation, differentiation of Pre-B cell)

—> Immature B cells: Cell surface expression
- Monomeric ***IgM (2 μ heavy + 2 light) molecules on cell surface (as antigen receptor)

—> ready to leave bone marrow (central lymphoid organs) to secondary lymphoid organs (spleen, LN)

—> Mature B cells (Naive B cell): Monomeric **IgM + **IgD on cell membrane

—> Plasma cells (Effector cells): Ab production —> ***Humoral immunity

5-15% circulating lymphoid pool are B cells, defined by ***presence of surface Ig

Question 2

Phenotypic changes on cell surface during B cell differentiation

Answer 2

Main markers (surface molecules):
—> Cluster designation / Clusters of Differentiation (CD molecules: leukocyte cell-surface molecules recognised by clusters of mAbs, each cluster of mAb bind specifically to a particular CD molecule —> ∴ some mAB can attack human leukocyte)
—> Identification of immune cells in lymphoid organs

Examples:

1. CD34 (early progenitor B cell)
2. ***CD19 (throughout the development in bone marrow, also in peripheral B cell)
3. CD20 (B cell, T cell)
4. CD22
5. CD40 (throughout the development in bone marrow)

Question 3

Microenvironmental factors

Answer 3

Stromal cell-derived cytokines:

1. ***SCF (stem cell derived factors)
2. ***IL-7 (enhance B cell survival) —> clinical manifestation: leukaemia / lymphoma changes

Pro-B cells bind to stromal cells via
- **CD44 + **c-kit (on B cell surface)
- **Hyaluronic acid + **SCF (on stromal cells)
—> activate Tyrosine kinase and stimulate proliferation

Late Pro-B cell, Pre-B cell:
- ***IL-7 receptors (bind to IL-7 to promote further maturation)

Question 4

B cell deletion during development in bone marrow

Answer 4

• 5-10% survive (successfully rearrange Ig gene), 80% lost
  —> Reason: Ig gene rearrangement is an error-prone process (only 3-5% successfully complete gene rearrangement)
• By Apoptosis
  —> condensed
  —> apoptotic bodies without releasing intracellular contents
  —> no inflammation

(Necrosis: release intracellular contents, trigger inflammation
Necroptosis (programmed necrosis) for infected cells)

**Allelic exclusion:
- productive Ig gene rearrangement can only occurs on **one chromosome (H chain locus for H chain; κ, λ light chain locus for L chain)
- one chromosome fail —> other one activated
- one chromosome succeed —> proceed
- suppression of non-expressed H chain loci and non-expressed L chain loci
—> **ensure each cell only expresses Ig of a **single H chain isotype and V region specificity, and a ***single L chain isotype and V region specificity

Question 5

***B cell selection during development

Answer 5

***ONLY Negative selection

When **immature B cell express functional surface **IgM but not IgD
—> ready for selection by self-antigens (self MHC molecules: cell bound / soluble)
—> occurs in ***Bone marrow
—> successful in selection
—> proceed to express IgD
—> peripheral secondary lymphoid organs

1. Autoreactive B cells to multivalent self molecule (**Cell-bound self MHC)
   —> Deletion by Apoptosis (Negative selection)
   —> Immune **tolerance induction
2. Autoreactive B cells to **soluble self MHC molecule
   —> migrates to periphery
   —> **Anergy (suppression; cell life prolonged)
   —> Deletion by Apoptosis (Negative selection)
3. No self reaction
   —> migrates to periphery
   —> Proliferation and Maturation
4. **Receptor editing in autoreactive B cell
   —> strong ligation of IgM by self antigen
   —> trigger arrest of B-cell development and continue **light-chain rearrangement (再黎過)
   —> a new receptor specificity expressed
   —> if still reactive —> Apoptosis
   —> if no longer self-reactive —> migrates to periphery —> Proliferation and Maturation (rescue some autoreactive B cells)

Question 6

Clinical significance of B cell development

Answer 6

1. Defective B cell development —> Immunodeficiency
2. Autoreactive B cells escape selection —> Autoimmune disease
3. B cells with aberrant Ig gene rearrangement —> B cell tumours

Question 7

***T cell development process in Thymus

Answer 7

HSC in bone marrow
—> progenitors migrate to Thymus for differentiation

• Undergo gene rearrangement
  —> produce unique antigen receptor of ***different isotypes (T cell receptor)
  —> stage and nature of gene rearrangement an indication of maturation level
• Unlike B cells:
  1. Gene rearrangement: **2 different sets of receptor genes (TCR is a Heterodimer)
  2. Gene rearrangement give rise to **2 T cell lineages (α:β and γ:δ)
  3. Rate of T cell production greatest ***before puberty
  —> after puberty, Thymus shrink and production of new T cell is very low
  —> T cells very long-lived (unlike B cell)
• Thymus ***fully developed before birth
  —> young individuals: numerous developing T cell embedded in Thymic stroma (epithelial network)
  —> unique microenvironment for T cell development

**3 stages (migrate from capsule —> cortex of thymic lobules —> medulla of Thymus: 8-10 days):
**Double negative thymocytes (CD4-, CD8-) + no surface marker CD3 (associated with TCR)
—> **Double positive thymocytes (CD4+, CD8+, CD3+) (Cortico-medullary junction)
—> **Positive selection + **Negative selection
—> **Single positive (CD4+ or CD8+) + CD3+ (during migration through medulla)
—> leave thymus and migrate to peripheral lymphoid organs
—> **Cell-mediated immunity + **Induction of Humoral immune responses

CD4: co-receptor for MHC class II molecule, occurs on helper T cells
CD8: co-receptor for MHC class I molecule, occurs on cytotoxic T cells

Question 8

***T-cell subset

Answer 8

2 types of T cells:

1. α:β T cells (expressing α:β TCRs) (95%)
2. γ:δ T cells (expressing γ:δ TCRs) (5%)

α:β T cells:
—> CD4+ cells (Th): Help / induce immune responses —> Th1 (IL-2, IFN) + Th2 (IL-4, IL-5, IL-6, IL-10)
—> CD8+ cells (Tc): Cytotoxic

Question 9

***T cell selection in Thymus

Answer 9

***Positive selection —> Negative selection —> Only T cells that do not recognise self antigens will leave Thymus

Positive selection (要bind到self-MHC + foreign peptide):
- double-positive T cell are screened for their ability to recognise **foreign peptides when presented to TCR by **self-MHC
—> T cell has opportunity to test several **consecutive α chain rearrangements
—> recognise peptide bound to self-MHC class I molecules
—> programmed to express CD8 co-receptors (vice versa)
- Mediated by **Thymic cortical epithelial cells
- **Ensures mature T cells can recognise antigens **in context of self-MHC molecules
- T cells with TCRs having moderate affinity for self-MHC are allowed to further develop

Negative selection (唔可以bind self peptide):
- T cells with receptors that recognise **self peptides:self MHC complex too well will be destroyed
—> eliminating potentially **self-reactive cells
- Mediated by ***Dendritic cells, Macrophages

Question 10

Clinical significance of T cell development

Answer 10

Primary T-cell deficiency
- **DiGeorge syndrome (1 in 5 per 100,000)
—> arises from defect in Thymus **embryogenesis
—> ***non-functional Thymus
—> impaired cell immunity + cannot support B cell responsiveness
—> critically susceptible to infectious diseases
—> die at young age

Question 11

Clinical significance of lymphocyte development and selection

Answer 11

1. Self from Non-self
   - Autoreactive lymphocytes —> Autoimmune disease
2. Immune surveillance
   - Abnormal lymphocyte —> Lymphocyte malignancy

Question 12

B cell-targeted therapy for RA, Lymphoma, SLE

Answer 12

Rituximab
- monoclonal Ab against CD20 (expressed mainly in B cell)
—> deplete most B cell in immune system
—> treatment of RA, Lymphoma, SLE

Question 13

B cell heterogeneity

Answer 13

B2 cells (conventional B cells)

B1 cells (10% of B cells) (IgM+, IgD-)

• produce natural Ab
• located in body cavity
• generated by HSC in fetus
• may have role in autoimmunity