B cells Flashcards
Describe the major steps of B cell development
B cells develop from hematopoietic stem cells
- after birth - the bone marrow is the site of B cell development and maturation
B cell development is co-ordinated by a complex genetic program in the bone marrow.Failure of, or defects in, this developmental program can be catastrophic
B cell development is tightly regulated by specific signals from the extracellular environment and by signals from the B cell ntigen receptors
the formation of a mature B cell involved a number of phenotypically distinct precursor cells :
- HSC – Early B — pro B — preBI—Large Pre BII ( Pre B receptor is formed inside cytosol at this stage) — small Pre-BII– immature B ( IGM is expressed on the surface) — mature B ( IgD and IgM is expressed on the surface)
mature B vells exit the bone marrow and continually recirculate.
upon activation with an ANtigen B cells can go into two types of B cells
- Antibody ( Ig) secreting cells - plasma cells
- memory B cells
B cells detect antigen through their antigen specific BCR and produce protective, antigen-specific antibodies
Demonstrate good understanding of the key mechanisms and signals that control B cell development
the cytokines involved are : CAMs FLT3 ligand SCF IL7
FLT3 ligand: - important for survival of HSCs and progenitor Cells
- a growth factor secreted by BM stromal cells ( binds to tyrosine kinase receptor - Flt3- expresed on early progenitor cells)
- important for the commitment to the Pro-B cell stage of B cell development
Stem cell factor- survival of HSCs and Proliferation of early pro- B cells
- secreted by BM stromal cells and binds to the tyrocine kinase receptor - cKit
- increases the survival of HSCs in vitro and contributes to the self - renewal and maintainence of HSCs in vivo
- plays important role from the Pro-B cell stage - induces IL7 - receptor expression
Interleukin 7 - proliferation of Pro B and pre B cells
( also important for regulation of thymocyte development and survival of mature T cells)
( for IL7)
Proliferation
Activation of STAT5
Survival-
Transcription of pro-survival genes (BCL2, BCLXL, MCL1)
Differentiation -
Promotes Ig HC gene rearrangements
PI3 Kinase signaling induces RAG gene expression
Prevents light chain gene rearrangements
What are B cells
B cells are the antibody producing subset of lymphoid cells
Account for 5–15% of all circulating lymphoid cells
B cells also are antigen presenting cells
Link the humoral and cell mediated adaptive immune responses
B cells originate and mature in the bone marrow
What are the five main immunoglobulin classes?
- IgG
-IgM
IgD
IgA1
IgE
heavy chain determines the class of antibody !
IgA and IgG also have subclasses
Diversity in the B cell (i.e. the antibody) population is due to Ig chain gene rearrangements
potential antibody diversity is 10^11
Different B cells produce Ig of different antigen specificities
What serves as the B cell antigen receptor (BCR) ?
membrane Bound Ig
Each B cell produces Ig’s of a single antigen specificity
Ig gene rearrangement is tightly controlled ! - ensures only one heavy and one light chain is expressed.
Different B cells produce Ig’s of different antigen specificities
What is the relevance of mutations in genes that are important for B cell development ?
they can cause :
Immunodeficiency diseases
B cell malignancies
Alterations in B cell tolerance may underlie some autoimmune diseases
Give some examples of B cell mutation diseases?
- X-linked Agammaglobulinemia
Bruton’s tyrosine kinase (Btk)
Encoded by gene on X chromosome
Essential for B cell maturation
Mutation of the Btk gene
B cells maturation blocked
Defect in antibody production
Immunodeficiency results in increased susceptibility to opportunistic (recurrent) bacterial infections
Streptococcus pneumoniae
Haemophilus influenzae
Therapy:
prophylactic antibiotics and replacement
gamma globulin (IVIg)
- Burkitt’s Lymphoma
High transcriptional activity and DNA breakage and repair at Ig gene loci during Ig gene rearrangements
Mistakes can result in deregulated cell growth
Burkitt’s lymphoma:
Ig gene segment mistakenly fused to a proto-oncogene called MYC that regulates the cell cycle; along with additional mutation(s) this leads to malignancy
Therapy:
cytotoxic chemotherapy
Rituximab (a monoclonal antibody drug that depletes B cells)
Why is the spleen very important? and what would happen without a spleen ?
The spleen is the site for the final maturation of developing B cells
The spleen also serves as a site for the development of new RBCs
The spleen also acts as a filter for the blood
Removal of damaged RBCs
Capture blood-borne pathogens
individuals without a spleen are extremely susceptible to infections with encapsulated bacteria - e.g. s. pneumoniae, H.influenza
the therapy would include vaccinations against the relevant pathogens
What environmental signals control B cell development ?
The Bone Marrow provides a maturation and differentiation microenvironment for B cell development
- Bone marrow Stromal cells nurture developing B cells -
( the function of these stromal cells is: - Specific cell-cell contacts between stromal cells and developing B cells through cell adhesion molecules (CAM’s)
- secretion of cytokines by stromal cells as growth and differentiation factors for B cells
( Different types of cytokines and cell-cell contacts needed at each stage of differentiation)
- e.g. IL7 - used for pre B cell…
Summary of B cell development and cytokine involvement?
- pluripotent potential
- self renewal capacity
- linease specific genes = off
( induction of cytokines - CAMs, FLT3 ligand , SCF, IL7) - lineage commitment
- differentiation , proliferation and survival
- B cell lineage specific genes= ON
- other lineage specific genes= OFF
How do BCR ( B cell receptors ) control B cell development ?
Expression of a functional pre-BCR leads to a transient downregulation of RAG1/2 expression
What are the checkpoints for B cell development?
During their early development in the bone marrow, B cells have to successfully pass two major checkpoints
These checkpoints test the quality of Ig heavy and light chain gene rearrangements:
Checkpoint 1
Occurs at the late pro-B cell stage
Mediated by a functional pre-BCR
- selection by the Pre - BCR ( allelic exclusion , signals cells to survive and become Pre- B cells (50% of cells)
Checkpoint 2
Occurs at the small pre-B cell stage
Mediated by a functional mature BCR
Why are the checkpoints for B cell development important?
Gene rearrangement process is imprecise : Unproductive Not translated into Ig chain B-cell dies Productive Translated into Ig chain Development proceeds
Immunoglobulin loci
Each B cell has 2 copies on homologous chromosomes
Rearrangements potentially made on both – deleterious effect antigen specificity of the B cell
What is Allelic exclusion?
Mechanism used by lymphocytes to prevent any one individual cell from expressing more than one specific antigen receptor
What is the structure of the preBCR?
The Igm heavy chain combines with surrogate light chain (SLC) to form a functional pre-BCR (analogous to BCR)
The SLC proteins VpreB and l5 look similar to Ig light chains but
Do not contain hypervariable regions
Have unique extended N/C-teminal tail regions – functionally important for preBCR signaling
Interactions between SLC tails and ligands thought to stimulate preBCR signaling
The pre-BCR signals the transition to next stage of development
( it signals in an antigen independent manner)
What are the steps of ligation of the pre- B cell receptor?
- unconfirmed ligand of pre B cell receptor ( and stromal cell) binds to large Pre- B
- this suppresses further H chain rearrangement - this triggers entry into the cell cycle
- this ensures only one specificity of Ab expressed per cell.
- this expands only the pre B cells with a functional preBCR - allelic exclusion can occur ( an expression of a gene on one chromosome prevents expression of the allele on the second chromosome.
( with answer from lecture !!!) Be able to describe the major steps of B lymphocyte (B cell) development?
HSC Pro-B cell Pre-B cell Immature B cell Mature B cell
Defined by Ig gene rearrangement and expression of the preBCR and the BCR
Developmental progression is regulated by key cytokines and preBCR/BCR expression
( with answer from lecture !!)
Demonstrate good understanding of the key receptors, signals and mechanisms that control B lymphocyte (B cell) development
Stromal cell derived cytokines and cell adhesion molecules play key roles in regulating early B cell development (HSC Pre-B cell stage)
Transient preBCR expression regulates the transition from the Pro-B cell to the Pre-B cell stage of development (Checkpoint 1)
Ig heavy chain allelic exclusion, cell cycle entry, BCR expression
Expression of the BCR regulates the transition from the Pre-B cell to the Immature B cell stage of development (Checkpoint 2)
Self-tolerance is mediated by Negative Selection of Immature B cells
Deletion
Anergy
Receptor editing
What is the pre - B cell receptor responsible for ?
- Ig heavy chain allelic exclusion
- Stimulates cell cycle entry
Generates a pool of pre-B cells that express identical Igm heavy chains (limited number of cycles) - Loss of preBCR expression
Down-regulation of l5 and VpreB expression - Expression of the mature BCR
Re-expression of RAG1/2
Igk/l light chain gene rearrangements
What is negative selection of immature B cells?
Acquisition of antigen specificity creates a need to check for recognition of self antigens
Up to 75% of Immature B cells in bone marrow have specificity for self antigen
Small pre-B cell
No antigen receptor at cell surface
Unable to sense Ag environment
!! May be self-reactive !!
Immature B cell
Cell surface Ig expressed
Able to sense Ag environment
Can now be checked for self-reactivity
What are some important mechanisms for self tolerance ( negative selection) ?
- Physical removal from the repertoire - Deletion
- Paralysis of function- Anergy
- Alteration of specificity - Receptor editing
After successful completion of negative selection
self tolerant B cells are exported to the periphery
What are some key points about BCR signalling?
The effect of BCR signaling is dependent on the developmental stage:
Pro-B cells + pre-BCR signal: developmental progression
Immature B cell + antigen: inactivation or death of the cell
Mature B cell + antigen: activation of the cell
(but needs additional signals–described in subsequent lectures)
Be able to discuss the major events that occur as B cells respond to antigen?
..
Be able to describe the major differences between T dependent and T independent antigens and discuss how they can each activate B cells?
…
Demonstrate good understanding of the spatial regulation of B cell activation in secondary lymphoid tissues?
…
What are the three major types of mature B cells ?
Conventional B cells (B2 cells, Follicular B cells)
Circulate between lymph node follicles and blood
Size of population determined by BAFF levels
Marginal zone (MZ) B cells
Reside in marginal zone of spleen where they can respond to particulate antigens in blood (bacteria, etc.)
Also dependent on BAFF for survival.
B1 B cells Prominent in peritoneal and pleural cavities, Produce “natural IgM antibodies” Respond to T-independent antigens Less dependent on BAFF
What is the importance of Allelic exclusion?
Prevents unwanted responses
Needed for efficient clonal selection
Needed to prevent holes in the repertoire
What is somatic hypermutation?
B cells that make high affinity antibodies are selected by the process of affinity maturation in the Germinal Centre
Goal: enhance production of antibodies that can strongly bind to an antigen
Process is mediated by Activation Induced Deaminase (AID)
introduces random single nucleotide mutations into Ig heavy & light chain gene hypervariable regions
Mutations that increase the affinity of the BCR for antigen result in a selective advantage
Low affinity BCR takes up and presents antigen to T cells inefficiently (no CD40L) no signal 2 B cells die by neglect
What is Ig isotype switching ( class switching ) ?
biological mechanism that changes a B cell’s production of immunoglobulin (antibodies) from one type to another, such as from the isotype IgM to the isotype IgG. During this process, the constant-region portion of the antibody heavy chain is changed, but the variable region of the heavy chain stays the same
this is an irreversible change in DNA - and depends on the action of the enzyme called - activation induced cytidine deaminase ( AID)
