L22: Cell mediated Immunity and Lymphocyte Ontogeny

Question 1

What is the difference between polyclonal vs monoclonal antiserum/ antibodies : production and characteristics

Answer 1

Polyclonal antisera:
-produced by injecting antigens in an animal and purify the antibodies within the serum.

-These antibodies would have a range of epitope recognised and range of affinities

Monoclonal
- Antibody producing B cell fused with immortal B cell tumour cell (myeloma).

• produced just one class of antibody with single affinity and single epitope

Question 2

What immune response is triggered in first and second skin graft rejection (non HLA matched)

Answer 2

First time
1. Precursors to CD8 cytotoxic T cell recognises the antigen/ incompatible HLA complex on the graft cells with their TCRaB receptor.

2. Becomes activated with further co-stimulation from H-Tcell cytokines
3. Proliferate and differentiate into
   a) cytotoxic effector population which seek out and destroy cells
   b) memory population

Second time
- Mostly antibody mediated rejection due to memory B& T cells activated more rapidly.

Question 3

What are the 2 killing mechanism used by cytotoxic T cell effector population

Answer 3

After using the CD8 and TCR to bind to target cell

1. Release of Perforin and enzymes for polymerisation leading to formation of pores in the target cell membrane
   - Release hydrolytic enzymes (granzymes) which enter and digest target cell
2. Release cytokines (TNF-a, B and IFN-y) which induce apoptosis

Question 4

Describe the steps in B cell ontogeny (5)

Answer 4

1. uncommitted stem cell in the bone marrow
2. Pre B cell rearrangement and deletions in genes that code for Heavy chain, then Light chain
3. Immature B cell (that can produce surface IgM)
4. IgM is tested for receptor binding against self antigens ONLY in the bone marrow, if self-reactive then clonal deletion of immature B cell (central tolerance)
5. Mature B cell expresses both surface IgM and IgD. Exported from the bone marrow then live in the 2ndary lymphoid organs

Question 5

Describe the process of Immunoglobulin gene rearrangements in B cell ontogeny (of heavy chain, K light chain and lamba light chain locus) in the Bone marrow

Answer 5

1. @ the Heavy chain locus, random selection of each of the V,D,J exons and deletion of the rest to form the variable region which is all added together
2. this is transcribed, translated and spliced with together with the constant regions that code for different types of antibodies. Cmu is first so it will make IgM first.
3. @ the Light chain locus random selection of each of VJ kappa exons and is transcribed, translated and spliced together with the one constant region
   - If kappa exons fail to translate on both alleles, then lambda exons will be used.

Question 6

Why can we produce a wide range of b cells/t cells with different variabilities despite a limited amount of DNA

(the same variable region is used with different constant regions to make different antibodies)

Answer 6

1. For Variable region, more than one exon (eg. VDJ) and a random selection of 1/multiple versions of a eg. V exon leads to more possible combinations than if the variable region was only coded by one exon
2. Joining the different exons (eg. VDJ) together is sloppy leading to base deletion/addition in the overall sequence.

Question 7

What is the gene mechanism behind class switching in B cells

Answer 7

Initially mature B cell will be using IgM, however

After a B cell has been activated and divided a bit, it will receive a signal from helper T cell to delete the Cmu and Cdelta regions of gene and select Cgamma to make IgG whilst still using the same variable region

Question 8

Describe the steps in T cell ontogeny (moving from the cortex to the medulla) - two paths

Answer 8

1. Uncommitted thymocyte (came from the bone marrow)

Path 1:

1) Within the Cortex Rearrangement of TCR gamma/delta chains) and surface expression
2) death or export

Path 2

1. Within the Cortex Rearrangement of TCR Beta and Alpha chain and surface expression
   2) TCR express both CD4 and CD8 accessory molecules. Then combo of TCR + CD4 , TCR + CD8 is tested against presentation from dendritic cells.
   3) If HLA class 1 or 2 is not recognised by either combo, then it is killed = Positive selection
   4) Downregulation of the CD that was not in the combo which recognised the HLA.
   5) Within the medulla, Receptor combo is tested against self peptides with HLA. If strong self reactivity then killed = negative selection
   7a) Cells exported to 2ndary lymphoid organs

Question 9

Describe the process of TCR gene rearrangements (alpha, beta chain) in the Thymus

The antigen binding site

Answer 9

1. Beta chain rearrangement: VDJ and Alpha chain rearrangement VJ, through random selection, deletion of others to generate one a and B variable region
2. Transcription, translation and splicing together with constant regions to form a TCR

Question 10

How does HLA haplotype influence susceptibility of disease

Answer 10

The specificity of T cells produced depends on the positive and negative selection which relates to the individuals HLA.

This might lead to TCR in the repertoire that has lower affinity for some pathogen peptide compared to someone else because of the HLA haplotype

Question 11

Where are TCRgd cells found and their role

Answer 11

Mucosa of the gut and in the skin

Maybe early defence against pathogens as they respond to heat shock proteins which may be expressed by invading microorganisms in response to stress