Cell Mediated Cytotoxicity

Question 1

Cells that are CD25+. . .

Answer 1

. . .have high affinity IL-2 receptors!!

Question 2

CD40L/CD40 interaction:

Answer 2

T cell giving instructions back to the APC

Question 3

Where does IL-2 bind?

Answer 3

CD25!!

Question 4

What activates CTLs?

Answer 4

APC

Question 5

What sequence signals are required for the activation of CTL effector cells?

Answer 5

1. Antigen-specific signal, transmitted by the TCR upon recognition of the proper peptide/MHC Class I complex presented by a licensed APC
2. Co-stimulatory signal is transmitted by CD28/CD80/86 interaction between the CTL-P cell and licensed APC
3. IL-2 secreted by Th1 or Th17 CD4+ cell or the CTL itself result in the proliferation and differentiation of the antigen-activated CTL-P cell to fully active CTL.

Question 6

What effector molecules are produced by effector CTLs?

Answer 6

• High affinity IL-2R (CD25) and synthesizes IL-2
• Low levels of L-selectin and CCR7 (so they can escape lymph node - opposite naive CTL-P)
• High levels of CD44 and LFA-1 (homing and retention to sites of inflammation)
• Exhibits cytotoxicity (starts to produce perforin and granzyme)

Question 7

What is NK cells major role?

Answer 7

Killing virus-infected cells, intracellular pathogen-infected cells and tumor cells

Question 8

What stimulates NK cell activity?

Answer 8

IFN-alpha (Type I interferon), IFN-beta (Type II interferon), IFN-gamma, TNF-alpha, IL-15
-NK cells are recruited within 3 days of viral infection, before CTLs

Question 9

What two things do NK cells do?

Answer 9

• Recognize lack of self

- Recognize self + foreign antigen

Question 10

What do NK cell release/produce?

Answer 10

IFN-gamma

• It tilts the immune response toward Th1 cells by inhibiting Th2 cells and inducing IL-12 production by macrophages and dendritic cells
• IFN-gamma can activate macrophages (M1 - angry) and NK cells

Question 11

What are NK cells like in the bloodstream?

Answer 11

• Make up 5-10% of circulating lymphocytes
• 90% of blood NK cells are CD56low –> most effective killers of target cells (10% CD56high - no granules - release cytokines like IFN-gamma)

Question 12

What cell surface proteins of CD4+ T cells and APCs are important in the process of “licensing” the APC for cross-presentation of antigen?

Answer 12

TCR on Th1 or Th17 binds to MHC Class II on APC.

CD40 on APC binds CD40L on Th1 or Th17. This interaction gives the APC instructions about what cytokines it can release.

Question 13

What happens in Licensing APCs?

Answer 13

• Licensing APCs requires interaction with CD4+ Th1 or Th17 or direct interaction with a pathogen - TLR molecule
• Presentation of antigen to the CTL-P through MHC Class I can only take place after the APC has “found” a pathogen or has been “told” that is has found a pathogen. This acts as a control to prevent self-recognitation by CTLs.
• Infection of a dendritic cell by a virus can also assist in the presentation of antigen associated with MHC Class I molecules.

Question 14

How do CD8+ T cells bind target cells?

Answer 14

• TCR-CD3 complex on CTL recognizes the peptide/MHC Class I complex on the target cell
• LFA-1 on the CTL binds to ICAMs on the target cell
• Antigen activation converts the LFA-1 from a low affinity state to a high affinity state for better binding
• After about 5 to 10 minutes, LFA-1 returns to a low-affinity state, resulting in the dissociation of the CTL from the target cell.

Question 15

What are the mechanisms by which CTLs kill?

Answer 15

1. Perforin & granzyme secretions:
   - -Perforin molecules form a pore on target cell membranes
   - -Granzyme molecules activate apoptosis by cleavage of caspases (granzymes flow through perforin pore)
2. Fas ligand protein on the cell membrane surface:
   - -Membrane-bound FasL binds to Fas on the membrane of the target cells and initiates killing
   - -Activates apoptosis by cleavage of caspases
3. CTLs can also kill by TNF production and secretion

Question 16

What are the two pathways to activate apoptosis?

Answer 16

1. FasL binds Fas and activates Caspase-8 (through FADD and procaspase-8) –> Bid gets stimulated which causes mitochondria to release cytochrome C which binds to Apaf-1 and caspase-9 –> This complex activates Caspase-3 which then activates apoptotic effectors –> Caspase-8 also directly activates caspase-3
2. Granzyme gets into the cell and activates Bid and produces procaspase-3 (inactive) –> same progression as FasL

Question 17

What cell surface proteins do NK cells express?

Answer 17

CD16 (FcgammaRIIIA) and NKRs

Question 18

Do NK cells need to be educated in the thymus?

Answer 18

NO

Question 19

Do NK cells undergo rearrangement of receptor genes?

Answer 19

NO - they don’t make TCR or CD3

Question 20

Do NK cells have memory?

Answer 20

They may have. . .?

Question 21

Like CD8+ T cells, is NK killing MHC restricted?

Answer 21

NO - no CD4 or CD8 corrector binding requirement

Question 22

How is NK cell killing similar to CTL killing?

Answer 22

1. FasL expressed not the surface can kill cells expressing Fas
2. Perforin and granzyme released from granules
3. TNF expressed on the surface and secreted

Question 23

What is the process of NK cell recognition of cell targeted for killing?

Answer 23

• NK cells kill cells that mask the presence of foreign antigen on MHC Class I.
• If cell’s ligand binds the NK cells activating receptor and there is no ‘negative signal from MHC Class I (no MHC Class I present!) binding to Ker ligand, then the NK cell will kill!
• If the NK cell binds Class I MHC to its inhibitory receptor/Ker ligand and binds the cells ligand to the NK cell’s activating receptor, it will not kill.

Question 24

What effector cells are associated with ADCC (antibody-dependent cell mediated cytotoxicity)?

Answer 24

• NK cells
• Macrophages
• Monocytes
• Neutrophils
• Eosinophils

Question 25

How does ADCC happen?

Answer 25

1. Effector cells bind antigen via the antibody, through the Fc receptor.
2. Killing is mediated by cytolytic enzyme release by macrophages, neutrophils, and eosinophils.
3. TNF release by NK cells, monocytes and macrophages.
4. Perforin release by NK cells and eosinophils.
5. Granzyme release by NK cells.

Question 26

What does ADCC look like? What cell surface receptors are involved?

Answer 26

ADCC is a category of Ab effector function. It leads to NK-induced apoptosis. The Fc receptor FcgammaR and IgG allows the NK cell to bind antigen on the surface of a ‘bad cell’ (ex: tumor cell, infected cell)