Lecture 16 - CTLs and NK cells

Question 1

What is a CTL?

Answer 1

Cytotoxic T Lymphocyte

A CD8+ lymphocyte that has recognize its antigen and been activated.

Part of adaptive immunity.

Question 2

What is an NK cell?

Answer 2

Natural Killer cell

Contains granules like a CTL but is not antigen specific.

Part of innate immunity.

Question 3

Identify this cell type

Answer 3

Probably an NK cell or a CTL.

Note the large granules in the cytoplasm. Together, NK cells and CTLs are called “large granular lymphocytes.”

Question 4

What is the major function of CTLs and NK cells?

Answer 4

Protect against intracellular pathogens

Question 5

What process is occurring in this image?

Answer 5

Cell with pointer is undergoing apoptosis.

Note the chromatin clumping and membrane blebbing that are characteristic of a cell undergoing programmed death.

Question 6

Learning Objective 1

Describe, in general terms, how apoptosis in induced and what happens in a cell that is induced to undergo apoptosis.

[Wow, that’s just terrible wording]

Answer 6

Extrinsic pathway

• FasL (for Fas ligand) on the killer cell binds to the Fas receptor (the death receptor) on the target cell, inducing apoptosis.
• TNF molecule binds to TNF receptor on target cell, inducing apoptosis.

Intrinsic pathway

• Release of cytochrome C induces apoptosis.

Common to both pathways: activation of caspase 9.

Question 7

What is a granzyme?

Answer 7

Proteolytic enzyme released by a killer cell and taken up by a target cell.

Initiates the intrinsic apoptotic pathway.

Question 8

What is a caspase and how does initiate cell breakdown?

Answer 8

Caspases are “executioner” molecules initiated by both apoptotic pathways.

• Cleave proteins, causing breakdown of the cytoskeleton.
• Activate endonucleases, fragmenting DNA and causing chromatin clumping.

Question 9

Why does apoptosis not cause inflammation?

Answer 9

The process of apoptosis causes formation of cytoplasm blebs, which contain surface molecules for phagocytic cells. The blebs are digested by phagocytic cells with no leakage, so inflammation is not initiated.

Question 10

Learning Objective 2

Explain the differences between apoptosis and necrosis.

Answer 10

Apoptosis

• Organized
• Cell degrades into membrane-bound blebs, which are phagocytosed. No inflammation.
• Individual or clusters of cells

Necrosis

• Violent and uncontrolled
• Membrane degrades, releasing cytoplasmic contents. Inflammation.
• Larger area involved

Question 11

What is the function of perforin?

Answer 11

An activated CTL will release granules of perforin onto its target cell. These polymerize and insert into the target cell membrane, creating a hole (similar to the MAC or defensins).

This allows granzymes to enter the target cell.

Question 12

What is the Fas receptor?

Answer 12

Fas is described as the “death receptor.”

Fas is expressed on somatic cell surfaces. FasL (Fas ligand) is expressed on the surface of CTLs. If a CTL is targeting a somatic cell, it will upregulate the amount of FasL expressed in its membrane. Binding of FasL to Fas triggers apoptosis.

Question 13

What is TNF?

Answer 13

Tumor necrosis factor

Binding of TNF to the TNF receptor on a target cell will initiate apoptosis.

Question 14

Learning objective 3

List 3 mechanisms that CTLs use to kill target cells. For each, include the molecules and functions involved - for binding and killing.

Answer 14

1. Perforin and granzyme pathway - perforin creates a hole in the cell membrane and granzymes enter to initiate the intrinsic apoptosis cascade.
2. Fas/FasL pathway - FasL expressed on the CTL binds with Fas on the target cell, inducing the extrinsic apoptosis cascade.
3. TNF produced byt the CTL binds with TNF receptor on the target cell, initiating the extrinsic apoptosis cascade.

Question 15

CTLs bind to which molecule on the target cell?

Answer 15

TCR (t cell receptor) binds to target peptide presented on MHC I.

CD8 molecule binds to MHC I itself.

Question 16

Describe the activation process of a CTL

Answer 16

CTL leaves the blood stream in response to upregulated inflammatory markers.

CTL binds to MHC I on target cell and recognizes its target peptide.

CTL redistributes its cytoskeleton to polarize killing granules toward the target cell.

CTL releases its granules onto target cell (the immunological synapse), initiating apoptosis.

Question 17

What is the difference in surface molecules between a CTL and NK cell?

Answer 17

Recall that an NK cell is part of innate immunity and cannot recognize specific peptides. So, NK cells do not express CD3, CD4, or CD8.

(CTLs express CD3 and CD8)

Question 18

True or False

NK cells have similar granule content to CTLs

Answer 18

TRUE

Thought they do not recognize specific antigens, NK cells have similar killing mechanisms to CTLs.

Question 19

What is MICA receptor?

Answer 19

MICA receptor is and MHC I - like molecule that is expressed by stressed cells.

If the NK cell binds to the MICA receptor on a target cell, it will induce apoptosis.

Question 20

What is the Fc receptor?

Answer 20

Recall that Fc is a component of antibody. Like many other leukocytes, NK cells have Fc receptors to recognize when a molecule is opsonized by antibody.

If a target cell is opsonized, an NK cell will induce apoptosis in the target cell.

Question 21

Which cytokines activate NK cells?

Answer 21

IFNalpha and IL12 - produced by cells infected with virus.

Th1 cytokines (IL2, IFNgamma, TNFalpha)

These cytokines make NK cells better at killing.

Question 22

What is antibody-dependent cell-mediated cytotoxicity (ADCC)?

Answer 22

If a cell is coated by antibody, this will be detected by the Fc receptor of the NK cell. The NK cell will then initiate apoptosis in the opsonized cell.

Question 23

Describe the MHC I inhibitory receptor mechanism in NK cells.

Answer 23

When an NK cell binds to MHC I on the target cell, it is inhibited from killing the target cell.

If the target cell is not expressing MHC I, it will be killed by the NK cell.

Question 24

Suppose a tumor cell is not expressing MHC I normally. What will the result be?

Answer 24

If an NK cell fails to bind to MHC I on a target cell, it will initiate apoptosis.

Question 25

Some viruses prevent antigen presentation on MHC molecules, so viral particles cannot be recognized by CTLs. Can the infected cell still be killed by the immune system?

Answer 25

YES

NK cells act as a “backup” for CTLs

If the virally-infected cell is not expressing MHC I in the normal fashion, it will be killed by the NK cell (even though the virus was not recognized by the CTL).

Question 26

Learning Objective 4

Describe how an NK cell recognizes a cell to kill and describe the killing mechanisms of the NK cell.

Answer 26

NK cells do not recognize specific peptides.

1. When an NK cell binds to an MHC I molecule on the target cell, it is inhibited from killing. If the cell is not expressing MHC I, it will be killed.
2. If the target cell is expressing the MICA receptor (a stress protein), it will be killed.
3. If a cell is coated in antibody, it will be killed.

Killing mechanism: NK cells have granules containing perforins and granzymes, just like CTLs. They also possess the Fas ligand. NK cells can be made more effective at killing by cytokine activation.

Question 27

Learning Objective 5

Explain how the CTL and NK cell are complimentary, including timing, targets, and the benefit to the host to have both cell types.

Answer 27

Both protect against intracellular pathogens.

CTLs recognize antigenic peptides presented on MHC I. They are important in acting against specific infections.

NK cells are part of innate defense. They respond to antibodies, stress proteins, and cells not correctly expressing MHC I.

In the case of a viral infection, a CTL could recognize specific viral proteins to respond (NK cell could not recognzie specific proteins). In the case of a tumor that stopped expressing MHC I, NK cells could induce apoptosis (CTL could not).

Question 28

Learning Objective 6

Compare and contrast the various killing mechanisms used by the immune system.

Answer 28

Neutrophils

• Release of toxic granules, oxidative burst, netosis

Macrophages

• Phagocytosis, nitric oxide

NKs and CTLs

• Induction of apoptosis

Others

• Defensins, complement, antibody inactivation

Question 29

Learning Objective 7

Give an example of an infection that CTLs would be important in controlling.

Answer 29

CTLs are important for controlling intracellular infections. Peptides in the cytoplams are processed and presented on MHC I through the endogenous pathway.

Examples: viral infections, intracellular bacteria

Question 30

Learning Objective 8

List a few important properties of a vaccine that would stimulate CTLs.

Answer 30

Remember that CTLs respond to peptides presented through the endogenous pathway. This means that the pathogen must be located intracellularly.

Vaccine properties: modified live virus that replicates intracellularly.

Can also use an ISCOM (immune-stimulatory complex) - antigen encased in lipid can enter into the cytoplasm.

Question 31

Which antibodies are typically found in circulation?

Answer 31

IgM and IgG

Question 32

Which antibodies are typically found at mucosal surfaces?

Answer 32

IgA and IgE

Question 33

Which cell type is IgE most closely associated with?

Answer 33

Mast cell

Very little IgE is usually found in circulation because most IgE is bound to mast cells on mucosal surfaces.

Question 34

Name the major components of cell-mediated immunity (3 components)

Answer 34

Cytokine secretion

Cytotoxic T lymphocytes

Gamma Delta T cells

Question 35

If you want to stimulate antigen presentation through both the endogenous and exogenous pathways with a vaccine, what kind of vaccine should you use?

Answer 35

Modified live vaccine

Viral particles will be phagocytosed by APCs and presented on MHC II through the exogenous pathway.

Viral particles replicating intracellularly will be processed through the endogenous pathway and presented on MHC I.

Question 36

Learning Objective 9

Use information given to you about pathogenesis of an infectious disease or about a vaccine and predict the protective immune mechanisms.

Answer 36

Question 37

Learning Objective 10

Explain the difference between measuring humoral immunity and cell-mediated immunity

Answer 37

Humoral immunity - measure antibody levels in the serum

Cell-mediated immunity

• In vivo: inject antigen into animal’s skin and observe the cellular response (think TB test)
• In vitro: collect lymphocytes from animal and add antigen to evaluate memory T cell action. The cytokines produced are indicative of what kind of infection is present.

Question 38

You collect lymphocytes from an animal and add antigen. The main cytokine found in the supernatant is IFNgamma. What T helper response is likely present?

Answer 38

Th1

IFNgamma is a Th1 cytokine

Likely indicates an intracellular infection

Question 39

You collect lymphocytes from an animal and add antigen. The main cytokines produced in the supernatant are IL4 and IL10. Which T helper response is active in this infection?

Answer 39

Th2

IL4 and IL10 are Th2 cytokines.

Likely indicates a parasitic infection.