DCs

Question 1

E - Why are DCs a bridge between innate and adaptive immunity?

Answer 1

Uptake pathogens (upon recognition via receptors of the innate immune system) and present them to the cells of the adaptive immune system in lymphatic organs.

Question 2

E - Name an example where the cross-presentation on DCs is required to generate CTL immunity.

Answer 2

Viruses that inhibit MHCI presentation - prevents CTL stimulation by direct MHCI pathway.
But: X-presentation of antigens that are derived from infected cells by uninfected DCs is likely to result in CTL immunity despite inhibitory mechanisms.

Question 3

E - Name 2 receptors/molecules which are expressed on mature (and immature) DCs.

Answer 3

Immature: PRR, MHCI, ICAM-1,

Mature in lymphoid tissue: MHCI/II, ICAM-1, CCR7, B7.1/2

Question 4

E - Which receptor on DC is necessary for the migration to the lymph nodes?
Name the ligand (interaction + origin) and the events which lead to the expression of the “migration” receptor.

Answer 4

Receptor: CCR7
Ligand: CCL19, CCL21

CCR7+ DC respond to a chemotactic gradient of CCL19 and/or CCL21 that originates from the lymphatic vessel

Receptor expression on DCs is induced by:

• DC maturation signals
• uptake of apoptotic material without maturation

Question 5

E - One major role of DCs is to promote T cell immunity.

Explain how a mature DC induces immunity?

Answer 5

• tissue inflammation induced maturation of DCs
• mature DCs migrate to draining lymph nodes
• express peptide:MHC complexes and costimulatory molecules to prime T cells, activate B cells and initiate adaptive immune response
  (also activate Tregs to control the immune response)

Question 6

E - Explain the role of DCs in the peripheral T cell tolerance.

Answer 6

Immature DCs induce tolerance:

• in the absence of inflammation, DCs remain immature
• still present antigens in lymph nodes, but without costimulation
• -> leads to either clonal T cell deletion or iTregs generation

Question 7

E - What is meant by LICENSING signal of DCs?

Answer 7

licensing = activation of a DC (e.g. TLR signaling) so that it is able to present antigen to naive T cells and activate them

signal through CD40:CD40L when MHCII:TCR on CD4 T cell recognise
–> this signal is costimulatory signal which leads to naive CTL priming

Licensing signal can be mediated by pathogen too (?).

Question 8

E - What does the term ‘migratory’ DC mean?

Answer 8

That DCs are in peripheral non-lymphoid tissues.

Question 9

E - Shortly explain the types of DC effector functions and name 3 specific roles.

Answer 9

Mature DC inducing:
CTL priming
deletional tolerance
de novo regulatory T cells
Th1 cell response

Types: pDCs and cDCs

Question 10

E - Describe DC checkpoint immunology - from the point of the progenitor cell up to a functional matured DC; name 3 features of DC maturation.

Answer 10

immature DC -> uptake of Ag in periphery after recognition with PRRs -> mature DC that goes into lymph node

maturation:

• not phagocytosing anymore
• upregulated MHCs
• upregulation of homing receptors (CCR7 for lymph nodes)
• upregulation of costimulatory molecules and cytokine secretion -> able to activate T cells

Question 11

E - What are checkpoint inhibitors, what is their role in melanoma immunotherapy?

Answer 11

immune checkpoints normally mediate immune tolerance to stop collateral tissue damage = negatively regulate immune response to stop T cell proliferation

e. g. CTLA-4:
- binds B7 more avidly than CD28 and delivers inhibitory signals to activated T cells
- distinct binding orientation allows clustering

e. g. PD-1:
- binds B7-family member ligand PD-L1

therapy: melanoma treatment to increase antitumor response (side effect = autoimmunity)

Question 12

E - Explain 2 examples for the role of DCs in autoimmunity.

Answer 12

Depending on the inflammatory contex and the expression of cell intrinsic regulators: DC presentation of self-antigens might promote or inhibit autoimmune response

a) presentation to T cells and interaction via PD1 –> anergy of self-reactive T cells
b) if context of pro-inflammatory mediators (e.g. IL-6, I-12) –> promote development of self-reactive CD4+ T cells and CTLs

Question 13

Ž - How do you get DCs with FACS and how do you separate pDCs and cDCs?

Answer 13

1) exclude the cells with following markers:
- CD3 (T)
- CD19 (B)
- CD14 (monocytes)
- CD56 (NK)

2) sort DCs on CD11c vs HLA-DR
- take HLA-DR positive cells
- CD11c high = cDCs
- CD11c low: additional CD123 vs HLA-DR plot -> pDCs are CD123 high

Question 14

Ž - How can you generate DCs in cell culture?

Answer 14

from bone marrow by adding GM-CSF -> BMDCs

Question 15

Ž - What is the major difference between immature and mature DCs?

Answer 15

only immature are very good at antigen uptake, mature do not do it anymore

Question 16

Ž - Which types of DCs are known and what is their role in immunity?

Answer 16

pDCs: type I IFNs release
cDCs 1: tumor priming
cDCs 2: major MHCII presenting cells

Question 17

Ž - How do DCs get to prime T cells? (How is the initial contact between these two cells established?)

Answer 17

T cells initially bind APC through LOW affinity LFA-1
recognition of MHC:peptide complex with TCR leads to conformational change to high affinity LFA-1:ICAM-1 binidng = prolonged contact

Question 18

Ž - Which APC can activate naive T cells besides DCs?

Answer 18

none

Question 19

Ž - What will determine if DC will induce tolerance or immunity when presenting the antigen?

Answer 19

it’s maturity:

if there was no inflammation present in the tissue, it will not mature and express costimulatory signals
so when it presents AG in the secondary lymphoid organ in absence of co-stimulation (MHC:p only), it will induce regulatory phenotype of T cells

if there was inflammation, it matured after antigen uptake and will present Ag with the costimulatory signal which will lead to effector T cell differentiation (of different classes, including Tregs, but in this case only to control the immune response)

Question 20

Ž - Which marker is upregulated upon PRR activation on DCs?

Answer 20

CD86 (and MHCII)

Question 21

Ž - Costimulatory signals provided by DCs can be positive or negative. What happens in presence of each? Name some.

Answer 21

positive = facilitate activation of T cells
negative = turn activation off

positive = B7, ICOS, 4-IBB, OX40
negative = CTLA-4, PD-1, BTLA

Question 22

Ž - Why was the first DC vaccine unsuccessful? What was improved?

Answer 22

first: isolated immature DCs from patients, reinjecting them back after maturation = weren’t migratory anymore!
better: isolated DC precursors, exposed them to fusion protein (GM-CSF + prostate cancer Ag), reinject, undergo maturation in vivo