Dendritic cells
Emmanuel STEPHEN-VICTOR (PhD)
INSERM U 1138 - Centre de Recherche des Cordeliers, Paris
Pathogens
Bacteria, virus, fungi
The pathogen-derived antigens need to be transported
adptaive immune response needs to be initiated
Innate immune cells
NET
Adaptive immunity
DC
Neutrophils
Mф
Priming adaptive
immune response
Immunity and Anti-immunity – Host Vs Pathogen
Bacteria, virus,
fungi
Pathogens
CD4 TTregs
Th1
Th17
Th2
Priming adaptive
immune response
Dendritic cells -Discovery
1868 Paul Langerhans => Branched skin cells at epidemis resembling neurons
Dendritic cells -Discovery
1868 Paul Langerhans
=> Branched skin cells at epidemis resembling neurons
1973 Ralph Steinman
=> Dendritic cells from the spleen of mice
One of the first views of DCs in mouse spleen
Steinman RM et al. J Exp Med, 1973
Critical for primary immune response
Large array of endocytic receptors (not B cells)
Large array of antigen presenting and co-stimulatory molecules (not MØ)
Ability to migrate to T cell areas for presentation (not other APCs)
Dendritic cells: Major professional antigen presenting cells
Some features of dendritic cells
Adhesion and co-stimulation
ICAM 1,3; B7-1, B7-2; CD58/LFA3
Receptors for pathogens
CD4, DC-SIGN, chemokine
receptors
Secretion
IL-12, IL-10, TNF, chemokines
High levels of Ag presenting
molecules
HLA I, II; CD1
Receptors for Ag uptake
MMR, DC205, Langerin, integrins,
FcγR TLR, CR
Signaling of DC
TNF-R, CD40, TLR
Cytokine R: GM-CSF, IL-4,
IL1, TGF-b, IL-10
Dendritic cells in health and disease
Immunological Reviews
Volume 219, Issue 1, pages 118-142, 10 SEP 2007 DOI: 10.1111/j.1600-065X.2007.00551.x
http://onlinelibrary.wiley.com/doi/10.1111/j.1600-065X.2007.00551.x/full#f1
Ontogeny of dendritic cells
Origin and development of dendritic cells
MP: Myeloid progenitor, MDP: monocyte dendritic cell progenitor, CDP: committed DC progenitor
Liu & Nussenzweig, Immunol Rev 2010
Flt3L: fms-like tyrosine kinase 3 ligand
LC pro
LC
Subsets of dendritic cells
Dendritic cell subsets in health and disease
Immunological Reviews
Volume 219, Issue 1, pages 118-142, 10 SEP 2007 DOI: 10.1111/j.1600-065X.2007.00551.x
http://onlinelibrary.wiley.com/doi/10.1111/j.1600-065X.2007.00551.x/full#f4
Collin et al., Nature Rev Immunol 2011
Human dendritic cell deficiency
Purification / preparation of dendritic cells
DCs – Low population <1% Spleenocytes
<1% blood leukocytes
<1% tissues (skin, lungs)
Obtaining DC
1992 J Banchereau et C Caux (Lyon)
1992 R Steinman (NY)
K Inaba (Kyoto)
1994 A Lanzavecchia (Bellinzona, Suisse)
Bone marrow cells DC human
Splenic cells DC murine
Blood monocytes DC or macrophages
Pathogens
Dendritic cell CD4+ Th cell
B cell Antibodies
Humoral/antibody response
Bacteria, fungi
Host immune response to pathogens
invariant chain (Ii)
MHC class II compartment (MIIC)
class II-associated Ii peptide (CLIP) Neefjes et al 2011
MHC Class II presentation by Dendritic cells
Pathogens
Dendritic cell
CD8+ Tc cell
B cell Antibodies
Humoral/antibody response
CTL response
Intracellular bacteria , virus
CD4+ Th cell
Host immune response to pathogens
TAP:transporter associated with antigen presentation,
ER: endoplasmic reticulumNeefjes et al 2011
MHC Class I presentation by Dendritic cells
PRR
APC
PRR
PRR
Antigen
Antigen
CD4+T cell immune response to pathogens
AIR
SOIL
Up to 109/m3
Airborne fungal
spores
Innate
immune cell
Elimination by
phagocytosisInvasive
aspergilosis
Healthy Immunocompromised
A. fumigatus
Rodlet
layer
Immune response –
Inert
Immune response –
Activated
Enlarged or
Germinated conidia
Innate cell
Rodlet layer
degraded
Dormant conidia
A. fumigatus sensing by innate cells – Role of Rodlet
Aimanianda*, Bayry* et al Nature. 2009
A. fumigatus – structural organization of the conidia
Polysaccharides
Proteins
Lipids
Rodlet layer
Electron micrograph of fixed conidia
Courtesy: Beauvais A
Chitin
Exposure of polysaccharides during
germination
PAMPS
PRRs on dendritic cells
Chitin
Exposure of polysaccharides during
germination
PRRS and DCs – 2011 Noble prize
Bruce
BeutlerJules
Hoffmann
Ralph Seinman
Toll receptors Dendritic cells
Immature DC
Mature DC
Changes during dendritic cell maturation
Immature DC
High intracellular HLA II
High endocytosis,
Low CD54, 58, 80, 86
Low CD40,
Low CD83
Low granule antigens
High surface HLA II
Low endocytosis
High CD54, 58, 80, 86
High CD40,
High CD83
Production of inflammatory
cytokines
Mature DC
Pathogens
Pro-inflammatory cytokines
T cells
Changes during dendritic cell maturation
Methodology
CD14
Monocytes
MoDC
PBMC
GM-CSF & IL-4
5 days
DC responses
48 hrs
A. fumigatus
(morphotypes)
Dormant, Enlarged,
Germinated conidia
Enlarged and germinated conidia but not dormant conidia
induce maturation and activation of human dendritic cells
6000
0
4000
2000
* *
Ctr
DO
C
EC
GC
1800
0
1200
600
* *
CD
80
MF
I
Ctr
DO
C
EC
GC
1600
0
2400
800
3200 * *
HL
A-D
R M
FI
4000
0
6000
2000
8000
* *
CD
40 M
FI
CD
86
MF
I
IL-8
(p
g/m
l)
0
**
1000
15000
30000
45000
60000
IL-1b
(pg
/ml)
0
*
*
10150
300
450
600
IL-6
(p
g/m
l)
0
*
*
303000
6000
9000
12000 60
0
40
20
IL-1
2 p
70
(p
g/m
l)
*
*
Migration of dendritic cells to draining lymph nodes
CD54 (ICAM-1) CD11a/CD18 (LFA-1)
CD40 CD40L
MHC II + Ag Peptide TCR CD3 complex
CD80/86 CD28/CTLA-4
CD4
CD58 (LFA-3) CD2
APC CD4 T cellSIGNAL 1
SIGNAL 2
SIGNAL 2
SIGNAL 3
SIGNAL 3Cytokines
Cytokines
Cytokine receptor
Cytokine receptor
Th1
Th2
Th17
iTreg
T-bet
STAT 4
STAT 1
GATA3
STAT 6
RORC2
STAT 3
FoxP3
STAT 5
IFN-IL-2
IL-4
IL-5
IL-13
IL-17
IL-17F
IL-21
IL-22
TGF-b
IL-10
Transcription
Factors Cytokines
Effector
cells
suppresors
CD4+T cell immune response to pathogens
Dendritic cells in health and disease
Immunological Reviews
Volume 219, Issue 1, pages 118-142, 10 SEP 2007 DOI: 10.1111/j.1600-065X.2007.00551.x
http://onlinelibrary.wiley.com/doi/10.1111/j.1600-065X.2007.00551.x/full#f1
Regulation of Dendritic cell functions by
immune compartment
DC
Effector CD4+ T cells
B cells / Immunoglobulins
Cytokines
Regulatory T cells
Regulation of Dendritic cell functions by
immune compartment
DC
Effector CD4+ T cells
B cells / Immunoglobulins
Cytokines
Regulatory T cells
Regulation of Dendritic cell functions by
immune compartment
MFI
CD80 CD86
800
600
400
200
0
1200
900
600
300
0
RO
RA
No
ne
RO
RA
No
ne
Co-stimulatory molecules
T cells induce activation of dendritic cells
pg
/ml
0
30
20
0
TNF-a IL-8IL-6
900
600
300
10
ng
/ml
RO
RA
RO
RA
No
ne
No
ne
RO
RA
No
ne
pg
/ml
0
900
600
300
Inflammatory cytokines
T cells induce activation of dendritic cells
HLA-DR CD86 CD80 CD40CD11c
Stimulation of dendritic cells by CD40L (CD154) on T
cells leads to maturation
Isotype CtrlAnti-CD154/CD40L
DC
Effector CD4+ T cells
B cells / Immunoglobulins
Cytokines
Regulation of Dendritic cell functions by
immune compartment
Regulatory T cells
•Represent ~2-4% of total PBMCs
•Express CD25
•FoxP3 (transcription factor forkhead)
•Intracellular CTLA-4 Expression
•Immunosuppressors
CD4+CD25+ regulatory T cells in human circulation
Dendritic cells and regulatory T cells
CD4+CD25+ regulatory T cells: Mechanisms of suppression
Vignali et al., 2008; Nat Rev Immunol
CD4+CD25+ regulatory T cells: Mechanisms of suppression
Vignali et al., 2008; Nat Rev Immunol
**
MFI
**
**800
600
400
200
0
1200
900
600
300
0
Tre
g
RO
RA
No
ne
Tre
g
RO
RA
No
ne
CD80 CD86
CD4+CD25+ regulatory T cells inhibit TLR-mediated
activation of human dendritic cells
Co-stimulatory molecules
J Immunol 2004
J Immunol 2007
Arthritis Rheum 2007
Nature Rev Immunol 2008
Nature 2008
Arthritis Rheum 2009
Nature Rev Rheumatol 2009
Am J Pathol 2009
Vaccine 2011
pg
/ml
0
40
30
20
0
TNF-a IL-8IL-6
*
**
900
600
300
10
**
ng
/ml
Tre
g
RO
RA
Tre
g
RO
RA
No
ne
No
ne
Tre
g
RO
RA
No
ne
pg
/ml
0
900
600
300
Inflammatory cytokines
J Immunol 2004
J Immunol 2007
Arthritis Rheum 2007
Nature Rev Immunol 2008
Nature 2008
Arthritis Rheum 2009
Nature Rev Rheumatol 2009
Am J Pathol 2009
Vaccine 2011
CD4+CD25+ regulatory T cells inhibit TLR-mediated
activation of human dendritic cells
CTLA-4 is the major molecule implicated in the CD4+CD25+
regulatory T cell-mediated inhibition of dendritic cells
CTLA-4 (1µg) - + CTLA-4 (1µg) - +
CTLA-4 (1µg) - + CTLA-4 (1µg) - +
CTLA-4 (1µg) - + CTLA-4 (1µg) - +
CD83
0
200
400
600
MF
I
CD80
0
600
1200
1800
2400
MF
I
CD86
0
400
800
1200
1600
MF
I
HLA-DR
0
7000
14000
21000
28000
MF
I
CD40
0
7000
14000
21000
28000
MF
I
CD1a
0
6000
12000
18000
24000
MF
I
CFSE
Poly IC DC-Treg
100 101 102 103 104
Poly IC DC-RO
100 101 102 103 104
Bayry et al., J Immunol. 2007
CD4+CD25+ regulatory T cells inhibit T cell
proliferation mediated by TLR-activated human
dendritic cells
IFN- IL-2
ng
/ml
RO
Treg
Poly IC DC
0
65
130
0
150
300
TNF-ap
g/m
l
0
125
250
0
600
1200
IL-5
0
100
200
IL-4
0
40
80
IL-10
pg
/ml
Bayry et al., J Immunol. 2007
CD4+CD25+ regulatory T cells inhibit DC-mediated IFN-
responses
DC
Effector CD4+ T cells
B cells / Immunoglobulins
Cytokines
Regulation of Dendritic cell functions by
immune compartment
Regulatory T cells
Regulation of Dendritic cell functions by immune
compartment
** IL-8 IL-6 A
CCL22 CCL17 CCL19
GM-CSF + IL-4 GM-CSF + IL-4 + IFNb1a
B
* *
*
TNF-a IL-1b
*
CCL3 CCL4
Type I interferon:
Promotion of activation of DC
Maddur et al., Arthrits Rheum 2010
Regulation of Dendritic cell functions by immune
compartment
TGF-β: inhibit the
DC maturation
TGF-β (10ng)- + TGF-β (10ng)- +
TGF-β (10ng) - + TGF-β (10ng) - +
TGF-β (10ng) - + TGF-β (10ng) - +
CD83
0
250
500
750
MF
I
CD80
0
2000
4000
6000
MF
I
HLA-DR
0
2000
4000
6000
MF
I
CD86
0
800
1600
2400
3200
MF
I
CD40
0
10000
20000
30000
40000
MF
I
CD1a
0
5000
10000
15000
20000
25000
MF
I
DC
Effector CD4+ T cells
B cells / Immunoglobulins
Cytokines
Regulation of Dendritic cell functions by
immune compartment
Regulatory T cells
©2012 by American Society of Hematology
Morva A et al. Blood 2012;119:106-114
Maddur M S et al. Blood 2012;119:3863-3864
B cell stimulation:
CD40L+CpG-ODN 2006
Regulatory B cells slow down the complete maturation of iDCs
into mDCs.
HLA-DR CD80 CD86
Activated B cells induce dendritic cell maturation and induce
Th2 polarization by inducing OX-40 ligand
Resting B cells
Activation
Maturation of DCs
OX40L
Immature DCs
Mature DCs
Activated
B cells
Maddur et al., Nature Commun 2014
Activated B cells induce dendritic cell maturation and induce Th2 polarization by
inducing OX-40 ligand
Resting B cells
Activation
Maturation of DCs
Differentiation
OX40L
Th2 cells
B cell help
GATA3IL-4
IL-13 IL-5
Naïve
CD4+ T cells
Immature DCs
Mature DCs
Activated
B cells
Maddur et al., Nature Commun 2014
Regulation of Dendritic cell functions by
immune compartment
IgG immunoglobulins at
therapeutic high doses (IVIg)
DC
Effector CD4+ T cells
B cells / Immunoglobulins
Cytokines
Regulatory T cells
• Therapeutic preparation of pooled normal polyspecific human
IgG obtained from large numbers of healthy donors
Contains
Antibodies to non-self (foreign antigens)
Antibodies to self antigens
Antibodies to antibodies (anti-ids)
Regulation of dendritic cells by circulating
antibodies: Intravenous Immunoglobulin (IVIg)
Intravenous Immunoglobulin (IVIg)
Polyclonal IgG – plasma pool – thousands of healthy donors
Intravenous Immunoglobulin (IVIg)
Autoimmune and
inflammatory diseases
Immune thrombocytopenic purpura (ITP)
Guillain-Barré syndrome (GBS)
Kawasaki disease (KD)
CIDP
Dermatomyositis (DM)
Polymyositis (PM)
Psoriasis
Systemic lupus erythematosus (SLE)
ANCA-associated vasculiits
Myasthenia gravis (MG)
Transplant rejection
Primary Immunodeficiencies
CVID,
XLA
Secondary Immunodeficiencies
BM transplantation,
Multiple myeloma
Wide - divergent therapeutic
applications
Regulation of maturation and function of dendritic
cells by IVIg at therapeutic concentration
60(80)
15(21) CD1a
43
(39)1
(27)CD83
IVIg
IVIg
Immature Mature
70
140
*
0
pg/m
L
*40
0
20
Ctl
IVIgHSA
IL-12
IL-10
Blood 2003, Arthritis Rheum 2003, PNAS 2009
Contribution of DC to autoimmunity
Genetic
Epigenetic
Environmental
Cytokines/chemokines
Ag presentation
Co-stimulatory molecules
Migratory properties
Effector /pathogenic T cells
Treg number and function
Dendritic cells prevent autoimmunity by
expansion of regulatory T cells
Treg
Transforming growth factor-β (TGF-β)
Cyclooxygenase-2 (COX-2)
Indoleamine 2,3-dioxygenase (IDO)
Inducible costimulatory ligand (ICOS-L)
CD252 (OX40L)
Programmed death ligands
Regulatory T cell expansion by dendritic cells
Pallotta et al., Nat Immunol. 2011
Trinath et al., Blood 2013
Gopisetty et al., Immunology 2013
Fantini et al., Immunology 2004
Sakaguchi et al., Cell, 2008
Dejaco et al., Immunology, 2007
Trinath et al. J Infect Dis 2012
Regulatory T cell expansion by dendritic cells: Role of Programmed death ligands
0
25
50
75
100
PD-L2 PD-L1 PD-1
Ctr Mtb Ctr Mtb Ctr Mtb
*
% p
osit
ive d
en
dri
tic c
ells
Trinath et al. J Infect Dis 2012
Regulatory T cell expansion by dendritic cells: Role of Programmed death ligands
0
5
10
15
Ctr Mtb Mtb+anti-PD-L1
* *
% C
D4+
CD
25+
Fo
xP
3+
T c
ells
Regulatory T cell expansion by dendritic cells: Role of PGE2
Trinath et al Blood 2013
Ctr
IVIg
10
mg
IVIg
15
mg
HS
A
COX-2
! -actin
Ctr
IVIg
10 m
g
IVIg
15 m
g
HS
A
0
1000
2000
3000
4000
5000* *
PG
E2 (
pg
/ml)
Ctr IVIg
10 mg
IVIg
15 mg
HSA Ctr IVIg
10 mg
IVIg
15 mg
HSA
COX-2
β-actin
Regulatory T cell expansion by dendritic cells: Role of PGE2
Ctr
IVIg
NS
-39
8 +
IV
Ig
0
2
4
6
8**
% C
D4
+F
oxP
3+ T
cells
Ctr IVIg IVIg +
COX-2 inhibition
Ctr
IVIg
NS
-39
8 +
IV
Ig
0
1000
2000
3000
4000
*
**
PG
E2 (
pg
/ml)
Ctr IVIg IVIg +
COX-2 inhibition
Trinath et al Blood 2013
Comparison of different cell populations for the induction of CD4+CD25+ FOXP3+ Treg's.
Banerjee D K et al. Blood 2006;108:2655-2661
Differentiation of iTregs by dendritic cells
Immune tolerance
Prevention of autoimmunity
Induction of Tregs
Maintenance/homeostasis of Tregs
T and B cell response
to pathogens
Clearance of pathogens
Anti-tumor immunity
Take home message
Immature/tolerogenic DC Activated DC
Immune tolerance
Prevention of autoimmunity
Induction of Tregs
Maintenance/homeostasis of Tregs
T and B cell response
to pathogens
Clearance of pathogens
Anti-tumor immunity
Take home message
Immature/tolerogenic DC Activated DC
Autoimmunity
Inflammatory diseases
Genetic
Epigenetic
Environmental
Thank you
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