Transcript DC intro - Univerzita Karlova v Praze

Disturbances in the homeostasis of Th17 lymphocytes
in patients with hyper IgE syndrome (HIES) and
chronic granulomatous disease (CGD)
Horvath R.1, Lastovicka J.1, Polouckova A.1, Sedlacek P.2, Bartunkova J.1, Sediva A.1, Spisek R.1
1Department
of Immunology, Charles University, 2nd Medical School and Faculty Hospital Motol,
Prague, Czech Republic
2Department
of Pediatric Hematology and Oncology, BMT unit, Charles University, 2nd Medical
School and Faculty Hospital Motol, Prague, Czech Republic
Th-cell phenotypes
Th17 cell lineage development
APC
IL23-APC
Pre-Th17
Naive CD4
IL-6
TGF-β
RORγT
STAT-3
Th17
IL-17
IL-22
IL-21
Th 17 cell functions

Production of IL-17 cytokines family (IL-17, IL-21, IL-22) which leads to
the chemoattraction of neutrophils

Accumulating data suggest that Th17 are highly pro - inflammatory and
that Th17 cells with specificity for self-antigens lead to severe
autoimmunity- (psoriasis, Crohn´s disease, multiple sclerosis)
Physiological role of Th17 cells in humans



Initially, from studies in mice, Th17 cells were thought to play an important
role in host defense against extracellular pathogens, which are not
efficiently cleared by Th1-type and Th2- type immunity
However, identity of pathogens cleared by Th17 was unknown
Direct evidence for understanding physiological target of Th17 cells came
from studies of patients with mutations in STAT-3, a critical transcription
factor for the differentiation of Th17 cells
Defective Th17 cells in Hyper IgE syndrome

Primary immunodeficiency
caused by mutations in STAT-3
transcription factor, NEJM 2007

Dermatitis, boils, cyst-forming
pneumonias, retained primary
dentition, bone abnormalities and
elevated serum IgE levels
specific stim. with candida


Abnormal and devastating
susceptibility to a narrow spectrum
of infections, most commonly
Staphylococcus aureus and
Candida albicans
nonspecific stimulation
Abrogated numbers of Th17 cells,
Nature 2008, JEM 2008
specific stim. with stp.aureus
Defective Th17 cells in Hyper IgE syndrome

Studies in Hyper IgE point to a critical role of Th17 cells in the response
against candidal and staphylococcal infections

However, there are other diseases with similar spectrum of dominant
pathogens where the characteristics of Th17 have not been tested

We thus decided to test Th17 cells compartment in chronic granulomatous
disease
Clinical mimicry of HIES with other primary
immunodeficiencies – Chronic granulomatous disease (CGD)

Primary immunodeficiency

Mutations in the NADPH oxidase
system

Profound defect of respiratory burst in
myeloid cells

Recurrent infections, organ
granulomas

Dominant susceptibility to
staphylococcal and candidal
infections
Aim of the study

Analyze and compare the characteristics of Th17 compartment in
HIES and CGD patients
Patients and methods



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4 patients from 3 families with HIES
7 patients with CGD (2 patients underwent allo-BMT)
Mutations in STAT-3 and NADPH oxidase - genetics
FACS, ELISA
Results – Th17 numbers in CGD and HIES
A
10
104
3
103
2
10
2
10
0.51
0
2
0 10
IFN gamma - PE
104
10
10
5
10
3
10
4
10
5
4
0.13
2
0 10
10
3
10
4
2
0 10
10
3.5
5
10
10
4
104
103
103
103
102
0.49
0
2
0 10
5
10
3
10
4
10
5
6.51
2
5
3
10
4
5
10
6.97
0 10
5
10
104
103
103
103
102
102
102
0.76
2
3
10
4
10
5
10
Controls
0.36
0
2
0 10
3
10
4
10
HIES
5
10
3,5
p<0,05
3
p<0,05
2,5
3
10
4
10
1,5
1
0,5
0
Controls HIES
1.55
2
0 10
3
10
4
10
CGD
5
10
1.91
0
p<0,05
2
2.87
2
10
104
0 10
5
10
1.25
0
104
0
4
10
2
0 10
10
3
10
10
0.078
0
10
1.98
0
5
10
10
102
2.32
2
0
5
10
5.91
10
104
3
10
5
1.67
5
10
CGD
IL-17 A647
• absent Th17 cells in HIES
• high frequencies of Th17 in CGD
% of IFN gamma+ CD4+ cells
5
4.12
% of IL-17+ CD4+ cells
B
5
10
14
p<0,05
12
p=0,39
p=0,05
10
8
6
4
2
0
Controls
HIES CGD
Results – cytokine production
Th17 effector cytokines
A
p<0,05
1000
p<0,05
2500
2000
IL-23
p=0,06
1200
p<0,05
p=0,29
1000
p<0,05
p=0,14
p<0,05
800
B
IL-21
IL-17
1200
Polarization cytokine
800
600
pg/ml
pg/ml
pg/ml
1500
1000
400
p<0,05
600
400
500
200
200
0
0
0
Controls
HIES
CGD
Controls
HIES
CGD
Controls
HIES
CGD
•low levels of IL-17 and IL-21 in HIES
•extremely elevated levels of IL-23 in HIES
•high levels of IL-17 and IL-21 in CGD
•elevated levels of IL-23 in CGD
Correction of defect in Th17 cells in two CGD patients after
successful BMT
A
p#1
1.83
104
102
105
p=0,52
3
2,5
2
1,5
1
,5
0
0.12
0
10
2
3
10
10
4
10
0
pre-BMT
5
8.64
104
103
102
0
0.84
0
102
post-BMT
IFN-g
% of CD4 pos. T cells
IFN gamma - PE
103
p#2
Th17
B
% of CD4 pos. T cells
105
10 3
104
10 5
IL-17 A647
14
p=0,24
12
Controls
10
HIES
8
6
CGD
4
CGD after BMT
2
0
pre-BMT
post-BMT
• normalized numbers of Th17 cells after BMT
Results – cytokine production after allo-BMT
A
B
Th17 effector cytokines
Polarization cytokine
p=0,24
1400
IL-17
1200
1000
IL-23
800
1000
pg/ml
600
400
800
200
0
600
post-BMT
pg/ml
pre-BMT
IL-21
2000
400
p=0,12
200
1750
p=0,33
0
1500
pre-BMT
1250
post-BMT
pg/ml
1000
750
500
Controls
250
HIES
0
pre-BMT
post-BMT
CGD
CGD after BMT
•decreased production of IL17,21 and 23 after BMT
Possible theoretical explanations
Eradication
Healthy
Candida
S.aureus
IL-23
APC
Lympho
IL-17,22,21 etc
Neutro
Th17
Mono
STAT3 mutation
HIES
Candida
S.aureus
IL-23
APC
IL-17,22,21 etc
In-efficient immune cells attraction
Th17
Pathogen persistence
In-efficient eradication
CGD
Candida
S.aureus
IL-23
APC
IL-17,22,21 etc
Lympho
Th17
NADPH mutation
Neutro
Mono
Pathogen persistence
Conclusions

We identified disturbances in the homeostasis of Th17 lymphocytes
in HIES and CGD patients

Absent Th17 cells in STAT-3 deficient HIES patients

Significantly higher frequencies of Th17 cells in CGD

Increase of Th17 cells in CGD is likely to be secondary as a result of
defect in neutrophils

BMT leads to the normalization of elevated Th17 cell numbers and
coresponding IL-17 production

Positive pro-inflammatory loop caused by Th17 cells contributes to
the formation of granulomas

These findings confirm the critical role of Th17 lymphocytes in the
elimination of candidal and staphylococcal infections
Acknowledgements
Dpt. of Immunology, Charles University,
Prague, Czech Republic
Lastovicka Jan
Polouckova Andrea
Rozkova Daniela
Kayserova Jana
Budinsky Vit
Sochorova Klara
Kytka
Minarik Ivos
Fucikova Jitka
Jaresova Irena
Sediva Anna
Bartunkova Jirina
Spisek Radek
Dpt. of Pediatric Hematology and Oncology,
BMT Unit, Charles University, Prague, Czech
Republic
Formankova Renata
Keslova Petra
Stary Jan
Sedlacek Petr