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Universidade Federal do Rio de JaneiroCentro de Ciências da Saúde
Instituto de Biofísica Carlos Chagas Filho
The importance of apoptosis for immune regulation in Chagas’ disease
George A. DosReis
Fabrício Montalvão. IBCCF-UFRJLandi V. Guillermo. IBCCF-UFRJ
Geisy M. de Almeida. IBCCF-UFRJElizabeth M. Silva. IBCCF-UFRJ
Marise P. Nunes. IOC-Fiocruz-RJJuliana de Meis. IOC-Fiocruz-RJ
Christina M. Takiya. Dept. Histologia-UFRJRichard Siegel. NIAID-NIH, USA
Celio Freire-de-Lima. IBCCF-UFRJMarcela F. Lopes. IBCCF-UFRJ
Lopes MF et al. Immunol. Today 21; 489-494, 2000Freire-de-Lima CG et al. Nature 403; 199-203, 2000
Phagocytosis of apoptotic cells (efferocytosis) inactivates macrophages and exacerbates parasite replication through TGF-β, ODC and polyamine synthesis
X
Efferocytosis
T. cruzi infection: Consequences of apoptosis
Efferocytosis
Apoptosis in T. cruzi infection
Modify efferocytosis by:
1. Caspase inhibition2. Anti-FasL treatment3. Antibodies to apoptotic cells
Efferocytosis
Blocking apoptosis in T. cruzi infection
Caspase inhibition
Efferocytosis
Caspase inhibition reduces lymphocyte apopotosis in vivo
Silva, E. M. et al. Caspase inhibition reduces lymphocyte apoptosis and improves host immune responses to Trypanosoma cruzi infection. Eur. J. Immunol. 2007, 37 (3), 738-746.
Treatment with zVAD reduces parasitemia
10 15 20 25 30 35 400
10
20
30
40
50
60
70
80
90
100
zVAD
zFA
control
Days post infection
Par
asit
es x
10
4/m
l
10 15 20 25 30 35 400
10
20
30
40
z-VAD
z-FAcontrol
Days post infectionP
aras
ites
x 1
04/m
l
Silva, E. M. et al. Caspase inhibition reduces lymphocyte apoptosis and improves host immune responses to Trypanosoma cruzi infection. Eur. J. Immunol. 2007, 37 (3), 738-746.
Treatment with zVAD in vivo increases macrophage activation
The general caspase inhibitor zVAD
reduces apoptosis of T lymphocytes in vitro
and in vivo
Treatment with zVAD increases immunity
to T. cruzi infection
Blocking apoptosis in T. cruzi infection
Anti-FasL
Efferocytosis
Increased immunity to T. cruzi after treatment with anti-FasL in vivo
Guillermo, L.V. et al. The Fas death pathway controls coordinated expansions of type 1 CD8 and type 2 CD4 T cells in Trypanosoma cruzi infection. J. Leukoc. Biol. 2007, 81, 942-951.
Anti-FasL in vivo accelerates and increases type 1 cytokine responses
Anti-FasL reduces apoptosis of CD4 and CD8 T cells
Anti-FasL increases type 1 cytokine responses, increases macrophage NO production and reducesparasitemia
Lymphocyte apoptosis is deleterious for T. cruziInfection in vivo
Modifying immune regulation throughopsonization of apoptotic cells
Antibodies
Efferocytosis
Differential reaction of chagasic IgG with apoptotic lymphocytes
1 10 100 1000
0
20
40
60
80
100
% o
f M
ax
Annexin V+
CHACT - 26.1%
CHA - 36.3%
IgG (H+L)
A
10000
OD (Arbitrary Units)
Migration Distance (Arbitrary Units)
CT
400 600 800 1000
50
40
30
20
10
0
B
C
Position 403
Pea
k va
lues
(A
U)
Position 901 Position 1055Position 1022
Cha
0
2
4
6
8
10Parasite number (x104)
CT CHA CT CHA
None IgG IgM
None APO
**
Opsonization of apoptotic cells with chagasic IgG reduces the effect of efferocytosison replication of T. cruzi
IgG purified from T. cruzi infection(90 days) or age matched serum
Control of parasite replication is mediated by increased TNF-α production
Apoptotic cells opsonized with chagasic IgGincrease TNF-α production
02468
101214
NoneAnti-TNF-Rat IgG1
Parasite number (x10 5)
CT CHA None IgG
None APO
** **
0
200
400
600
800
1000
1200
NoneAPOAPO/CT IgGAPO/CHA IgG
TNF- (pg/ml)
**
A
B
Control of parasite replicationrequires engagement of FcγRs
0
3
6
9
12
15
ControlChagasic
Parasite number (x104)
IgG IgG F(ab') 2
None APO
**
0
4
8
12
16
20Control FabCD16/CD32 Fab
Parasite number (x104)
Control Chagasic
None IgG APO
**
A
B
Previous immunization with apoptotic cells reduces parasitemia in vivo
0 5 10 15 200
1020304050607080
Saline/Al(OH)3APO/Al(OH)3
Parasitemia (x104/mL)
**
Dm28c
dpi
Conclusions Chagasic IgG opsonizes apoptotic cells and reduces the effect of efferocytosis on parasite replication
The protective effect is mediated by increased TNF-α production
The protective effect requires FcγRs
Generation of autoantibodies against apoptotic cell antigens could play a protective role against parasite infection