Limits...
A human type 5 adenovirus-based Trypanosoma cruzi therapeutic vaccine re-programs immune response and reverses chronic cardiomyopathy.

Pereira IR, Vilar-Pereira G, Marques V, da Silva AA, Caetano B, Moreira OC, Machado AV, Bruna-Romero O, Rodrigues MM, Gazzinelli RT, Lannes-Vieira J - PLoS Pathog. (2015)

Bottom Line: Therapeutic vaccination increased survival and reduced electrical abnormalities after the prime (analysis at 160 dpi) and the boost (analysis at 180 and 230 dpi).Post-therapy mice exhibited less heart injury and electrical abnormalities compared with pre-therapy mice. rAdVax therapeutic vaccination preserved specific IFNγ-mediated immunity but reduced the response to polyclonal stimuli (anti-CD3 plus anti-CD28), CD107a+ CD8+ T-cell frequency and plasma nitric oxide (NO) levels.Moreover, therapeutic rAdVax reshaped immunity in the heart tissue as reduced the number of perforin+ cells, preserved the number of IFNγ+ cells, increased the expression of IFNγ mRNA but reduced inducible NO synthase mRNA.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Biologia das Interações, Instituto Oswaldo Cruz/Fiocruz, Rio de Janeiro, Rio de Janeiro, Brazil.

ABSTRACT
Chagas disease (CD), caused by the protozoan Trypanosoma cruzi, is a prototypical neglected tropical disease. Specific immunity promotes acute phase survival. Nevertheless, one-third of CD patients develop chronic chagasic cardiomyopathy (CCC) associated with parasite persistence and immunological unbalance. Currently, the therapeutic management of patients only mitigates CCC symptoms. Therefore, a vaccine arises as an alternative to stimulate protective immunity and thereby prevent, delay progression and even reverse CCC. We examined this hypothesis by vaccinating mice with replication-defective human Type 5 recombinant adenoviruses (rAd) carrying sequences of amastigote surface protein-2 (rAdASP2) and trans-sialidase (rAdTS) T. cruzi antigens. For prophylactic vaccination, naïve C57BL/6 mice were immunized with rAdASP2+rAdTS (rAdVax) using a homologous prime/boost protocol before challenge with the Colombian strain. For therapeutic vaccination, rAdVax administration was initiated at 120 days post-infection (dpi), when mice were afflicted by CCC. Mice were analyzed for electrical abnormalities, immune response and cardiac parasitism and tissue damage. Prophylactic immunization with rAdVax induced antibodies and H-2Kb-restricted cytotoxic and interferon (IFN)γ-producing CD8+ T-cells, reduced acute heart parasitism and electrical abnormalities in the chronic phase. Therapeutic vaccination increased survival and reduced electrical abnormalities after the prime (analysis at 160 dpi) and the boost (analysis at 180 and 230 dpi). Post-therapy mice exhibited less heart injury and electrical abnormalities compared with pre-therapy mice. rAdVax therapeutic vaccination preserved specific IFNγ-mediated immunity but reduced the response to polyclonal stimuli (anti-CD3 plus anti-CD28), CD107a+ CD8+ T-cell frequency and plasma nitric oxide (NO) levels. Moreover, therapeutic rAdVax reshaped immunity in the heart tissue as reduced the number of perforin+ cells, preserved the number of IFNγ+ cells, increased the expression of IFNγ mRNA but reduced inducible NO synthase mRNA. Vaccine-based immunostimulation with rAd might offer a rational alternative for re-programming the immune response to preserve and, moreover, recover tissue injury in Chagas' heart disease.

Show MeSH

Related in: MedlinePlus

Shift of the immune response through rAdVax immunotherapy in chronically T. cruzi-infected mice.(A) Chronically Colombian-infected mice were primed-boosted with 2 × 108 plaque-forming units (PFU) of rAdCtrl or a mixture of 108 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax) and analyzed at the endpoints, as indicated. (B) Numbers of IFNγ-secreting cells in the spleens of immunized mice were determined using ELISpot after 20 hours of stimulation with anti-CD3 plus anti-CD28. (C) Representative histograms showing the R1-gated CFSEhigh-based lymphoproliferative response after 72 hours of stimulation with anti-CD3 plus anti-CD28. (D) Representative dot plot showing the R1-gated CFSELow-based lymphoproliferative response after 72 hours of treatment with anti-CD3 plus anti-CD28. After stimulation, the spleen cells were stained with APC-conjugated anti-CD8. Cycling CFSELowCD8+ cells were detected in gates R3, R4 and R5. (E) Numbers of CD8+IFNγ-secreting cells in the spleens of immunized mice were determined by ELISpot using the H-2Kb-restricted VNHRFTLV peptide (3 and 10 μg/mL). The results represent three to five mice per experimental group of non-infected (NI) controls and T. cruzi-infected mice immunized with rAdCtrl or rAdVax and analyzed at 190 dpi (70 days post-therapy). * P <0.05, ** P <0.01 and ***P <0.001, experimental groups compared with NI controls. †P <0.05, non-stimulated compared with stimulated condition in an experimental group. § P <0.05, rAdVax-immunized compared with rAdCtrl-injected T. cruzi-infected mice.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4305326&req=5

ppat.1004594.g004: Shift of the immune response through rAdVax immunotherapy in chronically T. cruzi-infected mice.(A) Chronically Colombian-infected mice were primed-boosted with 2 × 108 plaque-forming units (PFU) of rAdCtrl or a mixture of 108 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax) and analyzed at the endpoints, as indicated. (B) Numbers of IFNγ-secreting cells in the spleens of immunized mice were determined using ELISpot after 20 hours of stimulation with anti-CD3 plus anti-CD28. (C) Representative histograms showing the R1-gated CFSEhigh-based lymphoproliferative response after 72 hours of stimulation with anti-CD3 plus anti-CD28. (D) Representative dot plot showing the R1-gated CFSELow-based lymphoproliferative response after 72 hours of treatment with anti-CD3 plus anti-CD28. After stimulation, the spleen cells were stained with APC-conjugated anti-CD8. Cycling CFSELowCD8+ cells were detected in gates R3, R4 and R5. (E) Numbers of CD8+IFNγ-secreting cells in the spleens of immunized mice were determined by ELISpot using the H-2Kb-restricted VNHRFTLV peptide (3 and 10 μg/mL). The results represent three to five mice per experimental group of non-infected (NI) controls and T. cruzi-infected mice immunized with rAdCtrl or rAdVax and analyzed at 190 dpi (70 days post-therapy). * P <0.05, ** P <0.01 and ***P <0.001, experimental groups compared with NI controls. †P <0.05, non-stimulated compared with stimulated condition in an experimental group. § P <0.05, rAdVax-immunized compared with rAdCtrl-injected T. cruzi-infected mice.

Mentions: Next, we investigated whether the beneficial effect of the prime-boost immunotherapy with rAdVax in chronically infected C57BL/6 mice was associated with reduction in the abnormal polyclonal activation observed in chronically infected mice and/or shift of the immune response to a protective profile. To this end, chronically infected mice received the homologous prime-boost vaccine rAdVax and were analyzed at 190 and 230 dpi, corresponding to 70 and 110 days post-therapy (Fig. 4A). As depicted in Fig. 4B, the potent IFNγ recall response after stimulation of mononuclear spleen cells with anti-CD3 plus anti-CD28, previously described as a hallmark of chronically T. cruzi-infected mice [22], was reproduced in the present study in the Colombian-infected rAdCtrl-immunized mice. In contrast, this response was abrogated by rAdVax immunotherapy (Fig. 4B). Moreover, the intense anti-CD3 plus anti-CD28-triggered lymphoproliferative response observed in total splenic T-cells from T. cruzi-infected mice injected with rAdCtrl was also inhibited by rAdVax immunotherapy (Fig. 4C). Notably, the increased anti-CD3 plus anti-CD28-triggered CD8+ T-cell proliferation observed during chronic infection was also reversed by rAdVax inoculation (Fig. 4D), whereas CD8+ T-cell recognition of the H-2Kb-restricted VNHRFTLV ASP2 peptide was preserved in rAdVax-immunized mice (Fig. 4E).


A human type 5 adenovirus-based Trypanosoma cruzi therapeutic vaccine re-programs immune response and reverses chronic cardiomyopathy.

Pereira IR, Vilar-Pereira G, Marques V, da Silva AA, Caetano B, Moreira OC, Machado AV, Bruna-Romero O, Rodrigues MM, Gazzinelli RT, Lannes-Vieira J - PLoS Pathog. (2015)

Shift of the immune response through rAdVax immunotherapy in chronically T. cruzi-infected mice.(A) Chronically Colombian-infected mice were primed-boosted with 2 × 108 plaque-forming units (PFU) of rAdCtrl or a mixture of 108 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax) and analyzed at the endpoints, as indicated. (B) Numbers of IFNγ-secreting cells in the spleens of immunized mice were determined using ELISpot after 20 hours of stimulation with anti-CD3 plus anti-CD28. (C) Representative histograms showing the R1-gated CFSEhigh-based lymphoproliferative response after 72 hours of stimulation with anti-CD3 plus anti-CD28. (D) Representative dot plot showing the R1-gated CFSELow-based lymphoproliferative response after 72 hours of treatment with anti-CD3 plus anti-CD28. After stimulation, the spleen cells were stained with APC-conjugated anti-CD8. Cycling CFSELowCD8+ cells were detected in gates R3, R4 and R5. (E) Numbers of CD8+IFNγ-secreting cells in the spleens of immunized mice were determined by ELISpot using the H-2Kb-restricted VNHRFTLV peptide (3 and 10 μg/mL). The results represent three to five mice per experimental group of non-infected (NI) controls and T. cruzi-infected mice immunized with rAdCtrl or rAdVax and analyzed at 190 dpi (70 days post-therapy). * P <0.05, ** P <0.01 and ***P <0.001, experimental groups compared with NI controls. †P <0.05, non-stimulated compared with stimulated condition in an experimental group. § P <0.05, rAdVax-immunized compared with rAdCtrl-injected T. cruzi-infected mice.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4305326&req=5

ppat.1004594.g004: Shift of the immune response through rAdVax immunotherapy in chronically T. cruzi-infected mice.(A) Chronically Colombian-infected mice were primed-boosted with 2 × 108 plaque-forming units (PFU) of rAdCtrl or a mixture of 108 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax) and analyzed at the endpoints, as indicated. (B) Numbers of IFNγ-secreting cells in the spleens of immunized mice were determined using ELISpot after 20 hours of stimulation with anti-CD3 plus anti-CD28. (C) Representative histograms showing the R1-gated CFSEhigh-based lymphoproliferative response after 72 hours of stimulation with anti-CD3 plus anti-CD28. (D) Representative dot plot showing the R1-gated CFSELow-based lymphoproliferative response after 72 hours of treatment with anti-CD3 plus anti-CD28. After stimulation, the spleen cells were stained with APC-conjugated anti-CD8. Cycling CFSELowCD8+ cells were detected in gates R3, R4 and R5. (E) Numbers of CD8+IFNγ-secreting cells in the spleens of immunized mice were determined by ELISpot using the H-2Kb-restricted VNHRFTLV peptide (3 and 10 μg/mL). The results represent three to five mice per experimental group of non-infected (NI) controls and T. cruzi-infected mice immunized with rAdCtrl or rAdVax and analyzed at 190 dpi (70 days post-therapy). * P <0.05, ** P <0.01 and ***P <0.001, experimental groups compared with NI controls. †P <0.05, non-stimulated compared with stimulated condition in an experimental group. § P <0.05, rAdVax-immunized compared with rAdCtrl-injected T. cruzi-infected mice.
Mentions: Next, we investigated whether the beneficial effect of the prime-boost immunotherapy with rAdVax in chronically infected C57BL/6 mice was associated with reduction in the abnormal polyclonal activation observed in chronically infected mice and/or shift of the immune response to a protective profile. To this end, chronically infected mice received the homologous prime-boost vaccine rAdVax and were analyzed at 190 and 230 dpi, corresponding to 70 and 110 days post-therapy (Fig. 4A). As depicted in Fig. 4B, the potent IFNγ recall response after stimulation of mononuclear spleen cells with anti-CD3 plus anti-CD28, previously described as a hallmark of chronically T. cruzi-infected mice [22], was reproduced in the present study in the Colombian-infected rAdCtrl-immunized mice. In contrast, this response was abrogated by rAdVax immunotherapy (Fig. 4B). Moreover, the intense anti-CD3 plus anti-CD28-triggered lymphoproliferative response observed in total splenic T-cells from T. cruzi-infected mice injected with rAdCtrl was also inhibited by rAdVax immunotherapy (Fig. 4C). Notably, the increased anti-CD3 plus anti-CD28-triggered CD8+ T-cell proliferation observed during chronic infection was also reversed by rAdVax inoculation (Fig. 4D), whereas CD8+ T-cell recognition of the H-2Kb-restricted VNHRFTLV ASP2 peptide was preserved in rAdVax-immunized mice (Fig. 4E).

Bottom Line: Therapeutic vaccination increased survival and reduced electrical abnormalities after the prime (analysis at 160 dpi) and the boost (analysis at 180 and 230 dpi).Post-therapy mice exhibited less heart injury and electrical abnormalities compared with pre-therapy mice. rAdVax therapeutic vaccination preserved specific IFNγ-mediated immunity but reduced the response to polyclonal stimuli (anti-CD3 plus anti-CD28), CD107a+ CD8+ T-cell frequency and plasma nitric oxide (NO) levels.Moreover, therapeutic rAdVax reshaped immunity in the heart tissue as reduced the number of perforin+ cells, preserved the number of IFNγ+ cells, increased the expression of IFNγ mRNA but reduced inducible NO synthase mRNA.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Biologia das Interações, Instituto Oswaldo Cruz/Fiocruz, Rio de Janeiro, Rio de Janeiro, Brazil.

ABSTRACT
Chagas disease (CD), caused by the protozoan Trypanosoma cruzi, is a prototypical neglected tropical disease. Specific immunity promotes acute phase survival. Nevertheless, one-third of CD patients develop chronic chagasic cardiomyopathy (CCC) associated with parasite persistence and immunological unbalance. Currently, the therapeutic management of patients only mitigates CCC symptoms. Therefore, a vaccine arises as an alternative to stimulate protective immunity and thereby prevent, delay progression and even reverse CCC. We examined this hypothesis by vaccinating mice with replication-defective human Type 5 recombinant adenoviruses (rAd) carrying sequences of amastigote surface protein-2 (rAdASP2) and trans-sialidase (rAdTS) T. cruzi antigens. For prophylactic vaccination, naïve C57BL/6 mice were immunized with rAdASP2+rAdTS (rAdVax) using a homologous prime/boost protocol before challenge with the Colombian strain. For therapeutic vaccination, rAdVax administration was initiated at 120 days post-infection (dpi), when mice were afflicted by CCC. Mice were analyzed for electrical abnormalities, immune response and cardiac parasitism and tissue damage. Prophylactic immunization with rAdVax induced antibodies and H-2Kb-restricted cytotoxic and interferon (IFN)γ-producing CD8+ T-cells, reduced acute heart parasitism and electrical abnormalities in the chronic phase. Therapeutic vaccination increased survival and reduced electrical abnormalities after the prime (analysis at 160 dpi) and the boost (analysis at 180 and 230 dpi). Post-therapy mice exhibited less heart injury and electrical abnormalities compared with pre-therapy mice. rAdVax therapeutic vaccination preserved specific IFNγ-mediated immunity but reduced the response to polyclonal stimuli (anti-CD3 plus anti-CD28), CD107a+ CD8+ T-cell frequency and plasma nitric oxide (NO) levels. Moreover, therapeutic rAdVax reshaped immunity in the heart tissue as reduced the number of perforin+ cells, preserved the number of IFNγ+ cells, increased the expression of IFNγ mRNA but reduced inducible NO synthase mRNA. Vaccine-based immunostimulation with rAd might offer a rational alternative for re-programming the immune response to preserve and, moreover, recover tissue injury in Chagas' heart disease.

Show MeSH
Related in: MedlinePlus