RESUMO
BACKGROUND: Interruption of benznidazole therapy due to the appearance of adverse effects, which is presumed to lead to treatment failure, is a major drawback in the treatment of chronic Chagas disease. METHODS: Trypanosoma cruzi-specific humoral and T cell responses, T cell phenotype and parasite load were measured to compare the outcome in 33 subjects with chronic Chagas disease treated with an incomplete benznidazole regimen and 58 subjects treated with the complete regimen, during a median follow-up period of 48 months. RESULTS: Both treatment regimens induced a reduction in the T. cruzi-specific antibody levels and similar rates of treatment failure when evaluated using quantitative PCR. Regardless of the regimen, polyfunctional CD4+ T cells increased in the subjects, with successful treatment outcome defined as a decrease of T. cruzi-specific antibodies. Regardless of the serological outcome, naive and central memory T cells increased after both regimens. A decrease in CD4+ HLA-DR+ T cells was associated with successful treatment in both regimens. The cytokine profiles of subjects with successful treatment showed fewer inflammatory mediators than those of the untreated T. cruzi-infected subjects. High levels of T cells expressing IL-7 receptor and low levels of CD8+ T cells expressing the programmed cell death protein 1 at baseline were associated with successful treatment following benznidazole interruption. CONCLUSIONS: These findings challenge the notion that treatment failure is the sole potential outcome of an incomplete benznidazole regimen and support the need for further assessment of the treatment protocols for chronic Chagas disease.
Assuntos
Doença de Chagas , Nitroimidazóis , Tripanossomicidas , Trypanosoma cruzi , Doença de Chagas/tratamento farmacológico , Humanos , Nitroimidazóis/uso terapêutico , Tripanossomicidas/uso terapêuticoRESUMO
In a pilot study, we showed that the intermittent administration of benznidazole in chronic Chagas disease patients resulted in a low rate of treatment suspension and therapeutic failure, as assessed by quantitative PCR (qPCR) at the end of treatment. Here, a 3-year posttreatment follow-up study of the same cohort of patients is presented. The treatment scheme consisted of 12 doses of benznidazole at 5 mg/kg of body weight/day in two daily doses every 5 days. Parasite load, Trypanosoma cruzi-specific antibodies, and serum chemokine levels were measured prior to treatment and after a median follow-up of 36 months posttreatment by DNA minicircle kinetoplastid and nuclear DNA satellite sequence qPCR methods, conventional serological techniques, a Luminex-based assay with recombinant T. cruzi proteins, and a cytometric bead array. At the end of follow-up, 14 of 17 (82%) patients had negative qPCR findings, whereas three of 17 (18%) had detectable nonquantiï¬able findings by at least one of the qPCR techniques. A decline in parasite-specific antibodies at 12 months posttreatment was confirmed by conventional serological tests and the Luminex assays. Monocyte chemoattractant protein 1 levels increased after treatment, whereas monokine induced by gamma interferon levels decreased. New posttreatment electrocardiographic abnormalities were observed in only one patient who had cardiomyopathy prior to treatment. Together, these data strengthen our previous findings by showing that the intermittent administration of benznidazole results in a low rate of treatment suspension, with treatment efficacy comparable to that of a daily dose of 5 mg/kg for 60 days.
Assuntos
Doença de Chagas , Nitroimidazóis , Tripanossomicidas , Trypanosoma cruzi , Doença de Chagas/tratamento farmacológico , Seguimentos , Humanos , Nitroimidazóis/uso terapêutico , Projetos Piloto , Tripanossomicidas/uso terapêuticoRESUMO
Background: In contrast to adults, Trypanosoma cruzi-infected children have more broadly functional Trypanosoma cruzi-specific T cells, and the total T-cell compartment exhibits fewer signs of immune exhaustion. However, not much is known about the link between immunocompetence and the treatment efficacy for human Chagas disease. Methods: Using cytokine enzyme-linked immunosorbent spot (ELISPOT) polychromatic flow cytometry, cytometric bead assay, multiplex serological assays and quantitative PCR, we evaluated T. cruzi-specific T-cell and antibody immune responses, T-cell phenotypes and parasitemia in children in the early chronic phase of Chagas disease undergoing anti-Trypanosoma cruzi treatment. Results: Treatment with benznidazole or nifurtimox induced a decline in T. cruzi-specific IFN-γ- and IL-2-producing cells and proinflammatory cytokines and chemokines. T-cell responses became detectable after therapy in children bearing T-cell responses under background levels prior to treatment. The total frequencies of effector, activated and antigen-experienced T cells also decreased following anti-T. cruzi therapy, along with an increase in T cells expressing the receptor of the homeostatic cytokine IL-7. Posttreatment changes in several of these markers distinguished children with a declining serologic response suggestive of successful treatment from those with sustained serological responses in a 5-year follow-up study. A multivariate analysis demonstrated that lower frequency of CD4+CD45RA-CCR7-CD62L- T cells prior to drug therapy was an independent indicator of successful treatment. Conclusions: These findings further validate the usefulness of alternative metrics to monitor treatment outcomes. Distinct qualitative and quantitative characteristics of T cells prior to drug therapy may be linked to treatment efficacy.
Assuntos
Doença de Chagas , Quimiocinas/imunologia , Nitroimidazóis/administração & dosagem , Parasitemia , Linfócitos T/imunologia , Trypanosoma cruzi/imunologia , Adolescente , Doença de Chagas/tratamento farmacológico , Doença de Chagas/imunologia , Doença de Chagas/patologia , Criança , Pré-Escolar , Feminino , Seguimentos , Humanos , Masculino , Parasitemia/tratamento farmacológico , Parasitemia/imunologia , Parasitemia/patologia , Linfócitos T/patologiaRESUMO
The parasite Trypanosoma cruzi is the causative agent of Chagas disease, a potentially life-threatening infection that represents a major health problem in Latin America. Several characteristics of this protozoan contribute to the lack of an effective vaccine, among them: its silent invasion mechanism, T. cruzi antigen redundancy and immunodominance without protection. Taking into account these issues, we engineered Traspain, a chimeric antigen tailored to present a multivalent display of domains from key parasitic molecules, combined with stimulation of the STING pathway by c-di-AMP as a novel prophylactic strategy. This formulation proved to be effective for the priming of functional humoral responses and pathogen-specific CD8+ and CD4+ T cells, compatible with a Th1/Th17 bias. Interestingly, vaccine effectiveness assessed across the course of infection, showed a reduction in parasite load and chronic inflammation in different proof of concept assays. In conclusion, this approach represents a promising tool against parasitic chronic infections.
RESUMO
BACKGROUND: Subjects are considered infected with Trypanosoma cruzi when tested positive by at least two out of three serological tests, whereas a positive result in only one of up to three tests is termed "serodiscordant" (SD). Assessment of parasite-specific T-cell responses may help discriminate the uninfected from infected individuals among SD subjects. METHODS: Peripheral blood mononuclear cells from SD and seropositive (SP) subjects, who were born in areas endemic for T. cruzi infection but living in Buenos Aires city, Argentina, at the time of the study, and seronegative unexposed subjects were included for analysis. The function and phenotype of T cells were assessed by interferon-γ (IFN-γ) and interleukin (IL)-2 enzyme-linked immunospot assay and multiparameter flow cytometry. T. cruzi-specific antibodies were quantified by conventional serology and a multiplex assay format. RESULTS: SD subjects exhibited immunity cell responses to T. cruzi but in contrast to SP subjects, T cells in SD subjects more often display the simultaneous production of IFN-γ and IL-2 in response to T. cruzi antigens and have a resting phenotype. SD individuals also have higher IFN-γ spot counts, polyfunctional CD4+ T-cells enriched in IL-2 secreting cells and low levels of antibodies specific for a set of T. cruzi-derived recombinant proteins compared with the SP group. Long-term follow-up of SD individuals confirmed that humoral and T-cell responses fluctuate but are sustained over time in these subjects. T cells in SD subjects for T. cruzi infection did not recognize Leishmania antigens. CONCLUSION: Both T-cell and humoral responses in most subjects assessed by conventional tests as SD for T. cruzi infection indicate prior exposure to infection and the establishment of immunological memory suggestive of a resolved infection.
RESUMO
BACKGROUND: Chagas disease is the highest impact parasitic disease in Latin America. We have proposed that changes in Trypanosoma cruzi-specific immune responses might serve as surrogate indicators of treatment success. Herein, we addressed in a long-term follow-up study whether cure achieved after treatment can be predicted by changes in non-conventional indexes of anti-parasite serological and T cell activities. METHODOLOGY/PRINCIPAL FINDINGS: T. cruzi-specific T cell responses, as measured by interferon-γ ELISPOT and T. cruzi-specific antibodies assessed by ELISA, hemagglutination and immunofluorescence tests as well as by a multiplex assay incorporating 14 recombinant T. cruzi proteins were measured in 33 patients at 48-150 months post-benznidazole treatment. Cure - as assessed by conventional serological tests - was associated with an early decline in T. cruzi-specific IFN-γ-producing T cells and in antibody titers measured by the multiplex serological assay. Changes in the functional status and potential of T. cruzi-specific T cells, indicative of reduced antigen stimulation, provided further evidence of parasitological cure following benznidazole treatment. Patients showing a significant reduction in T. cruzi-specific antibodies had higher pre-therapy levels of T. cruzi-specific IFN-γ- producing T cells compared to those with unaltered humoral responses post-treatment. CONCLUSIONS/SIGNIFICANCE: Monitoring of appropriate immunological responses can provide earlier and robust measures of treatment success in T. cruzi infection.
Assuntos
Linfócitos B/imunologia , Doença de Chagas/tratamento farmacológico , Monitoramento de Medicamentos/métodos , Nitroimidazóis/uso terapêutico , Linfócitos T/imunologia , Trypanosoma cruzi/imunologia , Adulto , Anticorpos Antiprotozoários/sangue , Doença de Chagas/imunologia , ELISPOT , Seguimentos , Humanos , Interferon gama/metabolismo , Pessoa de Meia-Idade , Prognóstico , Resultado do Tratamento , Adulto JovemRESUMO
Vaccines to prevent Trypanosoma cruzi infection in humans or animals are not available, and in many settings, dogs are an important source of domestic infection for the insect vector. Identification of infected canines is crucial for evaluating peridomestic transmission dynamics and parasite control strategies. As immune control of T. cruzi infection is dependent on humoral and cell-mediated immune responses, we aimed to define a serodiagnostic assay and T cell phenotypic markers for identifying infected dogs and studying the canine T. cruzi-specific immune response. Plasma samples and peripheral blood mononuclear cells (PBMCs) were obtained from forty-two dogs living in a T. cruzi-endemic region. Twenty dogs were known to be seropositive and nine seronegative by conventional serologic tests two years prior to our study. To determine canine seroreactivity, we tested sera or plasma samples in a multiplex bead array against eleven recombinant T. cruzi proteins. Ninety-four percent (17/18) of dogs positive by multiplex serology were initially positive by conventional serology. The frequency of IFNγ-producing cells in PBMCs responding to T. cruzi correlated to serological status, identifying 95% of multiplex seropositive dogs. Intracellular staining identified CD4+ and CD8+ T cell populations as the sources of T. cruzi lysate-induced IFNγ. Low expression of CCR7 and CD62L on CD4+ and CD8+ T cells suggested a predominance of effector/effector memory T cells in seropositive canines. These results are the first, to our knowledge, to correlate T. cruzi-specific antibody responses with T cell responses in naturally infected dogs and validate these methods for identifying dogs exposed to T. cruzi.
Assuntos
Doença de Chagas/veterinária , Doenças do Cão/diagnóstico , Linfócitos T/citologia , Trypanosoma cruzi/fisiologia , Animais , Argentina , Doença de Chagas/sangue , Doença de Chagas/diagnóstico , Doença de Chagas/parasitologia , Citocinas/genética , Doenças do Cão/sangue , Doenças do Cão/parasitologia , Cães , Imunofenotipagem/veterinária , Fenótipo , Testes Sorológicos/métodos , Testes Sorológicos/veterinária , Linfócitos T/parasitologia , Estados UnidosRESUMO
BACKGROUND: Adults with chronic Trypanosoma cruzi exhibit a poorly functional T cell compartment, characterized by monofunctional (IFN-γ-only secreting) parasite-specific T cells and increased levels of terminally differentiated T cells. It is possible that persistent infection and/or sustained exposure to parasites antigens may lead to a progressive loss of function of the immune T cells. METHODOLOGY/PRINCIPAL FINDINGS: To test this hypothesis, the quality and magnitude of T. cruzi-specific T cell responses were evaluated in T. cruzi-infected children and compared with long-term T. cruzi-infected adults with no evidence of heart failure. The phenotype of CD4(+) T cells was also assessed in T. cruzi-infected children and uninfected controls. Simultaneous secretion of IFN-γ and IL-2 measured by ELISPOT assays in response to T. cruzi antigens was prevalent among T. cruzi-infected children. Flow cytometric analysis of co-expression profiles of CD4(+) T cells with the ability to produce IFN-γ, TNF-α, or to express the co-stimulatory molecule CD154 in response to T. cruzi showed polyfunctional T cell responses in most T. cruzi-infected children. Monofunctional T cell responses and an absence of CD4(+)TNF-α(+)-secreting T cells were observed in T. cruzi-infected adults. A relatively high degree of activation and differentiation of CD4(+) T cells was evident in T. cruzi-infected children. CONCLUSIONS/SIGNIFICANCE: Our observations are compatible with our initial hypothesis that persistent T. cruzi infection promotes eventual exhaustion of immune system, which might contribute to disease progression in long-term infected subjects.
Assuntos
Doença de Chagas/imunologia , Linfócitos T/imunologia , Trypanosoma cruzi/imunologia , Adolescente , Adulto , Ligante de CD40/análise , Criança , Pré-Escolar , ELISPOT , Feminino , Citometria de Fluxo , Humanos , Imunofenotipagem , Interferon gama/metabolismo , Interleucina-2/metabolismo , Masculino , Pessoa de Meia-Idade , Linfócitos T/química , Fator de Necrose Tumoral alfa/metabolismoRESUMO
We had formerly demonstrated that subjects chronically infected with Trypanosoma cruzi show impaired T cell responses closely linked with a process of T cell exhaustion. Recently, the expression of several inhibitory receptors has been associated with T cell dysfunction and exhaustion. In this study, we have examined the expression of the cytotoxic T lymphocyte antigen 4 (CTLA-4) and the leukocyte immunoglobulin like receptor 1 (LIR-1) by peripheral T. cruzi antigen-responsive IFN-gamma (IFN-γ)-producing and total T cells from chronically T. cruzi-infected subjects with different clinical forms of the disease. CTAL-4 expression was also evaluated in heart tissue sections from subjects with severe myocarditis. The majority of IFN-γ-producing CD4(+) T cells responsive to a parasite lysate preparation were found to express CTLA-4 but considerably lower frequencies express LIR-1, irrespective of the clinical status of the donor. Conversely, few IFN-γ-producing T cells responsive to tetanus and diphtheria toxoids expressed CTLA-4 and LIR-1. Polyclonal stimulation with anti-CD3 antibodies induced higher frequencies of CD4(+)CTAL-4(+) T cells in patients with severe heart disease than in asymptomatic subjects. Ligation of CTLA-4 and LIR-1 with their agonistic antibodies, in vitro, reduces IFN-γ production. Conversely, CTLA-4 blockade did not improved IFN-γ production in response to T. cruzi antigens. Subjects with chronic T. cruzi infection had increased numbers of CD4(+)LIR-1(+) among total peripheral blood mononuclear cells, relative to uninfected individuals and these numbers decreased after treatment with benznidazole. CTLA-4 was also expressed by CD3(+) T lymphocytes infiltrating heart tissues from chronically infected subjects with severe myocarditis. These findings support the conclusion that persistent infection with T. cruzi leads to the upregulation of inhibitory receptors which could alter parasite specific T cell responses in the chronic phase of Chagas disease.
Assuntos
Antígenos CD/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Antígeno CTLA-4/metabolismo , Doença de Chagas/imunologia , Regulação da Expressão Gênica/imunologia , Coração/parasitologia , Receptores Imunológicos/metabolismo , Trypanosoma cruzi/patogenicidade , Adulto , Antígenos de Protozoários/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/parasitologia , Doença de Chagas/complicações , Doença de Chagas/tratamento farmacológico , Doença de Chagas/metabolismo , Doença Crônica , Regulação da Expressão Gênica/efeitos dos fármacos , Coração/efeitos dos fármacos , Humanos , Interferon gama/biossíntese , Receptor B1 de Leucócitos Semelhante a Imunoglobulina , Masculino , Pessoa de Meia-Idade , Miocardite/complicações , Miocardite/imunologia , Miocardite/metabolismo , Nitroimidazóis/farmacologia , Nitroimidazóis/uso terapêutico , Especificidade da Espécie , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/imunologiaRESUMO
BACKGROUND: Trypanosoma cruzi is a protozoan parasite that causes severe disease in millions of habitants of developing countries. Currently there is no vaccine to prevent this disease and the available drugs have the consequences of side effects. Live vaccines are likely to be more effective in inducing protection than recombinant proteins or DNA vaccines; however, safety problems associated to their use have been pointed out. In recent years, increasing knowledge on the molecular genetics of Trypanosomes has allowed the identification and elimination of genes that may be necessary for parasite infectivity and survival. In this sense, targeted deletion or disruption of specific genes in the parasite genome may protect against such reversion to virulent genotypes. METHODS AND FINDINGS: By targeted gene disruption we generated monoallelic mutant parasites for the dhfr-ts gene in a T. cruzi strain that has been shown to be naturally attenuated. In comparison to T. cruzi wild type epimastigotes, impairment in growth of dhfr-ts(+/-) mutant parasites was observed and mutant clones displayed decreased virulence in mice. Also, a lower number of T. cruzi-specific CD8(+) T cells, in comparison to those induced by wild type parasites, was detected in mice infected with mutant parasites. However, no remarkable differences in the protective effect of TCC wild type versus TCC mutant parasites were observed. Mice challenged with virulent parasites a year after the original infection with the mutant parasites still displayed a significant control over the secondary infection. CONCLUSION: This study indicates that it is possible to generate genetically attenuated T. cruzi parasites able to confer protection against further T. cruzi infections.
Assuntos
Doença de Chagas/parasitologia , Complexos Multienzimáticos/genética , Vacinas Protozoárias/genética , Vacinas Protozoárias/imunologia , Tetra-Hidrofolato Desidrogenase/genética , Timidilato Sintase/genética , Trypanosoma cruzi/genética , Trypanosoma cruzi/imunologia , Animais , Linfócitos T CD8-Positivos , Doença de Chagas/imunologia , Doença de Chagas/prevenção & controle , Técnicas de Inativação de Genes , Camundongos , Complexos Multienzimáticos/imunologia , Mutação , Estatísticas não Paramétricas , Tetra-Hidrofolato Desidrogenase/imunologia , Timidilato Sintase/imunologia , Trypanosoma cruzi/patogenicidade , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologiaRESUMO
Trypanosoma cruzi-specific immune responses were evaluated in a total of 88 subjects living in areas endemic of Chagas disease of Argentina by IFN-gamma ELISPOT assays and immunoblotting. Positive T. cruzi antigen-induced IFN-gamma responses were detected in 42% of subjects evaluated (15/26 positive by conventional serology and 22/62 seronegative subjects). Using immunoblotting, T. cruzi-specific IgG reactivity was detected in all seropositive subjects and in 11% (7/61) of subjects negative by conventional serology. Measurements of T cell responses and antibodies by immunoblotting, in conjunction with conventional serology, might enhance the capability of detection of exposure to T. cruzi in endemic areas.
Assuntos
Doença de Chagas/imunologia , Trypanosoma cruzi/imunologia , Adulto , Idoso , Anticorpos Antiprotozoários/sangue , Argentina , Doenças Endêmicas , Feminino , Humanos , Immunoblotting , Imunoglobulina G/sangue , Interferon gama/metabolismo , Leucócitos Mononucleares/imunologia , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: As many as 20 million people are living with Trypanosoma cruzi infection in Latin American, yet few receive any treatment. The major limitation in developing and evaluating potential new drugs for their efficacy is the lack of reliable tests to assess parasite burden and elimination. METHODS: Adults volunteers with chronic T. cruzi infection were evaluated clinically and stratified according to the Kuschnir classification. Individuals with group 0 and group 1 clinical status were treated with benznidazole (5 mg/kg per day for 30 days). The changes in T. cruzi-specific T cell and antibody responses, as well as in clinical status, were measured periodically over the 3-5-year follow-up period and were compared with pretreatment conditions and with values in an untreated control group. RESULTS: The frequency of peripheral interferon (IFN)-gamma-producing T cells specific for T. cruzi declined as early as 12 months after benznidazole treatment and subsequently became undetectable in a substantial proportion of treated subjects. In addition, decreases in antibody responses to a pool of recombinant T. cruzi proteins also decreased in many of these same subjects. The shift to negative IFN-gamma T cell responses was highly associated with an early increase in IFN-gamma-producing T cells with phenotypic features of effector/effector memory cells in a subset of subjects. Benznidazole treatment also resulted in an increase in naive and early differentiated memory-like CD8(+) T cells in a majority of subjects. CONCLUSIONS: Benznidazole treatment during chronic Chagas disease has a substantial impact on parasite-specific immune response that is likely indicative of treatment efficacy and cure.
Assuntos
Doença de Chagas/tratamento farmacológico , Doença de Chagas/imunologia , Nitroimidazóis/uso terapêutico , Tripanossomicidas/uso terapêutico , Trypanosoma cruzi/fisiologia , Adulto , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Citometria de Fluxo , Humanos , Interferon gama/metabolismo , Interleucina-2/metabolismo , Pessoa de Meia-Idade , Linfócitos T/imunologia , Linfócitos T/metabolismo , Resultado do Tratamento , Trypanosoma cruzi/imunologia , Adulto JovemRESUMO
Previously we found that the frequency of IFN-gamma-producing CD8(+) T cells specific for Trypanosoma cruzi inversely correlates with disease severity in chronic human Chagas disease along with low levels of IL-2-secreting CD8(+) T cells in all clinical stages. This impairment of the parasite-specific T cell responses was associated with phenotypic features of immune senescence of the CD8(+) T cell compartment. These data prompted us to address the question of whether the CD4(+) T cell compartment also experiences signs of exhaustion. Thus, we performed a functional and phenotypical characterization of T. cruzi-specific and overall CD4(+) T cells in chronically infected subjects with different degrees of cardiac dysfunction. The results show an inverse association between disease severity and the frequency of T. cruzi-specific IFN-gamma-producing CD4(+) T cells. The high expression of CD27 and CD28 with a relative low expression of CD57 found on CD4(+)IFN-gamma(+) T cells suggests that the effector T cell pool in chronic T. cruzi infection includes a high proportion of newly recruited T cells, but a low frequency of long-term memory cells. The total CD4(+) T cell compartment shows signs of senescence and later stages of differentiation associated with more severe stages of the disease. These findings support the hypothesis that long-term T. cruzi infection in humans might exhaust long-lived memory T cells.
Assuntos
Linfócitos T CD4-Positivos/patologia , Doença de Chagas/imunologia , Trypanosoma cruzi , Adulto , Idoso , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/parasitologia , Diferenciação Celular , Senescência Celular , Doença de Chagas/complicações , Doença de Chagas/patologia , Doença Crônica , Feminino , Cardiopatias/etiologia , Cardiopatias/patologia , Humanos , Memória Imunológica , Imunofenotipagem , Interferon gama/biossíntese , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de Doença , Fatores de TempoRESUMO
BACKGROUND: Previously, we identified a set of HLA-A020.1-restricted trans-sialidase peptides as targets of CD8+ T cell responses in HLA-A0201+ individuals chronically infected by T. cruzi. METHODS AND FINDINGS: Herein, we report the identification of peptides encoded by the same trans-sialidase gene family that bind alleles representative of the 6 most common class I HLA-supertypes. Based on a combination of bioinformatic predictions and HLA-supertype considerations, a total of 1001 epitopes predicted to bind to HLA A01, A02, A03, A24, B7 and B44 supertypes was selected. Ninety-six supertype-binder epitopes encoded by multiple trans-sialidase genes were tested for the ability to stimulate a recall CD8+ T cell response in the peripheral blood from subjects with chronic T. cruzi infection regardless the HLA haplotype. An overall hierarchy of antigenicity was apparent, with the A02 supertype peptides being the most frequently recognized in the Chagas disease population followed by the A03 and the A24 supertype epitopes. CD8+ T cell responses to promiscuous epitopes revealed that the CD8+ T cell compartment specific for T. cruzi displays a functional profile with T cells secreting interferon-gamma alone as the predominant pattern and very low prevalence of single IL-2-secreting or dual IFN-gamma/IL-2 secreting T cells denoting a lack of polyfunctional cytokine responses in chronic T. cruzi infection. CONCLUSIONS: This study identifies a set of T. cruzi peptides that should prove useful for monitoring immune competence and changes in infection and disease status in individuals with chronic Chagas disease.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Doença de Chagas/imunologia , Epitopos de Linfócito T/genética , Glicoproteínas/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Neuraminidase/genética , Adulto , Animais , América Central/epidemiologia , Doença de Chagas/epidemiologia , Doença de Chagas/genética , Ensaio de Imunoadsorção Enzimática , Epitopos de Linfócito T/imunologia , Antígenos HLA/genética , Antígenos HLA/imunologia , Antígenos HLA-A/genética , Antígeno HLA-A2 , Antígenos HLA-B/imunologia , Hemaglutinação , Humanos , Interferon gama/imunologia , Pessoa de Meia-Idade , América do Sul/epidemiologia , Linfócitos T/imunologia , Trypanosoma cruzi/imunologiaRESUMO
Para determinar el efecto del tratamiento con benznidazol sobre las células T de memoria específica para Trypanosoma cruzi, se seleccionaron 47 pacientes con tres reacciones serológicas positivas para T. cruzi, sin cardiopatía y edades comprendidas entre los 30 y los 50 años. El tratamiento se realizó con benznidazol en dosis de 5 mg/kg/d por 30 días. Se efectuó una evaluación serológica, inmunológica y clínica pretratamiento (tiempo 0) y a los 2, 6 y 12 meses postratamiento. Posteriormente, los controles se hicieron anualmente. La respuesta de linfocitos T frente a un lisado de amastigotas de T. cruzi se evaluó por la técnica de ELISPOT para IFN-ã. La frecuencia de linfocitos T de memoria productores de IFN-ã específicos para T. cruzi disminuyó significativamente en el grupo tratado (n = 33) versus el no tratado (n = 14) 12 meses después del seguimiento. Once de 25 (44%) pacientes que recibieron benznidazol negativizaron la respuesta para IFN-ã. Cuatro de los 11 (36%) pacientes con ELISPOT (+) que negativizaron la respuesta por ELISPOT para IFN-ã también negativizaron la serología convencional a los 2 años postratamiento. Durante el seguimiento no se observaron alteraciones clínicas. Estos hallazgos muestran que el benznidazol es capaz de modular la respuesta celular T de memoria específica para T. cruzi. La medición de la frecuencia de linfocitos T de memoria productores de IFN-ã podría constituir un ensayo más sensible y precoz para determinar el impacto/eficacia del tratamiento específico contra este parásito.
To determine the effect of benznidazol therapy on memory T cells specific for Trypanosoma cruzi, 47 patients between 30 and 50 years old and three positive serological tests for T. cruzi without cardiopathy were selected. Benznidazol was administered in a dose of 5 mg/kg/d during 30 days. Serological, immunological and clinical assessment was performed at basal (time 0) and at 2, 6 and 12 months following treatment, and once a year thereafter. IFN-ã ELISPOT assay was used to evaluate T cell responses against a T. cruzi lysate obtained from amastigotes. The frequency of IFN-ã - producing memory T lymphocytes specific for T. cruzi was significantly lower in the treatment group (n=33) compared to the control group (n=14) 12 months after the therapy. IFN- ã response became negative in 11 patients in the treatment group (44%). Among these 11 patients, conventional serology also became negative in 4 patients (36%) after 2 years of treatment. No clinical manifestations occurred during follow-up. These findings show that benznidazol is capable of modulating T cell responses specific for T. cruzi. Measuring the frequency of memory T lymphocytes producing IFN-ã might become a more sensitive test to determine earlier the impact and/or efficacy of the specific treatment against this parasite.
Assuntos
Antiparasitários/uso terapêutico , Cardiomiopatia Chagásica/tratamento farmacológico , Doença de Chagas/tratamento farmacológico , Produção de Droga sem Interesse Comercial , Vacinas Protozoárias , Doenças Raras/tratamento farmacológico , Trypanosoma cruzi/imunologia , Animais , Cardiomiopatia Chagásica/diagnóstico , Cardiomiopatia Chagásica/epidemiologia , Cardiomiopatia Chagásica/prevenção & controle , Doença de Chagas/diagnóstico , Doença de Chagas/epidemiologia , Doença de Chagas/prevenção & controle , Doença de Chagas/transmissão , Controle de Doenças Transmissíveis , Vetores de Doenças , Humanos , Doenças Raras/diagnóstico , Doenças Raras/epidemiologia , Doenças Raras/prevenção & controle , América do Sul , Trypanosoma cruzi/patogenicidadeRESUMO
CD8+ T cells are crucial for control of a number of medically important protozoan parasites, including Trypanosoma cruzi, the agent of human Chagas disease. Yet, in contrast to the wealth of information from viral and bacterial infections, little is known about the antigen specificity or the general development of effector and memory T-cell responses in hosts infected with protozoans. In this study we report on a wide-scale screen for the dominant parasite peptides recognized by CD8+ T cells in T. cruzi-infected mice and humans. This analysis demonstrates that in both hosts the CD8+ T-cell response is highly focused on epitopes encoded by members of the large trans-sialidase family of genes. Responses to a restricted set of immunodominant peptides were especially pronounced in T. cruzi-infected mice, with more than 30% of the CD8+ T-cell response at the peak of infection specific for two major groups of trans-sialidase peptides. Experimental models also demonstrated that the dominance patterns vary depending on the infective strain of T. cruzi, suggesting that immune evasion may be occurring at a population rather than single-parasite level.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Doença de Chagas/imunologia , Neuraminidase/genética , Trypanosoma cruzi/genética , Trypanosoma cruzi/imunologia , Adulto , Animais , Argentina , Brasil , Células Cultivadas , Citotoxicidade Imunológica , Modelos Animais de Doenças , Variação Genética , Genoma , Humanos , Isoenzimas/genética , Complexo Principal de Histocompatibilidade , Camundongos , Camundongos Endogâmicos C57BL , Trypanosoma cruzi/enzimologiaRESUMO
We present a cross-sectional analysis of the maturation and migratory properties of the memory CD8(+) T cell compartment, in relation to the severity of heart disease in individuals with chronic Trypanosoma cruzi infection removed from endemic areas for longer than 20 years. Subjects with none or mild heart involvement were more likely to mount T. cruzi-specific memory IFN-gamma responses than subjects with more advanced cardiac disease, and the T. cruzi-specific CD8(+) T cell population was enriched in early-differentiated (CD27(+)CD28(+)) cells in responding individuals. In contrast, the frequency of CD27(+)CD28(+)CD8(+) T cells in the total memory CD8(+) T cell population decreases, as disease becomes more severe, while the proportion of fully differentiated memory (CD27(-)CD28(-)) CD8(+) T cells increases. The analysis of CCR7 expression revealed a significant increase in total effector/memory CD8(+) T cells (CD45RA(-)CCR7(-)) in subjects with mild heart disease as compared with uninfected controls. Altogether, these results are consistent with the hypothesis of a gradual clonal exhaustion in the CD8(+) T cell population, perhaps as a result of continuous antigenic stimulation by persistent parasites.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Doença de Chagas/imunologia , Doença de Chagas/parasitologia , Memória Imunológica/imunologia , Trypanosoma cruzi/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Antígenos de Protozoários/sangue , Apoptose/imunologia , Doença de Chagas/sangue , Doença Crônica , Feminino , Humanos , Linfopoese , Masculino , Pessoa de Meia-IdadeRESUMO
This study sought to quantify CD8(+) T cell responses to Trypanosoma cruzi and to identify potential links between these responses and the severity of disease in humans. In the majority of patients with Chagas disease, staining with class I major histocompatibility complex tetramers and analysis of interferon (IFN)- gamma ELISPOT responses to a panel of known cytotoxic T lymphocyte target epitopes from T. cruzi failed to identify parasite-specific CD8(+) T cells. However, the frequency of individuals with positive ELISPOT responses was higher in areas of active transmission. Analysis of IFN- gamma ELISPOT responses to a parasite lysate revealed a very high frequency of responders among patients with mild clinical disease and a very low frequency of responders among those with the most severe form of the disease. These data suggest that the frequency of IFN- gamma -producing T cells in patients with chronic Chagas disease is associated with the history of recent exposure and with the clinical status of the patient.