RESUMEN
We recently evaluated the in vitro proliferative response and interferon (IFN)-gamma production of peripheral blood mononuclear cells from a group of 25 people who were treated for Chagas' disease during the acute phase of Trypanosoma cruzi infection and followed up for a period of 14-30 years. On the basis of the parasitological and serological tests, the individuals were classified as cured (C), dissociated, or not cured (NC). Members of group C (the group without cardiac alterations) presented significantly stronger proliferative response against the parasite antigens, with secretion of high levels of IFN-gamma in comparison with the NC group, raising a question about the role of this cytokine in the curing of human T. cruzi infection. Severe cardiac alterations were observed only in 1 of 25 patients, which suggests that treatment benefited the patients.
Asunto(s)
Enfermedad de Chagas/tratamiento farmacológico , Enfermedad de Chagas/inmunología , Enfermedad Aguda , Adolescente , Adulto , Anciano , Enfermedad de Chagas/complicaciones , Enfermedad Crónica , Estudios de Seguimiento , Humanos , Interferón gamma/análisis , Activación de Linfocitos , Persona de Mediana EdadAsunto(s)
Enfermedad de Chagas/tratamiento farmacológico , Nitroimidazoles/uso terapéutico , Tripanocidas/uso terapéutico , Adolescente , Adulto , Animales , Anticuerpos Antiprotozoarios/aislamiento & purificación , Enfermedad de Chagas/inmunología , Enfermedad de Chagas/parasitología , Humanos , Persona de Mediana Edad , Pruebas Serológicas , Trypanosoma cruzi/aislamiento & purificaciónRESUMEN
An apparently paradoxical role for IFN-gamma in human Chagas' disease was observed when studying the pattern of cytokine production by peripheral blood mononuclear cells (PBMC) obtained from two groups of chagasic patients after specific stimulation with Trypanosoma cruzi-derived antigens. The groups studied were 1) patients treated with benznidazole during the acute phase of Trypanosoma cruzi infection and 2) chronically infected untreated patients. In the treated group, higher levels of IFN-gamma were produced by PBMC from individuals cured after treatment when compared to non-cured patients. In contrast, in the chronically infected group (not treated) higher levels of IFN-gamma were produced by PBMC from cardiac patients in comparison with asymptomatic (indeterminate) patients. This apparently paradoxical role for IFN-gamma in human Chagas' disease is discussed in terms of the possibility of a temporal difference in IFN-gamma production during the initial stages of the infection (acute phase) in the presence or absence of chemotherapy. The maintenance of an immune response with high levels of IFN-gamma production during the chronic phase of the infection may favor cure or influence the development of the cardiac form of the disease.
Asunto(s)
Enfermedad de Chagas/inmunología , Interferón gamma/fisiología , Enfermedad de Chagas/sangre , Humanos , Leucocitos MononuclearesRESUMEN
An apparently paradoxical role for IFN-gamma in human Chagas'disease was observed when studying the pattern of cytokine production by peripheral blood mononuclear cells (PBMC) obtained from two groups of chagasic patients after specific stimulation with Trypanosoma cruzi-derived antigens. The groups studied were 1) patients treated with bendnidazole during the acute phase of Trypanosoma cruzi infection and 2) chronically infected untreated patients. In the treated group, higher levels of IFN-gamma were produced by PBMC from individuals cured after treatment when compared to non-cured patients. In contrast, in the chronically infected group (not treated) higher levels of IFN-gamma were produced by PBMC from cardiac patients in comparison with asymptomatic (indeterminate) patients. This apparently paradoxical role for IFN-gamma in human Chagas'disease is discussed in terms of the possibility of a temporal difference in IFN-gamma production during the initial stages of the infection (acute phase) in the presence or absence of chemotherapy. The maintenance of an immune response with high levels of IFN-gamma production during the chronic phase of the infection may favor cure or influence the development of the cardiac form of the disease.
Asunto(s)
Humanos , Enfermedad de Chagas/inmunología , Enfermedad de Chagas/fisiopatología , Interferón gamma/fisiología , Enfermedad de Chagas/sangre , Leucocitos MononuclearesRESUMEN
The aim of this study is to evaluate the effects of parasite clearance on the immunological profile of peripheral blood mononuclear cells (PBMC) from chagasic patients submitted to specific drug therapy. PBMC were examined by flow cytometry and proliferative responsiveness to Trypanosoma cruzi-related stimuli. Three groups of patients were studied: not treated (NT), treated not cured (TNC) and cured (C). All data were compared to values from uninfected individuals (NI). NT displayed a lower percentage of CD3+ cells as compared to NI, while TNC and C had mean values that were between those from NI and NT. Infected patients had double the percent of CD3+ HLA-DR+ cells, independent of the efficacy of the treatment. Thus, absence of circulating parasites did not reduce T cell activation in Chagas' disease. NT displayed a higher percentage of CD5+ B cells as compared to NI, while TNC and C had mean values between those from NI and NT. In contrast to the phenotypic data, the in vitro mean proliferative responses to parasite-related stimuli of PBMC from C were reduced to the low mean levels observed in NI. These striking differences were statistically different from the high responses seen in NT and TNC. Our data suggest that proliferative responses of PBMC from C reflect immunological changes due elimination of parasite. However, successful treatment did not alter the levels of peripheral T cell activation.
Asunto(s)
Enfermedad de Chagas/tratamiento farmacológico , Enfermedad de Chagas/inmunología , Leucocitos Mononucleares/inmunología , Tripanocidas/uso terapéutico , Adulto , Anciano , Animales , Antígenos de Protozoos , Linfocitos B/inmunología , Antígenos CD5/metabolismo , Enfermedad de Chagas/parasitología , Humanos , Técnicas In Vitro , Activación de Linfocitos , Persona de Mediana Edad , Fenotipo , Linfocitos T/inmunología , Trypanosoma cruzi/inmunología , Trypanosoma cruzi/aislamiento & purificaciónRESUMEN
A balanced host-parasite interaction during Trypanosoma cruzi infection allows for the establishment of a chronic infection that can last for many years. T cells are a major element responsible for parasite specific and non-specific immunity during the complex immune response of the host. However, the subpopulations of T cells involved in the response, as well as the exact mechanisms through which those cells are activated or rendered unresponsive, are not well defined. It is known that co-stimulatory signals, some of which are mediated via CD28, are of critical importance in the triggering of appropriate T cell responses. In this study the authors performed double-labelling studies to determine the frequency of expression of CD28 by CD4+ and CD8+ T lymphocytes in the peripheral blood of patients with Chagas' disease. The results show that chagasic patients throughout the spectrum of chronic clinical forms of the infection have significantly higher mean frequencies of CD4+CD28- and CD8+CD28-T cells, as compared with non-chagasic individuals. Considering the importance of CD28 for T-cell activation, the observed down-regulation or loss of CD28 during infection may indicate a possible basis for observed immunoregulatory events or distinct stages of T-cell activation in this infection. Recent evidence from patients with HIV/AIDS indicates that CD28- cell populations are more likely to undergo apoptosis, and increased apoptosis has been observed in experimental Chagas disease.
Asunto(s)
Antígenos CD28/biosíntesis , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Enfermedad de Chagas/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Anticuerpos Monoclonales , Citometría de Flujo , Humanos , Activación de Linfocitos , Persona de Mediana EdadRESUMEN
Sera from patients chronically infected with Trypanosoma cruzi display antibodies that bind to epitopes of living trypomastigotes, known as lytic antibodies (LA), and are detected by a complement-mediated lysis test. Conventional serology antibodies (CSA) are also present in sera from patients with chronic infections but, in contrast to LA, are unable to recognize viable trypomastigotes. The presence of LA has been used as an important element in the criterion of cure in human Chagas' disease. Using flow cytometry technology, we introduced a new and sensitive immunomethod for the detection of anti-live trypomastigote membrane-bound antibodies. On the basis of serological tests (LA and CSA detection) and parasitological assays such as hemoculture (HE), patients were classified into the following groups: chronically infected untreated patients (NT) and treated not-cured patients (TNC), with positive HE and both LA and CSA in their sera; "dissociated" HE-negative patients (DIS), in whom LA was not detected whereas CSA were present; a group of cured HE-negative patients (CUR), who were both LA and CSA negative; and, as control, a group of non-chagasic individuals (NC). Sera from these patients were assayed by incubation with live bloodstream trypomastigotes, which were subsequently exposed to fluorescein isothiocyanate-conjugated anti-human immunoglobulin G. The parasites were then fixed, run in the cytometer, and identified on basis of their size and granularity gain adjustments. On the basis of experience with the complement-mediated lysis test, a level of 20% of parasites being fluorescein isothiocyanate fluorescence positive was used as a cutoff between effective and ineffective treatments. With this criterion, our results indicated that sera from NT and TNC patients were antibody positive whereas all sera from DIS, CUR, and NC patients did not contain membrane-bound antibodies. This new approach is a tool to easily identify anti-live T. cruzi membrane-bound antibodies that can be used to monitor the efficacy of Chagas' disease treatment.
Asunto(s)
Anticuerpos Antiprotozoarios/análisis , Enfermedad de Chagas/sangre , Enfermedad de Chagas/inmunología , Citometría de Flujo , Trypanosoma cruzi/inmunología , Adulto , Anciano , Animales , Anticuerpos Antiprotozoarios/sangre , Antígenos de Protozoos/inmunología , Enfermedad de Chagas/tratamiento farmacológico , Proteínas del Sistema Complemento/inmunología , Citometría de Flujo/métodos , Técnica del Anticuerpo Fluorescente Indirecta , Humanos , Inmunidad Innata , Masculino , Ratones , Persona de Mediana Edad , Pruebas Serológicas/métodos , Trypanosoma cruzi/crecimiento & desarrolloRESUMEN
A 24-kDa recombinant protein from Trypanosoma cruzi (rTc24) was evaluated by enzyme-linked immunosorbent assay (ELISA) and Western blot (immunoblot) tests to identify treated chagasic patients considered parasitologically cured on the basis of persistently negative tests of hemocultures and lytic antibodies. Some of these patients were termed dissociated because their sera, although negative by the complement-mediated lysis test, were positive by conventional serology. The negative lysis test indicates the absence of active infection after specific treatment, but this assay requires live and infectious parasites and cannot be used easily in a laboratory routine. Here we tested rTc24 by ELISA and Western blotting as an alternative for the complement-mediated lysis test. For the group of patients with active infection despite the treatment (uncured patients), all the sera tested recognized rTc24 in both tests. For the dissociated patients, approximately 80% of the sera did not react with rTc24 in the ELISA or in Western blots, in agreement with the negative complement-mediated lysis tests. Thus, the 24-kDa T. cruzi recombinant antigen, when used for initial trials to evaluate cure of chagasic patients submitted to specific treatment, will allow the identification of most, but not all, cases.
Asunto(s)
Enfermedad de Chagas/tratamiento farmacológico , Enfermedad de Chagas/inmunología , Proteínas Protozoarias/inmunología , Trypanosoma cruzi/inmunología , Animales , Anticuerpos Antiprotozoarios/sangre , Antígenos de Protozoos/química , Antígenos de Protozoos/genética , Western Blotting , Enfermedad de Chagas/parasitología , Enfermedad Crónica , Ensayo de Inmunoadsorción Enzimática , Estudios de Evaluación como Asunto , Humanos , Peso Molecular , Proteínas Protozoarias/química , Proteínas Protozoarias/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/inmunología , Tripanocidas/uso terapéutico , Trypanosoma cruzi/genéticaRESUMEN
Two soluble antigens isolated from T. cruzi were evaluated by ELISA for the diagnosis of chronic infection and assessment of cure after specific chemotherapy, namely, supernatants from parasite cell-cultures (SpAg) and trypomastigote excretory/secretory antigens (ESAg). Among the treated patients, a group defined as "dissociated" had been monitored for 3 to 10 years and displayed negative hemocultures and tests of trypomastigote complement-mediated lysis persistently negative although positive by conventional serology (indirect fluorescence with epimastigote, mainly). A negative lysis test indicates parasite elimination by the patient. Our ELISA results showed that among the non-treated chagasic controls the SpAg e ESAg detected 93% and 100%, of the cases, respectively. However, only 28% of sera from the dissociated group, considered as cured, were positive by ELISA using SpAg whereas all of such sera were negative using ESAg. Therefore ELISA with ESAg seems to be ideal to replace the complement-mediated lysis test with the aim to define cure after treatment of the chronic infection by T. cruzi.
Asunto(s)
Antígenos de Protozoos/sangre , Enfermedad de Chagas/diagnóstico , Trypanosoma cruzi/inmunología , Animales , Enfermedad de Chagas/sangre , Enfermedad de Chagas/tratamiento farmacológico , Ensayo de Inmunoadsorción Enzimática/métodos , Epítopos , Inducción de RemisiónRESUMEN
Immunoprecipitation of the purified 160-kDa complement regulatory protein of Trypanosoma cruzi by Chagas' disease patient sera was examined as a possible correlate of the complement-mediated lysis test and as an indicator of parasite clearance. The results presented demonstrate that assessment of the humoral response to this antigen is a useful indicator of parasite clearance and may be particularly helpful in the assessment of some patients for whom other serological tests produce ambiguous results.
Asunto(s)
Anticuerpos Antiprotozoarios/sangre , Enfermedad de Chagas/inmunología , Proteínas del Sistema Complemento/inmunología , Proteínas Protozoarias/inmunología , Trypanosoma cruzi/inmunología , Animales , Enfermedad de Chagas/parasitología , HumanosRESUMEN
The sensitivity of hemocultures, performed once or three times, was investigated in 52 patients in the chronic phase of Chagas disease. Modifications were introduced in the technique such as, processing the blood more rapidly, gently homogenizing the cultures and examining them after 120 days of culture. Our results show a high percent age of positivity i.e. 79% and 94% of patients submitted, respectively, to one or three tests. No significant differences related to the patients age were detected, which varied from 14 to 82 years old. Our results demonstrate that hemoculture is a sensitive method for the parasitological diagnosis of Chagas disease and is ideal for monitoring cure in treated patients.
Asunto(s)
Sangre/parasitología , Enfermedad de Chagas/parasitología , Trypanosoma cruzi/aislamiento & purificación , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Enfermedad Crónica , Femenino , Humanos , Masculino , Persona de Mediana Edad , Sensibilidad y EspecificidadRESUMEN
Whole blood preparations from patients with either the indeterminate (asymptomatic) or cardiac clinical forms of chronic Trypanosoma cruzi infection were analyzed by flow cytometry using double-labeling to identify subsets of circulating lymphocytes. Several significant differences were demonstrated between the blood lymphocyte profiles of chagasic patients and non-chagasic controls. Clear increase in the percentages and actual numbers of double-positive cells of the phenotype CD3+/HLA-DR+, as well as decrease in the percentage of CD45RA+/CD4+ and CD45RA+/CD8+ T cells, indicate greater numbers of activated T cells circulating in the blood of infected patients. Consistent parallel increases were seen also in the B lymphocyte subset which stained double-positive for CD19/CD5. There were no significant differences in the circulation of these chronic chagasic patients in the CD4:CD8 ratios. Also, no substantive phenotypic differences were observed in the lymphocyte populations between the two ends of the clinical spectrum (indeterminate versus cardiac) in chronic human Chagas' disease. These observations demonstrate that increased levels of activated T cells and CD5+ B cells are present in the circulation of people with chronic Chagas' disease. These are cell phenotypes that have been associated in other conditions with autoimmune, polyclonal, and hyperimmune responses. The specificities of these activated cells and the roles they may play in resistance or pathogenesis during chronic Chagas' disease need now to be determined.
Asunto(s)
Subgrupos de Linfocitos B/inmunología , Enfermedad de Chagas/inmunología , Activación de Linfocitos/fisiología , Subgrupos de Linfocitos T/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Antígenos CD/sangre , Relación CD4-CD8 , Enfermedad Crónica , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Antígenos HLA-DR/inmunología , Humanos , Persona de Mediana EdadRESUMEN
With the exception of assays for the detection of antibodies promoting complement-mediated lysis of Trypanosoma cruzi, serologic tests have generally failed to assess the effectiveness of chemotherapy for Chagas' disease. Conventional serology, although useful for the diagnosis of infection, is not capable of determining which patients have been cured. Here we demonstrate that a high proportion of antibodies detected by conventional serology (using fixed epimastigotes or trypomastigotes or crude extracts obtained therefrom) are directed against the carbohydrate residue galactosyl alpha 1- > 3 galactose (Gal alpha 1- > 3 Gal), a determinant also recognized by antibodies from noninfected healthy volunteers. In a study of 14 cured patients with long-term followup, we found that the persistently positive reactions detected using conventional serology were largely eliminated following immunoadsorption with melibiose. Because of their wide distribution among microorganisms of intestinal and pulmonary microflora, these carbohydrate determinants may keep stimulating lymphocytes previously primed by T. cruzi Gal alpha 1- > 3 Gal epitopes, thereby accounting for false-positive results in cured patients. Consistent with this proposition, enzyme-linked immunosorbent assays performed with two distinct T. cruzi antigen preparations that lack the Gal alpha 1- > 3 Gal epitope, namely purified GP57/51 and trypomastigote-shed antigens, were indeed capable of determining a cure after chemotherapy, albeit to a different degree. Collectively, the data indicate that conventional immunoassays prepared with highly specific T. cruzi antigens can be useful in the assessment of a cure after chemotherapy.
Asunto(s)
Anticuerpos Antiprotozoarios/sangre , Antígenos de Protozoos/inmunología , Enfermedad de Chagas/diagnóstico , Glicoproteínas/inmunología , Trypanosoma cruzi/inmunología , Enfermedad Aguda , Adsorción , Adulto , Animales , Especificidad de Anticuerpos , Enfermedad de Chagas/tratamiento farmacológico , Niño , Enfermedad Crónica , Reacciones Cruzadas , Cisteína Endopeptidasas , Ensayo de Inmunoadsorción Enzimática , Técnica del Anticuerpo Fluorescente , Estudios de Seguimiento , Humanos , Persona de Mediana Edad , Nifurtimox/uso terapéutico , Nitroimidazoles/uso terapéutico , Resultado del Tratamiento , Tripanocidas/uso terapéutico , Trypanosoma cruzi/químicaRESUMEN
Several studies have been done to analyse the relationship between the characteristics of the electrocardiogram (ECG) and mortality in the several stages of the disease, using different methods like multiple case and longitudinal studies. We analysed the ECG from the acute stage up to twenty years of follow-up (+/- 9 years) in 42 patients with Chagas' disease to determine their evolution and it's value like an index for therapeutic evaluation. The 42 patients (18 female, 24 males) were originally from the north of the State of Minas Gerais (Brazil) and the initial stage was mainly in the first two decades of age. All bad cardiac involvement and received full specific treatment. We utilized the following criteria for the ECG analyses: Modified Minnesota Code for Chagas' disease; WHO/ISFC TASK FORCE for inter ventricular conduction disturbances and Pieretti criteria for inactive electrical areas. We conclude that: a) The electrocardiogram changes tend to get worse with evolution into the chronic stage; b) The electrocardiogram is not a good index for therapeutic valuation.
Asunto(s)
Cardiomiopatía Chagásica/diagnóstico , Electrocardiografía , Enfermedad Aguda , Adolescente , Adulto , Distribución por Edad , Brasil/epidemiología , Cardiomiopatía Chagásica/epidemiología , Cardiomiopatía Chagásica/terapia , Niño , Preescolar , Enfermedad Crónica , Electrocardiografía/estadística & datos numéricos , Femenino , Estudios de Seguimiento , Humanos , Lactante , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Población Rural/estadística & datos numéricosRESUMEN
A complement-mediated lysis test (CoML) using living trypomastigotes was compared with conventional serological methods and with haemoculture. Over a 10 years follow-up period evidence was obtained which supported the view that chagasic patients, treated with nitroheterocyclic drugs, in whom CoML had reverted to negative, might be considered cured despite conventional serology remaining positive.
Asunto(s)
Anticuerpos Antiprotozoarios/análisis , Enfermedad de Chagas/inmunología , Trypanosoma cruzi/inmunología , Adulto , Animales , Enfermedad de Chagas/tratamiento farmacológico , Ensayo de Actividad Hemolítica de Complemento , Quimioterapia Combinada , Estudios de Seguimiento , Humanos , Nifurtimox/uso terapéutico , Nitroimidazoles/uso terapéutico , Tripanocidas/uso terapéuticoRESUMEN
Ketoconazole, an azole antifungic drug which is already in the market has also been demonstrated to be active against Trypanosoma cruzi experimental infections. In this paper we confirmed the drug effect and investigated its range of activity against different T. cruzi strains naturally resistant or susceptible to both standard drugs Nifurtimox and Benznidazole used clinically in Chagas disease. Moreover, we have shown that the association of Ketoconazole plus Lovastatin (an inhibitor of sterol synthesis), which has an antiproliferative effect against T. cruzi in vitro, failed to enhance the suppressive effect of Ketoconazole displayed when administered alone to infected mice. Finally, administration in chronic chagasic patients of Ketoconazole at doses used in the treatment of deep mycosis also failed to induce cure as demonstrated by parasitological and serological tests. The strategy of identify and test drugs which are already in the market and fortuitously are active against T. cruzi has been discussed.
Asunto(s)
Enfermedad de Chagas/tratamiento farmacológico , Cetoconazol/uso terapéutico , Adulto , Animales , Humanos , Cetoconazol/administración & dosificación , Lovastatina/uso terapéutico , Ratones , Nifurtimox/uso terapéutico , Nitroimidazoles/uso terapéutico , Trypanosoma cruzi/efectos de los fármacosRESUMEN
Patients with chronic Trypanosoma cruzi infections have peripheral auto-anti-idiotype (Id) T cells that proliferate on exposure to immunoaffinity-purified antibodies against T. cruzi epimastigote antigens (EPI). The responses of some patients' (group 1) peripheral blood mononuclear cells (PBMC) to anti-EPI antibodies from sera of patients with the cardiac form of Chagas' disease (Id-C) were inhibited by chloroquine, but responses of other patients' (group 2) PBMC to Id-C were not inhibited. PBMC responses of both group-1 and -2 patients to anti-EPI antibodies from asymptomatic (indeterminate) patients (Id-I) were inhibited by chloroquine, as were their responses to the antigens in EPI. Most patients (69%) in group 1 had indeterminate Chagas' disease, and 100% of the patients in group 2 had severe, cardiac or digestive Chagas' disease. Both the direct (chloroquine-insensitive) and indirect (processed) modes of stimulation by anti-EPI antibodies required adherent cells. In group 2 (direct stimulation), this requirement was met by exogenous IL-1, and neither anti-HLA-DR,DP(DQ) monoclonal antibody (mAb) nor sodium azide inhibited T-cell proliferation. Indirect Id stimulation of group-1 cells by Id-I or Id-C, and group-2 cells by Id-I or EPI, was inhibited by anti-HLA-DR,DP(DQ) mAb or sodium azide, and exogenous IL-1 alone did not support this processed, MHC-mediated T-cell stimulation, but live adherent cells did. The mode of activation of auto-anti-Id T cells from patients with Chagas' disease depends on the clinical form of infection of both the cell donor and the donor of the stimulating anti-EPI antibodies.
Asunto(s)
Anticuerpos Antiidiotipos/inmunología , Cardiomiopatía Chagásica/inmunología , Enfermedad de Chagas/inmunología , Activación de Linfocitos/inmunología , Linfocitos T/inmunología , Adulto , Animales , Anticuerpos Antiprotozoarios/inmunología , Células Presentadoras de Antígenos/inmunología , Antígenos de Protozoos/farmacología , Cloroquina/farmacología , Humanos , Interleucina-2/farmacología , Leucocitos Mononucleares/efectos de los fármacos , Trypanosoma cruzi/inmunologíaRESUMEN
Trypanosoma cruzi antigenic specificities involved in human T-cell and antibody responses were compared in chronic chagasic patients affected with cardiomyopathy (C) or with the indeterminate form (I), the asymptomatic form of chronic Chagas' disease. T-cell Western blotting (immunoblotting) was performed to identify the most active antigens in epimastigote extracts (EPI-Ag). The patterns of peripheral blood mononuclear cell (PBMC) and T-cell proliferative responses induced by fractions blotted to nitrocellulose were heterogeneous, but the computation of their frequency distribution disclosed some important antigen specificities. Molecules ranging from 100 to 150 kilodaltons (kDa) were frequently stimulatory to PBMC from I patients (5 of 8 cases) and were less so when confronted with C patient (1 of 7 cases) lymphocytes. In contrast, both groups of patients actively responded to fractions ranging from 48 to 57 and 28 to 32 kilodaltons (kDa). The Western immunoblotting patterns of antibody reactivity displayed by 17 C and 15 I patients were also similar, yielding outstanding staining in the molecular mass ranges of 70 to 80 and 43 to 57 kDa. The latter antigen complex was recognized by 100% of the 32 chronic Chagas' disease serum specimens tested and closely corresponded to the migratory position recognized by T cells of most patients tested. The identification of the active molecules contained in the 43- to 57-kDa region was sought, with a focus on GP57/51, an antigen with well-established serodiagnostic properties. Immunoblotting analysis of EPI-Ag with a monoclonal antibody to GP57/51 confirmed its presence within the predicted molecular weight region. Highly purified GP51 was then used to demonstrate directly its capacity to promote specific PBMC proliferative responses in vitro. Data computed from a survey with 12 patients have shown a linear correlation (r = 0.93) between PBMC responses to EPI-Ag and to purified GP51, suggesting that the immune response to this particular glycoprotein may be an important component of human immune responses against T. cruzi.
Asunto(s)
Anticuerpos Antiprotozoarios/inmunología , Antígenos de Protozoos/inmunología , Enfermedad de Chagas/inmunología , Linfocitos T/inmunología , Trypanosoma cruzi/inmunología , Animales , Western Blotting , Glicoproteínas/inmunología , Humanos , Inmunidad Celular , Activación de Linfocitos , Peso MolecularRESUMEN
Hemoculture tests, a method for the detection of Trypanosoma cruzi, were used to investigate the effects of the anticoagulants heparin or EDTA on the parasite growth in culture medium (liver infusion tryptose, LIT). Hemocultures from 13 patients with positive serology for chronic Chagas' disease performed in parallel with both anticoagulants resulted in a total of seven (54%) positive hemocultures, three positive with blood samples collected with EDTA (23%), two with heparin (15%) and two with both anticoagulants (15%). There was no significant difference between the number of positive tubes in blood samples collected with either heparin (11%) or with EDTA (13%), an indication that heparin does not block the growth of T. cruzi. However, the simultaneous use of both anticoagulants may improve the positivity index of the hemocultures.