Manoel Otávio da Costa Rocha

Elevated Concentrations of CCL2 and Tumor Necrosis Factor—α in Chagasic Cardiomyopathy

Chronic myocarditis is the main pathological finding associated with Chagas disease-related morbidity. Concentrations of CCL2, CCL3, tumor necrosis factor (TNF)-alpha, and brain natriuretic peptide (BNP) were evaluated in plasma samples obtained from patients with different clinical forms of chronic chagasic cardiomyopathy. Patients with more-severe Chagas disease had elevated plasma concentrations of TNF-alpha, CCL2, and BNP, and there was a good correlation between levels of these proteins (especially TNF-alpha ) and the degree of heart dysfunction. Indeed, TNF-alpha level was an excellent predictor of heart failure. Peripheral blood mononuclear cell samples obtained from patients with mild or severe chagasic cardiomyopathy produced greater amounts of TNF-alpha and CCL2 than did those obtained from noninfected individuals. The elevation of TNF-alpha and CCL2 levels in the plasma of patients appears to be secondary to the degree of heart dysfunction, whereas spontaneous production of TNF-alpha and CCL2 by mononuclear cells is secondary not only to heart dysfunction, but also to the underlying inflammation in the heart of chagasic patients. Measurement of the TNF-alpha level could be a useful tool in the identification of patients with heart dysfunction who may benefit from further investigation and treatment.

Potential role of CD4+CD25HIGH regulatory T cells in morbidity in Chagas disease.

Several immunoregulatory mechanisms are proposed to be effective both in human and experimental Trypanosoma cruzi infection. However, the role of CD4+CD25high T cells in Chagas disease has not yet been elucidated. These cells are critical for the regulation of immune response to infectious agents and in the control of autoimmune diseases. In this study, the presence of CD4+CD25high regulatory T cells in the whole blood of non-infected individuals (NI), and patients with the indeterminate (IND) and cardiac form (CARD) of Chagas disease was evaluated. To further characterize this population of regulatory cells, the co-expression of CTLA-4, CD62L, CD45RO, CD45RA, HLA-DR, CD40L, CD69, CD54, IL-10R and the intracellular molecules FOXP3 and IL-10 on the CD4+CD25high T lymphocytes was examined. FOXP3 was expressed by the majority of CD4+CD25high when compared with the other CD4+ T cells subsets in patients with Chagas disease. Patients with the IND form of the disease had a higher frequency of circulating regulatory CD4+CD25high T cells than patients with the CARD form. Moreover, there was an increase in CD4+CD25highFOXP3+ cells that were also IL-10+ in the IND group whereas, in the CARD group, there was an increase in the percentage of CD4+CD25high FOXP3+ cells that expressed CTLA-4. These data suggest that IL-10 produced by regulatory T cells is effective in controlling disease development in patients with the IND form. However, in individuals with the CARD form of the disease, the same regulatory mechanism, mediated by IL-10 and CTLA-4 expression is not sufficient to control the progression of the disease. The data suggest that CD4+CD25highFOXP3+ regulatory T cells in patients with Chagas disease might play a role in the immune response against T. cruzi infection although with distinct effects in patients with the IND and CARD forms of disease.

Chemokine Receptor Expression on the Surface of Peripheral Blood Mononuclear Cells in Chagas Disease

We evaluated the expression of chemokine receptors (CCR1, CCR2, CCR5, and CXCR4) on the surface of peripheral blood mononuclear cells obtained from patients with chronic chagasic cardiomyopathy (CCC) and noninfected individuals. Only CCR5 and CXCR4 expression was different on the surface of the subsets (CD4, CD8, and CD14) evaluated. Patients with mild CCC had elevated leukocyte expression of CCR5, compared with noninfected individuals or those with severe disease. CXCR4 expression was lower on leukocytes from patients with severe CCC. The differential expression of both receptors on leukocytes of patients with CCC was consistent and clearly correlated with the degree of heart function such that the lower the heart function, the lower the expression of either CCR5 or CXCR4. These results highlight the possible participation of the chemokine system in early forms of chagasic cardiomyopathy and the relevance of heart failure-induced remodeling in modifying immune parameters in infected individuals.

Predictors of Mortality in Patients with Dilated Cardiomyopathy: Relevance of Chagas Disease as an Etiological Factor

INTRODUCTION AND OBJECTIVES Previous studies on the influence of Chagas disease on mortality in patients with heart failure were limited by the heterogeneity of the patient populations. Few data are available on the association between the underlying cause of dilated cardiomyopathy and long-term prognosis. The aims of this study were to identify risk factors for mortality in patients with dilated cardiomyopathy that was either secondary to Chagas disease or idiopathic and to determine the prognostic value of identifying Chagas disease as the underlying etiology of dilated cardiomyopathy. METHODS We investigated outcomes in 287 patients with heart failure secondary to dilated cardiomyopathy. Patients were divided into two groups according to the underlying etiology: Chagas cardiomyopathy (224 patients) and idiopathic dilated cardiomyopathy (63 patients). The study end-points were death and heart transplantation. RESULTS Over a median follow-up period of 39.5 months, 104 patients died and 9 underwent heart transplantation. Under multivariate Cox proportional hazards analysis, New York Heart Association functional class, left ventricular ejection fraction, right ventricular function and left atrial volume remained predictors of an adverse outcome. Chagas etiology was also independently associated with a poor prognosis (hazard ratio=2.48; 95% confidence interval, 1.28-4.78; P=.007) compared with idiopathic disease, after adjustment for other well-established predictive parameters in heart failure. CONCLUSIONS The identification of Chagas etiology in patients with dilated cardiomyopathy was of prognostic significance. Chagas cardiomyopathy was associated with poorer survival compared with idiopathic disease, irrespective of other clinical and echocardiographic parameters related to a poor prognosis in heart failure.

Prognostic Value of Signal-Averaged Electrocardiogram in Chagas Disease

Background: The value of signal‐averaged ECG (SAECG) in the risk stratification of Chagas disease (ChD), a potentially lethal illness prevalent in Latin America, remains controversial. The aim of this prospective longitudinal study was to determine the prognostic value of SAECG in ChD, using multivariate models with other established prognostic predictors, and to develop a simple prediction risk score.

Phenotypic and functional characteristics of CD28+ and CD28− cells from chagasic patients: distinct repertoire and cytokine expression

Chronic human Chagas’ disease ranges from an asymptomatic to a severe cardiac clinical form. The involvement of the hosts immune response in the development and maintenance of chagasic pathology has been demonstrated by several groups. We have shown that activated T‐cells lacking CD28 expression are increased in the peripheral blood of chagasic patients (CP), suggesting a relationship between these cells and disease. In order to better characterize this cell population, determining their possible role in immunoregulation of human Chagas’ disease, we evaluated the expression of TCR‐Vbeta regions 2, 3·1, 5, 8 and 17, as well as the expression of IFN‐γ, TNF‐α, IL‐4 and IL‐10 by CD28+ and CD28− cells from polarized indeterminate and cardiac CP. Flow cytometric analysis demonstrated equivalent TCR‐Vbeta usage between CD4+CD28+ and CD4+CD28− cells from all groups (chagasic and healthy controls). However, there was a predominance of Vbeta5 expression in the CD28+ and CD28− populations in the CP groups (indeterminate and cardiac). Interestingly, CD8+CD28− cells from CP, but not from nonchagasic individuals, displayed a reduced frequency of most analysed Vbetas when compared with the CD8+CD28+ subpopulation. Comparison of V‐beta expression in CD28+ or CD28− cell populations among individuals from different groups also showed several interesting differences. Functionally, cardiac CP displayed a higher frequency of IFN‐γ, TNF‐α and IL‐4 producing lymphocytes than indeterminate CP. Correlation analysis between the frequency of cytokine expressing cells, and the frequency of CD4+ T‐cells with differential expression of CD28 demonstrated that CD4+CD28− T‐cells were positively correlated with TNF‐α in cardiac and with IL‐10 in indeterminate CP, suggesting that these cells might have an important regulatory role in human Chagas’ disease.

Foxp3+CD25high CD4+ regulatory T cells from indeterminate patients with Chagas disease can suppress the effector cells and cytokines and reveal altered correlations with disease severity

Immunoregulatory mechanisms are important to control the intense immune activity induced in Chagas disease. We evaluated the phenotypic profile and the mechanisms by which Treg cells function in patients with the indeterminate (IND) and cardiac (CARD) clinical forms of Chagas disease. The frequency of Foxp3(+)CD25(high) CD4(+)-T cells is augmented and correlated with the maintenance of a better cardiac function in IND. Treg cells from IND present suppressive activity, although the mechanism is not IL-10 or CTLA-4 dependent and are able to produce augmented levels of IL-17, IL-10 and granzyme B being its frequency correlated with percentage of Annexin V(+) CD4(+)-cells. In contrast, CARD presents higher frequency of IL-6(+), IFN-gamma(+), TNF-alpha(+) and CTLA-4(+) Treg-cells than IND. Thus, our data suggest that Treg cells have an important role in controlling the exacerbated immune response and morbidity in Trypanosoma cruzi infection, probably modulating the cytokine environment and/or killing effector cells.

Plasma cytokine expression is associated with cardiac morbidity in chagas disease.

The expression of immune response appears to be associated with morbidity in Chagas disease. However, the studies in this field have usually employed small samples of patients and statistical analyses that do not consider the wide dispersion of cytokine production observed in these patients. The aim of this study was to evaluate the plasma cytokine levels in well-defined clinical polar groups of chagasic patients divided into categories that better reflect the wide cytokine profile and its relationship with morbidity. Patients infected with Trypanosoma cruzi (T. cruzi) were grouped as indeterminate (IND) and cardiac (CARD) forms ranging from 23 to 69 years of age (mean of 45.6±11.25). The IND group included 82 individuals, ranging from 24 to 66 years of age (mean of 39.6±10.3). The CARD group included 94 patients ranging from 23 to 69 years of age (mean of 48±12.52) presenting dilated cardiomyopathy. None of the patients have undergone chemotherapeutic treatment, nor had been previously treated for T. cruzi infection. Healthy non-chagasic individuals, ranging from 29 to 55 years of age (mean of 42.6±8.8) were included as a control group (NI). IND patients have a higher intensity of interleukin 10 (IL-10) expression when compared with individuals in the other groups. By contrast, inflammatory cytokine expression, such as interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), and interleukin 1 beta (IL-1β), proved to be the highest in the CARD group. Correlation analysis showed that higher IL-10 expression was associated with better cardiac function, as determined by left ventricular ejection fraction and left ventricular diastolic diameter values. Altogether, these findings reinforce the concept that a fine balance between regulatory and inflammatory cytokines represents a key element in the establishment of distinct forms of chronic Chagas disease.

Ischemic cerebrovascular events in patients with Chagas cardiomyopathy: A prospective follow-up study

BACKGROUND Chagas disease cardiomyopathy is a common form of dilated cardiomyopathy worldwide, and an important cause of stroke in Latin America. The long-term cumulative risk of ischemic cerebrovascular event (ICE) and its relation to left ventricular (LV) dysfunction have not been determined. The aims of this study were to describe the incidence and to evaluate the effect of LV ejection fraction on the risk for ICE in patients with Chagas cardiomyopathy. METHODS A total of 213 consecutive patients with Chagas disease and LV systolic dysfunction, 131 males, mean age 48+/-12 years, were prospectively enrolled. The use of anticoagulation was based on clinical indications. The end point was ICE, which included fatal or nonfatal stroke and transient ischemic attack. Risk factors for events were assessed by Cox proportional-hazards analysis. RESULTS Mean follow-up was 36 months; 69 patients died and seven underwent cardiac transplantation. The overall incidence of ICE was 2.67 events per 100 patient/years. Independent risk factors for ICE included LV ejection fraction (HR 0.95, 95% CI 0.91 to 0.99, p=0.009) and left atrial volume corrected for body surface area (HR 1.04, 95% CI 1.01 to 1.07, p=0.007), which persisted after adjustment for anticoagulation use. Patients with ejection fractions<or=35% had a relative risk of events of 3.41, as compared with the other patients. The presence of apical aneurysm or thrombus, age, and history of previous stroke were not an independent long-term risk for ICE. CONCLUSIONS In patients with Chagas cardiomyopathy, the annual incidence of ICE is low. Left ventricular ejection fraction and indexed left atrial volume appear to be independently associated with these events.

Brain natriuretic peptide and left ventricular dysfunction in chagasic cardiomyopathy

Global left ventricular (LV) systolic dysfunction is the strongest predictor of morbidity and mortality in Chagas disease. Echocardiography is considered the gold standard for the detection of LV dysfunction, but not always available in endemic areas where chagasic cardiomyopathy is most common. Brain natriuretic peptide (BNP) is a neurohormone that has been recently described as a simple and inexpensive diagnostic and prognostic marker for patients with congestive heart failure. Chagasic patients (n = 63) and non-infected healthy individuals (n = 18) were recruited prospectively and underwent complete clinical examination, echocardiography and 24-h Holter monitoring. BNP was measured from thawed plasma samples using the Triage BNP test. We observed high levels of BNP in association with depression of LV ejection fraction, with increase of LV end-diastolic diameter and with LV premature complexes. An elevated concentration of BNP, defined as a concentration of 60 pg/ml or more, had a sensitivity of 91.7%, specificity of 82.8%, positive predictive value of 52.4%, and negative predictive value of 98% for detecting LV dysfunction (LV ejection fraction < 40%).BNP measurement using a simple, relatively inexpensive and rapid test has a promising role in identifying LV dysfunction associated with chagasic cardiomyopathy. Equally important, patients with Trypanosoma cruzi infection who have low levels of BNP level in plasma have a very low likelihood of severe cardiac involvement, and echocardiography is probably not necessary.