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Many studies have demonstrated that monocyte-derived macrophages display critical activities in immunity to parasites. The ability of these cells to process and present antigens, produce cytokines, and provide costimulatory signals demonstrates their pivotal role in initiating immune responses. Although potential modulatory function has been attributed to monocytes from patients with Chagas' disease, a systematic phenotypic and functional analysis of these cells has not been performed. In this work, we analyzed the ex vivo expression of important surface molecules (CD11b and HLA-DR) and immunoregulatory cytokines (interleukin-10 [IL-10], IL-12 and tumor necrosis factor alpha [TNF-␣]) in CD14؉ and CD14 ؊ monocytes from Chagas' disease patients with polar clinical forms of the disease: indeterminate or severe cardiac. We also evaluated the influence of in vitro infection with T. cruzi in the expression of such molecules. We observed that monocytes from indeterminate-disease patients display lower levels of HLA-DR than those from noninfected individuals both ex vivo and after in vitro infection with T. cruzi. Although ex vivo expression of CD11b was similar among the groups, in vitro infection led to decreased expression of this molecule by monocytes from Chagas' disease patients but not from noninfected individuals. Analysis of the expression of immunoregulatory cytokines showed that while monocytes from indeterminate-disease patients are committed to IL-10 expression, a higher percentage of monocytes from cardiac-disease patients express TNF-␣ after exposure to live parasites. These results suggest that monocytes from indeterminate-disease patients display modulatory characteristics related to low HLA-DR and high IL-10 expression whereas monocytes from cardiac-disease, patients may be committed to induction of inflammatory responses related to high TNF-␣ expression.

Interactions between macrophages and lymphocytes through costimulatory molecules and cytokines are essential for mounting an efficient immune response and controlling its pathogenic potential. Here we demonstrate the immunomodulatory capacity of Trypanosoma cruzi, the causative agent of Chagas' disease, through its ability to induce differential expression of costimulatory molecules and cytokines by monocytes and T cells. Costimulatory molecule and cytokine modulation was evaluated using cells from noninfected individuals and from patients with the asymptomatic indeterminate form and those with the severe cardiac clinical form of Chagas' disease. Our results show that while exposure of monocytes to live T. cruzi leads to an increase in the frequency of CD80 ؉ monocytes in all groups, it decreases both the frequency and intensity of CD86 expression by monocytes from patients with the cardiac form but not from those with the indeterminate form. Conversely, exposure of lymphocytes to monocytes infected with T. cruzi increased the surface expression of cytotoxic-Tlymphocyte-associated antigen 4 (CTLA-4) by T cells from indeterminate but not from cardiac patients, compared to that from control patients. These data suggest that T. cruzi induces a potentially down-regulatory environment in indeterminate subjects, which is associated with higher CD80 and CTLA-4 expression. To test the functional importance of this modulation, we evaluated the expression of cytokines after in vitro infection. Although exposure of lymphocytes to parasite-infected monocytes induced high expression of inflammatory and anti-inflammatory cytokines by T cells in all groups, indeterminate patients displayed a higher ratio of monocytes expressing interleukin 10 than tumor necrosis factor alpha following infection than did controls. These data show the ability of T. cruzi to actively change the expression of costimulatory molecules and cytokines, suggesting molecular mechanisms for the differential clinical evolution of human Chagas' disease.

A discrete atomistic solid-on-solid model is proposed to describe dissolution of a crystalline solid by a liquid. The model is based on the simple assumption that the probability per unit time of a unit cell being removed is proportional to its exposed area. Numerical simulations in one dimension demonstrate that the model has very good scaling properties. After removal of only about 10(2) monolayers, independently of the substrate size, the etched surface shows almost time-independent short-range correlations and the receding surface presents the Family-Vicsek scaling behavior. The scaling parameters alpha=0.491+/-0.002 and beta=0.330+/-0.001 indicate that the system belongs to the Kardar-Parisi-Zhang universality class. The imposition of periodic boundary conditions on the simulations reduces the effective system size by a factor of 0.68 without changing the exponents alpha and beta. Surprisingly, the periodic condition changes drastically the statistics of the surface height fluctuations and the short-range correlations. Without periodic conditions, that statistics is, up to 3 standard deviations, an asymmetric Lévy distribution with mu=1.82+/-0.01, and outside this region the statistics is Gaussian. With periodic conditions, that statistics is Gaussian, except for large negative fluctuations.

No abstract

SUMMARYChronic human Chagas' disease ranges from an asymptomatic to a severe cardiac clinical form. The involvement of the host's 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-g , TNF-a , 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-g , TNF-a 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-a in cardiac and with IL-10 in indeterminate CP, suggesting that these cells might have an important regulatory role in human Chagas' disease.

Pretreatment of macrophages with Toll-like receptor (TLR)2 or TLR4 agonists leads to a stage of cell hyporesponsiveness to a second stimulation with TLR agonists. This tolerance state is accompanied by the repression of IL-1 receptor-associated kinase-1, mitogen-activated protein kinases, and IκB phosphorylation and expression of genes encoding proinflammatory cytokines, like IL-1β and TNF-α. In this report, we demonstrated that mucin-like glycoprotein (tGPI-mucin) of Trypanosoma cruzi trypomastigotes (TLR2 agonist) and LPS (TLR4 agonist) induce cross-tolerance in macrophages and we addressed the role of phosphatase activity in this process. Analysis of the kinetic of phosphatase activity induced by tGPI-mucin or LPS revealed maximum levels between 12 and 24 h, which correlate with the macrophage hyporesponsiveness stage. The addition of okadaic acid, an inhibitor of phosphatase activity, reversed macrophage hyporesponsiveness after exposure to either LPS or tGPI-mucin, allowing phosphorylation of IL-1R-associated kinase-1, mitogen-activated protein kinases, and ΙκB and leading to TNF-α gene transcription and cytokine production. Furthermore, pretreatment with either the specific p38/stress-activated protein kinase-2 inhibitor (SB203580) or the NF-κB translocation inhibitor (SN50) prevented the induction of phosphatase activity and hyporesponsiveness in macrophage, permitting cytokine production after restimulation with LPS. These results indicate a critical role of p38/stress-activated protein kinase-2 and NF-κB-dependent phosphatase in macrophage hyporesponsiveness induced by microbial products that activate TLR2 and TLR4.

Here, we analysed the use of Vb-TCR regions by CD4 and CD8 T cells from acute and chronic chagasic patients using¯ow cytometry. We determined the Vb expression in cells freshly isolated from patients, as well as after in vitro stimulation with antigens derived from epimastigote (EPI) or trypomastigote (TRYPO) forms of Trypanosoma cruzi. Analysis of Vb-TCR expression of T cells freshly isolated from patients showed a decrease in Vb5 expression in the CD4 T-cell population from acutely infected individuals, whereas CD4 Vb5 T cells were found to be increased in chronic patients with the cardiac, but not indeterminate, clinical form. After culturing peripheral blood mononuclear cells (PBMC) from chronic patients with EPI or TRYPO, we found that both antigenic preparations led to a preferential expansion of CD4 Vb5 T cells. EPI stimulation also led to the expansion of CD8 Vb5 T cells, whereas TRYPO led to the expansion of this cell population only if PBMC were from cardiac and not indeterminate patients. We observed that TRYPO stimulation led to an increase in the frequency of CD4 Vb17 T cells in cultures of PBMC from indeterminate patients, whereas an increase in the frequency of CD8 Vb17 T cells was found upon TRYPO stimulation of PBMC from cardiac patients. Despite this increase in the frequency of Vb17 T-cell populations upon TRYPO stimulation, the same antigenic preparation led to a much higher expansion of Vb5 T cells. These results show a differential expression of Vb5-TCR in cells freshly isolated from chagasic patients in different stages of the disease and that parasite-speci®c antigens stimulate a portion of the T-cell repertoire with preferential usage of Vb5-TCR.