Chronic human infection with Trypanosoma cruzi drives CD4+ T cells to immune senescence

Abstract

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.

FIGURE 1

A decrease in the frequencies of T. cruzi-specific IFN-γ-secreting CD4⁺ and CD8⁺ T cells is associated with a more severe clinical status in chronic T. cruzi infection. PBMCs were cultured for 16–20 h with T. cruzi lysate or medium alone. Intracellular and surface markers were stained after fixation and permeabilization of cells. Lymphocytes were gated in side scatter vs forward scatter light. The number of T. cruzi-specific T cells was determined by subtracting the percentage of IFN-γ⁺ T cells in unstimulated cultures from the percentage of IFN-γ⁺ cells upon stimulation with T. cruzi lysate. Bars represent the mean percentages of T. cruzi-specific CD4⁺IFN-γ⁺ and CD8⁺IFN-γ⁺ T cell responses; error bars represent SD. *, p < 0.05 compared with G1–G3 and controls.

FIGURE 2

T. cruzi-specific CD4⁺ T cells in chronically infected subjects are primarily recently recruited T cells. The CD4 IFN-γ double-positive compartment was analyzed for the expression of CD27, CD28, and CD57 by flow cytometry (A). Bars represent the mean frequencies of CD4⁺ IFN-γ⁺ T cells expressing CD27 and CD28 in chronically T. cruzi-infected subjects; error bars indicate SD. Number of subjects studied in each patient group: G0 = 7, G1 = 7, G2 = 2, and G3 = 4. B, CD57 expression on CD4⁺IFN-γ⁺ T cells in the different clinical stages of the disease. Median values are represented by horizontal lines. C and D, Correlation between the expression of CD27, CD28, and CD57 on T. cruzi-specific IFN-γ-producing CD4⁺ T cells was evaluated by the Spearman correlation test.

FIGURE 3

The total peripheral CD4⁺ T cell population displays features of immune senescence in subjects with chronic T. cruzi infection. PBMCs were stained for CD4, CD45RA, CD27, CD28, CD57, and caspase 3. Lymphocytes were gated in side scatter vs forward scatter light. Each point represents the expression of CD57 (A and C) and caspase 3 (B and D) on total terminally differentiated effector (CD45RA⁺CD27⁻CD28⁻) (A and B) and naive-like (CD45RA⁺CD27⁺CD28⁺ (C and D) CD4⁺ T cells. Median values are represented by horizontal lines.

FIGURE 4

IL-7R expression in effector CD4⁺ T cells in chronic Chagas disease patients. PBMCs were stained for CD4, CD45RA, CD28, and IL-7R. Lymphocytes were gated in side scatter vs forward scatter light and analyzed by flow cytometry. Each point represents the expression of IL-7R on the total CD4⁺CD45RA⁺CD28⁻ T cell population. Median values are represented by horizontal lines.