Generation of Trypanosoma cruzi-specific CD8+ T-cell immunity is unaffected by the absence of type I interferon signaling

Abstract

Trypanosoma cruzi is a protozoan parasite that causes human Chagas' disease, a leading source of congestive heart failure in Central and South America. CD8+ T cells are critical for control of T. cruzi infection, and CD8+ T cells recognizing the immunodominant trans-sialidase gene-encoded peptide TSKB20 (ANYKFTLV) account for approximately 30% of the total CD8+ T-cell population at the peak of infection in C57BL/6 mice. Type I interferons (IFN-I) are pleiotropic cytokines that play a critical role in both innate and adaptive immunity against a variety of infections, but their induction and their role in infection are dictated by the infectious agent. Because type I IFNs and IFN-responsive genes are evident early after T. cruzi infection of host cells, we examined the influence of IFN-I on the development of CD8+ T-cell responses during this infection. Mice lacking the receptor for IFN-I (IFNARKO) and their wild-type counterparts both developed chronic infections and generated similar frequencies of immunodominant TSKB20- and subdominant TSKB18-specific CD8+ T cells following T. cruzi infection. In contrast, peak TSKB20-specific CD8+ T-cell responses generated during infection with vaccinia virus engineered to express TSKB20 were approximately 2.5-fold lower in IFNARKO mice than B6 mice, although after viral clearance, the frequencies of TSKB20-specific CD8+ T cells stabilized at similar levels. Together, these data suggest that IFN-I induction and biology are dependent upon the microbial context and emphasize the need to investigate various infection models for a full understanding of CD8+ T-cell development.

FIG. 1.

Type I interferons (IFN-I) are not required for generation of T. cruzi-specific CD8⁺ T cells. Eight-week-old B6 (triangles) and IFNARKO (circles) mice were infected intraperitoneally (i.p.) with 1,000 Brazil strain tissue culture trypomastigotes. Whole blood from serially bled mice was stained with H-2K^b MHC-I tetramers bearing TSKB20 (closed symbols) or TSKB18 (open symbols), anti-CD8, and an exclusion channel containing PE-Cy5.5-labeled anti-CD4, anti-B220, and anti-CD11b. Values represent the mean of 8 mice per group with P > 0.1 for all time points.

FIG. 2.

Peptide-specific IFN-γ-producing CD8⁺ T cells are generated in the absence of IFN-I signaling following T. cruzi infection. Spleen cells were harvested from uninfected B6 (left column), wild-type B6 (middle column), and IFNARKO (right column) mice at 180 days after infection with 1,000 Brazil strain T. cruzi parasites. Frequencies of IFN-γ-producing CD8⁺ T cells were determined by intracellular cytokine staining after 5 h of in vitro stimulation with T. cruzi peptides as described in Materials and Methods. Values represent the percentage of IFN-γ⁺ cells among CD8⁺ T cells and are representative of a total of 4 mice. P = 0.83 for TSKB20-stimulated IFN-γ production, and P = 0.36 for TSKB18-stimulated IFN-γ production.

FIG. 3.

The phenotype of T. cruzi-specific CD8⁺ T cells is unaltered in IFNARKO mice. Spleen cells were harvested from uninfected B6 (left column), wild-type B6 (middle column), and IFNARKO (right column) mice at 180 days after infection with 1,000 Brazil strain T. cruzi parasites. Surface staining with MHC-I tetramers and fluorochrome-conjugated antibodies was performed as described in Materials and Methods. (A) MHC-I tetramer staining of CD8⁺ T cells from uninfected mice, T. cruzi-infected B6 mice, or T. cruzi-infected IFNARKO mice. For B6 mice, the percentage of CD8⁺ IFN-γ⁺ cells was 3.1% ± 1.7%; for IFNARKO mice, the percentage of CD8⁺ IFN-γ⁺ cells was 3.4% ± 1.8% (P = 0.83; n = 4). Data are representative of two experiments. (B) Expression of CD62L (y axis) and CD44 (x axis) on total CD8⁺ T cells (top) or TSKB20-specific CD8⁺ T cells (bottom). (C) Expression of KLRG-1 (y axis) and CD44 (x axis) on total CD8⁺ T cells (top) or TSKB20-specific CD8⁺ T cells (bottom). The data presented show phenotyping for representative mice for each group (n = 5 or 6 mice per group).

FIG. 4.

IFNARKO mice develop lower peak TSKB20-specific responses following TSKB20-VV infection. Eight week old female B6 (triangles), male B6 (open circles), and male IFNARKO (diamonds) mice were infected i.p. with 2 × 10⁶ PFU TSB20-VV. Because of the dearth of female IFNARKO mice in the colony, we used all males in the IFNARKO group and compared them to both male and female B6 mice. Mice were serially bled through the tail vein at the designated times, and whole blood was stained with H-2K^b MHC-I tetramers bearing TSKB20, anti-CD8, and an exclusion channel containing PE-Cy5.5-labeled anti-CD4, anti-B220-, and anti-CD11b. Values represent the mean of 4 to 6 mice per group and are representative of 2 experiments. P is <0.05 for time points before 30 days postinfection.

FIG. 5.

Peptide-specific IFN-γ-producing CD8⁺ T cells are generated in the absence of IFN-I signaling following TSKB20-VV infection. Spleen cells were harvested from uninfected B6 mice (left column) and B6 (middle column) and IFNARKO (right column) mice infected i.p. with 2 × 10⁶ PFU TSB20-VV. (A) MHC-I tetramer staining of CD8⁺ T cells from uninfected mice, T. cruzi-infected B6 mice, or T. cruzi-infected IFNARKO mice. For B6 mice, the percentage of CD8⁺ IFN-γ⁺ mice is 1.0% ± 0.8%; for IFNARKO mice, the percentage of CD8⁺ IFN-γ⁺ mice is 1.2% ± 0.6% (P = 0.83; n = 12 mice). The data in the figure show one representative mouse per group. (B) Frequencies of IFN-γ-producing CD8⁺ T cells determined by intracellular cytokine staining after 5 h of in vitro stimulation with T. cruzi peptides as described in Materials and Methods. Values represent the percentage of IFN-γ⁺ cells among CD8⁺ T cells and are representative of a total of 4 mice (P > 0.1). (C) Expression of CD62L (y axis) and CD44 (x axis) on total CD8⁺ T cells (top) or TSKB20-specific CD8⁺ T cells (bottom). The data presented show phenotyping for representative mice for each group (n = 5 mice per group).