FIG 3.

The location of OVA in blood-stage parasites influences the development of OVA-specific T-cell responses in the spleen. A total of 10,000 naive CD45.1⁺ OT-I and 250,000 naive CD45.1⁺ OT-II cells were adoptively transferred into CD45.2⁺ C57BL/6 mice 1 day prior to infection (10⁴ pRBCs i.v.) with different P. berghei ANKA lines (GIMO_ANKA [control], OVA_hsp70 [OVA], OVA::mCherry_hsp70 [OVA::mC], and OVA::Hep17_hep17 [OVA::Hep17]). (A) Representative flow cytometric plots of total and donor CD4⁺ and CD8⁺ T cells in spleens of mice infected with different P. berghei ANKA lines at day 6 after infection. Hierarchal gating was performed on total splenic CD4⁺ and CD8⁺ T cells (top row), host CD45.1⁻ CD4⁺ T cells and donor OVA-specific CD45.1⁺ CD4⁺ OT-II cells (middle row), and host CD45.1⁻ CD8⁺ T cells and donor OVA-specific CD45.1⁺ CD8⁺ OT-I cells (bottom row). (B) Percentages and absolute numbers (means and standard errors of the means [SEM]) of total and donor (OT-I and OT-II) T cells in spleens of mice (n = 4) infected with different P. berghei ANKA lines at day 6 after infection. The magnitudes of the total splenic T-cell responses were comparable in mice infected with the different parasite lines (left 4 graphs). Stronger OVA-specific T-cell responses were observed in mice infected with OVA::Hep17 parasites than in mice infected with OVA::mC and OVA parasites (right 4 graphs). The results are representative of 2 separate experiments. ND, not detected; *, P < 0.05; **, P < 0.01; ***, P < 0.001. (C) Percentages and absolute numbers (means and SEM) of donor (OT-I and OT-II) T cells in spleens of mice (n = 4) infected with different P. berghei ANKA lines at day 6 after infection. The activation of CD4⁺ CD45.1⁺ OT-II cells (top row) and CD8⁺ CD45.1⁺ OT-I cells (bottom row) was determined by examining the expression of CD69, CD25, Ki67, CD44, and CD62L. The results are representative of 2 separate experiments. *, P < 0.05; **, P < 0.01; ***, P < 0.001 (by one-way ANOVA with Tukey's post hoc analysis).