Unique T cells with unconventional cytokine profiles induced in the livers of mice during Schistosoma mansoni infection

Abstract

During infection with Schistosoma, serious hepatic disorders are induced in the host. The liver possesses unique immune systems composed of specialized cells that differ from those of other immune competent organs or tissues. Host immune responses change dramatically during Schistosoma mansoni infection; in the early phase, Th1-related responses are induced, whereas during the late phase Th2 reactions dominate. Here, we describe unique T cell populations induced in the liver of mice during the period between Th1- and Th2-phases, which we term the transition phase. During this phase, varieties of immune cells including T lymphocytes increase in the liver. Subsets of CD4(+) T cells exhibit unique cytokine production profiles, simultaneously producing both IFN-γ and IL-13 or both IFN-γ and IL-4. Furthermore, cells triply positive for IFN-γ, IL-13 and IL-4 also expand in the S. mansoni-infected liver. The induction of these unique cell populations does not occur in the spleen, indicating it is a phenomenon specific to the liver. In single hepatic CD4(+) T cells showing the unique cytokine profiles, both T-bet and GATA-3 are expressed. Thus, our studies show that S. mansoni infection triggers the induction of hepatic T cell subsets with unique cytokine profiles.

Figure 1. Schistosoma mansoni infection induced robust increase of immune competent cells in the liver.

(A) Hepatic cells isolated from 3 mice were pooled and the cell number was calculated. (B and C) Flowcytometric analysis was conducted with the liver lymphocytes prepared in (A) or the splenocytes. The percentages in (B) represent the proportions in CD3-positive population, and those in (C) express the proportions in population of (B). (A-C) each shows one representative result of three independent experiments.

Figure 2. Th1 cells were induced in early and Th2 cells in late phase in the liver.

Hepatic lymphocytes were isolated from S. mansoni-infected BALB/c mice at indicated time points, and their potential for producing IFN-γ, IL-4, or IL-13 was analyzed by ICS upon TCR ligation. Insets at the top represent one example using liver lymphocytes prepared at 4 weeks PI are shown. The percentages represent the proportions in CD4-positive population. Similar results were obtained in three independent experiments.

Figure 3. S. mansoni infection-induced hepatic T cells exhbited the potential to produce uncommon combinations of cytokines.

Hepatic lymphocytes were isolated from S. mansoni-infected mice at indicated time points, and the proportions and absolute numbers of γ4 and γ13 cells were investigated by ICS. (A) One example using hepatic lymphocytes prepared at 6 weeks PI is displayed. (B) The percentages represent the proportions in CD4-positive population. This experiment is representative of three independent experiments.

Figure 4. The unique hepatic T cells exhbited the ability to produce IFN-γ, IL-13, and IL-4 simultaneouly.

Hepatic lymphocytes were isolated from S. mansoni-infected mice at 6 weeks PI and ICS was conducted after TCR stimulation. γ4 cells within IL-13-producing CD4⁺ T cell population (A), γ13 cells within IL-4-producing CD4⁺ T cell population (B), and IL-4- and IL-13-secreting cells within IFN-γ-producing CD4⁺ T cell population (C) were analyzed. Data shown are a representative of five independent experiments.

Figure 5. CD4⁺ T cells doubly producing IFN-γ and IL-5 were rarely induced after S. mansoni infection.

Hepatic lymphocytes were isolated from S. mansoni-infected mice at 6 weeks PI and ICS was conducted upon TCR ligation. (A) One representative result is shown. The numbers in the insets represent the percentages of γ4, γ13, or γ5 in CD4-positive population. (B) Data represent the mean values + SD of three independent experiments.

Figure 6. Hepatic αβ T cells are the responsible cells showing the unconventional cytokine profiles.

Hepatic lymphocytes were isolated from S. mansoni-infected mice and flowcytometry was conducted after TCR ligation. (A) Expression levels of γδ TCR on CD4⁺ γ4 cells or γ13 cells were analyzed with the hepatic lymphocytes isoleted at 6 weeks PI. The values in the right insets indicate percentages of γδ TCR-positive or –negative population in CD4⁺ γ4 or γ13 cells. This experiment is a representative of four independent experiments. (B) The absolute numbers of γ4 cells (upper graph) or γ13 cells (lower graph) in αβ TCR- or γδ TCR-potitive population were investigated. Similar results were obtained in three independent experiments.

Figure 7. DX5-negative as well as –positive cells displayed the unique cytokine production patterns.

(A and B) Hepatic lymphocytes were isolated from S. mansoni-infected mice and flowcytometric analysis was conducted upon TCR ligation at 6 weeks PI. The numbers in the insets represent percentages of γ4 (A) or γ13 (B) cell population in CD4⁺ DX5-positive or -negative population. This experiment is representative of five independent experiments. (C) Data represent the mean + SD of five independent experiments.

Figure 8. Both T-bet and GATA-3 were expressed in a single hepatic γ4 or γ13 cell.

(A and B) Hepatic lymphocytes were isolated from S. mansoni-infected mice and the expressions of T-bet and GAT-3 were analyzed by flowcytometry after TCR stimulation at 6 weeks PI. The numbers in the upper, large insets represent percentages of each population divided by the expressions of T-bet and GATA-3 in γ4 (A) or γ13 (B) cells. The lower, small insets represent the data using isotype control antibodies. Similar results were obtained in three independent experiments.