Figure 1.
Figure 1. CD103+ and CD103− TM persist in the brain after cerebral viral infection. (A) Representative brain sections of WT mice immunostained for LCMV nucleoprotein (arrows) at indicated days after rLCMV i.c. infection. Bars: 100 µm; (inset) 20 µm. (B–F) Flow cytometric analysis of brain-derived CD8+ T cells after rLCMV i.c. infection (as in Fig. S1). (B) Quantification of brain CD8+ T cell numbers and (C) frequencies of TM and short-lived effectors (SLE) at indicated days (d) after infection. (D) CD69+ brain TM (bTM) were analyzed for surface expression of CD103, binding of Db-NP396-404-tetramer, and intracellular expression of Bcl-2 and Granzyme B (GzmB; filled histograms, antibody stainings; open histograms, staining controls) 6 wk after infection. Numbers indicate frequencies of positive cells (mean ± SEM; n = 3 mice). (E) Frequencies of CD103+ bTM at indicated days (d) after infection. (F and G) P14 or OT-1 TCR transgenic CD8+ T cells were adoptively transferred into naive WT mice 24 h before infection with indicated viruses. CD69 and CD103 surface expression on the adoptively transferred T cells (CD45.1+) and on endogenous MVA-specific CD8+ T cells (Kb-B8R20-27-Tet+) in brains was analyzed by flow cytometry at 14 (F) or 6 wk (G) after infection. The frequencies of cells in the different gates are indicated (mean ± SEM; n = 4–5 mice per group). (H) 3 mo after infection with rLCMV i.c., anti–CD8b-PE was injected i.v. and PE-labeled CD8b+ T cells were measured in the blood and the brain by flow cytometry. Representative plots of CD8b-positive (intravascular, red) and -negative (extravascular, blue) CD8a+ T cells in relation to CD69 expression are shown. The frequencies of cells in the different gates are indicated (mean ± SEM; n = 4–5 mice per group). (I) rLCMV i.c. infected WT mice were treated with anti-CD8a depleting antibody i.p. 6 wk after infection. Frequencies and numbers of CD103+ and CD103− brain TM (CD8+CD44+CD69+) were analyzed by flow cytometry 6 wk later. (B, E, and I) Bars represent mean + SEM (n = 5–6 mice per group). One representative of (A, D, H, and I) or data pooled from (B, C, and E) at least two independent experiments is shown. (F and G) Experiments were performed once. For flow cytometric gating strategy see Fig. S1.

CD103+ and CD103− T M persist in the brain after cerebral viral infection. (A) Representative brain sections of WT mice immunostained for LCMV nucleoprotein (arrows) at indicated days after rLCMV i.c. infection. Bars: 100 µm; (inset) 20 µm. (B–F) Flow cytometric analysis of brain-derived CD8+ T cells after rLCMV i.c. infection (as in Fig. S1). (B) Quantification of brain CD8+ T cell numbers and (C) frequencies of TM and short-lived effectors (SLE) at indicated days (d) after infection. (D) CD69+ brain TM (bTM) were analyzed for surface expression of CD103, binding of Db-NP396-404-tetramer, and intracellular expression of Bcl-2 and Granzyme B (GzmB; filled histograms, antibody stainings; open histograms, staining controls) 6 wk after infection. Numbers indicate frequencies of positive cells (mean ± SEM; n = 3 mice). (E) Frequencies of CD103+ bTM at indicated days (d) after infection. (F and G) P14 or OT-1 TCR transgenic CD8+ T cells were adoptively transferred into naive WT mice 24 h before infection with indicated viruses. CD69 and CD103 surface expression on the adoptively transferred T cells (CD45.1+) and on endogenous MVA-specific CD8+ T cells (Kb-B8R20-27-Tet+) in brains was analyzed by flow cytometry at 14 (F) or 6 wk (G) after infection. The frequencies of cells in the different gates are indicated (mean ± SEM; n = 4–5 mice per group). (H) 3 mo after infection with rLCMV i.c., anti–CD8b-PE was injected i.v. and PE-labeled CD8b+ T cells were measured in the blood and the brain by flow cytometry. Representative plots of CD8b-positive (intravascular, red) and -negative (extravascular, blue) CD8a+ T cells in relation to CD69 expression are shown. The frequencies of cells in the different gates are indicated (mean ± SEM; n = 4–5 mice per group). (I) rLCMV i.c. infected WT mice were treated with anti-CD8a depleting antibody i.p. 6 wk after infection. Frequencies and numbers of CD103+ and CD103− brain TM (CD8+CD44+CD69+) were analyzed by flow cytometry 6 wk later. (B, E, and I) Bars represent mean + SEM (n = 5–6 mice per group). One representative of (A, D, H, and I) or data pooled from (B, C, and E) at least two independent experiments is shown. (F and G) Experiments were performed once. For flow cytometric gating strategy see Fig. S1.

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