Figure 3.
Turnover, Survival, and Expression of Inhibitory Receptors on Sustained Memory CD8+ T Cell Pools
C57BL/6 mice were infected i.v. with 1 × 106 pfus MCMV or immunized i.v. with 1 × 109 pfus Ad-LacZ.
(A) Longitudinal analysis of the percentage of cells expressing Ki67 on MCMV-specific (left panel) and βgal-specific CD8+ T cells (right panel) in the spleen on days 0, 7, 21, 50, and 100 postinfection or postvaccination, measured by flow cytometry (n = 6–8; ±SEM).
(B) Representative flow cytometry plots show the expression of Ki67 in the spleen on days 7 and 50 postinfection or d21 and d100 postvaccination gated on live M38- or M45-specific (left panel) and βgal96- or βgal497-specific (right panel) CD8 T cells.
(C) Heatmap showing mRNA expression levels of proteins involved in apoptosis measured by microarray analysis of naive, M38-, and M45-specific CD8+ T cells days 7 and 50 postinfection (left panel) and naive CD8+ T cells, βgal96-, and βgal497-specific CD8+ T cells days 21 and 100 postvaccination (right panel). Shades of red indicate upregulated genes, and shades of blue indicate downregulated genes. Filter criteria of at least 2-fold changes with p ≤ 0.05, compared to naive CD8+ T cells, are shown.
(D) Longitudinal analysis of the percentage of cells expressing Bcl2 on MCMV-specific (left panel) and βgal-specific CD8+ T cells (right panel) in the spleen on days 0, 7, 21, 50, and 100 postinfection or postvaccination, measured by flow cytometry (n = 6–8; ±SEM).
(E) Flow cytometry analysis showing the percentage or the MFI of cells expressing the inhibitory receptors PD1, TIM3, and BTLA on M38- (red) and M45-specific CD8+ T cells (blue; n = 6–8; ±SEM) and on βgal96- (red) and βgal497-specific (blue) CD8+ T cells in the spleen (n = 6–8; ±SEM). Histograms show expression of PD-1, TIM-3, and BTLA and are gated on live naive (green) M38/βgal96-specific (red) and M45/βgal497-specific CD8+ T cells (blue).