HVEM-deficient CD8 T cells have intact activation and early division in response to VACV infection or lymphopenic environment.

<p>(A) WT or HVEM^−/− mice were infected i.p. with VACV-WR (3×10⁴ PFU/mouse). On day 6-post infection, splenocytes were harvested and stained for CD8, CD44, and B8R or A8R tetramer. <i>Top panel</i>, Representative plots of tetramer staining, gating on CD8 cells. Percentages of CD44^high expressing B8R-tetramer (<i>left panels</i>) or A8R-tetramer (<i>right panels</i>) positive CD8 T cells are indicated. CD8 T cell activation was assessed by up-regulation of CD25, and down-regulation of CD62L and IL-7R (CD127) on tetramer-positive CD44^high cells. Naive (CD44^low tetramer-negative) CD8 T cells were used as controls. Percentages that stained positive for each marker are indicated. (B) One×10⁵ WT or HVEM −/− CFSE-labeled OT-I CD8 T cells were adoptively transferred into naïve WT mice. One day later, mice were infected i.p. with VACV-WR-OVA. After 2 or 3 days, OT-I CD8 T cells were analyzed. <i>Top</i>, Representative contour plots of co-staining for Vα2 and Vβ5 after gating on CD8⁺ T cells. Numbers indicate the percentage of CD8⁺Vα2⁺Vβ5⁺-positive cells. <i>Middle</i>, Representative dot plots of CFSE dilution after gating on CD8⁺Vα2⁺Vβ5⁺ cells. <i>Bottom</i>, Representative histogram of CFSE dilution gating on CD8⁺Vα2⁺Vβ5⁺ cells. (C) One×10⁵ highly purified CFSE-labeled polyclonal WT or HVEM^−/− CD8 T cells were adoptively transferred into RAG^−/− mice. After 4 days, splenocytes were harvested and stained with anti-CD3, anti-CD8, and anti-CD44. <i>Top</i>, Representative dot plots of CFSE dilution after gating on CD3⁺CD8⁺ cells. <i>Bottom</i>, Representative histogram of CFSE dilution gating on CD3⁺CD8⁺ cells. Results are the mean number ± SEM (<i>n</i> = 4 mice/group) from one experiment. Similar results were obtained in two separate experiments.