Extended Data Fig. 1. Naïve CD8⁺ T cells that recognize hepatocellular Ag are activated locally and expand, but fail to develop effector function.

(a) Schematic representation of the experimental setup. 5 x 10⁶ Env28 T_N were transferred into C57BL/6 x Balb/c F1 (WT) or HBV replication-competent transgenic (HBV Tg, C57BL/6 x Balb/c F1) recipients. Livers were collected and analysed five days after Env28 T_N transfer and sera from the same mice were collected every day from day 0 to day 5 after Env28 T_N transfer. (b-c) Absolute numbers (b) and frequency of IFNγ-producing (c) Env28 T cells in the livers of the indicated mice. (d) ALT levels detected in the sera of the indicated mice at the indicated time points. n = 4. Results are expressed as mean ± SEM. Means among groups were compared with two-tailed t test (e) Schematic representation of the experimental setup. 5 x 10⁶ Cor93 T_N were transferred into C57BL/6 (WT) or MUP-core recipients. Mice were splenectomized and treated with anti-CD62L 48 hours and 4 hours prior to cell transfer, respectively. Untreated WT mice that received 5 x 10⁶ Cor93 T_N were used as controls. Where indicated, mice were injected with 2.5 x 10⁵ infectious units of non-replicating rLCMV-core 4 hours prior to Cor93 T_N transfer. Liver-draining lymph nodes⁶⁸ (dLN) and non-draining inguinal lymph nodes (ndLN) were collected at four hours and one day after Cor93 T_N. (f) Representative flow cytometry plot at four hours upon Cor93 T_N transfer. Numbers indicate the percentage of cells within the indicated gate. (g-h) Quantification of the absolute numbers of cells recovered from the ndLN (g) and dLN (h) of the indicated mice four hours and one day upon Cor93 T_N transfer. n = 3. Results are expressed as mean ± SEM. Means among groups were compared with one-way ANOVA with Bonferroni post-test (i) Confocal immunofluorescence micrographs of liver sections from WT mice (WT), rLCMV-core-injected WT mice (WT + rLCMV-core), MUP-core mice and R26-ZsGreen mice injected with 2.5 x 10⁵ infectious units of non-replicating rLCMV-cre (R26-ZsGreen + rLCMV-cre). Scale bars represent 100 μm. Note that, because HBV core protein did not accumulate at detectable levels in KCs and hepatic dendritic cells [DCs] upon rLCMV-core injection, we confirmed the tropism of this vector by injecting rLCMV-cre into R26-ZsGreen mice – these mice express the fluorescent protein ZsGreen upon Cre-mediated recombination. (j) Mean Fluorescent Intensity (MFI) of CD69 expression on Cor93 T cells in the liver, blood, lung and bone marrow of the indicated mice four hours after Cor93 T_N transfer. n = 4. Results are expressed as mean ± SEM. Means among groups were compared with one-way ANOVA with Bonferroni post-test.

Data are representative of at least 3 independent experiments. *** p value < 0.001. Mouse drawings were adapted from reference ⁶⁹.