Fig. 4

Validation of the existence of PD-1⁺CD8⁺ T_RM cells in HBV⁺ samples. A Multiplex immunofluorescence staining of formalin-fixed paraffin-embedded TB tissues of enrolled HCC patients in CyTOF analyses, stained with different markers, and the colocalized cells were colored by white, scale bar represents 20 μm. B Comparisons of the relative counts of PD-1⁺CD8⁺ T_RM cells across sample groups by multiplex immunofluorescence images as in (A). C Frequency comparisons of CD8⁺ T cells in CD45⁺ cells (left) and PD-1⁺CD8⁺T_RM cells in CD8⁺ T cells (right) of different sample groups, colored by HBV infections. D The representative flow cytometry plots of HBV tetramer pool and surface marker staining and gating strategies of different cell subsets. E and F Frequency comparisons of PD-1⁺ T cells in CD8⁺ T cells and HBV tetramer⁺CD8⁺ T cells (E) and PD-1⁺CD8⁺ T_RM cells in PD-1⁺CD8⁺ T cells and PD-1⁺HBV tetramer⁺CD8⁺ T cells (F). G Frequency comparison of CD8⁺CD137⁺ T cells in CD8⁺ T cells stimulated with DMSO or HBV-specific antigen peptide pools, colored by treatment groups. H and I Frequency comparison of PD-1⁺ T cells (H) and PD-1⁺CD8⁺T_RM cells (I) in paired CD8⁺ and CD8⁺CD137⁺ T cells of HBV-specific antigen peptide pools stimulated T cells. J Frequency comparison of IFNγ⁺CD8⁺ T cells in stimulated CD137⁺CD8.⁺ T cells treated with HBV-specific antigen peptide pools in the presence or absence of of anti-PD-L1 mAb. Unpaired student’s t-test was used in (B, C, E, and F), Paired t-test was used in (G-J), with *p < 0.05, **p < 0.01, and ***p < 0.001