Fig. 5

PRV-LAV treatment effectively relieves the immunosuppressive state of T cells in a Hepa1-6 mouse model. A Heatmap displaying normalized marker expression of each CD45⁺ cell cluster. B t-SNE plot derived from CyTOF analysis of tumor immune infiltrates obtained from each treatment group. Cells are colored by the clusters identified by PhenoGraph. C-D Density t-SNE plots of equal numbers of tumor immune infiltrates from vehicle and PRV-LAV treatment group. Merged (C) and separate (D) images are shown. Mø, macrophage; DC, dendritic cell; Mono, monocytes. E Quantitative analysis of each cell cluster as a percentage of CD45⁺ cells. P values were calculated by paired t test. Cluster 1: B cells; cluster 4: exhausted CD8⁺ T cells; cluster 3: activated CD8⁺ T cells; cluster15: naïve neutrophils; cluster11: activated neutrophils. F–H Tumor immune infiltrates were analyzed by flow cytometry the day after mice received the final dose of 4 doses of mock (n = 12) or PRV-LAV (n = 15) treatment. The numbers of tumor-infiltrating CD4⁺ T cells, CD8⁺ T cells, neutrophils, dendritic cells, macrophages, monocytes, and NK cells per gram tumor were calculated (F). The mean fluorescence intensity of CD86 on DC cells (G), CTLA4, PD-1, TIM3, and LAG3 on CD4⁺ and CD8⁺ T cells (H) were counted. The data are presented as the mean ± s.d. values. The black bars indicate the mean values. Statistical analysis was performed by t test. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001. I-J The knockout of CD4⁺ and CD8⁺ T cells during PRV-LAV treatment. Changes in the tumor volume curves (I) for mice bearing Hepa1-6 tumors treated with PRV-LAV (1 × 10.⁷ PFUs, intratumorally). Changes in the tumor volumes are expressed as the mean ± SEM values (n = 9). In (I), statistical analysis was performed by repeated measure ANOVA. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001. Changes in the tumor volumes curves of each mouse were shown in (J)