Extended Data Fig. 12: Pdcd1 deletion enhances anti-tumour immunosurveillance in livers of TGAlb-cre+ mice, Related to Fig. 5.
From: Activated ATF6α is a hepatic tumour driver restricting immunosurveillance
(a) Breeding strategy of TG:Pdcd1−/− mice (top) and representative FACS scatter plot showing PD1-knockout efficiency (bottom). (b) Liver-to-body weight of 9-month-old TGAlb-cre+ (n = 19; 9 males, 10 females) and TG:Pdcd1−/− mice (n = 18; 8 males, 10 females). (c) Serological parameters (c, ALT, AST, ALP and cholesterol) of 9-month-old TGAlb-cre+ (n = 19; 9 males, 10 females) and TG:Pdcd1−/− mice (n = 16; 7 males, 9 females). (d) NMR metabolomics PLS − DA with coloured 95% confidence intervals, showing no distinct separation in metabolic profile between livers of 9-month-old TGAlb-cre+ and TG:Pdcd1−/− mice (n = 5/group). (e) NMR-based metabolic analysis of lactate or glucose levels in livers of 9-month-old TGAlb-cre+ versus TG:Pdcd1−/− mice (n = 5/group). (f-g) Representative PAS staining and IHC for BiP, CHOP, Ki67, γ-H2AX, and cl-Caspase 3 (f; scale bar = 100 µm), with quantification (g) in non-tumour livers from TGAlb-cre+ (n = 5) and TG:Pdcd1−/− (n = 6) mice. (h) Representative IHC for HA (scale bar = 200 µm) in tumour (T) and non-tumour (NT) liver of TGAlb-cre+ and TG:Pdcd1−/− mice. (i) Representative in-situ hybridization of Tnf mRNA (left; scale bar = 100 µm) and related quantification (right) in livers from TGAlb-cre+ (n = 3) and TG:Pdcd1−/− (n = 4) mice. Arrowheads indicate immune cell clusters. (j) IHC quantification for intratumoral CD8+ and PD1+ cells in Fig. 5o from livers of TGAlb-cre+ (n = 5) and TG:Pdcd1−/− (n = 6) mice. (k-m) Liver immune cells were isolated and stimulated (with PMA and ionomycin) for cytokine production, cytokine-secreting CD8+ T-cells (k), CD4+ T-cells (l) and NKT cells (m) were measured by flow cytometry in 9-month-old TGAlb-cre+ (n = 4) and TG:Pdcd1−/− mice (n = 4). (n-p) Liver immune cells isolated from 9-month-old TGAlb-cre+ (n = 4) and TG:Pdcd1−/− (n = 4) mice with naïve (n), CD8+PD1+ (o), and effector (p) CD8+ T-cell populations measured by FACS. (q-r) Contour plot (q) and KDE plot (r) for LDH expression in CD8+ TILs from livers of 9-month-old TGAlb-cre+ and TG:Pdcd1−/− mice by MIBI (n = 3/group). (s) Schematic depiction of prolonged ATF6α activation driving early liver injury and DNA damage through cellular stress and metabolic reprogramming that dampens immunosurveillance efficacy, perpetuating oncogenic signalling for progression to HCC in mice and humans. Scatter dot plot data are presented as mean values ± SEM. Data in 12b,c,e,g,i,j-p were analysed by two-tailed unpaired t-test or Mann-Whitney test based on data normality distribution. Data in 12q were analysed by Mann-Whitney test. The distribution of LDH expression in 12r was visualized using overlaid KDE plots. Differences in cumulative distributions were evaluated using the Kolmogorov-Smirnov test. Mouse icons and schematic depiction were created in BioRender. Heikenwälder, M. (2026) https://BioRender.com/lgjnsy9.
