Fig. 6: PTGIR signaling promotes metabolic and transcriptional features of T_ex cells.

a,b, Schematic and volcano plot of WT CD8⁺ T cells transduced with PTGIR (WT + PTGIR) (a) or Keap1^−/− CD8⁺ T cells treated with vehicle (veh.; DMSO) or iloprost (ilo.) 5 µM for 24 h (b). DEGs are shown as log₂(fold-change) (x axis) versus −log₁₀(FDR) (y axis). Shaded regions denote significant enrichment (FDR < 0.05) in iloprost- or vehicle-treated cells. c,d, Normalized enrichment scores (NESs; q < 0.05) of the top ten Hallmark pathways downregulated by iloprost treatment in WT + PTGIR (c) and Keap1^−/− T cells (d). e,f, NESs of the top ten Hallmark pathways upregulated by iloprost in WT + PTGIR (e) and Keap1^−/− CD8⁺ T cells (f). g, Relative growth and viability of activated WT and Keap1^−/− CD8⁺ T cells cultured with vehicle or iloprost (5 µM) for 24 h. Cell counts for growth assays were normalized to initial seeding density (n = 3–4 mice per group). h, Bioenergetic profiling of WT and Keap1^−/− CD8⁺ T cells after 24 h of vehicle (+veh) or iloprost (+ilo) treatment. OCR (left) and ECAR (right) for T cells over time are shown (n = 8–12 technical replicates per group). Time of addition of oligomycin (O), fluorocarbonyl cyanide phenylhydrazone (FCCP) (F), rotenone and antimycin A (R/AA) and monensin (M) are indicated. i, NESs of T_ex cell transcriptional signatures (accession no. GSE188670, ref. ²³) in iloprost-treated Keap1^−/− T cells using DEG values from b. Heatmap shows expression of T_ex^ISG genes. Inset highlights top contributing genes to the T_ex^ISG signature in iloprost-treated Keap1^−/− T cells. j,k, Cytokine production by WT and Keap1^−/− CD8⁺ T cells after 24 h of treatment with vehicle control or iloprost (5 μM). j, Representative flow cytometry plots for IFNγ versus TNF expression for CD8⁺ T cells. k, Quantification of IFNγ⁺/TNF⁺ T cells after treatment (n = 4 mice per group). The data in h are indicated as mean ± s.d. and display technical replicates from a single representative experiment. The data in g, j and k are indicated as mean ± s.e.m. Statistical significance was calculated via one-way ANOVA with Tukey’s test. NS, P ≥ 0.05; ^*P = 0.05; ^**P = 0.01; ^***P = 0.001; ^****P = 0.0001. Cholest. homeost.; cholesterol homeostasis; FA, fatty acid; Inflamm., inflammatory; rej., rejection; resp., response; sig., signaling.