Transitory-Tex-CNI maintained responsiveness to PD-1 blockade. (A-L) Allo-HCT was performed as in Figure 1A. Recipients were administered with CSP or vehicle from days 0 to 14 followed by intraperitoneal injection of αPD-L1 on day 14 after allo-HCT. (A) A schematic overview of experiments. (B-G) FCM of donor T cells in the spleen were performed on day 16. Contour plots (B-C) and proportions of Ki-67 (D) and GZMB (E) expression in 2C-Tg CD8⁺ donor T cells. Absolute numbers of PD-1⁺TIGIT^– (F) and PD-1⁺TIGIT⁺ (G) in 2C-Tg CD8⁺ donor T cells (n = 5 per group). (H) MFI of Ki-67 and GZMB in Ly6C⁺ and Ly6C^– 2C-Tg CD8⁺ donor T cells from CSP-treated recipients with or without injection of αPD-L1 on 16 days (n = 5 per group). (I-L) OCR of purified donor T cells were evaluated using a Seahorse XF analyzer. OCR curves before (day 14; I) and after (day 16; J) αPD-L1. Basal respiration (K) and spare respiratory capacity (L) on day 16 after allo-HCT. (M-Q) Recipient mice were administered with CSP or vehicle from days 0 to 14 and intraperitoneally injected with αPD-L1 on day 28 after allo-HCT. FCM analysis of donor T cells in the spleen were performed on day 30. (M) A schematic overview of the experiments. Proportions of Ki-67 (N) and GZMB (O) expression, and absolute numbers of PD-1⁺TIGIT^– (P) and PD-1⁺TIGIT⁺ (Q) in 2C-Tg CD8⁺ donor T cells (n = 5 per group). (D-G, K, L, N-Q) Data from 2 independent experiments were combined and shown as means ± SEM. (H-J) Data from one of 3 similar experiments were shown as means ± SEM. ∗P < .05; ∗∗P < .01; ∗∗∗P < .005. MFI, mean fluorescence intensity; ns, not significant; OCR, oxygen consumption rate.