Acute MCMV infection impairs generation of Gr1^HI-MDSCs. (A) Kinetics of circulating CD11b⁺Gr1^HI MDSCs in ECDI-SP–treated, either uninfected or Δm157-infected (on day 0), transplant recipients. Total live CD11b⁺Gr1^HI cells were enumerated by FACS in 50 µL of blood drawn on the indicated days. (B) Quantitative analysis of total CD11b⁺Gr1^HI cells in 50 µL of blood collected on day 10 posttransplantation from recipients of the indicated groups. Data shown in panels A and B were from 2 to 3 independent experiments with a total of 4 to 6 mice in each group. *P < .05. (C) Representative histograms of expression of C5aR and FcγRII/III on circulating Gr1^HI-MDSCs on day 10 posttransplantation from naïve or transplant recipients with or without day 0 Δm157 infection. (D) Mean fluorescence intensities (MFI) of C5aR and FcγRII/III of groups shown in panel C. Data presented in panels C and D were obtained from 2 independent experiments with a total of 4 mice in each group. (E) In vitro suppression assay using Gr1^HI-MDSCs sorted from the spleen of the indicated groups 10 days posttransplantation. Sorted Gr1^HI-MDSCs were cocultured with CFSE-labeled syngeneic CD8 T cells stimulated with anti-CD3/CD28 coated beads (at a ratio of 1:1:1). Proliferation of CD8 T cells was measured by CFSE dilution by FACS. (F) Quantification of CD8 T-cell proliferation in the presence of Gr1^HI-MDSCs sorted from the indicated groups shown in panel E. Data shown in panels E and F were obtained from 2 independent experiments with a total of 4 mice in each group. Data were presented as mean ± SD. *P < .05. (G) Representative FACS plot depicting the purity of sorted Gr1^HI cells pooled from the BM and the spleen of naïve B6 mice used for adoptive transfers. (H) Schematic treatment plan and percent graft survival with adoptive transfer of sorted Gr1^HI cells. Two cohorts of ECDI-SP–treated transplant recipients were infected with Δm157 on day 0. The first cohort additionally received ∼30 × 10⁶ sorted Gr1^HI cells on the indicated days, while the other cohort did not receive any cells (N = 3-4, compiled from 2 independent experiments). *P < .05 (log-rank test).