Figure 4.

Effect of combination BM irradiation and PD-1 treatment on regulatory immune cell populations in distant flank tumors. A. Tissue from flank D4M tumors were harvested at the conclusion of the experiment described in Figure 3, dissociated by collagenase/hyaluronidase digestion, and stained for regulatory T cell markers before being quantified by flow cytometry. Regulatory T cells were defined as CD25^hi Foxp3⁺ cells among CD3⁺ CD4⁺ T cells. B. Based on the percentages of the flow stain, the numbers of Treg per tumor and the ratio of these cells to the number of CD8⁺ T cells was determined (B, C). D. The percentage of myeloid derived suppressor cells (MDSCs) was determined in the tumors by anti-body staining and flow cytometry. Cells were gated on CD45⁺ Gr1⁺ CD11b⁺ populations. *p ≤ 0.5 for the indicated comparisons to the anti-PD-1 plus irradiation group.