Phenotypic MDSCs are relatively expanded after PTCy in mice and in patients. (A-D) Ten- to 12-week-old female nonthymectomized recipient mice underwent irradiation (10.5 Gy) and transplantation with TCD BM cells and splenocytes from 10- to 12-week-old female donors. Cell doses were 10 × 10⁶ TCD BM cells and 40 × 10⁶ splenocytes for the B6C3F1→B6D2F1 and C3H→AKR models and 5 × 10⁶ TCD BM cells and 50 × 10⁶ splenocytes for the C3H→B6D2F1 model.² PBS vehicle or 25 mg/kg per day PTCy was administered intraperitoneally on days +3 and +4. (A-B) Data are shown for the B6C3F1→B6D2F1 MHC-haploidentical HCT model using CD45.1⁺CD45.2⁺ B6C3F1 donors and WT B6D2F1 (CD45.1⁻CD45.2⁺) recipients. Donor cells were gated based on CD45.1 vs CD45.2 expression. (A) Donor MDSC content in the allograft and at day +3 (before PTCy treatment). (B) PTCy increased the percentages of donor G-MDSCs (CD11b⁺Ly6G⁺Ly6C^int/low) and M-MDSCs (CD11b⁺Ly6G⁻Ly6C^high) in various tissues at days +7 and +21. (C) M-MDSCs also were increased at day +6 in the C3H→B6D2F1 MHC-disparate HCT model. Donor cells were gated based on H2k^k-positivity. (D) Both G-MDSCs and M-MDSCs were increased at day +7 in the C3H→AKR MHC-matched HCT model. (E) This increase in M-MDSCs also was seen in early posttransplantation samples from patients (n = 5) treated with T-cell–replete, HLA-haploidentical BM transplantation using PTCy. G-MDSCs in patients were not reported because differentiating true G-MDSCs from normal neutrophils contaminating the buffy coat layer of Ficolled blood may not be reliable. For panels A-D, combined results from 2 independent experiments are shown with n = 4 per group per experiment. ∗P < .05, ∗∗P < .01, ∗∗∗P < .001, ∗∗∗∗P < .0001, and NS = not significantly different on unpaired t test.