Low-dose decitabine-treated MDSCs ameliorated thrombocytopenia and enhanced bone marrow–derived MDSC metabolic function in active-ITP model mice. ITP models were established in irradiated SCID mice by engraftment with 2 × 10⁴ splenocytes from CD61-knockout mice immunized against wild-type C57 mouse platelets. The mice were divided into 3 groups: the control group did not receive any treatment, the other 2 groups of active-ITP mice were given PBS- or decitabine-treated MDSC transfer, respectively, and platelet counts were monitored weekly for 5 weeks (mean ± standard error of the mean). (A) The lines represent the platelet counts of ITP mice (mean ± standard error of the mean). On days 28 and 35, the decitabine-treated MDSC group had significantly higher platelet counts than the control and PBS-treated MDSC groups. Significance among groups were determined by 2-way ANOVA (∗∗∗∗P_Time < .0001, ∗∗∗P_platelets = .0001, ∗P_interaction = .0418; multiple comparisons on day 28: ∗∗∗P_{Ctrl vs Dec-MDSC} = .0005, ∗P_{PBS-MDSC vs Dec-MDSC} = .0146; day 35: ∗∗∗P_{Ctrl vs Dec-MDSC} = .0001, ∗P_{PBS-MDSC vs Dec-MDSC} = .0455). (B) The lines represent the anti-CD61 antibody level of ITP mice (mean ± standard error of the mean). The star indicates significant differences between groups emerged on day 14. (C) MDSCs were sorted from the bone marrow for OCR assessment. (D) Mitochondrial basal respiration (unpaired t tests, ∗∗∗P_{Ctrl vs Dec-MDSC} = .0001, ∗P_{PBS-MDSC vs Dec-MDS} = .0327), ATP production (unpaired t tests,∗∗∗P_{Ctrl vs Dec-MDSC} = .0007, ∗P_{PBS-MDSC vs Dec-MDSC} = .0436), and maximal respiration (unpaired t tests,∗∗P_{Ctrl vs Dec-MDSC} = .0042, ∗P_{PBS-MDSC vs Dec-MDSC} = .0194) were higher in the decitabine-treated group than in the other 2 groups. (E) Intracellular ATP levels were significantly improved after decitabine-treated MDSC transfer (unpaired t tests,∗∗P_{Ctrl vs Dec-MDSC} = .0066, ∗P_{PBS-MDSC vs Dec-MDSC} = .0445). (F-H) mRNA expression of ACADM, HADHA, and PGC1β was significantly lower in the control and PBS-treated groups than in the decitabine-treated group (unpaired t tests, ACADM: ∗∗P_{Ctrl vs Dec-MDSC} = .0033, ∗P_{PBS-MDSC vs Dec-MDSC} = .0413; HADHA, ∗∗P_{Ctrl vs Dec-MDSC} = .0076, ∗P_{PBS-MDSC vs Dec-MDSC} = .0313; PGC1β: ∗∗P_{Ctrl vs Dec-MDSC} = .0016, ∗P_{PBS-MDSC vs Dec-MDSC} = .0252). (I) TGF-β, IL-10, IL-4, IL-2 serum levels were significantly higher in the decitabine-treated MDSC group than in the control or PBS-treated groups (unpaired t tests: IL-10: ∗P_{Ctrl vs Dec-MDSC} = .0225, ∗P_{PBS-MDSC vs Dec-MDSC} = .0421; IL-4, ∗∗∗P_{Ctrl vs Dec-MDSC} = .0007, ∗P_{PBS-MDSC vs Dec-MDSC} = .0488; TGF-β: ∗∗P_{Ctrl vs Dec-MDSC} = .0082, ∗P_{PBS-MDSC vs Dec-MDSC} = .0409). TNF-α, IL-1β, IL-6, IL-17a, IFN-γ, IL-12 p70 serum levels were significantly lower in the decitabine-treated MDSC group than in the control or PBS-treated groups (unpaired t tests: TNF-α: ∗∗∗P_{Ctrl vs Dec-MDSC} = .0085, ∗∗P_{PBS-MDSC vs Dec-MDSC} = .0005; IL-1β: ∗∗P_{Ctrl vs Dec-MDSC} = .0066, ∗P_{PBS-MDSC vs Dec-MDSC} = .0356; IL-17a: ∗∗P_{Ctrl vs Dec-MDSC} = .0035, ∗P_{PBS-MDSC vs Dec-MDSC} = .0470; IFN-γ: ∗∗P_{Ctrl vs Dec-MDSC} = .0096, ∗∗P_{PBS-MDSC vs Dec-MDSC} = .0077; IL-12 p70: ∗∗P_{Ctrl vs Dec-MDSC} = .0035, ∗P_{PBS-MDSC vs Dec-MDSC} = .0151. Mann-Whitney U tests: IL-6: ∗P_{Ctrl vs Dec-MDSC} = .0260, ∗P_{PBS-MDSC vs Dec-MDSC} = .0152). The levels of VEGF, IL-2, and IL-13 did not vary significantly among the groups.