Figure 6.

CD47 or Sirpα blockade abrogates mitochondria transfer–mediated erythroid recovery from acute anemic stress. (A) Schematic illustration of the experiment to validate the function of CD47 in mitochondria transfer–mediated erythroid recovery. CD47 blocking antibody (anti-CD47) or Sirpα blocking antibody (anti-Sirpα) was administered in parallel with EBI macrophages from mito-Dendra2 mice into PHZ-treated mice. (B) Spleen morphology before and after PHZ treatment (upper), with and without CD47 blockade (lower). (C and D) FACS analysis of reticulocytes/RBC fraction in (C) spleen and (D) BM of PHZ-treated mice administered either IgG (control), CD47 Ab (anti-CD47), and/or EBI macrophages (MΦ). (E) Peripheral blood analysis of PHZ-treated mice administered either IgG (control), CD47 Ab (anti-CD47), and/or EBI macrophages (MΦ). (F) Frequency of mitochondria transfer, as indicated by the percentage of mito-Dendra2⁺ events, among early erythroblasts in spleen and BM; in the presence or absence of CD47 blockade. (G) Expression level of Sirpα mRNA and protein in control (F4/80⁺CD106⁻CD169⁻) and EBI (F4/80⁺CD106⁺CD169⁺) macrophages. (H and I) Peripheral blood (H) and (I) FACS analysis of reticulocytes/RBC fraction in PHZ-treated mice administered either IgG (control), Sirpα blocking antibody (anti-Sirpα), and/or EBI macrophages (MΦ). (J) Frequency of mitochondria transfer among splenic and BM early erythroblasts in the presence or absence of Sirpα blockade. For all quantification, mean ± SEM; *, P < 0.05; **, P < 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001 by Student’s t test. (K) Frequency of mitochondria transfer from cocultured mito-Dendra2⁺ EBI macrophages to erythroblasts isolated from PHZ treated tdTomato-Vav-Cre mice, in the presence or absence of CD47 and/or Sirpα blockade. Schematic illustration of the experimental design is shown. For all quantification, mean ± SEM; **, P < 0.01; ***, P ≤ 0.001 by Student’s t test.