Fig. 5. Rapamycin acts through ULK1 to alleviate clinical phenotypes of β-thalassemia in mice.

WT mice were transplanted with fetal liver HSCs of the indicated genotypes, as in Fig. 1B. Twelve weeks later, rapamycin (Rap; 4 mg/kg) or vehicle was administered intraperitoneally, once daily for 30 days. (A) Protein synthesis rates in splenic and bone marrow erythroblasts, determined by O-propargyl-puromycin incorporation. Ery.A, Ery.B, and Ery.C represent increasingly mature Ter119⁺ erythroblast populations designated according to forward scatter and CD71 expression (see fig. S3): vehicle, n = 3; rapamycin, n = 4. (B) Erythroid indices (y axis) according to HSC genotype (x axis) after treatment with rapamycin or vehicle. Hbb^Th3/+Ulk1^+/+, n = 15; Hbb^Th3/+Ulk1^−/−, n = 12. (C) Sulfo-NHS-biotin was injected intravenously on day 13 of rapamycin treatment, and the fraction of biotinylated RBCs was quantified serially by streptavidin labeling and flow cytometry. (D) Spleens of β-thalassemic mice (Hbb^Th3/+Ulk1^+/+) after rapamycin or vehicle treatment. Scale bars, 0.5 cm. (E) Summary of spleen weights in multiple Hbb^Th3/+Ulk1^+/+ mice after treatment with rapamycin (n = 12) or vehicle (n = 12). Data are mean values ± SDs; ****P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05; ns, not significant.