Tfr2 haploinsufficiency in combination with Tmprss6-ASO significantly ameliorates erythropoiesis and splenomegaly of β-thalassemic mice. At the end of the treatment with Tmprss6-ASO or CTRL-ASO, Tfr2^{BM hetero}/Hbb^th3/+ and tHbb^th3/+ mice were euthanized, and we evaluated kidney mRNA expression of EPO (Epo) relative to hypoxanthine phosphoribosyltransferase 1 (Hprt1) (A), serum EPO levels (B), Erfe mRNA levels in the BM (C) and spleen (D), spleen size relative to body weight (E), percentage of Ter119⁺ cells on alive cells, and subpopulation composition (gated clusters: proerythroblasts [I], basophilic erythroblasts [II], polychromatic erythroblasts [III], orthochromatic erythroblasts and immature reticulocytes [IV], and mature red cells [V]) based on Ter119/CD44 expression and forward scatter (reflecting cell size) both in the BM (F) and in the spleen (SP) (G). Dotted red line indicates mean values for untreated WT mice. Bars represent SD. Asterisks refer to statistically significant differences between age-matched tHbb^th3/+ and Tfr2^BMKO/Hbb^th3/+ mice (***P < .005; **P < .01; *P < .05). Complete statistical analysis relative to panels F and G (1-way analysis of variance with Bonferroni’s multiple comparison test) is reported in supplemental Tables 1 and 2.