Fig. 2

Overexpression of bdh2 or siderophore supplementation is sufficient to partially rescue hemoglobinization and erythroid maturation block in bdh2 morphants. (A) Morphological assessment of embryos injected with control or bdh2-specific MOs and bdh2-overexpressing bdh2 morphants. O-dianisidine (O-das) staining confirmed partial restoration of hemoglobinization in bdh2-overexpressing bdh2 morphants. (B) Analysis of circulating erythrocytes from embryos injected with control or bdh2-specific MOs and bdh2-overexpressing bdh2 morphants demonstrating decreased erythrocyte immaturity and partial restoration of hemoglobinization in bdh2-overexpressing bdh2 morphants. (Top) May-Grunwald/Giemsa stain. (Scale bars: 5 µM.) (Bottom) O-das staining. Erythrocytes are representative of the mean groups depicted in C. (C) (Left) The N:C ratio is an indicator of one aspect of a range of morphological features reflecting erythrocyte maturity. Tabulation of the N:C area ratio for erythrocytes isolated from embryos injected with control or bdh2-specific MOs and bdh2-overexpressing bdh2 morphants. Replenishment of bdh2 decreased the N:C ratio in bdh2 morphants. Data are mean ± SD for 300 cells. P < 0.05 was considered significant. (Right) Quantification of O-dianisidine-positive erythrocytes isolated from embryos injected with control or bdh2-specific MOs and bdh2-overexpressing bdh2 morphants. Replenishment of bdh2 restored hemoglobinization in bdh2 morphants. Data are mean ± SD for 300 cells. P < 0.05 was considered significant. (D-F) Supplementation with siderophore but not its chemical paralogs rescues hemoglobinization defect and erythrocyte maturation block in bdh2 morphants. (D) Analysis of whole of embryos. (E and F) Analysis of isolated erythrocytes. Image and data analyses are as in A-C.