Figure 7. Vitamin C rescues dyserythropoiesis due to IDH1 downregulation.

(A) Late-stage erythroblast differentiation was monitored as a GLUT1⁺CD49d⁻ phenotype, and representative dot plots are presented (top). The percentages of GLUT1⁺CD49d⁻ erythroblasts were evaluated following transduction with shIDH1–1 or shIDH1–2 vectors, and mean levels ± SEM relative to control cells are presented (n = 7, bottom).

(B) Enucleation of shControl- and shIDH1-transduced erythroblasts was assessed on GFP+ cells at day 14, and representative histograms are presented (top). Enucleation levels, relative to control cells, were quantified (n = 8, bottom).

(C) The impact of αKG and DHA on erythroblast enucleation following IDH1 knockdown was evaluated between days 7 and 10, and representative histograms are presented (top). Enucleation relative to control conditions was quantified (n = 4, bottom).

(D) The impact of vitamin C on enucleation of control and IDH1-downregulated erythroblasts was evaluated between days 7 and 10. Representative histograms (top) and quantifications (bottom) are presented (n = 5). Changes in mitochondrial superoxide were evaluated by MitoSOX staining, and MFIs are presented (n = 4).

(E) TET2 was downregulated in erythroid progenitors by shTET2 lentivirus (day 4), and enucleation was monitored at day 10 in the presence or absence of vitamin C. Means ± SEM are presented (n = 5, one-way ANOVA with Tukey’s post hoc test).

(F) Nuclear morphology of orthochromatic erythroblasts differentiated from shControl- and shIDH1-transduced progenitors in the absence or presence of vitamin C was monitored. Representative cytospins are shown (left), and the quantification of orthochromatic erythroblasts with abnormal nuclei was determined by counting 500–1,000 cells (n = 3, right).

ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.