MO activates the HIF-1α-EPO signaling pathway and enhances erythroid regulatory networks. (A) Binarized regulon activity heat map of the top 50 transcription factors in E13.5 and E17.5 fetal liver, colored by cell type and treatment group. (B, D, H-I) UMAP visualization of Gata1 (B) and Tal1 (D) regulon activities in erythroid cells. Quantification of Gata1 (H) and Tal1 (I) regulon activities (AUC scores) in CT and MO groups. (C and E) RT-qPCR analysis shows upregulation of the mRNA levels of Hif1a, Epo, Epor, Gata1, Tal1, Klf1 and Trfc in E13.5 (C) and E17.5 (E) fetal livers. (F, G, J, K) Western blot bands showing HIF-1α protein levels in E13.5 (F) and E17.5 (G) fetal livers. Quantification of HIF-1α expression relative to β-tubulin in E13.5 (J) and E17.5 (K) fetal livers. The scRNA-seq analyses were based on pooled embryos from 3 pregnancies per group (3 embryos per pregnancy). RT-qPCR and western blot analyses were conducted using embryos from 4 to 6 pregnancies per group. Data are presented as mean ± SEM; each dot represents an individual embryo. Statistical significance was determined by unpaired 2-tailed t test. ∗P < .05; ∗∗P < .01. UMAP, uniform manifold approximation and projection.