Figure 4.
Samd1 inhibits murine erythropoiesis and megakaryopoiesis. (A) Schematic outlining 4-day erythroid and megakaryocyte (Mk) differentiation. (B) Samd1 mRNA in shRNA-expressing (GFP+) bone marrow at differentiation day 4 in shControl– and shSamd1_exon2–infected cells (n = 9). Normalized to 18s ribosomal RNA (rRNA). (C) Western blotting of SAMD1 in control and shSamd1-1– and shSamd1-2–infected primary mouse bone marrow (day 4 culture). (D) Representative flow cytometry plot of anti-CD71 PE and anti-Ter119 APC at day 4 of erythroid differentiation. (E) Quantitation of erythroid differentiation stages as determined by flow cytometry. Early erythroid progenitors (CD71+) and late erythroid progenitors (CD71+ Ter119+)/mature red blood cells (Ter119+; n = 12). (F) Representative images of Wright-Giemsa–stained day-4 GFP+ erythrocytes. (G) Quantitation of CFU-E (day 3) and BFU-E colonies (day 7) from GFP+ bone marrow (n = 27). (H) mRNA levels of erythroid and signaling genes Gata2, Kit, Hba (hemoglobin α), Trim10, Kdm7a, and Dusp2 in day-4 GFP+ erythroid cells (n = 9). Normalized to 18s rRNA. (I) Quantitation of mature Mks (CD41+ CD42d+) at day 4 of differentiation as determined by flow cytometry (n = 9). Error bars represent SEM and SD. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001; ∗∗∗∗P < .0001 (2-tailed unpaired Student t test). CFU-E, CFU-erythroid.

Samd1 inhibits murine erythropoiesis and megakaryopoiesis. (A) Schematic outlining 4-day erythroid and megakaryocyte (Mk) differentiation. (B) Samd1 mRNA in shRNA-expressing (GFP+) bone marrow at differentiation day 4 in shControl– and shSamd1_exon2–infected cells (n = 9). Normalized to 18s ribosomal RNA (rRNA). (C) Western blotting of SAMD1 in control and shSamd1-1– and shSamd1-2–infected primary mouse bone marrow (day 4 culture). (D) Representative flow cytometry plot of anti-CD71 PE and anti-Ter119 APC at day 4 of erythroid differentiation. (E) Quantitation of erythroid differentiation stages as determined by flow cytometry. Early erythroid progenitors (CD71+) and late erythroid progenitors (CD71+ Ter119+)/mature red blood cells (Ter119+; n = 12). (F) Representative images of Wright-Giemsa–stained day-4 GFP+ erythrocytes. (G) Quantitation of CFU-E (day 3) and BFU-E colonies (day 7) from GFP+ bone marrow (n = 27). (H) mRNA levels of erythroid and signaling genes Gata2, Kit, Hba (hemoglobin α), Trim10, Kdm7a, and Dusp2 in day-4 GFP+ erythroid cells (n = 9). Normalized to 18s rRNA. (I) Quantitation of mature Mks (CD41+ CD42d+) at day 4 of differentiation as determined by flow cytometry (n = 9). Error bars represent SEM and SD. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001; ∗∗∗∗P < .0001 (2-tailed unpaired Student t test). CFU-E, CFU-erythroid.

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