Bclaf1 deficiency in hematopoietic cells reduces fetal HSC expansion but does not alter adult HSCs at steady state. (A-B) Western blot of BCLAF1 in sorted CD45.2+ hematopoietic cells from E14.5 fetal liver (A) or total adult bone marrow (BM) cells (B) of indicated genotypes. GAPDH shown as a loading control. (C) HSC numbers quantified by flow cytometry across developmental ages. (D-O) HSC, MPP3/4, and total cell numbers quantified by flow cytometry in E14.5 fetal liver (Bclaf1f/f, n = 15; Vav-Cre:Bclaf1f/f, n = 5) (D-F), E15.5 fetal liver (Bclaf1f/f, n = 15; Vav-Cre:Bclaf1f/f, n = 8) (G-I), E17.5 fetal liver (Bclaf1f/f, n = 17; Vav-Cre:Bclaf1f/f, n = 15) (J-L), and adult BM (Bclaf1f/f, n = 19; Vav-Cre:Bclaf1f/f, n = 34) (M-O). (P) Peripheral blood counts and red blood cell indices from adult Bclaf1f/f and Vav-Cre:Bclaf1f/f mice (n = 5 per genotype). (Q) Mature B220+ B, CD3+ T, and CD11b+ myeloid cell numbers in the spleen of adult Bclaf1f/f and Vav-Cre:Bclaf1f/f mice (n = 5 per genotype). (R) Adult (8-week-old) Bclaf1f/f and Mx-Cre:Bclaf1f/f mice were treated with pIpC, and BM hematopoietic cells were assessed 4 weeks after final dose of pIpC. HSC and MPP3/4 cell numbers were quantified by flow cytometry (Bclaf1f/f, n = 9; Mx-Cre:Bclaf1f/f, n = 11). Data in panels C-R are mean ± standard deviation (SD), and statistical significance was determined by unpaired 2-tailed Student t test. ∗P ≤ .05; ∗∗P ≤ .01; ∗∗∗P ≤ .001; ∗∗∗∗P ≤ .0001. GAPDH, glyceraldehyde-3-phosphate dehydrogenase; ns, not significant.
