<i>Nfix</i> blocks B cell differentiation in vitro in favour of myelopoiesis.

<p><b>(A)</b> Total FL was transduced with control MigR1 or <i>Nfix</i>, plated on OP9 cells and analyzed by flow cytometry on day 16. FACS plots of GFP⁺ B cells (B220⁺CD19⁺, top panels) and GFP⁺ myeloid cells (CD11b⁺Gr-1⁺, lower panels) representative of 3 replicates from 2 independent experiments. <b>(B)</b> Graph of average percentage of B220⁺CD19⁺ (left) and CD11b⁺Gr-1⁺ (right) from (A). Error bars denote +/- SD of 2 independent experiments (n = 3). Graph shows a statistically significant decrease in <i>Nfix</i> derived B220⁺CD19⁺ cells (p = 0.02) and a significant increase in <i>Nfix</i> derived CD11b⁺Gr-1⁺ cells (p = 0.01) when compared with MigR1 controls. <b>(C)</b> E14.5 FL HSPCs (Lin^-Sca1⁺cKit⁺Flt3^-) transduced with control MigR1 or <i>Nfix</i> retrovirus were plated on OP9 cells (Day 0) and analysed by flow cytometry on day 12 with FACs plots of GFP expression (left histograms) and GFP^- and GFP⁺ B220⁺CD19⁺ cells (right dot plots) representative of 2 independent experiments. <b>(D)</b> Graph of average percentage of B220⁺CD19⁺ cells from (C), error bars denote +/- SD of 2 independent experiments. Experiment shows significant decrease in <i>Nfix</i> expressing B cells compared with control MigR1 (p = 0.01) <b>(E)</b> Total BM cells were sorted for MigR1 and <i>Nfix</i> expression 24 hr post transduction and qRT-PCR was performed. Graphical presentation of relative mRNA expression of <i>Nfix</i>, E2A, ID2 and ID3 genes and presented relative to control MigR1 cells. Error bars denote +/- SD of 3 technical replicates. Data representative of 2 biological replicates.