BM cellularity decreases after pIpC-mediated NF-Ya deletion in the BM because of cell death and apoptosis. (A) Four days after pIpC treatment control and ko BM cells were isolated. After erythrocyte lysis cells were resuspended in equal volumes for analysis by FACS for equal amounts of time. Representative dot plots are shown. The percentage of cells within the gate is shown on the upper right corner of each plot, and the total amount of cells counted in equal periods of time is shown in the lower right. (B) BM cells from panel A were stained with PI and analyzed by FACS. One representative histogram overlay is shown (left; control [gray], ko [black]). On the basis of this analysis the scatter plot (right) shows the relative amount of dead cells for control and ko samples at indicated time points after pIpC treatment. Single data points and the averages, including SDs, are shown (n ≥ 3); *P < .05 and **P < .01. (C) Ten days after ko induction femur bones of control and ko BM chimera were fixed in formalin for 48 hours, decalcified in 0.5M EDTA for 5 days, paraffin embedded, sectioned, and H&E stained. The images show representative control and ko femur bones at a magnification of ×200, Nikon Eclipse 80i, Qimage Micropublisher 5.0 camera and software, 20×/0.50 NA objective. (D) Twenty-four hours after pIpC treatment BM cells were isolated from control and ko mice, erythrocytes were lysed, cells were stained with PI and annexin V and analyzed by FACS. Annexin V staining was determined among PI-negative cells (left). The histogram overlay (middle) shows a representative annexin V staining of live cells of control (gray) and ko (black). Results of 4 independent experiments are shown in the scatter plot (right). Single data points and the averages, including SDs, are shown (n = 4); **P < .01. (E) Twenty-four hours after pIpC treatment BM cells were isolated from control and ko mice, stained with DAPI, and analyzed by FACS. The histogram overlay shows a representative control and ko sample of the cell cycle analysis according to DNA content. The bar graph (right) shows the relative amount of cells in the respective phase of the cell cycle 24 hours after pIpC-induced NF-Ya deletion (n = 4); **P < .01.