Increased numbers and viability of c-kit⁺ and Sca-1⁺ cells in Bcl-2-overexpressing AGM and liver. E11 AGM, aorta, and liver tissues from Bcl-2 transgenic and nontransgenic embryos were dissected and cultured as explants for 3 to 4 days, during which time the embryos were genotyped. Tissues from embryos with a similar genotype were pooled, and single-cell suspensions were stained with antibodies specific for c-kit or Sca-1 and with annexin V and 7AAD. (A) Representative FACS plots for c-kit- and Sca-1-stained cells (top and bottom rows, respectively) within the 7AAD^- fraction of wild-type (WT) and Bcl-2-overexpressing AGM and aorta tissues, respectively. Side scatter (SSC) is shown on the x-axis and fluorescent antibody staining on the y-axis. Percentages of c-kit⁺ and Sca-1⁺ cells are shown for the gated region (HSCs are found within the nongranular side scatter). (B) Within the c-kit⁺ and Sca-1⁺ fractions of AGM and aorta tissues, the percentage of annexin V⁺ cells was determined (indicating the percentage of cells entering the apoptotic pathway). Representative histogram plots are shown with indicated percentages of annexin V⁺ cells in the c-kit⁺ (top row) or Sca-1⁺ (bottom row) cell fractions, respectively, in the AGM or subdissected aorta. Bcl-2-overexpressing tissues (right panels) contain less annexin V⁺ cells within the c-kit⁺ and Sca-1⁺ fraction. (C) Overview of the fold changes (increases) in percentages of c-kit⁺ and Sca-1⁺ cells and (D) fold changes (decreases) in percentages of annexin V⁺ cells within the c-kit⁺ and Sca-1⁺ fractions of Bcl-2 transgenic E11 AGM, aorta, or liver cells as compared with wild-type cells. Percentages were determined as indicated in panels A and B. Results are shown for 3 experiments with Sca-1 and annexin V, 3 experiments with c-kit and annexin V, and 2 experiments with E11 aorta cells. Each bar in the graph represents an independent experiment containing embryos from 1 to 3 litters. From each experiment, several samples (each contained a pool of cells from 1 to 3 tissues) were measured, and the average of these samples is shown. Total number of events analyzed ranges from 8 × 10⁴ to 2 × 10⁵.