The effect of MEK inhibition on megakaryocyte endomitosis. Murine bone marrow cells were cultured as described in the legend to Fig 6. Cells were then stained with FITC-conjugated anti-CD41, solubilized, and stained with PI. In the left-hand panels, the abscissa represents log FITC intensity and the ordinate represents cell size, as determined by forward light scatter. Megakaryocytes, identified by CD41 positivity, were gated to analyze for DNA content by PI staining (right-hand panels). The dot plot of cells stained with the isotype control for anti CD41-FITC is also shown ([A] isotype). The log intensity of PI is on the abscissa and the numbers of cells are on the ordinate. The bars mark the DNA contents of 2N, 4N, 8N, 16N, 32N, 64N, and 128N, respectively. Cells grown in DMSO (A) are compared with cells grown in PD 98059 (B). These data are the representative results from 3 separate experiments. Cell numbers after 72-hour cultures are shown in the table (C). The total numbers of viable cells were determined using a hemocytometer. The percentage of CD41⁺cells in each population was determined by immunofluorescent staining and flow cytometry. Megakaryocyte numbers were calculated as follows: total viable cells multiplied by the percentage of CD41⁺cells. The results represent the mean of 3 to 4 experiments.