Cre;Scl^fl/fl MKs display several maturation defects. (A) Bone marrow cells derived from 5-FU–treated mice (Cre;Scl^wt/wt and Cre;Scl^fl/fl, as indicated) were cultured in TPO-containing medium for 3 days. The morphology of cells at day 3 of culture is shown. The arrow points to a wild-type MK shading platelets. Images of MK culture were taken using an inverted IX51 Olympus microscope with a Jenoptik C14 camera. Openlab, Version 3 software (Improvision) was used for image acquisition, and images were exported into Adobe Photoshop, Version CS2 (Adobe Systems). Scale bars represent 50 μm. (B) The cultures shown in panel A are stained with May-Grunwald-Giemsa (left) and for AChE activity (right). The photographs were taken using an Olympus BX60 microscope with a Qimaging camera. Openlab, Version 3 software (Improvision) was used for image acquisition, and images were exported into Adobe Photoshop, Version CS2 (Adobe Systems). Scale bars represent 20 μm. (C) CD41 expression was analyzed by flow cytometry in MKs derived from 5-FU–treated mice (top) and peripheral blood (bottom). Genotypes are as indicated. (D) Real-time quantitative PCR expression analysis of MK-specific genes. MKs were derived from Cre;Scl^wt/wt (black bars) and Cre;SCL^fl/fl (gray bars) 5-FU–treated mice. The y-axis represents enrichment in cDNA sequences normalized to Gapdh gene control sequences. Data are the mean ± SD of 7 independent experiments. *P < .01. (E) Analysis of the ploidy of Cre;Scl^wt/wt and Cre;Scl^fl/fl MKs. Bone marrow cells derived from 5-FU–treated mice were stained using propidium iodide and ploidy analyzed by flow cytometry on the CD41⁺ population. Peaks representing each ploidy class are labeled. Hatched histogram represents Cre;Scl^fl/fl; and empty histogram, Cre;Scl^wt/wt. One representative experiment of 3 is shown; the mean ploidy ± SD of all 3 experiments is shown. P < .01. (F) Cell-cycle analysis of CD41⁺ cells. Cre;Scl^w/wt, Cre;Scl^wt/fl, and Cre;Scl^fl/fl bone marrow MkPs were purified, cultured for 4 days, and cell-cycle phases determined by analysis of BrdU incorporation and 7-amino-actinomycin D staining. The percentage of CD41⁺ cells in each culture is shown on the left. The percentage of cells in G₀/G₁, S, and G₂M phases is shown on the right. One representative experiment of 3 is shown. The percentage (mean ± SD) of the cells in S phase in the 3 experiments is in parentheses. P = .006.