Pdk1-deficient BM MKs show an abnormal ultrastructure with disrupted DMS and abrogated MK polarization. (A) Representative TEM images of Pdk1^fl/fl (upper) and Pdk1^−/− (lower) MKs in the BM showing decreased presence of demarcation membranes and large granula-free zones upon Pdk1 deficiency. Overview (left; scale bar equals 3 μm). Detail (right; scale bar equals 1 μm). Boxes mark the section in the illustrations which are highlighted in a higher magnification in the right panels. (B) Representative confocal images (maximum projection) of BM-derived Pdk1^fl/fl and Pdk1^−/− MKs cultured in vitro for 5 days and stained for F-actin (Phalloidin-Atto647N, red), nuclei (DAPI, blue), and GPIX (GPIX-Alexa488, green). Three-dimensional surface rendering of respective z-stacks (right; Imaris 9.2). White asterisks indicate polarized MKs. Scale bar equals 40 μm. Boxes mark the section in the illustrations which are highlighted in a higher magnification in panel C. (C) Detailed visualization of DMS polarization of individual cultured Pdk1^fl/fl and Pdk1^−/− BM MKs. (D) Quantification of the percent of polarized and nonpolarized Pdk1^fl/fl and Pdk1^−/− BM MKs (left) and the MK diameter (right). Arithmetic means ± SD (n = 6 mice, at least 100 MKs per genotype were analyzed, unpaired 2-tailed Student t test). **P < .01 indicates statistically significant difference.