In situ immunostaining of MK subpopulations in murine BM. (A) Relative expression levels of marker genes, including Mylk4, Lsp1, Cd53, and Arntl, in mouse MK subpopulations. (B) Immunofluorescence of MYLK4 (green) co-stained with CD41 (red) in mouse BM. Arrows indicate MYLK4⁺CD41⁺ cells. (C) Immunofluorescence of LSP1 (green) co-stained with CD41 (red) in mouse BM. Arrows indicate LSP1⁺ CD41⁺ cells. (D) Immunofluorescence of ARNTL (green) co-stained with CD41 (red) in mouse BM. Arrows indicate ARNTL⁺CD41⁺ cells. The images in panels B-D represent 3 independent experiments . (E) Bar plot showing the frequency of MYLK4⁺CD41⁺ (668 MKs), LSP1⁺CD41⁺ (1221 MKs), and ARNTL⁺CD41⁺ (748 MKs) in total MKs (CD41⁺) in murine BM. (F) Box plot showing the distribution of cell diameters of MYLK4⁺CD41⁺ (87 cells), LSP1⁺CD41⁺ (49 cells), and ARNTL⁺CD41⁺ (185 cells). Data are from one-way analysis of variance (ANOVA). Scale bars, represent 20 μm. Error bars indicate SEM. *P ≤ .05; ****P ≤ .0001. DAPI, 4′,6-diamidino-2-phenylindole.