Figure 3.

Impaired cell proliferation due to cell-cycle defects is observed in NECs upon aberrant licensing

(A) BrdU (green) and MKI67 (red) staining in brain sections of embryos at E9.5. Dotted lines demarcate the cortical walls. Boxed areas are represented as higher-magnification images. White arrows point to double-positive cells.

(B) The graph presents the percentage of MKI67⁺ cells that incorporated BrdU. Mean ± SEM (n = 3/genotype).

(C) pH3 (green) staining in coronal brain sections derived from E9.5 embryos. Dotted lines delineate the cortical walls. Boxes indicate higher magnifications of positive cells. Note the spotted pattern of pH3 staining and the characteristic interphase nuclei representative of G2 cells (box 1) and the nuclear staining observed in condensed nuclei representative of mitotic cells (box 2).

(D) The bar graph depicts the number of pH3⁺ cells in a 200-μm-wide column assigned as G2 or M (mitotic) according to pH3 pattern. Mean ± SEM (n = 4/genotype).

(E) Whole telencephalic vesicles of embryo brains immunostained with antibodies for pericentrin (red) and β-catenin (green) or α-tubulin (green). Arrows point to the multiple centrosomes of one mitotic nucleus. The table shows the percentage of mitotic cells (mitotic index) and the percentage of mitoses with more than two centrosomes. n represents the number of cells analyzed per genotype. Data are derived from two independent litters. DNA is stained with Hoechst or Draq5 (blue). Scale bars: 10 μm. ^∗p ≤ 0.05, ^∗∗p ≤ 0.01; t test.