The loss of Sad kinases leads to an increase in the mitotic index.

<p>(A, E) Coronal sections from the brains of E13, E15 or E17 embryos with the indicated genotypes were analyzed by staining with anti-Ki67 (A, proliferating cells, red) and anti-PH3 antibodies (A, cells in M-phase, green) or with an anti-PCNA antibody (E, green) and Hoechst 33342 (blue). (B—D) A significant decrease in the number of proliferating Ki67⁺ cells per 10⁴ μm² (B) was observed at E15 and E17 when comparing the <i>Sada</i>^<i>+/-</i><i>;Sadb</i>^<i>+</i>/- (magenta (B, C) and blue (D), respectively) and <i>Sada</i>^<i>-/-</i><i>;Sadb</i>^<i>-</i>/- cortex (green). The number of mitotic PH3⁺ cells per section remained constant (C) but the mitotic index (D, ratio of PH3⁺ and Ki67⁺cells in %) increased in the <i>Sada</i>^<i>-/-</i><i>;Sadb</i>^<i>-</i>/- cortex (green) at E15 and E17 compared to <i>Sada</i>^<i>+/-</i><i>;Sadb</i>^<i>+</i>/-controls since the number of Ki67⁺cells was reduced. (F—G) The number of PCNA⁺ cells in S-phase (PCNA⁺ cells with a punctate staining pattern (inset)) per 10⁴ μm² (F) and the proportion of cells in S-phase (G, ratio of PCNA⁺ S-phase and Ki67⁺cells in %) in the VZ/SVZ of the cortex were quantified in the <i>Sada</i>^<i>-/-</i><i>;Sadb</i>^<i>-</i>/- cortex (green) and <i>Sada</i>^<i>+/-</i><i>;Sadb</i>^<i>+</i>/-controls (magenta (F) and blue (G), respectively). Values are means ± s.e.m., n = 3 different embryos, two-way ANOVA compared to heterozygous control; *, p<0.05; **, p<0.01; ***, p<0.001, n. s., not significant). The Scale bars are 20 μm.