Fig. 3: Neural progenitors in the cortical plate show mitotic block with increase in cell death and centrosome defects indicating role for multipolar spindles. | Nature Communications

Fig. 3: Neural progenitors in the cortical plate show mitotic block with increase in cell death and centrosome defects indicating role for multipolar spindles.

From: Mitotic block and epigenetic repression underlie neurodevelopmental defects and neurobehavioral deficits in congenital heart disease

Fig. 3

A–H Comparison of the brain cortex from E14.5 Ohiam/m mutant (D–F) vs. wildtype littermate embryos (A–C) showed more pH3 and TUNEL staining (G) in the ventricular (VZ) and subventricular (SVC) zones in the mutants. A decrease was also noted in mitotic cells in anaphase-telophase (H, n = 3, Ohiam/m mutants, n = 3 littermate controls). I–K Centrosomes were visualized by confocal microscopy in E14.5 brain tissue sections from 4 Ohiam/m mutant (n = 766) and 3 wildtype control (n = 1394) embryos using γ-tubulin (green) antibody staining. Coimmunostaining with Tbr2 was conducted to track cells in the intermediate zone that will give rise to cortical layers II–IV. 3D serial confocal imaging stacks were collected to investigate centrosome distribution. Commonly observed were paired centrosomes of similar size, predicting normal bipolar spindles I. Also observed were abnormal juxtapositioning of three or more centrosomes of varying sizes that would predict multipolar spindle defects J. The insets show magnified views of the same regions indicated by the arrowheads. Quantification showed a significant increase of the abnormal centrosomes in the mutant brain tissue (J, K). L–R. Confocal imaging was conducted using MEFs from wildtype (n = 3) (L-N) and Ohiam/m mutant embryos (n = 3) (O–Q) with α-tubulin and γ-tubulin used to visualized the mitotic spindles. >500 cells analyzed per embryo. Mutant MEFs showed increase in multipolar spindles (R). S–U. Ohiam/m MEFs showed reduction in the proportion of mitotic cells in anaphase/telophase (S) and an increase in lagging chromosomes during anaphase (T, U) Q, R. CP cortical plate, IZ intermediate zone, SVZ subventricular zone, VZ ventricular zone, P/M prophase/metaphase, A/T anaphase/telophase. Scale bars = 100 µm. All graphs represent mean ± SD. p-values were obtained using 2-way, unpaired t-test. Source data are provided as a Source Data file.

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