Fig. 6

Defective CNS development in Wwox knockout mice. a Cerebellum, hippocampus and cerebral cortex protein samples from Wwox^+/+ and Wwox^−/− mice at postnatal day 14 or 20 were examined for the expression levels of an early neuronal differentiation marker DCX by western blotting. β-actin was used as an internal control. Quantitative densitometry of the immunoblots was performed and the numbers depict the ratio of DCX to β-actin protein level in the brain tissues. b Immunohistochemistry was performed to determined DCX expression in the cerebral cortex of Wwox^+/+ and Wwox^−/− mice at postnatal day 14 (N = 5). Scale bars = 50 μm. The numbers of DCX-positive cells in five representative subareas of Wwox^+/+ and Wwox^−/− cerebral cortex tissue sections were quantified (right panel). The results are expressed as means ± SEM. *P < 0.05. c Sagittal brain sections of Wwox^+/+ and Wwox^−/− mice at postnatal day 20 were immunostained with anti-NeuN. Compared with the age-matched control littermates, a large proportion of neurons in the hippocampal dentate gyrus (DG) of Wwox^−/− mice showed absent expression of a mature neuronal marker NeuN (black star). Dispersed distribution of NeuN-positive neurons in the hippocampal CA1 region of Wwox^−/− brain was observed (black arrow). The lower panel pictures of dentate gyrus and CA1 regions are the magnified images from the boxed area in the upper panel. The representative images of six independent experiments are shown. Scale bars = 200 (upper) or 100 μm (lower). DG, dentate gyrus. d Cresyl violet staining of sagittal brain sections revealed a decreased cell density in the subgranule zone (yellow arrow) and a less orderly arrangement of granule neurons (yellow arrowhead) in the hippocampal dentate gyrus of Wwox^−/− mice at postnatal day 20. The right panel pictures are the magnified images from the boxed area in the left panel. The representative images of five independent experiments are shown. Scale bars = 50 (left) or 20 μm (right)