Fig. 3: Histological characterization and quantification of periventricular ependymal cells and their cilia.

a Representative images of H&E stained ependyma, comprising ependymal cells (Epn) and interspersed basal cells (arrowheads), lining the 4th ventricle (Vent) in trip6^+/− (Control) and trip6^−/− mice. Note the poorly differentiated epithelial layer comprising atrophic ependymal cells that exhibit “endothelial” (flattened) cell morphology and aberrant ciliation, as well as the relative sparsity of basal cells in the trip6^−/− brain. b Semi-schematic depiction of the segmentation method used in cell number and ciliation index quantification of the ependyma. Individual ependymal cells were determined by their nuclei (circles) and inspected for the presence of cilia on their apical site (lines). c Representative images depicting native, mild and severe phenotypes for cell number and ciliation of control and trip6^−/− mice. Quantification demonstrated that although the mean number of ependymal cells was not altered (d), ciliation is significantly affected (e). [The box plot (e) indicates the median (middle line), the interquartile range from 1st to the 3rd quartile (box) and the range of the minima to maxima (whiskers)]. f Semi-schematic depiction of the method used for quantification of cilia length and coverage of the ependyma. Individual segments of 10 µm were determined (horizontal full and dotted lines) and maximum cilia “lawn” height was measured at each second segment (vertical lines) for quantification and found to be reduced in trip6^−/− mice (g). The coverage of cilia (indicated as the fraction of unciliated segments per 100 µm) is significantly reduced in trip6^−/− mice (h). The length of cilia (determined as maximum cilia “lawn” height per 10 µm ciliated segment) was also significantly reduced (i). Further information on quantification methodology and statistical tests performed can be found in “Methods” section (including Supplementary Table 7). The statistical significance of the analyses is represented by the p-values indicated in the graphs (e, d, g, b, h, i). Source data are provided as a Source Data file. Chp choroid plexus. Asterisk (*) indicates unciliated areas. Scale bars: 10 µm.