Extended Data Fig. 9 |. Characterization of ependymal cilia and choroid plexus in hydrocephalic Trim71 mutant mice.

a) Coronal brain sections from control and hydrocephalic mice were stained for ciliary markers (acetyl-α-tubulin and ARL13B), ependymal cell markers (S100B and FOXJ1), and choroid plexus epithelial cell markers (E-Cadherin, OTX2).b-d) Immunostaining for molecular correlates of choroid plexus hypersecretion in WT controls and hydrocephalic Trim71^R595H/+ P0 mice. Immunostaining of phosphorylated SLC12A2/NKCC1 (pSLC12A2) (b) and phosphorylated STK39/SPAK (pSTK39) (c) in the choroid plexi of a WT control and hydrocephalic Trim71^R595H/+ P0 mouse. d) Immunostaining of SLC12A2/NKCC1 and STK39/SPAK in the choroid plexi of a WT control and hydrocephalic Trim71^R595H/+ P0 mouse. e-f) Validation of OCT imaging platform to characterize ependymal cilia-driven CSF flow ex vivo. e). Representative flow polarity maps demonstrating bead trajectories (by temporal color coding) over time in an adult WT mouse brain explant in toxin-naive condition, with ciliobrevin, and after washout of toxin. Color bar represents color versus corresponding frame in the color-coded image. f) Quantitation of local CSF flow speed at the ventricular wall in brain explants from adult WT mouse brains in the different experimental conditions. g-i) CSF flow directionality in WT littermate controls and hydrocephalic Trim71^R595H/+ mice at P7. g) Post-Gaussian processing CSF flow maps in a WT littermate control and hydrocephalic Trim71^R595H/+ mice at P7. h,i) Quantitation of CSF flow directionality in WT littermate controls and hydrocephalic Trim71^R595H/+ mice at P7. Directionality is represented as the distance between the start point and end point (d) divided by the bead pathway (d), see panel (i). Significance was tested by a two-sided paired t-test (f) or two-sided unpaired t-test (h): *: P<0.05, **: P<0.01, ns (not significant): P>0.05. Data represented as boxplots (f,h): median (line), 25^th and 75^th percentiles (box), whiskers go down to the smallest value and up to the largest, overlaid with individual data points. For detailed statistical information (f, h), see Supplementary Table 13.