Supplementary Figure 7: Characterization of microglia phenotype in conditional Rab7 knockout mice

(a) Confocal image showing microglia with shorter and less branched processes in Rab7^ΔMG at the age of 10 weeks compared to control mice. Scale bar: 30µm. Quantification of area of microglia processes in µm² in Rab7^ΔMG mice as compared to control (n=3 mice per group, 6 weeks after tamoxifen injection, mean +/- s.d., *P=0.0162, t=2.475, df=2, Student's two-tailed t test). 40 cells per each brain slice were analysed. Each dot represents the mean value of 3 brain slices per mouse (b) To determine the time-course of FITC-Dextran distribution, we performed injections into the cortex of 10 weeks old wild-type mice and the number of FITC-Dextran-positive microglia was determined at different time points post-injection (n=4-5 mice per time point). No decay of FITC-Dextran signal was seen within 96 hours. (c) FITC-Dextran was injected into the cortex (6 and 18 weeks after tamoxifen injection of P21 mice) of Rab7^ΔMG and control mice and uptake was assessed 7 hours post-injection. Quantification of FITC-Dextran positive microglia (n=3 mice per group, mean +/- s.d., two-way ANOVA, genotype effect: *P= 0.0192, F=14.37, followed by Bonferroni’s post hoc test, *P=0.0255, t=4.436). (d) Electron microscopic visualization of myelin fragments (arrows) in corpus callosum of 12 month old Rab7^ΔMG and control mice (48 weeks after tamoxifen injection) in extracellular space (a), in the cell (b) and attached to axon (c). Quantification of number of extracellular myelin fragments (n=3 mice per group, mean +/- s.d., two-way ANOVA, genotype effect: *P= 0.0175, F=8.425, followed by Bonferroni’s post hoc test, *P<0.05). Each dot represents the mean value of 3 brain slices per mouse.