Figure 4.

Reduction of Microglial Activation in Naglu^−/− Mice Treated with Neural Stem Cells Overexpressing NAGLU

(A) Schematic illustration of ICV injection (green) at the bregma (1.54 mm) and PAR injection (blue) at the bregma (−0.58 mm), demonstrating regions of analysis (red squares) relative to the site of engraftment (ICV and PAR), including the motor cortex, striatum, and ventral forebrain. (B) Representative bright-field images taken at a low magnification of half-brain coronal sections of immunohistochemical staining of CD68 in a mouse injected with N-iNSCs (−/− treated) compared with unaffected heterozygous (unaffected) and vehicle-injected Naglu^−/− mice (−/− vehicle). Scale bar, 1 mm. The locations of ICV and intraparenchymal injections are outlined by red dotted lines. The blue dotted lines outline regions where CD68 staining is less pronounced, indicating a reduction of microglial activation. (C) Representative bright-field images of the motor cortex (1), striatum (2), and ventral forebrain (3) of immunohistological staining for CD68 in −/− vehicle compared with unaffected mice and in −/− treated mice upon ICV and intraparenchymal injection. Scale bars, 100 μm. The blue dotted lines highlight the difference in CD68 immunoreactivity close to the ventricle (yellow box) versus farther away from the ventricle (dark blue box) in ICV-injected brains. Inset images were taken at higher magnification to highlight the differences in morphology of CD68-stained microglia. Scale bars, 20 μm. Post-acquisition processing was applied to all images and included adjustments to brightness and contrast and RGB curves using Adobe Photoshop CS6 to improve visibility and consistency in color tone. The histograms on the right show the area of CD68 immunoreactivity for each group, measured. *p < 0.05, **p < 0.01, ***p < 0.001; two-tailed, unpaired parametric t test. Values are shown as mean ± SEM (n = 3 mice per group). Mean with SEM. For p values, see Tables 1 and 2.