Figure 7.
Restoration of CNS remyelination capacity by ARL6IP1 gene transfer. (A) After AAV9 gene delivery, Arl6ip1 mRNA levels were confirmed using RT-qPCR for gene transfer efficiency in the cerebral cortex of Arl6ip1−/− mice. mRNA levels of glial or pro-inflammatory marker genes after AAV9 gene delivery compared with that in Arl6ip1+/+. Data represent mean ± SEM of n = 4 mice/group. (B) Change in protein levels of neural and glial markers (left) and of apoptosis- (middle) or autophagy-related genes (right) from pooled lysates in mouse cortex after AAV9 gene delivery (n = 4 mice/group). (C) Changes in mouse cytokine and chemokine levels in CSF samples after AAV9 gene delivery. A bar graph shows the quantification of cytokine array analysis. Data represent mean ± SD of n = 8 mice/group. (D and E) Mouse coronal section image from Allen Adult Mouse Brain Atlas showing internal capsule and corpus callosum area (left, marked in purple). Representative image of MBP staining of the IC and CC from Arl6ip1−/− after ARL6IP1 gene delivery (40×; scale bar: 500 μm). CTX, cerebral cortex; HIPP, hippocampus, STR, striatum; IC, internal capsule; CC, corpus callosum. (F and G) Quantification of myelin thickness (MBP-positive fibers) in IC and CC region. Data represent mean ± SD of 10 images/group and are calculated in GraphPad Prism 7.0 (n = 5 mice/group; *P < 0.05; **P < 0.01). (H) Representative image of MBP fluorescence staining in the spinal cord from Arl6ip1−/− after AAV9 gene delivery, and highlighted in corticospinal tract of mouse spinal cord (40×; scale bar: 500 μm, 400×, scale bar: 50 μm). (I) Quantification of myelin (MBP staining) of white matter in the spinal cord. Data represent mean ± SD of 10 images/group, calculated in GraphPad Prism 7.0 (n = 5 mice/group; **P < 0.01). (J) Representative TEM images of spinal cord white matter in Arl6ip1−/− after AAV9 gene delivery (n = 3 mice/group, 15,000×; scale bar: 800 nm). (K) Quantitative data of the percentage of myelinated axons, G ratio, and myelin sheath thickness. Data represent mean ± SD of 10 images/group and are calculated in GraphPad Prism 7.0. *P < 0.05; **P < 0.01. Data represent averages of three independent biological replicates with two technical replicates for each. Source data are available for this figure: SourceData F7.

Restoration of CNS remyelination capacity by ARL6IP1 gene transfer. (A) After AAV9 gene delivery, Arl6ip1 mRNA levels were confirmed using RT-qPCR for gene transfer efficiency in the cerebral cortex of Arl6ip1−/− mice. mRNA levels of glial or pro-inflammatory marker genes after AAV9 gene delivery compared with that in Arl6ip1+/+. Data represent mean ± SEM of n = 4 mice/group. (B) Change in protein levels of neural and glial markers (left) and of apoptosis- (middle) or autophagy-related genes (right) from pooled lysates in mouse cortex after AAV9 gene delivery (n = 4 mice/group). (C) Changes in mouse cytokine and chemokine levels in CSF samples after AAV9 gene delivery. A bar graph shows the quantification of cytokine array analysis. Data represent mean ± SD of n = 8 mice/group. (D and E) Mouse coronal section image from Allen Adult Mouse Brain Atlas showing internal capsule and corpus callosum area (left, marked in purple). Representative image of MBP staining of the IC and CC from Arl6ip1−/− after ARL6IP1 gene delivery (40×; scale bar: 500 μm). CTX, cerebral cortex; HIPP, hippocampus, STR, striatum; IC, internal capsule; CC, corpus callosum. (F and G) Quantification of myelin thickness (MBP-positive fibers) in IC and CC region. Data represent mean ± SD of 10 images/group and are calculated in GraphPad Prism 7.0 (n = 5 mice/group; *P < 0.05; **P < 0.01). (H) Representative image of MBP fluorescence staining in the spinal cord from Arl6ip1−/− after AAV9 gene delivery, and highlighted in corticospinal tract of mouse spinal cord (40×; scale bar: 500 μm, 400×, scale bar: 50 μm). (I) Quantification of myelin (MBP staining) of white matter in the spinal cord. Data represent mean ± SD of 10 images/group, calculated in GraphPad Prism 7.0 (n = 5 mice/group; **P < 0.01). (J) Representative TEM images of spinal cord white matter in Arl6ip1−/− after AAV9 gene delivery (n = 3 mice/group, 15,000×; scale bar: 800 nm). (K) Quantitative data of the percentage of myelinated axons, G ratio, and myelin sheath thickness. Data represent mean ± SD of 10 images/group and are calculated in GraphPad Prism 7.0. *P < 0.05; **P < 0.01. Data represent averages of three independent biological replicates with two technical replicates for each. Source data are available for this figure: SourceData F7.

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