Fig. 4: CD mitigates demyelination-related pathology in GA-Nes mice.

a Gene ontology of genes rescued by CD with |LFC| > 0.5 shows that upregulated genes in Oligo_mat1 and neuroblast cluster are contributing to myelination and axonal development. The number of genes per cell type is indicated in parenthesis. b Heatmap of gene expression of major myelin protein components demonstrates downregulation of myelination in all oligodendrocyte clusters as the result of transgene expression (left block), an effect which is significantly rescued with CD treatment (right block). Asterisks indicate significant changes (adjusted p < 0.05). c A myelin DAM signature is detected in various models of ALS as well as patient spinal cord. Microglia cluster also strongly shows this signature, while CD administration partially alleviates this signature, shifting it more towards the amyloid DAM signature. Asterisks indicate significant changes (adjusted p < 0.05). d, e SEM overview and quantification of corpus callosum shows dramatic loss of myelinated axons upon transgene expression, which is partially but significantly rescued by CD treatment (ANOVA, post hoc by Tukey’s HSD). n from independent biological replicates: WT Vehicle = 3, WT CD = 3, Tg Vehicle = 4, Tg CD = 5. Scale bars = 3 µm. f Measurements of cholesteryl ester species by mass spectrometry demonstrate significant upregulation in transgenic condition and CD-mediated rescue of CE 16:1, CE 18:1, and CE 22:4. CE 18:2, CE 20:4, and CE 22:6 were not significantly modulated (ANOVA with Fisher’s LSD post-hoc). n from independent biological replicates: WT Vehicle = 4, Tg Vehicle = 3, Tg CD = 5. g Total unmodified cholesterol and cholesteryl esters (from peak area sum) confirm downregulation of cholesterol in transgenic condition, which is not affected by CD. Cholesteryl esters are upregulated in GA-Nes mice, which is rescued by CD (ANOVA with Fisher’s LSD post-hoc. n from independent biological replicates: WT Vehicle = 4, Tg Vehicle = 3, Tg CD = 5).