Extended Data Fig. 8: Validation of skewed MS heterogeneity and oligodendrocyte gene expression profiling in control and NAWM tissues.

a, Validation of BCAN-expressing OPCs in combination with OLIG1 and OLIG2 IHC. Red arrowhead denotes BCAN⁺OLIG1/2⁺ OPC; blue arrowhead denotes BCAN⁻OLIG1/2⁺ oligodendrocyte. Scale bar, 20 µm. t-SNE overlay of BCAN expression in the snRNA-seq dataset in control and MS samples. Scale bar, 20 µm. Data are mean ± s.e.m. n = 4 samples from different control individuals, n = 6 NAWM samples and n = 5 MS lesion samples from different patients with MS. P values determined by ANOVA. b, KLK6-expressing oligodendrocytes in control white matter, NAWM and MS lesions. Scale bar, 50 µm. Data are mean ± s.e.m. n = 4 samples from different control individuals; n = 5 different MS individuals. P values determined by ANOVA. t-SNE overlay of KLK6 expression in the control and MS snRNA-seq dataset. c, Violin plots showing the normalized expression counts of genes enriched in ImOLG in the snRNA-seq dataset (OPC n = 352; COP n = 242; ImOLG n = 207; Oligo1n = 1,129; Oligo2 n = 1,839; Oligo3 n = 775; Oligo4 n = 1,579; Oligo5 n = 1,167; Oligo6 n = 1,484). Violin plots are centred on the median with interquartile ranges, and the shape represents cell distribution. d, MRF IHC in control white matter, NAWM and MS lesions. Scale bar, 50 µm. Data are mean ± s.e.m. n = 6 samples from different control individuals, and n = 7 different patients with MS. P values determined by ANOVA. t-SNE overlay of MRF expression in the snRNA-seq dataset. e, t-SNE overlay of MBP expression in the control and the MS snRNA-seq dataset (n = 4,037 oligodendrocytes in control; n = 4,737 oligodendrocytes in MS). f, Western blot of the MRF antibody on human brain lysate to validate the specificity of the antibody. For gel source data, see Supplementary Fig. 1. g, Combination of MRF mRNA and protein labelling to confirm the specificity of the MRF antibody in control white matter. Scale bar, 10 µm. h, Heat maps representing the average gene expression of a subset of genes, including myelin-related genes, in control versus MS samples in OPCs (control versus MS and control versus NAWM) and mature oligodendrocytes (control versus NAWM). In a, b and d, each experiment was performed in two (three for d) independent batches, and P values are only displayed compared to control; in f and g, each experiment was performed twice on independent samples.