Fig. 1

BTK and innate immune pathway gene expression in human microglia and across MS lesion types. A Uniform Manifold Approximation and Projection plot showing expression of BTK (right) in an snRNA-seq dataset of human brain samples. BTK is highly expressed in microglia in the human CNS. B snRNA-seq dataset showing broad expression of BTK across microglial subclusters from the human brain. Dots correspond to “pseudobulk” values for clusters, i.e. the sum of counts across all cells in one cluster, in one sample. Counts are normalized by the TMM-normalized library size. Values are only shown where there are at least 10 cells in the combination of cluster and sample. C Immunofluorescent images of postmortem MS brain tissue from subcortical WM containing an active lesion. Low-magnification overview, with DAPI identifying cell nuclei (blue), CD45 identifying immune cells (green) and MBP identifying myelin (magenta), with dotted white boxes containing higher magnification images showing the active lesion (a) and nonlesion WM (b). Scale bar is 2 mm. D The active lesion area shows prominent infiltration of microglia and macrophages labeled with HLA-DR (white) that are closely associated with BTK staining (red), as shown in the merged panels. Scale bars are 100 μm. E Heat map showing expression of BTK and associated innate immune pathway genes in microglia across MS lesion types. Differential expression relative to control WM and GM is shown. Expression of certain FcγR-encoding genes, including FCGR2A and FCGR2B, is significantly increased in WM active lesions and chronic active lesions. Significance is indicated by ****P < 0.0001, ***P < 0.001, **P < 0.01 and *P < 0.05. R package glmmTMB was used, and the Benjamini–Hochberg procedure was used to control the false discovery rate. AL active lesion, BTK Bruton’s tyrosine kinase, CAL chronic active lesion, CIL chronic inactive lesion, CNS central nervous system, COP committed oligodendrocyte precursor, DAPI 4′,6-diamidino-2-phenylindole, DE differential expression, endo endothelial, FcγR Fc gamma receptor, GM gray matter, GML gray matter lesion, HLA-DR human leukocyte antigen-DR variant, MBP myelin basic protein, MS multiple sclerosis, NAGM normal-appearing gray matter, NAWM normal-appearing white matter, OPC oligodendrocyte precursor cell, peri pericyte, prolif proliferating, PVM perivascular macrophage, RL remyelinating lesion, snRNA-seq single-nuclei ribonucleic acid sequencing, WM white matter