Fig. 1: ATV:4D9 induces temporally dynamic microglial states distinct from amyloid pathology by single-cell analysis.

a, Antibodies were detected by human IgG ELISA in whole brain lysates 1 day after IV dose of 10 mg kg⁻¹ ATV:4D9 or 4D9 (n = 5 mice). b, Designs for WT;TfR^mu/hu and App^SAA;TfR^mu/hu studies. Mice were injected with a single IV 10 mg kg⁻¹ dose of ATV:4D9 or ATV:ISO and sacrificed at indicated timepoints (n = 3 WT; TfR^mu/hu mice and n = 4 App^SAA;TfR^mu/hu mice). c, Integrated UMAP projection of 49,000 total cells from all mice in both studies. The WT;TfR^mu/hu dataset consisted of 102,043 cells, and the App^SAA;TfR^mu/hu dataset consisted of 74,758 cells (Extended Data Fig. 1a,b). Five distinct clusters of microglia were identified. d, Stacked bar plots of clusters distributed per group for both studies. Clusters are shown as percentages of the whole microglial compartment averaged for each biological replicate. e, UMAP projection split by group for cluster distribution. Data from WT;TfR^mu/hu mice are boxed with solid lines, and data from App^SAA;TfR^mu/hu mice are boxed with dashed lines. f, Heat map of average log₂FC in each cluster compared to the ‘homeostatic’ cluster. Pseudobulk expression was generated by summing every counts per gene. Each mouse was treated as a biological replicate, and differential expression (DE) was performed for each cluster versus ‘homeostatic’ using limma. g, Scatter plots of the comparison of versus ‘homeostatic’ log₂FC for every gene between clusters 3 and 2 (top) and clusters 3 and 5 (bottom). In the top-right quadrant, genes falling below the dashed line are more upregulated in cluster 3, whereas genes above the dashed line are further upregulated in cluster 2 (top) or cluster 5 (bottom). Canonical DAM genes and other genes of interest are highlighted in orange. h, Dot plot showing GSEA for each cluster using DE versus ‘homeostatic.’ Signatures were taken from the hallmark gene signatures collection. Dot size is inversely proportional to log₁₀(corrected P value), and color indicates direction. No dot indicates a non-significant result. FC, fold change.

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