Fig. 5: Reactive microglia are spatially associated with vulnerable neurons and plaques.

Average number of CD68⁺GAD1⁺ cells (cluster_15) per sample group in sections from non-disease control or AD donors homozygotic for the TREM2 common allele or heterozygotic for TREM2 R62H or R47H risk variants with the latter analysed together (a; CtrlCV, n = 6; CtrlTREM2, n = 6; AlzCV, n = 18; AlzTREM2, n = 13) or separately (b; CtrlCV, n = 6; CtrlTREM2, n = 6; AlzCV, n = 18; AlzR62H, n = 7; AlzR47H, n = 6). c Triple immunostaining for Iba1 (red), GAD1 (green), CD68 (grey) and DAPI (blue) in CtrlCV and AlzTREM2 donor sections with showing triple positive Iba1⁺CD68⁺GAD1⁺cells (yellow arrowhead). d Orthogonal projection of the region indicated by the white dashed square in (c). e Cell-cell interaction analyses between microglia and inhibitory or excitatory neuronal clusters from all samples analysed were performed with the buildSpatialGraph function. The calculated interaction score “sum_sigval” indicates the relative interactions between (>0) or relative proximity avoidance (<0) of the selected cell types. f Densities of CD68⁺GAD1⁺ cells by cortical layer (cluster_15) in CtrlCV (n = 6), CtrlTREM2 (n = 6), AlzCV (n = 18) and AlzTREM2 (n = 13) samples. g Layer-specific density of Aβ⁺ mask (shown in Fig. S8.i) generated from the 4G8 Aβ channel to identify plaques and their total areas within the sections of CtrlCV (n = 6), CtrlTREM2 (n = 6), AlzCV (n = 18), and AlzTREM2 (n = 13) samples. Quantification was performed on three ROIs acquired from a single section of each sample and pooled together before performing statistical analyses between groups. Statistical significance estimates were calculated with Dirichlet regression (a, b) or either ANOVA and two-sided Tukey tests or Kruskal–Wallis and two-sided Wilcoxon signed-rank test depending on whether groups showed normal or non-normal distributions, respectively (f, g). Boxplots show median (middle line), interquartile range (box) and variability outside of the first and third quartiles (lines extending from box). P values are indicated as: non-significant, ns, p > 0.05; *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001. Source data are provided as a Source Data file. Scale bars in (c) represent 100 μm.