Fig. 4: Spotiphy reveals microglia regional specification in AD mouse brain tissue.

a, UMAP of 1,058 microglia extracted from iscRNA data generated by Spotiphy to mouse brain Visium data. Clusters are labeled according to their corresponding topographic regions. b, Transcriptomic spots from the Visium data. Spots are color coded according to their constituent microglia’s subclusters, annotated in a. The opacity of the spots is representative of cell density. Spots without microglia were removed for clarity. Black boxes represent the corresponding FOVs of the CosMx data (FOV1: AD CTX; FOV 2: AD CTX; FOV6: AD HPF; FOV8: AD TH; FOV11: WT CTX; FOV15: WT HPF; FOV19: WT TH); scale bar, 500 μm. c,d, Violin plots of the microglia and DAM scores for each microglia subcluster identified from the iscRNA (c; the P value for the C&H region is <\({2.2\times 10}^{-16}\), and the P value for the TH region is \({1.8\times 10}^{-4}\)) and CosMx (d; P < \({2.2\times 10}^{-16}\)) data. e, Visualization of signature genes in selected FOVs of the CosMx data. Blue and red dots represent the signature genes used to calculate the microglia and DAM scores, respectively. f,g, Violin plots of the C&H and TH scores for the microglia subclusters present in the FAD sample iscRNA (f; all P values are <\({2.2\times 10}^{-16}\)) and CosMx (g; top CTX–TH: P = \({5.5\times 10}^{-4}\), HPF–TH: P = \({7.8\times 10}^{-5}\); bottom CTX–TH: P = \({8.1\times 10}^{-3}\), HPF–TH: P = \({3.7\times 10}^{-3}\)) data. h, Visualization of signature genes in selected FOVs of the CosMx data. Yellow and green dots represent the signature genes used to calculate the TH and C&H scores, respectively. Cell boundaries are depicted with gray lines; microglia are outlined in white. CosMx data in e and h include one biological replicate each for WT and AD samples (Supplementary Fig. 2d); scale bars, 100 μm (e and h). Data in c, d, f and g were analyzed by two-sided t-tests; ***P < 0.01.

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