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. 2022 Sep 30;25(10):1379–1393. doi: 10.1038/s41593-022-01167-6

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Extended Data Fig. 2 — a: Schematic of the bootstrapping effects on the distance between tree structures from the same population (within-population distance, green arrows) and two distinct populations (distance between average persistence images, purple lines). Increase of bootstrap-to-population size ratio (x/n) reduces within-population distance and increases distance between average persistence images. b: UMAP plots of MorphOMICs-analyzed microglia for frontal cortex (orange) and dentate gyrus (yellow) for different bootstrap-to-population size ratios. Left: x = 1, allows no segregation. Middle: x/n = 0.3. Right: x/n = 1 causes accentuation. c, d: Line plot ± SD displays how within-population distance (c, top) and mixing entropy (c, bottom), and the ratio between mixing entropy and within-condition distance (d) varies by enhancing bootstrap-to-population size ratio (x/n). An empirical threshold of 0.3 was selected (red dashed line). Data are presented as mean values ± SD. e: UMAP plots of MorphOMICs-analyzed microglia for frontal cortex (green) and dentate gyrus (purple) for different bootstrap-to-population size ratios and varying male-to-female ratios within the population size. f: UMAP plots of MorphOMICs-analyzed microglial morphology across seven brain regions as shown for Fig. 1f with examples of tested hyperparameters for number of neighbors (n_neighbors), minimum distance (min_dist), and spread. Olfactory bulb (OB), frontal cortex (FC), dentate gyrus (DG), somatosensory cortex (S1), substantia nigra (SN), cochlear nucleus (CN), and cerebellum (CB). g: tSNE plot of MorphOMICs-analyzed microglia across seven brain regions as shown in Fig. 1f. h: UMAP plots of stable ranks representation of microglial morphology (see Methods: Stable Ranks) across seven brain regions. n_samples = 500 per condition (see also Methods: Average and bootstrapped persistence images). SD: standard deviation.