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. 2021 Feb 24;12:1158. doi: 10.1038/s41467-021-21428-5

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© The Author(s) 2021

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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Fig. 6 — a EM analysis of microglia acutely isolated from pre-symptomatic WT and Npc1^−/− mice from three independent experiments (n = 3) shows the significant intracytoplasmic accumulation of MVBs in Npc1^−/− cells. Boxed regions are depicted at higher magnification in right panels (MVBs and lysosomes are pseudocolored in red and green, respectively). Scale bars: 2 μm (left panels) and 1 μm (right panels). b Representative high magnification images of Npc1^−/− cell shown in (a) (arrowheads 1 and 2) revealing accumulation of MVBs (1). Undigested lipid material (lamellar structures) could be detected within MVBs (2). Scale bars: 0.2 μm. c Transport of phagocytosed myelin (green) was followed over 1 h in P7 WT and Npc1^−/− microglia from three independent experiments (n = 3) using DQ-BSA to visualize lysosomal compartments (red) and fluorescently labeled cholera toxin (CTX, white) to visualize endocytic vesicles. Scale bar: 10 μm. d Quantification of the in vitro myelin transport assay shows that, in contrast to WT control, phagocytosed myelin did not reach lysosomal compartments in Npc1^−/− microglia. Values are expressed as percentages of myelin signal colocalizing with DQ-BSA positive lysosomes and represent mean ± SEM from three independent experiments (n = 3, p = 0.0003, unpaired two-tailed Student’s t-test).