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. 2021 Feb 3;11:2890. doi: 10.1038/s41598-021-82346-6

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© Crown 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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/.

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NgR1 and NgR3 are localized to B-cells within lymphoid-like follicles of the spinal cord. (a) Western blot of spleen and spinal cord extracts from ngr1^+/+ and ngr1^−/− mice showcasing the expression of NgR1 and NgR3. β-actin was used as a loading control. (b) Semi-quantitative analysis of both NgR1- and NgR3-positive B-cells per unit area (mm^y) of spleen and spinal cord infiltrates for the ngr1^+/+ and ngr1^−/− mice (n = 34 ngr1^+/+ mice; 108 sections, n = 15 ngr1^−/− mice; 40 sections) during the onset of EAE. These B-cells demonstrated a highly inducible receptor expression (student’s t test with Bonferroni correction ***p < 0.001 and **p < 0.01) compared to controls. (c) Illustration of inflammatory infiltrates analyzed via immunofluorescence immunolabeling of follicle-like structures (FLS) within the lumbosacral spinal cords of EAE-induced mice in (d–k). (d) Representative NgR1 and B220 double-labeled images showing the expression on clusters of B-cells localized to infiltrates around the leptomeninges during the onset of EAE in ngr1^+/+ mice. (e) Representative NgR3 and B220 double-labeled images showing the expression on clusters of B-cells localized to infiltrates around the leptomeninges during the onset of EAE in ngr1^+/+ mice (hatched boxes). (f) Representative images for the ngr1^−/− mice, highlighting the absence of the NgR1 protein expression in clusters of B-cells in similar infiltrates of the leptomeninges during the onset of EAE (hatched boxes). (g) Representative images for the ngr1^−/− mice, highlighting the strong expression of the NgR3 protein expression in clusters of B-cells in similar infiltrates of the leptomeninges during the onset of EAE (hatched boxes). (h) Representative NgR1 and CD19 double-labeled images showing the expression on clusters of B-cells localized to infiltrates around the leptomeninges during the onset of EAE in ngr1^+/+ mice. (i) Representative NgR3 and CD19 double-labeled images showing the expression on clusters of B-cells localized to infiltrates around the leptomeninges during the onset of EAE in ngr1^+/+ mice. (j) Representative images for the ngr1^−/− mice, highlighting the absence of the NgR1 protein expression in clusters of CD19 + B-cells in similar infiltrates of the leptomeninges during the onset of EAE (hatched boxes). (k) Representative images for the ngr1^−/− mice, highlighting the strong expression of the NgR3 protein expression in clusters of CD19 + B-cells in similar infiltrates of the leptomeninges during the onset of EAE. Scale = 50 μm.