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. 2024 Jan 10;625(7996):768–777. doi: 10.1038/s41586-023-06899-4

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© The Author(s) 2024

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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Extended Data Fig. 15 — a, Diagram of the experimental sequence for intraperitoneal (i.p.) injection of 200 μg of IgG or αIFNAR1 into aged (70-88 weeks old) C57BL/6J or Prox1-GFP mice every 72 hr for 6 weeks followed by intracisternal (i.c.) infusion of TMR-Dextran at 1.0 μl/min for 3 min and measurement of TMR-Dextran fluorescence in deep cervical lymph nodes (dcLN) 30 min later. b,c, Fluorescence microscopic images comparing amount of TMR-Dextran fluorescence in dcLNs (outlined by green dashed lines) after IgG or αIFNAR1. Scale bars, 200 μm. Each dot is the value for one mouse; n = 8 mice/group in two independent experiments. Error bars indicate mean ± s.e.m. AU, arbitrary unit. AU (arbitrary unit) normalized to mean IgG value. P values were calculated by two-tailed Mann-Whitney test. d,e, Immunofluorescence images of nasopharyngeal plexus whole mounts stained for Prox1-GFP, VEGFR3, and LYVE1 after IgG or αIFNAR1 administered to aged mice for 6 weeks. Scale bars, 200 μm. No differences between the groups are evident in lymphatic area, LYVE1 intensity, or lymphatic valves. Each dot is the value for one mouse; n = 5 mice/group from two independent experiments. Bars indicate mean ± s.e.m. AU, arbitrary unit. AU (arbitrary unit) normalized to mean control value = 1.0. P values were calculated by two-tailed Mann-Whitney test. f,g, Diagram of the experimental sequence for validating inhibition of interferon type I signalling by i.p. injection of 200 μg of αIFNAR1 antibody into adult (8-12 weeks old) C57BL/6J mice. One hour after injection of IgG or αIFNAR1 antibody, 300 μg of cGAMP was injected i.p. to stimulate type I interferon signalling. At 4 hr, nasopharyngeal tissue was removed for real-time PCR analysis of interferon-stimulating genes, Oas2 and Mx2. The blocking antibody reduced Oas2 and Mx2 expression to the PBS baseline. Each dot is the value for one mouse; Oas2 mRNA level and Mx2 mRNA level (PBS+IgG, n = 4; cGAMP+IgG, n = 5; cGAMP+ αIFNAR1, n = 5) in two independent experiments. Error bars indicate mean ± s.e.m. AU, arbitrary unit. P values were calculated by Brown-Forsythe ANOVA and two-tailed Dunnett’s T3 multiple comparison test.