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. 2023 Jul 21;14:4416. doi: 10.1038/s41467-023-39934-z

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© The Author(s) 2023

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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Fig. 4 — a Representative images show (upper panel) exemplary 3D reconstruction of fibrillar collagen and (lower panel) volume reconstructions of both fibrillar collagen and elastic fiber networks in the arterial media as detected by SHG and autofluorescence, respectively, in the PA of a donor, an LHD w/o PH patient, and a PH-LHD patient. b Box-and-whisker plots show the number of disconnected collagen particles, total surface area, and total volume of fibrillar collagen normalized to control and to image volume in the PA of a donor, an LHD w/o PH patient, and a PH-LHD patient. c–c’ Immunoblots show protein levels of Col I (138 kDa), full length (145 kDa), and C-terminal truncation product (32 kDa) of Col V, and loading control GAPDH (37 kDa) in biologically independent PA samples from donors (n = 6), LHD w/o PH (n = 6), and PH-LHD (n = 6) patients. Box-and-whisker plots show quantitative densitometric data of Col I and Col V expression (normalized to GAPDH and to the mean of the donor controls) in biologically independent PA samples from donors (n = 6), LHD w/o PH (n = 6), and PH-LHD (n = 6) patients. Collagen expression differs between groups as follows: Col I (p = 0.026 for PH-LHD vs. donor), Col V 145 kDa (p = 0.004 for LHD w/o PH vs. donor, and p = 0.002 for PH-LHD vs. donor), and Col V 32 kDa (p = 0.002 for PH-LHD vs. donor). d Box-and-whisker plots show area of anti-AGE immunostaining normalized to area of the image and to donor controls in biologically independent PA samples from donors (n = 10), LHD w/o PH (n = 8), and PH-LHD (n = 8) patients (p = 0.0014 for LHD w/o PH vs. donor and p = 0.0004 for PH-LHD vs. donor). e Representative images of the arterial media in a PA sample from a PH-LHD patient show co-localization of anti-AGE immunostaining (right) with fibrillar collagen (center). Similar co-localization was confirmed in biologically independent PA samples from n = 11 LHD w/o PH or PH-LHD patients. Box-and-whisker plots overlaid with dot plots (b–d) show individual data points, mean (rectangle), median (line within the box), lower and upper 25% quartiles (limits of the box), 1.5 IQR (whiskers), and outliers (if applicable). Statistics: b, d Unpaired two-tailed Kruskal–Wallis one-way ANOVA on ranks followed by pairwise multiple comparisons (Dunn’s test); c Unpaired two-tailed Mann–Whitney U test (vs. donor). Source data are provided as a Source Data file. Full scan blots are provided in Suppl. Fig. 12.