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. 2021 Oct 11;11:20168. doi: 10.1038/s41598-021-99643-9

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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 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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Staining of nuclei, collagen and senescent cells of venous leg ulcer biopsies revealed a strong link between presence of senescent cells and collagen area. (a) Haematoxylin and eosin staining of nuclei (purple) and extracellular proteins (pink) in a venous leg ulcer. Montage of high-power images [1200 (W) × 1000 (H) μm] of an 8 μM cryosection thick—two from high senescence (HS) regions and two from low senescence (LS) regions of a venous leg ulcer taken using × 20 objective lens. (b) MT staining collagen (green), muscle and keratin (red) and cytoplasm (pink/red) in a sister section. Montage followed by (c) X-gal staining as a marker for senescent cells (senescent cells (blue), nuclei and cytoplasm (pink). Scale bar: 500 μm (montage) and 20 μm (high power). (d) SHG imaging of collagen (white) in × 20 for VLU and arm tissue. Scale bar: 200 μm. MT and X-gal images of the arm tissue are also shown. (e) Regression plots of collagen area (x-axis) against senescent cell number (y-axis) of individual patients of the VLU cohort. (f) Single regression plot of the VLU cohort.