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. 2022 Nov 18;14(12):888–907. doi: 10.1093/procel/pwac057

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©The Author(s) 2022. Published by Oxford University Press on behalf of Higher Education Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 1. — Aging-related phenotypes of the cynomolgus monkey testes. (A) Flow chart of experimental design of testicular aging study in cynomolgus monkeys. (B) Masson-trichrome staining analysis of testicular tissues from young and aged monkeys. Representative images are shown on the left. Quantitative data are shown as means ± SEM on the right. Fibrosis area is quantified as fold changes (Aged vs. Young). Young, n = 4; aged, n = 4 monkeys. Scale bars, 50 μm and 25 μm (zoomed-in image). (C) Sudan black B staining of testicular tissues from young and aged monkeys. Representative images are shown on the left. Quantitative data are shown as means ± SEM on the right. Lipofuscin-positive area is quantified as fold changes (Aged vs. Young). Young, n = 4; aged, n = 4 monkeys. Scale bars, 50 μm and 12.5 μm (zoomed-in image). (D) Oil Red O staining of testicular tissues from young and aged monkeys. Representative images are shown on the left. Quantitative data are shown as means ± SEM on the right. Oil Red O-positive area is quantified as fold changes (Aged vs. Young). Young, n = 4; aged, n = 4 monkeys. Scale bars, 50 μm and 12.5 μm (zoomed-in image). (E) Immunofluorescence analysis of type IV Collagen in basal membrane of seminiferous tubules. Representative images are shown on the left. Quantitative data are shown as means ± SEM on the right. Thickness of basement membrane (BM, marked by collagen IV) is quantified as fold changes (Aged vs. Young). Young, n = 4; aged, n = 4 monkeys. Scale bars, 20 μm and 5 μm (zoomed-in image). (F) TUNEL analysis of apoptotic cells in testicular tissues from young and aged monkeys. Representative images are shown on the left. TUNEL-positive cells are quantified as fold changes (Aged vs. Young) and shown on the right. Young, n = 4; aged, n = 4 monkeys. Scale bars, 20 μm and 5 μm (zoomed-in image). (G) SA-β-Gal staining of testicular tissues from young and aged monkeys. Representative images are shown on the left. SA-β-Gal-positive areas are quantified as fold changes (Aged vs. Young), and shown as means ± SEM on the right. Young, n = 4; aged, n = 4 monkeys. Scale bar, 300 μm and 50 μm (zoomed-in image). (H) SPiDER-βGal staining of testicular tissues from young and aged monkeys. Representative images are shown on the left. SPiDER-βGal-positive areas in the tissues are quantified as fold changes (Aged vs. Young), and shown as means ± SEM on the right. Young, n = 4; aged, n = 4 monkeys. Scale bar, 20 μm (zoomed-in image). (I) Co-stainings of SA-β-Gal, collagen IV, UTF1, and WT1 in testicular tissues from aged monkeys. A schematic of the germinal epithelium of seminiferous tubule is shown on the left. Representative images are shown on the right. Scale bars, 20 μm and 5 μm (zoomed-in image).