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. 2023 Dec 21;15(5):364–384. doi: 10.1093/procel/pwad063

Figure 4.

Figure 4.

Transcriptional changes of several canonical aging hallmarks in NHP ovarian aging. (A) Schematic diagram showing the ovarian aging features unveiled by spatial transcriptomics. (B) Ridge plots (left) and spatial visualization (right) showing the global distribution density of gene set score of inflammatory response-related genes in young and old groups. The corresponding dashed lines represent the median of each group. (C) Ridge plots (left) and spatial visualization (right) showing the global distribution density of gene set score of SASP-related genes in young and old groups. The corresponding dashed lines represent the median of each group. (D) S100A9 immunofluorescence staining of ovaries from young and old monkeys. Representative images are shown on the left. Among them, “a” represents the staining in the non-follicle region, and “b” represents the staining in the follicle region. Scale bars, 200 μm and 20 μm (zoomed-in images). The numbers of S100A9-positive cells were quantified as fold changes (old vs. young), and presented as mean ± SEMs on the right. n = 4 monkeys for each group. (E) NF-κB p65 immunofluorescence staining of ovaries from young and old monkeys. Representative images are shown on the left. Among them, “a” represents the staining in the non-follicle region, and “b” represents the staining in the follicle region. Scale bars, 200 μm and 20 μm (zoomed-in images). The relative intensity was quantified as fold changes (old vs. young), and presented as mean ± SEMs on the right. n = 4 monkeys for each group. (F) Ridge plots (left) and spatial visualization (right) showing the global distribution density of gene set score of fibrosis-related genes in young and old groups. The corresponding dashed lines represent the median of each group. (G) Masson’s trichrome staining of young and old monkey ovaries. Representative images are shown on the left. Scale bar, 200 μm and 100 μm. The percentage of fibrosis area was quantified (old vs. young), and presented as mean ± SEMs on the right. n = 4 monkeys for each group. (H) Ridge plots (left) and spatial visualization (right) showing the global distribution density of gene set score of lipid storage-related genes in young and old groups. The corresponding dashed lines represent the median of each group. (I) ORO staining of ovaries from young and old monkeys. Representative images are shown on the left. Scale bars, 200 μm and 100 μm (zoomed-in images). The percentage of ORO-positive area was quantified was quantified as fold changes (old vs. young), and presented as mean ± SEMs on the right. n = 4 monkeys for each group. (J) Ridge plots (left) and spatial visualization (right) showing the global distribution density of gene set score of apoptosis related-genes in young and old groups. The corresponding dashed lines represent the median of each group. (K) TUNEL staining of ovarian tissues from young and old monkeys. Representative images are shown on the left. Among them, “a” represents the staining in the non-follicle region, and “b” represents the staining in the follicle region. Scale bar, 200 μm and 20 μm (zoomed-in images). The numbers of TUNEL-positive cells in the tissues were quantified as fold changes in old ovaries versus in young counterparts, and shown as mean ± SEMs on the right. n = 4 monkeys for each group. (L) Ridge plots (left) and spatial visualization (right) showing the global distribution density of gene set score of ROS-related genes in young and old groups. The corresponding dashed lines represent the median of each group. (M) 4-HNE staining of ovary tissues from young and old monkeys. Representative images are shown on the left. Among them, “a” represents the staining in the non-follicle region, and “b” represents the staining in the follicle region. The immunofluorescence expression intensity in the tissues was quantified as fold changes (old vs. young), and shown as mean ± SEMs on the right. n = 4 monkeys for each group. Scale bar, 200 μm and 20 μm (zoomed-in images). (N) Ridge plots (left) and spatial visualization (right) showing the global distribution density of gene set score of DNA repair-related genes in young and old groups. The corresponding dashed lines represent the median of each group. (O) γH2A.X staining of ovary tissues from young and old monkeys. Representative images are shown on the left. Among them, “a” represents the staining in the non-follicle region, and “b” represents the staining in the follicle region. γH2A.X-positive cells in the tissues were quantified as fold changes (old vs. young), and shown as mean ± SEMs on the right. n = 4 monkeys for each group. Scale bar, 200 μm and 20 μm (zoomed-in images). (P) Ridge plots (left) and spatial visualization (right) showing the global distribution density of gene set score of senescence-related genes in young and old groups. The corresponding dashed lines represent the median of each group. (Q) P21 staining of ovary tissues from young and old monkeys. Representative images are shown on the left. Among them, “a” represents the staining in the non-follicle region, and “b” represents the staining in the follicle region. P21-positive cells in the tissues were quantified as fold changes (old vs. young), and shown as mean ± SEMs on the right. n = 4 monkeys for each group. Scale bar, 200 μm and 20 μm (zoomed-in images).