Fig 2: Single-nucleus transcriptomic atlas of the primate gingiva during aging. (A) Uniform manifold approximation and projection (UMAP) plots showing the cell types of monkey gingiva identified in single-nucleus RNA sequencing. Left, UMAP plots showing the 14 cell types of monkey gingiva. Right, UMAP plots showing the distribution of different cell types during aging. YM, young male (n = 4); MM, middle-aged male (n = 3); OM, old male (n = 4). (B) Heatmap showing the expression profiles of the top 50 marker genes of each cell type in the monkey gingiva with their enriched functional annotations on the right. (C) Box plot showing the cell identity score of each cell type in the monkey gingiva during aging. The P-values are indicated in the graphs, with the blue letter representing a significant downregulation during aging, the red letter representing a significant upregulation, and the black letter representing not significant. (D) Augur analysis showing prioritization of the most responsive cell types during gingival aging. The black arrows indicate epithelial cells. (E) Cell proportion of each cell type in the monkey gingiva during aging. (F–I) Immunofluorescence staining of CD45 (F), CD79B (G), IGKC (H), and CD163 (I) in gingival tissues of monkeys. Representative images are shown on the left and quantitative data are shown as means ± SEM on the right. The P-values are indicated in the graphs and ns indicates not significant. YM: n = 6; MM: n = 3; OM: n = 8. Scale bars, 100 μm and 20 μm (zoomed-in images).

Fig 3: Single-nucleus transcriptomics identifies major cell types in cynomolgus monkey livers. (A) UMAP plot showing the distribution of different cell types in liver from young and aged monkeys. (B) Feature plots showing the expression profiles of indicated cell-type-specific marker genes in monkey liver. The color key from gray to red indicates low to high gene expression levels. (C) Dot plot showing the expression level of representative marker genes across cell types. The color key from gray to red presents low to high gene expression levels. The size of dots indicates the percentage of cells with gene expression greater than zero. (D) Heatmap showing the expression profiles of the top 50 cell-type-specific marker genes for each cell type in monkey livers, with corresponding functional annotations on the right. The color key from blue to red represents low to high gene expression levels. (E) Sankey plots showing the cell number of each cell type and the proportion in young and aged monkey livers. The length of the bar indicates the cell number of each cell type, and the number was marked above the pie plot. The pie chart at bottom showing the ratios of each cell type in young and aged monkey livers. (F) Immunofluorescence staining of CD163 in liver tissues from young and aged monkeys. Representative images are shown on the left; quantitative data for the percentage of CD163-positive cells are shown on the right. Scale bars, 20 μm. (G) Immunofluorescence staining of CD247 in liver tissues from young and aged monkeys. Representative images are shown on the left; quantitative data for the percentage of CD247-positive cells are shown on the right. Scale bars, 20 μm. (H) Immunofluorescence staining of CD79B in liver tissues from young and aged monkeys. Representative images are shown on the left; quantitative data for the percentage of CD79B-positive cells are shown on the right. Scale bars, 20 μm. (I) Dot plot showing the log2 ratio of transcriptional noise between aged and young samples. The color key from gray to red corresponds to Log10 (adjusted P value) of transcriptional noise ratio from low to high. The size of dots indicates the number of genes with aging-related transcriptional noise. (J) Heatmap showing the row scaled expression levels of genes with high Pearson’s correlation coefficients (correlation coefficient > 0.6 and FDR < 0.05) between transcriptional noise and expression profiles in hepatocytes from young and aged monkeys. The bins are arranged based on the transcriptional noise ranking in each group. (K) Bar chart showing the enriched GO terms and pathways of genes with increased transcriptional noise during aging in hepatocytes. (F–H) Data are quantified as fold changes and shown as means ± SEM. Young, n = 8 monkeys; aged, n = 8 monkeys. *P < 0.05.