Journal: Aging Cell

Article Title: Spatial Transcriptomic Characteristics of the Aging Human Ovary

doi: 10.1111/acel.70288

Figure Lengend Snippet: Dynamic cell–cell interactions among ovarian cell types during aging. (A) Bar plot showing the overall number of cell–cell interactions across ages. (B) Sankey plot showing the altered percentage of cellular interactions in distinct cell types at each age. The percentage corresponding to age 12 and 42 is listed in brackets, respectively. (C) Chord plots showing the usage of ligand DLK1 and its receptor NOTCH3 . (D) Bar plots showing the expression level of DLK1 and NOTCH3 across cell types. Data are shown as mean ± SEM. (E) Left, immunofluorescence staining of DLK1 and STAR in follicular fluid samples from women of varying age. Right, scatter plot showing the correlation of the fluorescence density of DLK1 in STAR + TCs and the age of women. The Spearman's correlation coefficient (ρ) and correlation test value ( p ) are indicated. Scale bar, 50 μm. (F) Boxplots showing the expression level of DLK1 and NOTCH3 across ages. These ovarian RNA‐seq data from distinct ages are all obtained from GTEx database. (G) Left, spatial visualizations showing the cell type of STAR + TCs and CLDN5 + EDCs. Some regions are enlarged. Right, spatial visualizations showing the expression level of DLK1 and NOTCH3 , respectively. The spatial region is as same as the left panel. The scale of color bars is varied among ages and genes. (H) Immunofluorescence staining of DLK1 in STAR + TCs and NOTCH3 in CLDN + blood EDCs in the human ovary. Scale bar, 50 μm. (I) Bar plots showing the RT‐qPCR relative expression level of DLK1 (left), p16 ( CDKN2A ) (middle), and p21 ( CDKN1A ) (right) after overexpressing DLK1 within the human GC cell line. Data are shown as mean ± SEM. Two‐tailed t ‐test p values are calculated, in which * p < 0.05 and *** p < 0.01. (J) Bar plots showing the relative expression level of DLK1 (left) and NOTCH3 (right) using RT‐qPCR. Data are shown as mean ± SEM. Two‐tailed t ‐test p values are calculated, in which * p < 0.05 and ** p < 0.01. (K) Cartoon showing the main aging‐related findings revealed by our study.

Article Snippet: Overnight incubation with primary antibodies was performed at 4°C in a humidified chamber using the following antibodies: STAR (1:100; Santa Cruz, sc‐166821), DLK1 (1:100; Proteintech, 10636‐1‐AP), NOTCH3 (1:100; Absin, abs116318), NOTCH2 (1:100; Proteintech, KHC1062), CLDN5 (1:100; Thermofisher, 4C3C2), IgG (1:100; Proteintech, CL488‐10284), and PDGFRA (1:50; Santa Cruz, sc‐21789).

Techniques: Expressing, Immunofluorescence, Staining, Fluorescence, RNA Sequencing, Quantitative RT-PCR, Two Tailed Test