Spatial Atlas of Aging Human Ovary Reveals Key Inflammatory Driver
Researchers mapped gene expression across aging ovaries, uncovering a novel endothelial cell subtype and a DLK1-NOTCH3 signaling axis that accelerates ovarian decline.
Summary
Using single-nucleus RNA sequencing and spatial transcriptomics across 12 human ovaries (ages 12–54), researchers built a comprehensive aging atlas of the ovary. They identified a novel endothelial cell subtype (CLDN5+ blood EDCs) that acts as a semi-professional antigen-presenting cell and becomes increasingly inflammatory with age. Unlike other ovarian cell types that lose identity during aging, these cells grow more transcriptomically reactive. Aging also disrupted global cellular communication while amplifying a DLK1:NOTCH3 signaling axis between theca cells and CLDN5+ endothelial cells. Immunoglobulin-expressing cells accumulated at the ovarian periphery with age. Elevated DLK1 was also found in granulosa cells from women with primary ovarian insufficiency, pointing to shared mechanisms and potential therapeutic targets.
Detailed Summary
Ovarian aging is a key driver of female infertility and is linked to systemic conditions including osteoporosis, cardiovascular disease, and Alzheimer's disease. Despite its clinical importance, the spatial and cellular architecture of how the ovary ages has remained poorly characterized. This study addresses that gap with unprecedented resolution.
Researchers integrated single-nucleus RNA sequencing (snRNA-seq) and a high-resolution spatial transcriptomics method (scStereo-seq, chip size 10×10 mm) across 12 human ovarian tissue samples spanning ages 12 to 54. Samples were grouped into prepubertal, young (23–29), middle-aged (32–34), and older-aged (42–54) cohorts. Four samples were collected from breast cancer patients undergoing fertility preservation; eight came from a published dataset of donors who experienced sudden death or bilateral salpingo-oophorectomy.
A major finding was the identification of a previously uncharacterized endothelial cell subtype—CLDN5+ blood endothelial cells—that functioned as semi-professional antigen-presenting cells. While most ovarian cell types progressively lost transcriptional identity with age, CLDN5+ blood EDCs showed heightened sensitivity to aging, marked by enhanced antigen-presenting capacity and amplified inflammatory signaling. This counterintuitive gain of function in a specific immune-adjacent cell type suggests a mechanism by which the aging ovary generates chronic low-grade inflammation.
Spatial mapping revealed that immunoglobulin-expressing cells (IGHG1+/IGKC+) accumulated preferentially in the ovarian periphery (cortex) with advancing age, suggesting age-related immune infiltration is spatially organized rather than diffuse. Global cellular connectivity declined with aging across most cell types, but one intercellular signaling axis was paradoxically amplified: DLK1 ligand expressed by theca cells interacting with NOTCH3 receptor on CLDN5+ blood EDCs. This DLK1:NOTCH3 axis intensified with age and is proposed as a key contributor to the dysregulation of ovarian folliculogenesis and endocrine function. Critically, elevated DLK1 expression was also detected in granulosa cells from patients with primary ovarian insufficiency (POI), linking the aging mechanism to a clinical pathology of premature ovarian failure.
Additional aging-associated transcriptomic changes included impaired mitochondrial oxidative phosphorylation and disrupted reproductive structure development in older ovaries. The study provides a detailed spatiotemporal atlas that surpasses prior work in coverage, resolution, and age-group breadth, and it identifies concrete molecular targets—particularly the DLK1:NOTCH3 axis—that could be amenable to therapeutic intervention to extend female reproductive lifespan or treat POI.
Key Findings
- A novel CLDN5+ blood endothelial cell subtype acts as a semi-professional antigen-presenting cell and gains inflammatory activity with age.
- The DLK1:NOTCH3 signaling axis between theca cells and CLDN5+ endothelial cells is amplified with aging, potentially disrupting ovarian function.
- Immunoglobulin-expressing (IGHG1+/IGKC+) immune cells accumulate preferentially at the ovarian periphery as age advances.
- Aging impairs mitochondrial oxidative phosphorylation and reproductive structure development across multiple ovarian cell types.
- DLK1 is upregulated in granulosa cells from patients with primary ovarian insufficiency, linking normal aging to POI pathology.
Methodology
The study used snRNA-seq (10× Genomics) and scStereo-seq spatial transcriptomics on 12 human ovarian tissue samples spanning ages 12–54, including 4 newly collected samples and 8 from a published dataset. Samples were grouped into four age cohorts (prepubertal, young, middle-aged, older-aged) for comparative transcriptomic and cell-communication analyses.
Study Limitations
The study includes only one prepubertal sample, limiting conclusions about early developmental baselines. Sample collection involved different sources (fertility preservation patients vs. published datasets from sudden death or oophorectomy), introducing potential confounders. Sample sizes per age group are small (1–5 donors), and functional validation of the DLK1:NOTCH3 axis in vivo is not yet provided.
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