Senescence-Linked Fibrosis in the Aging Human Ovary Revealed by p16-Based Histological Profiling and Spatial Transcriptomics

Watson, M. A.; Devrukhkar, P. R.; Murad, N. F.; Wu, F.; Kim, M. J.; Anvari, H.; Tran, U.; Martin, N.; Tran, T.; Zaza, G.; Schneider, K.; Soygur, B.; Shanes, E. D.; Wirtz, D.; Pavone, M. G.; Melov, S.; Wu, P.-H.; Furman, D.; Duncan, F. E.; Schilling, B.

doi:10.64898/2025.12.03.692228

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bioRxiv · 10.64898/2025.12.03.692228

Senescence-Linked Fibrosis in the Aging Human Ovary Revealed by p16-Based Histological Profiling and Spatial Transcriptomics

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Abstract

Cellular senescence is implicated as a driver of ovarian aging, but senescent cells in the human postmenopausal ovary remain poorly defined. Using spatially resolved p16INK4a protein expression, a canonical senescence marker, we identified and mapped senescent cells in postmenopausal ovaries. We integrated p16 immunohistochemistry, multiplexed immunofluorescence, spatial transcriptomics, and AI-guided digital pathology to map senescent microenvironments. p16-positive cells formed discrete stromal, vascular, and cyst-associated clusters that increased with age and were enriched for macrophages and myofibroblast-like cells. Wholetranscriptome profiling of 92 spatial regions uncovered a 32-gene p16-associated signature, BuckSenOvary, that distinguished p16-positive regions across cortex and medulla. BuckSenOvary is characterized by suppression of cell-cycle regulators and activation of inflammatory and extracellular-matrix remodelling genes. AI-based collagen matrix analysis confirmed that p16-positive regions exhibit more architecturally complex collagen, demonstrating that focal senescent microenvironments are fibro-inflammatory. These findings position senescent ovarian niches as therapeutic targets to preserve ovarian function.

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Watson, M. A., Devrukhkar, P. R., Murad, N. F., Wu, F., Kim, M. J., Anvari, H., Tran, U., Martin, N., Tran, T., Zaza, G., Schneider, K., Soygur, B., Shanes, E. D., Wirtz, D., Pavone, M. G., Melov, S., Wu, P.-H., Furman, D., Duncan, F. E., Schilling, B.. 2025-12-08. Senescence-Linked Fibrosis in the Aging Human Ovary Revealed by p16-Based Histological Profiling and Spatial Transcriptomics. https://doi.org/10.64898/2025.12.03.692228

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