# Acute high-dose irradiation disrupts cell adhesion and Silk-Ovarioid formation in human primary ovarian cells

## TL;DR

This study shows that high-dose radiation affects ovarian cell adhesion and 3D structure formation, even without killing the cells.

## Contribution

The study reveals a new mechanism of radiation-induced ovarian damage unrelated to cell death.

## Key findings

- Irradiation altered DNA damage response, apoptosis, and cell cycle regulation in ovarian cells.
- Transcriptomic and proteomic changes indicated disrupted cell adhesion and signaling pathways.
- Irradiated cells had impaired ability to form 3D Silk-Ovarioids, suggesting compromised cell-cell adhesion.

## Abstract

Radiotherapy is a cornerstone of cancer treatment; however, its effects on healthy ovarian somatic cells remain largely unexplored. This study addresses this gap by investigating how human cortical and medullary primary ovarian cells (cPOCs and mPOCs, respectively) respond to acute, high-dose X-ray exposure in vitro.

Ovarian tissue was obtained from eight patients (aged 23–36 years) undergoing gender-affirming surgery at Karolinska University Hospital in Huddinge, Sweden. The tissue was separated into cortex and medulla and dissociated into cPOCs and mPOCs. Monolayer cultures of cPOCs and mPOCs were exposed to 10 Gy X-rays upon reaching confluency, or left unexposed as paired controls. Following irradiation, cells were assessed for ATP content and mitochondrial dehydrogenase activity, followed by immunofluorescence staining, bulk RNA sequencing (Illumina Stranded mRNA Prep Ligation protocol; sequencing on the Illumina NovaSeq 6000 platform), bulk proteomic analysis (liquid chromatography–tandem mass spectrometry), and a functional assay for assessing their ability to form 3D Silk-Ovarioids.

While irradiation did not significantly affect cell viability, immunofluorescence analyses revealed alterations in DNA damage response, apoptosis, and cell cycle regulation. Transcriptomic analysis showed minimal changes at 1 h post-irradiation in both cPOCs and mPOCs. However, marked shifts in transcriptomic profiles were observed at 4 h (2,810 and 2,540 DEGs in cPOCs and mPOCs, respectively) and at 24 h (2,462 and 2,802 DEGs, respectively), including upregulation of the p53 pathway and downregulation of MYC targets, E2F targets, the G2/M checkpoint, and the mTORC1 pathway. At the proteomic level, differentially expressed proteins associated with cell adhesion, focal adhesion, and cadherin binding were detected at 24 h post-irradiation. Functionally, irradiated cells demonstrated an impaired capacity to self-organize into 3D Silk-Ovarioids, indicating compromised cell–cell adhesion.

These findings reveal a novel mechanism by which radiotherapy may damage ovarian tissue independently of follicular loss, underscoring the need for targeted strategies to preserve somatic cell function in fertility preservation protocols.

The online version contains supplementary material available at 10.1186/s13048-025-01932-8.