KEY POINTS
- This preclinical study compared ultrahigh dose-rate X-ray irradiation at approximately 200 Gy/s with conventional irradiation at approximately 2 Gy/min in mice across 0-20 Gy, supported by spermatogonia cell experiments, transcriptomics, proteomics, and single-cell RNA sequencing.
- After 6 Gy, ultrahigh dose-rate irradiation preserved greater spermatogenic epithelial height than conventional irradiation on day 7 (p=0.0165), with more Ki-67-positive cells (p=0.0422) and fewer TUNEL-positive cells (p=0.0226).
- The histological advantage was transient and endpoint-dependent: no significant differences were observed in testicular weight or sperm count, while doses of 8 Gy or higher produced severe damage with both irradiation modalities.
- Glutathione S-transferase mu 3 knockdown worsened ultrahigh dose-rate-induced histological injury and reduced sperm counts, whereas overexpression improved tissue preservation and sperm counts. These effects were not significant after conventional irradiation.
- Glutathione S-transferase mu 3 regulated ferroptosis-associated markers, including GPX4, FTH1, ACSL4, and 4-HNE. Liproxstatin-1 partially reversed the injury caused by gene downregulation and improved sperm count (p=0.0491).
CLINICAL TAKEAWAY
The findings identify ferroptosis regulation through glutathione S-transferase mu 3 as a possible mechanism underlying early testicular sparing after ultrahigh dose-rate irradiation. However, the benefit was limited to selected histological endpoints in mice, without demonstrated preservation of fertility, making this hypothesis-generating rather than clinically actionable.