Cranial radiation-induced abscopal effects drive time-dependent testicular injury in mice

To investigate the time-dependent progression of cranial radiation-induced testicular injury in mice, focusing on apoptotic pathways and brain-testis axis involvement. Male mice were randomly divided into sham irradiation group (sham) and cranial irradiation group (IR) with 36 mice each group. The IR group received a single dose of 20 Gy cranial X-ray irradiation, while peripheral organs were shielded. Assessments of body weight, testis weight, and sperm parameters were conducted at 1, 2, and 4 weeks post-irradiation. Histological changes and apoptosis were evaluated via H&E staining and TUNEL assay, respectively. Cell-specific markers SOX9, WT1, SYCP3 and PRND were quantified by qRT-PCR, while apoptotic proteins Bcl-2, Bax and Cleaved-caspase 3 were examined by Western blot. Serum levels of brain injury biomarkers NSE and S100B were measured using ELISA. Compared with sham group, IR mice displayed distinct, time-dependent testicular and systemic alterations. At 1 week, no obvious testicular structural damage was observed, significant reductions in body and testis weight were accompanied by elevated serum NSE and S100B (NSE: sham: 102.1 ± 3.6, IR: 142.1 ± 16.5, t = 7.1, P < 0.05; S100B: sham: 123.9 ± 11.5, IR: 178.2 ± 23.8, t = 6.2, P < 0.05). By 2 weeks, disorganized spermatogenic cells, downregulated SYCP3 and PRND expression, and a marked increase in sperm abnormalities (sham: 33.46 ± 6.05, IR: 52.45 ± 3.35, t = 7.77, P < 0.05) were evident, alongside sustained weight loss and persistent elevation of NSE and S100B levels (NSE: sham: 110.6 ± 2.8, IR: 128.6 ± 3.2, t = 12.8, P < 0.05; S100B: sham: 131.5 ± 10.9, IR: 163.1 ± 5.9, t = 7.8, P < 0.05). At 4 weeks, severe testicular atrophy manifested, characterized by reduced seminiferous tubule diameter, further increased sperm abnormalities (sham: 42.64 ± 3.13, IR: 64.61 ± 8.18, t = 7.10, P < 0.05), decreased sperm count (sham: 8.55 ± 2.05, IR: 3.08 ± 1.36, t = 6.30, P < 0.05), upregulated Cleaved-caspase 3 protein expression, TUNEL-positive cells localized to spermatogonial stem cells (SSCs) niches. Concurrently, SOX9 and WT1 mRNA levels were significantly downregulated (SOX9: sham: 1.00 ± 0.31, IR: 2.12 ± 0.96, t =2.72, P < 0.05; WT1: sham: 1.00 ± 0.57, IR: 2.51 ± 1.34, t = 2.55, P < 0.05). Persistent reductions in body and testis weight, as well as sustained elevations in NSE and S100B, were observed throughout the study. Cranial radiation induces progressive, time-dependent testicular injury in mice via mechanisms mediated by the brain-testis axis, primarily targeting SSCs for apoptosis. These findings identify the brain-testis axis as a novel therapeutic target for mitigating reproductive toxicity in male patients undergoing cranial radiotherapy.