Mechanisms of cell senescence and apoptosis in cyclophosphamide-induced premature ovarian failure in rats
Jiaqi Wu, Yanmeng Wei, Qiangli Peng, Jing Zhu, Huacong Shi, Ting Zhao, Tao Yuan
Abstract
BACKGROUND: Premature ovarian failure (POF) is a clinical condition characterized by a diminished ovarian reserve occurring before the age of 40, significantly affecting female reproductive health. However, its exact pathogenesis remains unclear. This research aimed to examine the mechanisms of cyclophosphamide (CTX)-induced senescence and apoptosis in the ovarian and cerebral cortex tissues of rats to provide insights into delaying aging and protecting female reproductive health. METHODS: A rat model of POF was established by intraperitoneal injection of CTX. Model efficacy was evaluated by measuring ovarian volume, weight, estrogen, and anti-Müllerian hormone levels. Serum marker changes were detected via enzyme-linked immunosorbent assay (ELISA). Senescence and apoptosis in cerebral cortical and ovarian tissues were observed using β-galactosidase (SA-β-gal) staining and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. Western Blot and quantitative polymerase chain reaction (RT-qPCR) were employed to detect the expression levels of cell senescence- and apoptosis-related proteins and genes, verifying the correlation between POF and cellular senescence/apoptosis. RESULTS: High-dose CTX induced POF. In rats with POF, the levels of anti-Müllerian hormone (AMH), estradiol (E2), and vitamin D (VD) significantly decreased (P < 0.0001), whereas the levels of testosterone (T) and insulin (INS) significantly increased (P < 0.0001). The number of senescent and apoptotic-positive cells in the ovarian and cerebral cortex tissues of rats with POF was substantially augmented (P < 0.05; P < 0.01). Additionally, the expression of senescence-related proteins cyclin-dependent kinase inhibitor 1 A (CDKN1A), cyclin-dependent kinase inhibitor 2 A (CDKN2A), tumor protein p53 (P53), apoptosis-related protein BCL2-Associated X Protein (Bax), and cysteine-aspartic acid protease 3 (caspase 3) was upregulated. In contrast, the expression of the anti-apoptotic protein BCL-2 was downregulated. The changes ranged from 1.7- to 7.1-fold. These findings demonstrated that high-dose CTX injection leads to cellular senescence and apoptosis, resulting in ovarian pathology. CONCLUSION: High-dose CTX induced POF in rats, resulting in aging and apoptosis in the cerebral cortex and ovarian tissues. Therefore, inhibiting cellular senescence and apoptosis may be a potential approach for restoring ovarian reserve function in POF.