The accumulation of nanoparticles (NPs) in the female body has raised global concerns regarding potential effects on the reproductive system. This study aimed to investigate the toxic effects of nano-titanium dioxide (nano-TiO₂) exposure on the ovaries and the underlying mechanisms. By establishing a nano-TiO₂ accumulation model in mice, our research systematically evaluated the effects of different concentrations of nano-TiO₂ exposure on the development and reproductive endocrine functions of mice. The results showed that nano-TiO₂ exposure significantly reduced the littering rate, sex hormone levels, and ovarian index of mice, and the effects were dose-dependent. Studies on the mechanisms involved revealed that nano-TiO₂ induces an excessive production of reactive oxygen species (ROS), leading to the potential collapse of the mitochondrial membrane and an increase in the apoptosis rate of granulosa cells, thereby triggering oxidative stress and inhibiting the expression of ovarian-specific genes and granulosa-cell function genes. This study reveals the “dual blow” mechanism of nano-TiO₂-mediated ovarian morphology and function through oxidative stress in granulosa cells, namely directly disrupting cellular homeostasis and interfering with the reproductive-related gene network, ultimately leading to decreased ovarian function. This provides experimental evidence for assessing the reproductive risks of nanomaterials in women.

Ovarian tissue cryopreservation addresses critical challenges in fertility preservation for prepubertal female cancer patients, such as the lack of viable eggs and hormonal deficiencies. However, mitigating follicle and granulosa cell damage during freeze-thaw cycles remains an urgent issue. Luteinizing hormone (LH), upon binding to luteinizing hormone receptors (LHR) on granulosa cells, enhances estrogen synthesis and secretion, contributing to the growth of granulosa cells and follicles. This study examined mouse ovarian follicles and granulosa cells to identify optimal LH treatments using morphological assessments and LIVE/DEAD assays. The study found significant increases in the expression of Leucine-rich G-protein-coupled receptor 5 (Lgr5) and Forkhead box L2 (Foxl2) in mural and cumulus granulosa cells under LH influence, alongside marked reductions in active caspase-3 expression. Double immunofluorescence of Ki67 with Foxl2 and Lgr5 revealed ongoing proliferative activity in granulosa cells post freeze-thaw. In addition, LH treatment significantly boosted the expression of transforming growth factor (TGF-β) and its superfamily members in both granulosa cells and oocytes. These findings suggest that LH addition during cryopreservation can diminish damage to follicles and granulosa cells, offering new strategies to enhance the efficacy of mammalian ovarian cryopreservation.

While the mesonephros persists in the adult mouse ovary and contributes to folliculogenesis, its inductive role in oogonial stem cell (OSC) differentiation remains uncertain. In this study, we investigated whether mesonephros-derived cells direct OSCs toward follicle-like structures in vitro. OSCs were co-cultured with mesonephros cells, and morphological assessments and factor expression analyses were conducted at multiple time points. On day 14 of co-culture, 10× Genomics single-cell and bulk RNA sequencing were performed to profile transcriptomic changes, while renal capsule transplantation was conducted to evaluate the functional potential of co-cultured clusters in recipient mice. Transcriptomic analyses revealed a robust expression of mesonephros-specific genes and a significant enrichment of PI3K/AKT pathway-associated genes in co-cultured cells. Notably, transplanted clusters exhibited signs of estrogen secretion with maintained graft integrity. Collectively, these findings demonstrate that the mesonephros promotes OSC differentiation into follicle-like structures, with the PI3K/AKT pathway identified as a candidate mediator of this process. Our study provides novel insights into the developmental regulation of ovarian folliculogenesis and offers a valuable experimental model for future investigations.