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Mammalian fertility with aging is thought to decrease due to an increase in aneuploidy, abnormal mitochondrial function, and changes in the expression of oocytes. Although previous studies have reported that naturally aged over 8-month-old female mice have lower fertility, the mechanism of egg quality deterioration is still unknown. In this study, we focus on the effects of aging on ovarian tissue and fertility using young mice (2 months old) and aged mice (8 months old) and clarify the mechanism of age-related deterioration of egg quality. When we examined the fertility by natural mating, the number of pups produced by aged mice was significantly decreased. Histological analysis of ovaries displayed a decrease in the number of remaining follicles, and ovulated oocytes ovulated number by superovulation treatment was also significantly reduced in aged mice. Examination of the fertilization ability in vitro revealed a significant decrease in the fertilization rate in aged mice, whereas there was no difference in gene expression in MII eggs by RNA-seq. These results suggest that the age-related decrease in fertilization ability of mouse oocytes is not due to gene expression in the oocytes themselves, but to inhibition of sperm binding to surface of the zona pellucida. [ABSTRACT FROM AUTHOR]

Incorporation of bovine serum-derived albumin formulation (AlbuMAX) into a basic culture medium, MEMα, enables the completion of in vitro spermatogenesis through testicular tissue culture in mice. However, this medium was not effective in other animals. Therefore, we sought an alternative approach for in vitro spermatogenesis using a synthetic medium without AlbuMAX and aimed to identify its essential components. In addition to factors known to be important for spermatogenesis, such as retinoic acid and reproductive hormones, we found that antioxidants (vitamin E, vitamin C, and glutathione) and lysophospholipids are vital for in vitro spermatogenesis. Moreover, based on our experience with microfluidic devices (MFD), we developed an alternative approach, the PDMS-ceiling method (PC method), which involves simply covering the tissue with a flat chip made of PDMS, a silicone resin material used in MFD. The PC method, while straightforward, integrates the advantages of MFD, enabling improved and uniform oxygen and nutrient supply via tissue flattening. Furthermore, our studies underscored the significance of lowering the oxygen concentration to 10-15%. Using an integrated cultivation method based on these findings, we successfully achieved in vitro spermatogenesis in rats, which has been a long-standing challenge. Further improvements in culture conditions would pave the way for spermatogenesis completion in diverse animal species.