This study investigated the effect of recombinant human lactoferrin (rhLF) on the premature ovarian failure (POF) of rats. After cyclophosphamide treatments, the POF rats were divided into the following groups: normal control group (NC), low-dose group (LD), medium-dose group (MD) and high-dose group (HD) of rhLF. After drug administrations, the ovarian indexes and hormonal levels were detected. After follicle number count, the proliferation and apoptosis were analyzed with the expressions of genes related with oogenesis, reactive oxygen species (ROS) production and apoptosis detected, followed by the calculation of oxidative stress and protein expressions. After 4-hydroperoxy cyclophosphamide (4-HC) treatments, the effect of rhLF on the proliferation, ROS production and gene expressions of primary rat granulosa cells (GCs) cultured in vitro were detected. After mating, the fertilities of POF rats were recorded. The result showed that the rhLF administrations up-regulated the ovarian index with the number of developing follicles increased and the decreases of hormonal levels conferred. The Ki-67 intensities of the MD and HD groups were up-regulated with the Tunnel intensities decreased. The rhLF treatments significantly promoted the expression of oogenesis, antioxidant and anti-apoptosis related genes. The expression of Bax and Caspase 3 were decreased with the expression of Bcl-2 up-regulated after rhLF administrations. The in vitro treatments of rhLF effectively conferred the toxicity of 4-HC on primary rat GCs. The fertility assessment showed the rhLF treatments up-regulated the offspring’s’ folliculogenesis, which confirmed the ameliorative role of rhLF on the POF damages via the inhibition of ROS production in GCs.

Ovarian organoids, based on mouse female germline stem cells (FGSCs), have great value in basic research and are a vast prospect in pre-clinical drug screening due to their properties, but the competency of these in vitro-generated oocytes was generally low, especially, in vitro maturation (IVM) rate. Recently, it has been demonstrated that the 3D microenvironment triggers mitochondrial dysfunction during follicle growth in vitro. Therefore, therapies that protect mitochondria and enhance their function in oocytes warrant investigation. Here, we reported that exposure to 100 nM MitoQ promoted follicle growth and maturation in vitro, accompanied by scavenging ROS, reduced oxidative injury, and restored mitochondrial membrane potential in oocytes. Mechanistically, using mice granulosa cells (GCs) as a cellular model, it was shown that MitoQ protects GCs against H₂O₂-induced apoptosis by inhibiting the oxidative stress pathway. Together, these results reveal that MitoQ reduces oxidative stress in ovarian follicles via its antioxidative action, thereby protecting oocytes and granulosa cells and providing an efficient way to improve the quality of in vitro-generated oocytes.

Accumulating evidences revealed the connections between arsenic exposure and mitochondrial dysfunctions induced reproductive toxicology. Meanwhile, production declines were found in livestock suffering from arsenic exposure. However, the connections between arsenic exposure and livestock meiotic defects remain unclear. In this study, the effects of sodium arsenite (NaAsO₂) exposure during the in vitro maturation (IVM) on the meiotic potentials of ovine oocytes were analyzed. Furthermore, the effects of glutathione (GSH) supplementation on the meiotic defects of NaAsO₂ exposed ovine oocytes were investigated by the assay of nuclear maturation, spindle organization, chromosome alignment, cytoskeleton assembly, cortical granule (CGs) dynamics, mitochondrial dysfunctions, reactive oxygen species (ROS) accumulation, oxidative DNA damages, cellular apoptosis, epigenetic modifications and fertilization capacities. The results showed that the meiotic defects of NaAsO₂ exposed ovine oocytes were effectively ameliorated by the GSH supplementation via the inhibition of mitochondrial dysfunctions, which not only promoted the nuclear maturation, spindle organization, chromosome alignment, cytoskeleton assembly, CGs dynamic and fertilization capacities, but also inhibited the ROS accumulation, oxidative DNA damages and apoptosis of ovine MII oocytes. The abnormal expressions of 5mC, H3K4me3 and H3K9me3 in NaAsO₂ exposed ovine oocytes, indicating the abnormal epimutations of DNA methylation and histone methylation, were also effectively ameliorated by the GSH supplementation. Taken together, this study confirmed the connections between arsenic exposure and meiotic defects of ovine oocytes. Meanwhile, the effects of GSH supplementation on the developmental competence of livestock oocytes, especially for these suffering from arsenic exposure were also founded, benefiting the extended researches for the GSH applications.

Regardless of the essential role of copper (Cu) in the physiological regulation process of mammalian reproduction, excessive exposure to Cu triggers the meiotic defects of porcine oocytes via compromising the mitochondrial functions. However, the connections between the excessive Cu exposure and meiotic defects of ovine oocytes have not been reported. In this study, the effect of copper sulfate (CuSO₄) exposure on the meiotic potentials of ovine oocytes was analyzed. Subsequently, the ameliorative effect of glutathione (GSH) supplementation on the meiotic defects of CuSO₄ exposed ovine oocytes was investigated. For these purposes, the in vitro maturation (IVM) of ovine cumulus oocyte complexes (COCs) was conducted in the presence of 5, 10, 20 and 40 μg/mL of CuSO₄ supplementation. Subsequently, different concentrations of GSH (2, 4 and 8 mM) were added to the IVM medium containing CuSO₄ solution. After IVM, the assay, including nuclear maturation, spindle organization, chromosome alignment, cytoskeleton assembly, cortical granule (CGs) dynamics, mitochondrial function, reactive oxygen species (ROS) generation, apoptosis, epigenetic modification and fertilization capacity of ovine oocytes were performed. The results showed that excessive Cu exposure triggered the meiotic defects of ovine oocytes via promoting the mitochondrial dysfunction related oxidative stress damage. Moreover, the GSH supplementation, not only ameliorated the decreased maturation potential and fertilization defect of CuSO₄ exposed oocytes, but inhibited the mitochondrial dysfunction related oxidative stress damage, ROS generation, apoptosis and altered H3K27me3 expression in the CuSO₄ exposed oocytes. Combined with the gene expression pattern, the finding in the present study provided fundamental bases for the ameliorative effect of GSH supplementation on the meiotic defects of CuSO₄ exposed oocytes via inhibiting the mitochondrial dysfunctions, further benefiting these potential applications of GSH supplementation in the mammalian IVM system and livestock breeding suffering from the excessive Cu exposure.