The effect of Lycium barbarum polysaccharides (LBP) on the vitrification cryopreservation efficiency of bovine oocytes.

The vitrification cryopreservation of bovine oocytes is the cornerstone of modern animal breeding and genetic resource conservation. Cryogenic injury remains a major obstacle to efficient bovine oocyte vitrification, as it compromises both oocyte viability and subsequent developmental competence. In the present study, single-cell RNA sequencing (scRNA-seq) was employed to characterize transcriptional alterations in vitrified immature bovine oocytes. Moreover, the potential protective role of Lycium barbarum polysaccharides (LBP) during the vitrification process was further investigated. The results of single-cell RNA sequencing revealed the overall transcriptomic changes following vitrification treatment. The main features were mitochondrial and endoplasmic reticulum (ER) dysfunction, which led to energy metabolism disorders and dysregulation of key signaling pathways. To alleviate such damage, different concentrations of LBP (0, 200, 400 and 800 μg/mL) were added to the vitrification solution. The results showed that 400 μg/mL was the optimal concentration, which significantly increased the survival rate, maturation rate, cleavage rate and blastocyst rate after rewarming (P < 0.05). At the cellular and targeted molecular levels, LBP treatment was associated with lower intracellular reactive oxygen species levels, restored glutathione content, improved mitochondrial membrane potential and ATP levels, reduced endoplasmic reticulum stress markers, and a more favorable BCL2/BAX ratio (P < 0.05). Because transcriptomic analysis was not performed in LBP-treated oocytes, these findings indicate cellular responses associated with LBP supplementation rather than establishing a transcriptome-based mechanism. In conclusion, scRNA-seq identified vitrification-associated alterations involving mitochondrial and endoplasmic reticulum dysfunction. Under the current experimental conditions, LBP supplementation was associated with partial improvements in cellular homeostasis and developmental competence. Further transcriptomic analysis of LBP-treated oocytes is required to determine the molecular pathways directly involved in these effects.