Low-density small extracellular vesicles in bovine follicular fluid carrying let-7i target FASLG to inhibit granulosa cells apoptosis.

Apoptosis of granulosa cells is a key factor in mammalian follicular atresia. It has a significant impact on oocyte development and maturation. Extracellular vesicles (EVs) are a group of highly heterogeneous population. Previous studies have found that ovarian follicular fluid is rich in EVs. In the present study, the follicular fluid of 3-5 mm follicles from bovine ovaries without corpus luteum was collected, and a subtype of small EVs (sEVs), low-density small EVs (LD-sEVs), was successfully isolated by differential ultracentrifugation combined with iodixanol density gradient centrifugation. LD-sEVs were identified using transmission electron microscope, nanoparticle tracking analysis and Western blot. Flow cytometry, Quantitative reverse transcription PCR (RT-qPCR), Western blot, and other methods were used to detect the effect of LD-sEVs on follicular granulosa cell apoptosis. After that, let-7i, a highly expressed miRNA component in LD-sEVs, was screened and target validation was carried out in granulosa cells. The results showed that LD-sEVs could be taken up by granulosa cells and inhibited the apoptosis. Further research found that let-7i inhibits the apoptosis of granulosa cells by targeting FASLG. It plays an important role in regulating the apoptosis of follicular granulosa cells, which may affect follicular development.