A Long-Acting Recombinant Follicle-Stimulating Hormone Supports High-Quality Mouse Follicle Development And Oocyte Maturation In Vitro

In vitro folliculogenesis may represent a valid option to convey the large source of immature ovarian follicles in assisted reproductive technology(ART).In this study,a new type of recombinant FSH(KN015)with a prolonged half-life is proposed as an alternative to canonical FSH.KN015 supports the in vitro mice follicle development from primary to preovulatory stage with a higher efficiency than normal FSH.The use of KN015 also allows us to compare the dynamic transcriptome changes of the oocytes and granulosa cells at different stages in vivo and in vitro.KN015 facilitates mRNA accumulation in growing oocytes and prevents spontaneous luteinization of granulosa cells in vitro.Novel analyses of transcriptome changes in this study reveals that the oocytes in vivo were more efficient than oocytes in terms of maternal mRNA clearing during meiotic maturation.KN015 promotes the degradation of maternalmRNA during in vitro oocyte maturation,improves cytoplasmic maturation,and therefore enhances the embryonic developmental potential.These findings established new transcriptome data of oocyte and granulosa cells at the key stages of follicle development,and should help to widen the use of KN015 as a valid and commercially available hormonal support enabling optimized in vitro follicle and oocyte development.Objective:To explore whether a new type of recombinant FSH(KN015)can support follicle culture in vitro and oocyte maturation in vitro more efficiency than normal FSH,and its mechanism of action.Methods:The transcriptome analysis of gene expression during oocyte and granule cells of follicles in vivo and in vitro;RT-PCR verifies the mRNA expression levels of oocyte and granulosa cell-related genes;immunofluorescence analysis of oocyte nucleus and spindle morphology.Research contents:1.Quantification and qualification of follicle maturation rate and oocyte maturation under different hormonal treatments.Exploring the appropriate concentration of FSH for follicle culture in vitro.2.Comparing dynamic transcriptome changes in oocytes and granulosa cells at different stages in vivo and in vitro.3.Expansion of cumulus-oocyte complexes(COCs)cultured in vitro and gene expression of oocytes and cumulus cells under different follicle stimulating hormone treatments.Results:Compared with Gonal-F,KN015 supported high-quality mouse follicle development with a higher rate of COCs(p<0.001)and more nucleolus-surrounded(SN)oocytes(p<0.001).After ovulation induction by follicle culture in vitro,the KN015 group obtained more MII-stage oocytes(p<0.001),and their spindle morphology was close to that in vivo(p>0.05).Transcriptome results showed that KN015 promoted mRNA accumulation in growing oocytes and prevented spontaneous luteinization of granulosa cells in vitro.And it was found that the degradation efficiency of maternal mRNA during oocyte maturation in vivo was higher than that of oocytes matured in vitro,and KN015 could better promote the degradation of maternal mRNA during oocyte maturation in vitro,and the oocyte’s cytoplasmic maturation.Conclusion:The method of in vitro mouse follicle culture has been greatly improved,the used of KN015 promoted the accumulation and degradation of mRNA in cultured oocytes.This method could be widely used for clinical treatment and basic research in the future.