In the mouse, the use of the DNA-binding fluorochrome Hoechst 33342 allows the classification of fully-grown antral oocytes into two categories distinguished by their chromatin conformation: surrounding nucleolus (SN) and not-surrounding nucleolus (NSN) oocytes, the former capable of completing development, the latter unable to proceed beyond the 2-cell stage. In the present study, time-lapse observation of SN and NSN oocyte GV-to-MII transition highlighted differences in the timing of germinal vesicle breakdown (GVBD) and polar body I (PB-I) extrusion. PB-I extrusion, but not GVBD, revealed the presence of three main groups of significantly different oocytes: Group A (456-576 min) comprising mainly SN oocytes (91.4%), group B (584-728 min) entailing an almost equivalent percentage of SN (52.7%) and NSN (47.3%) oocytes, whereas group C (736-896 min) consisting of almost all NSN (94.4%) oocytes. In a further set of time-lapse experiments, GV oocytes were in vitro matured without Hoechst staining and, depending on the timing of PB-I extrusion, sorted into group A, B or C, inseminated with sperm and observed throughout preimplantation. The results show that 26.2 ± 12.3% of group A, 2.4 ± 5.0% of group B and none of group C MII oocytes developed to blastocyst. Overall, this study shows that SN oocytes that complete MI earlier are those with a better developmental competence. The possibility to avoid the use of the invasive DNA-binding fluorochrome Hoechst is relevant for future applications in human and domestic animal reproductive technologies.

IVM of mouse fully grown germinal vesicle oocytes upon a feeder layer of selected cumulus cells enhances their developmental competence

Cavalera F.;Zanoni M.;Merico V.;Garagna S.;Zuccotti M.
2019-01-01

Abstract

In the mouse, the use of the DNA-binding fluorochrome Hoechst 33342 allows the classification of fully-grown antral oocytes into two categories distinguished by their chromatin conformation: surrounding nucleolus (SN) and not-surrounding nucleolus (NSN) oocytes, the former capable of completing development, the latter unable to proceed beyond the 2-cell stage. In the present study, time-lapse observation of SN and NSN oocyte GV-to-MII transition highlighted differences in the timing of germinal vesicle breakdown (GVBD) and polar body I (PB-I) extrusion. PB-I extrusion, but not GVBD, revealed the presence of three main groups of significantly different oocytes: Group A (456-576 min) comprising mainly SN oocytes (91.4%), group B (584-728 min) entailing an almost equivalent percentage of SN (52.7%) and NSN (47.3%) oocytes, whereas group C (736-896 min) consisting of almost all NSN (94.4%) oocytes. In a further set of time-lapse experiments, GV oocytes were in vitro matured without Hoechst staining and, depending on the timing of PB-I extrusion, sorted into group A, B or C, inseminated with sperm and observed throughout preimplantation. The results show that 26.2 ± 12.3% of group A, 2.4 ± 5.0% of group B and none of group C MII oocytes developed to blastocyst. Overall, this study shows that SN oocytes that complete MI earlier are those with a better developmental competence. The possibility to avoid the use of the invasive DNA-binding fluorochrome Hoechst is relevant for future applications in human and domestic animal reproductive technologies.
File in questo prodotto:
Non ci sono file associati a questo prodotto.

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11571/1269606
Citazioni
  • ???jsp.display-item.citation.pmc??? 1
  • Scopus 3
  • ???jsp.display-item.citation.isi??? 3
social impact