J Smitz1, R Cortvrindt. 1. Reproductive Biology Laboratory, Academic Hospital and Medical School, Brussels Free University, Belgium. lriaszj@az.vub.ac.be
Abstract
OBJECTIVES: Cryostorage of ovarian cortical tissue, before devastating chemo- and/or radiotherapy for cancer, permits survival of primordial and early preantral follicles. This work aims for a system allowing the long-term maturation in vitro (IVM) of small immature oocytes up to fertilisable metaphase II oocytes. METHODS: A culture system allowing follicle attachment permitted the growth and maturation of isolated follicles (follicle diameter between 100 and 130 microns) from 14-day-old (prepuberal) mice. Follicle and oocyte development were observed under the inverted microscope and conditioned medium was used for biochemical analysis. Effects of recombinant gonadotrophins and oxygen tensions were studied for their specific effects on follicle development. RESULTS: A 12-day culture period yielded full-grown oocytes which were able to complete meiosis (metaphase II). Live young were obtained after IVF and intra-uterine transfer of in vitro matured oocytes. Growth and maturation were only successful when recombinant gonadotropins were added and when the incubator had a 20% oxygen tension. This system enabled the growth of early preantral follicles after cryopreservation: 80% of frozen and thawed follicles survived up to culture-day 12 and yielded a comparable blastocyst formation rate as in controls. CONCLUSIONS: The mouse model suggests that IVM is a valuable option after oocyte storage. The development of a comparable system for long-term culture of human follicles will imply the acquisition of non-invasive techniques to appreciate oocyte's maturity and developmental capacity.
OBJECTIVES: Cryostorage of ovarian cortical tissue, before devastating chemo- and/or radiotherapy for cancer, permits survival of primordial and early preantral follicles. This work aims for a system allowing the long-term maturation in vitro (IVM) of small immature oocytes up to fertilisable metaphase II oocytes. METHODS: A culture system allowing follicle attachment permitted the growth and maturation of isolated follicles (follicle diameter between 100 and 130 microns) from 14-day-old (prepuberal) mice. Follicle and oocyte development were observed under the inverted microscope and conditioned medium was used for biochemical analysis. Effects of recombinant gonadotrophins and oxygen tensions were studied for their specific effects on follicle development. RESULTS: A 12-day culture period yielded full-grown oocytes which were able to complete meiosis (metaphase II). Live young were obtained after IVF and intra-uterine transfer of in vitro matured oocytes. Growth and maturation were only successful when recombinant gonadotropins were added and when the incubator had a 20% oxygen tension. This system enabled the growth of early preantral follicles after cryopreservation: 80% of frozen and thawed follicles survived up to culture-day 12 and yielded a comparable blastocyst formation rate as in controls. CONCLUSIONS: The mouse model suggests that IVM is a valuable option after oocyte storage. The development of a comparable system for long-term culture of human follicles will imply the acquisition of non-invasive techniques to appreciate oocyte's maturity and developmental capacity.