Prematuration of human denuded oocytes in a three-dimensional co-culture system: effects on meiosis progression and developmental competence.
BACKGROUND
During conventional in vitro maturation (IVM), oocytes undergo nuclear maturation but do not attain full cytoplasmic maturity. The present study aimed to evaluate the quality of denuded in vitro matured human oocytes (= 'rescue IVM') following a prematuration culture (PMC) in a three-dimensional (3D) co-culture system with cumulus cells.


METHODS
Denuded, germinal vesicle (GV) oocytes retrieved from controlled ovarian hyperstimulation (COH) cycles were embedded in collagen (Type I) gel containing dissociated cumulus cells and a specific phosphodiesterase 3-inhibitor (PDE3-I) for 24 h. Afterwards, oocytes were removed from the gel and transferred to IVM inhibitor-free medium. The reversibility of PDE3 inhibition was assessed following IVM culture for a maximum of 48 h. Cytoplasmic maturation was assessed by the oocytes' capability to fertilize after ICSI and sustain further embryonic development. Conventional microdroplet cultures of denuded oocytes, with and without PMC, served as controls.


RESULTS
Oocytes embedded in collagen gel and exposed to PDE3-I were efficiently arrested at the GV stage. After removal from collagen and inhibitor, oocytes were capable of resuming meiosis: maturation rates (74.5%) were similar compared with non-prematured (68.5%) and prematured oocytes (73.4%) in microdroplets. Fertilization rates after PMC in 3D co-culture were significantly higher compared with non-prematured oocytes (72.2% versus 54.0%; P < 0.05), but similar to oocytes prematured in microdroplets (63.8%). Compared with both controls, the PMC in 3D co-culture improved the morphological quality and nuclear constitution of embryos on Day 3 post-ICSI.


CONCLUSIONS
Three-dimensional PMC co-culture using a PDE3-I may enhance cytoplasmic maturation of denuded human GV oocytes retrieved from COH cycles.
Vanhoutte
L
L
Department of Reproductive Medicine, Ghent University Hospital, De Pintelaan 185, 9000 Ghent, Belgium. leen.vanhoutte@ugent.be
Nogueira
D
D
De Sutter
P
P
D000818:Animals; D002478:Cells, Cultured; D018920:Coculture Techniques; D003094:Collagen / Q000378:metabolism; D003470:Culture Media / Q000378:metabolism; D054885:Cumulus Cells / Q000166:cytology; D003593:Cytoplasm / Q000378:metabolism; D046149:Embryo Culture Techniques; D005260:Female; D006801:Humans; D008540:Meiosis; D018613:Microscopy, Confocal / Q000379:methods; D009865:Oocytes / Q000166:cytology* / Q000378:metabolism / Q000473:pathology*; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D013256:Steroids / Q000378:metabolism
D003470:Culture Media; D010726:Phosphodiesterase Inhibitors; D013256:Steroids; D003094:Collagen