OBJECTIVE: To investigate the effect(s) of exogenous gonadotropin on the cytoplasmic and nuclear maturation of cumulus-partially enclosed immature human oocytes in vitro derived from ovarian stimulation cycles. DESIGN: Experimental human study. SETTING: University-based laboratory. PATIENT(S): Women, aged 26-35 years, with infertility secondary to male factors, underwent ovarian stimulation and intracytoplasmic sperm injection cycles using a long protocol of pituitary down-regulation. INTERVENTION(S): Cumulus-partially enclosed immature human oocytes that were retrieved from the stimulated cycles were collected at the time of intracytoplasmic sperm injection. The cumulus-partially enclosed immature human oocytes were allocated into two groups: [1] oocytes at the germinal vesicle (GV) stage; and [2] oocytes at the metaphase I (MI) stage. Each group was cultured in vitro with and without gonadotropin supplements. Some metaphase II (MII) oocytes derived from the two groups were parthenogenetically activated and exposed to subsequent embryonic development for 168 hours in vitro. Other MII oocytes were tested for meiotic apparatus analysis, including spindle morphology and chromosomal alignment, by immunofluorescence staining and scanning confocal microscopy. MAIN OUTCOME MEASURE(S): Oocyte maturation and activation rates, percentages of embryonic development, and spindle normalization were analyzed by χ(2) analysis, whereas oocyte maturation time was analyzed by one-way analysis of variance. RESULT(S): For GV oocytes the maturation and activation rates were significantly higher during in vitro maturation with supplementation with FSH/LH (68% vs. 60% and 82% vs. 62%, respectively). However, maturation time (22.78 ± 0.87 vs. 23.70 ± 0.94 hours), embryonic development (cleavage: 84% vs. 83%; four-cell: 72% vs. 66%; eight-cell: 48% vs. 43%; blastocyst: 5% vs. 7%), and meiotic apparatus normalization rates (55% vs. 61.1%) were similar. For MI oocytes there were no significant differences in the maturation rates (85% vs. 84%), maturation time (14.81 ± 0.65 vs.15.73 ± 0.58 hours), activation rates (77% vs. 80%), embryonic development (cleavage rates: 80% vs. 83%; four-cell: 68% vs. 72%; eight-cell: 56% vs. 51%; blastocyst: 7% vs. 6%), and meiotic apparatus normalization rates (52.4% vs. 54.5%). CONLUSION(S): Gonadotropin supplements to the maturation medium play an important role in cumulus-partially enclosed oocytes at the GV stage; however, MI stage-derived oocytes from stimulated cycles fail to acquire improved maturity after in vitro maturation. Furthermore, gonadotropin at the current concentration did not increase spindle or chromosomal abnormalities in MII oocytes maturated from either GV- or MI-stage oocytes in vitro.
OBJECTIVE: To investigate the effect(s) of exogenous gonadotropin on the cytoplasmic and nuclear maturation of cumulus-partially enclosed immature human oocytes in vitro derived from ovarian stimulation cycles. DESIGN: Experimental human study. SETTING: University-based laboratory. PATIENT(S): Women, aged 26-35 years, with infertility secondary to male factors, underwent ovarian stimulation and intracytoplasmic sperm injection cycles using a long protocol of pituitary down-regulation. INTERVENTION(S): Cumulus-partially enclosed immature human oocytes that were retrieved from the stimulated cycles were collected at the time of intracytoplasmic sperm injection. The cumulus-partially enclosed immature human oocytes were allocated into two groups: [1] oocytes at the germinal vesicle (GV) stage; and [2] oocytes at the metaphase I (MI) stage. Each group was cultured in vitro with and without gonadotropin supplements. Some metaphase II (MII) oocytes derived from the two groups were parthenogenetically activated and exposed to subsequent embryonic development for 168 hours in vitro. Other MII oocytes were tested for meiotic apparatus analysis, including spindle morphology and chromosomal alignment, by immunofluorescence staining and scanning confocal microscopy. MAIN OUTCOME MEASURE(S): Oocyte maturation and activation rates, percentages of embryonic development, and spindle normalization were analyzed by χ(2) analysis, whereas oocyte maturation time was analyzed by one-way analysis of variance. RESULT(S): For GV oocytes the maturation and activation rates were significantly higher during in vitro maturation with supplementation with FSH/LH (68% vs. 60% and 82% vs. 62%, respectively). However, maturation time (22.78 ± 0.87 vs. 23.70 ± 0.94 hours), embryonic development (cleavage: 84% vs. 83%; four-cell: 72% vs. 66%; eight-cell: 48% vs. 43%; blastocyst: 5% vs. 7%), and meiotic apparatus normalization rates (55% vs. 61.1%) were similar. For MI oocytes there were no significant differences in the maturation rates (85% vs. 84%), maturation time (14.81 ± 0.65 vs.15.73 ± 0.58 hours), activation rates (77% vs. 80%), embryonic development (cleavage rates: 80% vs. 83%; four-cell: 68% vs. 72%; eight-cell: 56% vs. 51%; blastocyst: 7% vs. 6%), and meiotic apparatus normalization rates (52.4% vs. 54.5%). CONLUSION(S): Gonadotropin supplements to the maturation medium play an important role in cumulus-partially enclosed oocytes at the GV stage; however, MI stage-derived oocytes from stimulated cycles fail to acquire improved maturity after in vitro maturation. Furthermore, gonadotropin at the current concentration did not increase spindle or chromosomal abnormalities in MII oocytes maturated from either GV- or MI-stage oocytes in vitro.