Ovarian stimulation for in vitro fertilization (IVF): a prospective randomized comparison of recombinant FSH alone or in combination with human menopausal gonadotropins.

affects oocyte ploidy and oocyte ploidy affects egg and embryo competence, we use digital polarized light microscopy (polscope) to non-invasively view meiotic spindles. The polscope images the inherent optical birefringence of microtubules. Previously, we showed that spindle integrity predicts fertilization and blastulation (Fertil Steril 2001). Now, we report the effects of cycle specific factors on spindles from human metaphase II (MII) oocytes imaged prior to ICSI. Design: Retrospective cohort study at a university-based IVF center. Materials/Methods: After retrieval, cumulus oocyte complexes were cultured for 5-6 hours in P1 medium with 6% synthetic serum substitute. Cumulus cells were removed for ICSI by pipetting in modified HTF containing 80 IU/ml hyaluronidase. For imaging, oocytes sat in drops of modified HTF under warm paraffin oil in a Bioptechs Delta T.C.O. Culture Dish at 37°C with stage and objective heaters. Oocytes were examined under a Zeiss Axiovert with a Neofluar 40⫻ strainfree objective and LC Polscope optics and controller (CRI, Woburn, Mass) with a computerized image analysis system (Metamorph, Universal Imaging, PA). Digital images were stored for analysis. Results: From 11/26/99-6/23/00, 183 fresh, non-donor, ICSI cycles led to retrieval of 2544 oocytes, of which, 2025 were at MII and imaged with the polscope. In 46 cycles ⬍70% of MII oocytes had spindles. In 62 cycles, 70-90% of MII oocytes had spindles and in 75 cycles ⬎90% of MII oocytes had spindles. Peak estradiol (PE2) levels were higher in cycles with 70-90% spindle-positive oocytes (SPOS) compared to cycles with ⬍ 70% or ⬎90% SPOS (2301 pg/ml vs 1843 pg/ml and 1890 pg/ml respectively; p ⫽ 0.016). Length of stimulation was less in cycles with 70-90% SPOS compared to cycles with ⬍ 70% SPOS (9.7 vs 10.6 days respectively; p ⫽ 0.003). Cycles with 70-90% SPOS had a greater percentage of MII oocytes than cycles with ⬍ 70% SPOS (85.7% vs 77%; p ⫽ 0.013). More follicles ⬎ 16 mm were observed in cycles with 70-90% SPOS than cycles with ⬍70% SPOS (7.2 vs 5.7; p ⫽ 0.03). GnRH flare cycles were 2.3⫻ more likely to have ⬍70% MII SPOS than late luteal phase GnRH down regulation cycles (RR ⫽ 2.33 95% CI: 1.23-4.41; p ⬍ 0.01). Neither mean age nor basal FSH levels differed among groups. Conclusions: Spindles tether the metaphase chromosomes and ensure their segregation at fertilization, and thus affect egg and embryo ploidy. Non-invasive imaging of meiotic spindles during IVF showed that stimulation and follicular dynamic factors associated with favorable IVF outcome i.e. PE2, number of lead follicles and late luteal GnRHa down regulation, affect spindle integrity. These results suggest that the polscope may help assess the impact of ovarian stimulation on developing oocytes and therefore help fine tuning IVF cycle management.

Wednesday, October 24, 2001 4:00 P.M. O-243 Ovarian stimulation for in vitro fertilization (IVF): a prospective randomized comparison of recombinant FSH alone or in combination with human menopausal gonadotropins. Z. A. Nassar, Z. Massad, G. Abdo, M. H. Fakih. IVF Michigan, Rochester Hills, MI. Objective: To compare the reproductive outcome in patients undergoing controlled ovarian hyperstimulation for IVF with recombinant FSH (r-FSH) alone or in combination with human menopausal gonadotropin (HMG) after pituitary desensitization with long acting depot GnRH agonist. Design: Prospective controlled trial. Materials/Methods: From January 2000 to October 2000, a total of 121 patients undergoing IVF cycles were recruited. Patients less than 36 years with infertility secondary to male factor (42.2%), ovulatory dysfunction (14.9%), endometriosis (14.9%) or unexplained (13.2%) were included. They were randomized into 2 groups: Group 1 (n ⫽ 61 patients) received a starting dose of 4 ampules/day of r-FSH. Group 2 (n ⫽ 60) received a starting dose of 2 ampules of r-FSH combined with 2 ampules of HMG daily. Dosage was adjusted according to patient’s response to stimulation. Women with azospermic partners, polycystic ovary syndrome, body mass index 35, severe endometriosis, poor response in previous cycles, more than 2 IVF failures and history of ovarian hyperstimulation syndrome were excluded. Downregulation was achieved with the depot form of Leuprolide acetate 1.25 mg in the late luteal phase. Statistical analysis was done with analysis of variance or Fischer Exact test as indicated with p ⬍ 0.05 considered statistically significant.

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Results: Mean female age was 31.3 ⫾ 2.8 in group 1 (r-FSH) vs 30.9 ⫾ 3.6 in group 2 (r-FSH ⫹ HMG) (p 0.05), days of stimulation 9.3 ⫾ 1.9 vs 9.9 ⫾ 1.7 (p 0.05), total ampules used 36.1 ⫾ 10.9 vs 41.9 ⫾ 13.9 (p ⫽ 0.02), LH day of HCG administration 1.9 ⫾ 1.5 vs 1.2 ⫾ 1.1 (p 0.05), peak estradiol level 2283.7 ⫾ 1108.1 vs 2054.0 ⫾ 724.1 (p 0.05), total eggs retrieved 18.6 ⫾ 9.9 vs 14.1 ⫾ 7.5 (p ⫽ 0.01), number of embryos transferred 3.9 ⫾ 1.0 vs 3.6 ⫾ 1.0 (p 0.05) in the 2 groups respectively. A total of 121 embryo transfer cycles were done (0% cancellation rate). Outcome of IVF in the 2 groups is shown in Table 1: Table 1. Outcome of IVF using 2 protocols for controlled ovarian hyperstimulation.

Fertilization rate* Implantation rate* Clinical pregnancy rate per embryo transfer* Ongoing pregnancy rate per embryo transfer*

r-FSH (n ⫽ 61)

r-FSH ⫹ HMG (n ⫽ 60)

80.5 17.7 50.8

83.2 13.4 46.7

45.9

43.4

* Results are reported in percentages. P ⬎ 0.05 (not statistically significant) in all parameters compared. Conclusions: In patients less than 36 years of age, following downregulation with depot leuprolide acetate, less ampules were required and more eggs retrieved with the use of r-FSH for ovarian hyperstimulation compared to the combined regimen. However, similar fertilization, implantation and pregnancy rates were achieved. Adding LH to the stimulation protocol does not seem to improve the reproductive outcome. Supported By: IVF Michigan, Michigan State University.

Wednesday, October 24, 2001 4:15 P.M. O-244 Controlled ovulation of the dominant follicle: a model for examining periovulatory events in the natural menstrual cycle. K. A. Young, C. L. Chaffin, T. A. Molskness, R. L. Stouffer. Div of Reproductive Science, Oregon Regional Primate Research Ctr, Oregon Health Science Univ, Beaverton, OR; Medical Coll of Georgia, Dept of Physiology, Augusta, GA. Objective: To develop a method controlling the timing of periovulatory events in the single dominant follicle during the spontaneous menstrual cycle. This model would complement existing protocols where periovulatory events are controlled in multiple follicles during ovarian stimulation cycles. Design: A prospective laboratory investigation (ORPRC IACUC approved) of the ovarian response in female rhesus monkeys after administration of GnRH antagonist and exogenous gonadotropins (Ares Advanced Technologies, Inc) during the late follicular phase of the menstrual cycle. Materials/Methods: Blood samples were obtained daily from 15 adult monkeys during the natural menstrual cycle. Serum samples were assayed for estradiol (E) and progesterone (P) using the Elecsys 2010 system; the treatment protocol was initiated when E concentrations reached 60-387 pg/ml. Treatment consisted of 4 sets of injections over 2 days. On Day 1 of treatment, Antide (3 mg/kg), r-hFSH and r-hLH (30 IU) were administered at 0800 h; gonadotropins alone were administered at 1600 h. On Day 2, gonadotropins and ⫾ Antide (0.5 mg/kg) were administered at 0800 h, and a final injection of either FSH ⫹ LH or an ovulatory bolus of 1000 IU r-hCG was administered at 1600 h. Seventy-two hours later, the ovaries were viewed by laparoscopy for evidence of follicle rupture (raised stigmata). Subsequently, serum levels of bioactive LH were assayed using a mouse Leydig cell bioassay; levels of ⬎150 ng/ml were considered indicative of an LH surge. Results: When treatment was initiated at E concentrations of ⬎ 120 pg/ml (mean serum levels ⫽ 152 ⫾ 10 pg/ml), 4 of 7 females experienced a spontaneous LH surge (459 ⫾ 178 ng/ml). Likewise, ovulation occurred in 3 of 5 females in this group without administration of an ovulatory hCG bolus. Normal luteal phases occurred in 6 of 7 females in this group, with peak concentrations of P averaging 10.3 ⫾ 2.8 ng/ml. In contrast, when

Vol. 76, No. 3, Suppl. 1, September 2001