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considering significant a difference more than 2 eggs retrieved. An interim analysis was planned. We report the preliminary results for the fifty two first patients enrolled. MATERIALS AND METHODS: Egg donors were prospectively enrolled in this study. All patients underwent the preliminary protocol studies for egg donation. Inclusion criteria were Body Mass Index (BMI) ⬍30 kg/m2 and regular menstrual cycles. Exclusion criteria were a personal history of Ovarian Hyperstimulation Syndrome (OHSS), oral contraceptive use in the previous cycle, and to have been included in the study before. All patients started (COS) on cycle day 2 with 225 UI alpha-FSHr. Multiple low-dose administration of Cetrorelix Acetate was introduced when at least one follicle ⱖ14mm and/or plasma estradiol was ⬎400 pg/ml. When ⱖ3 follicles were ⱖ18mm, patients were randomized to receive either GnRHa (Triptoreline 0.2 mg) or HCGr (250 micrograms) for triggering final oocyte maturation. Exclusion criteria at this moment were plasmatic estradiol ⬎4500 pg/mL and ⱖ20 follicles sized ⱖ14mm. Egg retrieval was performed 36 hours after triggering. Data are presented as mean ⫾ standard deviation. Differences between the study groups were examined by t- test and ␹2 test. A P value of ⬍ 0.05 was considered significant. RESULTS: A total of 52 oocyte donors (rHCG: 27, GnRHa: 25) were included in the analysis. Baseline characteristics (age, BMI, FSHr units administrated, plasmatic Estradiol at triggering) were similar. One (3,7%) versus five (20%) donors respectively were not assigned to a receptor due to less than 3 mature oocytes retrieved. No significant differences were observed in the total number of cumulus (13.6⫾8.1 vs 11.4⫾8.4), in the mature oocytes (8,5⫾5.6 vs 7,2⫾3.9), in the embryos obtained (5.6⫾4.4 vs 5.0⫾2.9), or in the fertilization rate (66,07⫾23.7 vs 64.96⫾28.1) between the two groups. Total number of receptors with an embryo transfer were 38 and 30 respectively (1.41 vs 1.2 per intended donor- NS) and 14 and 18 pregnancies were obtained (0.52 vs 0.72 per intended donor- NS). Pregnancy rate per embryo transfer was 36.84% and 60 respectively. Implantation rate per donor was 32.73⫾36.48 vs 39.58⫾31.86 respectively- NS) CONCLUSION: A similar laboratory outcome can be expected when GnRHa is used for triggering oocyte maturation instead of rHCG in egg donation cycles. Nevertheless, clinical differences in overall reproductive outcome may appear when completing the study. Supported by: None.

P-708 GNRH AGONIST (NAFARELIN) PLUS OVIDREL OR GNRH AGONIST ALONE FOR TRIGGERING OVULATION DURING GNRH ANTAGONIST OVULATION INDUCTION CYCLES. E. R. Rauch, T. Tsai. New York Presbyterian-Cornell, New York, NY; New York Hospital Queens, New York, NY. OBJECTIVE: To assess the endocrine profiles of trigerring final oocyte maturation with GnRH agonist “Synarel” nafarelin vs nafareline and Ovidrel after cotreatment with GnRH antagonist Ganerelix DESIGN: Interim data analysis from a prospective study evaluating hormone profiles of patients undergoing ovulation induction with gonadotropins and triggering of final oocyte maturation with nafarelin alone vs nafarelin and ovidrel MATERIALS AND METHODS: Hormone values were obtained from patients undergoing ovulation induction and triggering of final oocyte maturation using either nafarelin 0.15 mg administered by nasal spray in 3 divided doses, or coadminstration of ovidrel 250 mcg with the same nafarelin protocol. Pre and post trigger levels of progesterone were obtained. The change in the progesterone level was calculated and compared using the Mann Whitney U test. RESULTS: 22 patients had completed the triggering of ovulation at the time of interim analysis. Group A: 13 had combined triggering with ovidrel and nafarelin. Group B: 9 triggered with nafarelin alone. Median Age: Group A ⫽ 36 (32,39), B ⫽ 35 (33,37) p ⫽ 0.744c Median change in progesterone: Group A ⫽ 2 (1.4,8.6), B ⫽ 10.3 (0.8,35.2) p⫽ 0.593 CONCLUSION: Ovulation induction in patients undergoing GnRh antagonist downregulated cycles can be triggered with GnRh agonist as well as hcg/ovidrel. Previous studies have demonstrated the benefits of both methods, increased M2 oocytes in GnRH agonist and increased pregnancy rates in hcg triggered cycles. We have demonstrated that GnRH agonist plus ovidrel can be successfully used to trigger ovulation. This method of triggering ovulation using ovidrel/hcg plus nafarelin may lead to improved IVF outcomes. Supported by: None

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Abstracts

OVARIAN FUNCTION P-709 PREDICTIVE VALUE OF SERUM PROGESTERONE LEVELS ON THE DAY OF HUMAN CHORIONIC GONADOTROPIN ADMINISTRATION IN THE IVF OUTCOME. I. Rodriguez, C. Caligara, J. Ramos, L. Santana, F. Carranza, M. Fernandez-Sanchez. IVI, Seville, Spain. OBJECTIVE: To determine if the level of serum P4 on the day of HCG administration predict assisted reproductive technology (ART) outcome (implantation and pregnancy rates). DESIGN: Retrospective descriptive study. MATERIALS AND METHODS: Three hundred and seventy three IVF-ET cycles were evaluated. The cycles were divided according the serum P4 level and P4/E2 x 1000 level on the day of HCG administration. Fisher‘s exact test with the assisstance of Prism software considering p⬍0,05 statistically significant was performed. RESULTS: The average age of the patients were 35 years. The average doses of gonadotropins were: recombinant FSH:1350 UI, recombinant LH: 600 UI, HMG:825 UI, GnRHa (Leuprolid Acetate): 50,5% patients and Antagonists (Cetrorelix y Ganirelix) : 47,7%. The average of oocytes retrieved were 11,76 and embryos transfers were 93%. The average of estradiol, progesterone and P4/E2.x 1000 levels and number of oocytes on the day of HCG were similar in patients with/without pregnancy. There was a significant difference in impantation and pregnancy rates when progesterone level on the day of HCG ⬍0,5 VS ⬎0,5 (35% vs 60%; p⫽0,035) (13% vs 36%; P⫽0,013) (P⬍0,05),. Patients with less than five oocytes retrieved presented the same implantation and pregnancy rate, independently of P4 level. Patients with more than five oocytes retrieved and P4 level on the day fo HCG ⬍0,5, showed a worst implantation and pregnancy rates vs patients with a P4 level ⬎0,5 (difference was statistically significant). Patients with P4/E2 x 1000 level ⬎2 on the day of HCG, showed very bad results. CONCLUSION: Patients with more than five oocytes retrieved and P4 level on the day of HCG ⬍0,5 showed worst results than patients with less than five oocytes and the same P4 (difference statistically significant). Patients with P4/E2 x 1000 ⬎2 on the day of HCG administration (above all in poor responders), showed very bad results (a rise in the number of miscarriages and low pregnancy rate). Supported by: None

P-710 ANGIOTENSIN II REGULATES THE ANDROGEN PRODUCTION IN HUMAN OVARIAN FOLLICLE. D. Ko, H. Lee, J. Lee, W. Park. Eulji Univ. School of Medicine, Life Science Institute, Seoul, Republic of Korea; Eulji Univ. School of Medicine, Dept. of Ob/Gyn, Seoul, Republic of Korea. OBJECTIVE: The aim of this study is to assess the role of angiotensin II in steroidogenesis of ovarian follicle. DESIGN: Prospective cotrolled lavoratory study MATERIALS AND METHODS: Human follicular fluid was obtained from 25 follicles at the time of oocyte aspiration. The concentrations of pregnenolone, progesterone, DHEA and angiotensin II were measured. Human ovarian tissues were obtained from 11 patients at the time of oophorectomy for benign gynaecological conditions. Individual follicular tissue was dissected and cultured in the media containing pregnenolone with or without angiotensin II (1uM) or AII-receptor antagonist Saralasin (1uM). Testosterone, DHEA and progesterone concentrations in cultured media were measured by RIA. Expression of HSD3b2 and CYP17 mRNA in the tissues were measured by real time PCR RESULTS: There was positive correlation between angiotensin II concentration and DHEA/pregnenolone ratio in follicular fluid (R⫽0.69, p⬍0.05). The concentration of DHEA in the culture medium containing human follicular tissue was increased by angiotensin II (control: 6.21 ng/ml, angioteinsinII: 8.63 ng/ml, Saralasin: 4.61 ng/ml), and the concentration of testosterone was decreased by Saralasin (control: 28.16 ng/dl, angiotensinII: 30.22 ng/dl, Saralasin: 10.58 ng/dl). The expression of HSD3b2 mRNA was not changed by angiotensin II or Saralasin. The CYP17 mRNA expression

Vol. 86, Suppl 2, September 2006

was increased by angiotensin II, but the statistical significance was marginal (control: 1 fold, angiotensinII: 1.2 fold, Saralasin: 1.05 fold). CONCLUSION: These results suggest that angiotensin II increases the androgen production in human ovarian follicle probably by increasing CYP17 mRNA expression. Supported by: None

P-711 CORRELATION BETWEEN BASAL AND GNRH AGONIST STIMULATED 17␣ HYDROXYPROGESTERONE (17 OHP) SERUM LEVELS AND ANTRAL FOLLICLE COUNT (AFC) L. The´ron-Ge´rard. Jean Verdier Hospital, University Paris XIII, BONDY, France OBJECTIVE: Assessment of ovarian reserve is usually based on the measurement of AFC, as well as on hormonal parameters (day 3 serum levels of FSH, estradiol, inhibin B, AMH). Few attention has been paid to the assessment of thecal cell function. Nevertheless, serum androgens may predict the ovarian response to FSH. Furthermore, the 17 OHP response to a GnRH agonist has been proposed for diagnosing polycystic ovaries syndrom. No data have been published so far on the correlation between basal and post GnRH agonist stimulated 17OHP and AFC. The aim of this study was to find out a correlation between these parameters and to assess if the GnRH agonist testing should be included in the evaluation of ovarian reserve in order to identify patients with theca cell function deficiency. DESIGN: Prospective monocentric study performed in 43 women with ovulatory cycles. MATERIALS AND METHODS: Serum basal levels of 17␣ hydroxyprogesterone and testosterone were measured before and 24 hours after the subcutaneous administration of Triptorelin ( 0.1 mg) in the early follicular phase. Transvaginal ultrasonography was performed on the same day, by a single operator to assess the AFC. Patients were divided in two groups according to the AFC: group 1: ⱕ 10 follicles for both ovaries (low ovarian reserve, n⫽11) and group 2: 11 to 23 follicles (normal ovarian reserve, n⫽32). Significant relationships between hormonal values and AFC were evaluated by the Pearson’s correlation coefficient. Clinical and hormonal parameters between the two groups of AFC were determined statistically by Student’s t-test. RESULTS: Basal and post GnRH agonist levels of 17 OHP were positively correlated with the AFC (r ⫽ 0.3, p ⫽ 0.04 and r ⫽ 0.41, p ⫽ 0.006 respectively). By contrast, total testosterone levels before and after triptorelin were not significantly related to the AFC (r ⫽ 0.26, p ⫽ 0.08; r ⫽ 0.17, p ⫽ 0.28 respectively). Table 1 shows the results of the hormonal values according to the AFC. Triptorelin stimulated 17 OHP serum values were significantly reduced in women with ⱕ 10 follicles. However, basal 17 OHP levels, basal and triptorelin stimulated testosterone values were not significantly different between the two groups.

P-712 EXPRESSION OF FORKHEAD TRANSCRIPTION FACTORS IN HUMAN GRANULOSA CELLS. A. Ketefian, S. Khan, P. Zarrini, L. Kao, M. D. Pisarska. Cedars Sinai Medical Center, Los Angeles, CA; Cedars Sinai Medical Center/David Geffen School of Medicine at UCLA, Los Angeles, CA. OBJECTIVE: A number of forkhead transcription factors have recently been identified in the rodent ovary and are thought to play an important role in normal follicle development, maturation, ovulation, and possibly luteinization. For instance, the FOXO class of forkhead transcription factors (FOXO1, FOXO3, and FOXO4) are expressed in granulosa cells at various stages of follicle development. Thus, many of these transcription factors may play an important role in fertility and may ultimately be markers of follicle well-being. To date, there have been no studies looking at these transcription factors in the human ovary, particularly human granulosa cells. Thus, we set out to determine if the FOXO family of transcription factors are expressed in human granulosa cells. DESIGN: Laboratory experimental study. MATERIALS AND METHODS: This study was approved by the institutional review board (IRB) of our institute. Granulosa cells were obtained from consecutive patients undergoing in vitro fertilization (IVF). Granulosa cells were obtained from follicle aspirates after the cumulus-oocyte complexes were removed. The granulosa cells were separated from red blood cells by centrifugation and Percoll gradient prior to RNA extraction. Total RNA was extracted from the granulosa cells using the RNeasy Mini Kit (Qiagen) and cDNA was synthesized using the Sensiscript Reverse Transcription Kit (Qiagen). HotStar Taq DNA Polymerase Kit (Qiagen) was then used to perform the polymerase chain reaction (PCR) using the primers for the forkhead transcription factors FOXO1, FOXO3, and FOXO4. FOXP3, which has not been shown to have a role in the ovary, was also screened. Positive and negative controls were performed for each primer. After PCR, gel electrophoresis was performed to assess the presence of an amplified product. RESULTS: FOXO1, FOXO3, and FOXO4 were expressed in the granulosa cells of all patients undergoing IVF, regardless of their cause for infertility. In contrast, FOXP3 was not expressed in granulosa cells of any patients undergoing IVF. CONCLUSION: The FOXO family of transcription factors (FOXO1, FOXO3, and FOXO4) are expressed in granulosa cells at various stages of follicle development in the rodent. In particular, FOXO3 and FOXO4 are highly expressed in the corpus luteum, and mice null for FOXO3 have diminished fertility. We have been able to demonstrate that this class of forkhead transcription factors are expressed in luteinized granulosa cells in the human. Unlike the FOXO class, FOXP3, another forkhead transcription factor, was absent in human granulosa cells. Thus, the FOXO class of transcription factors may also play an important role in folliculogenesis and luteinization in the human, similar to the rodent, and may have potential implications for fertility. These markers may ultimately assist in identifying the best follicles, and the oocytes housed within, that may lead to a successful pregnancy. Supported by: None

P-713 SMAD3 MEDIATES FSH AND ACTIVIN STIMULATION OF AROMATASE EXPRESSION IN GRANULOSA CELLS. X. Gong, J. Brekosky, E. A. McGee. Magee-Womens Research Institute/Univ of Pittsburgh, Pittsburgh, PA. CONCLUSION: These preliminary data show that among parameters of theca cells, 17 OHP is more informative than the other androgens with a high correlation of stimulated values with AFC. They suggest that, measurement of 17 OHP following GnRH analog testing may be useful to assess the ovarian reserve and to identify patients with theca cell function deficiency. This study is still in progress to evaluate the predictive value of this test as regards ovarian response to stimulation. Theca cell function assessment could be used as a basis to select patients which could benefit from treatments increasing ovarian androgens in order to improve ovarian response to stimulation. Supported by: None

FERTILITY & STERILITY威

OBJECTIVE: FSH stimulation of estrogen production by follicles is essential to normal ovarian function. Paracrine factors have long been known to modulate FSH stimulation of granulosa cells. Smad3, a signal transduction molecule for the TGF? superfamily of growth factors, is necessary for normal folliculogenesis. The objective of this study is to determine the role of Smad3 on aromatase expression and activity following stimulation of granulosa cells with FSH and/or activin. DESIGN: In vitro experiments using mouse models that cannot express Smad3 protein (Smad3-/-) and their wild-type (Smad3⫹/⫹) litter-matched controls. MATERIALS AND METHODS: Granulosa cells were isolated from immature litter-mate Smad3-deficient mice (Smad3-/-) and wild-type mice.

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