Produção in vitro de embriões bovinos: utilização de diferentes fontes de gonadotrofinas na maturação dos oócitos

O presente trabalho foi conduzido com o objetivo de avaliar o efeito da utilização de diferentes fontes de gonadotrofinas para maturação in vitro dos oócitos bovinos fecundados e desenvolvidos in vitro sobre as taxas de clivagem (TC) e de blastocistos (TBL). Oócitos imaturos provenientes de ovários de vacas de abatedouro foram submetidos a maturação in vitro sob diferentes condições: meio TCM 199, acrescido de 10% de soro de vaca em estro (SVE), aditivos, hepes, NaHCO3, piruvato de sódio, antibióticos (meio B-199), 20 UI/mL de PMSG e 10 UI/mL de hCG (PMSG/hCG) ou meio B-199, acrescido de 5 mig/mL de FSH e 5 mig/mL de LH (FSH/LH). Seguidos 24 h de cultura a 38,5ºC em atmosfera com 5% de CO2, os oócitos maturos foram incubados com sêmen descongelado durante 18 a 21 horas. Após esse período, os oócitos foram transferidos para placas contendo microgotas de meio Ménezo suplementado com 10% de SVE e células epiteliais do oviduto bovino em suspensão, cobertas com óleo de silicone, os quais permaneceram em cultura por mais 9 dias. Os dados foram analisados pelo teste do Qui-quadrado. A TC e a TBL, para PMSG/hCG e FSH/LH, foram 60 e 13,9% e 61,2 e 10,6%, respectivamente. Não houve diferença entre os tratamentos com relação a TC ou a TBL. Esses resultados sugerem que ambas as fontes de gonadotrofinas podem ser utilizadas para maturação in vitro dos oócitos fecundados e desenvolvidos in vitro.

A curva de regressão da gonadotrofina coriônica humana é útil no diagnóstico precoce da neoplasia trofoblástica gestacional pós-molar?

OBJETIVO: avaliar a utilidade da curva de regressão normal da gonadotrofina coriônica humana (hCG) no diagnóstico precoce de neoplasia trofoblástica gestacional pós-molar (NTG). MÉTODOS: estudo longitudinal, incluindo 105 pacientes com mola hidatiforme completa (MHC) acompanhadas no Centro de Doenças Trofoblásticas de Botucatu, entre 1998 e 2005. Os títulos da hCG sérica foram mensurados quinzenalmente em todas as pacientes. Curvas individuais de regressão da hCG das 105 pacientes foram estabelecidas. A comparação entre a curva de regressão normal estabelecida em nosso serviço com as curvas individuais da hCG foi usada no rastreamento e diagnóstico (platô/ascensão) de NTG. O número de semanas pós-esvaziamento quando a hCG excedeu o limite normal foi comparado com o número semanas em que a hCG apresentou platô/ascensão. RESULTADOS: das 105 pacientes com MHC, 80 apresentaram remissão espontânea (RE) e 25 desenvolveram NTG. Das 80 pacientes com RE, 7 (8,7%) apresentaram, inicialmente, dosagem da hCG acima do normal, mas, no devido tempo, alcançaram a remissão. Todas as 25 pacientes com NTG apresentaram desvio da curva normal da hCG em 3,8±2,5 semanas e mostraram platô ou ascensão em 8,4±2,9 semanas (p<0,001). CONCLUSÕES: a curva de regressão normal da hCG pós-molar pode ser útil para diagnóstico de NTG.

Recombinant luteinizing hormone supplementation to recombinant follicle-stimulation hormone during induced ovarian stimulation in the GnRH-agonist protocol: A meta-analysis

Purpose: to compare the efficacy of recombinant LH supplementation for controlled ovarian stimulation in recombinant FSH and GnRH-agonist protocol.Methods: Search strategies included on-line surveys of databases. The fixed effects model was used for odds ratio and effect size (weighted mean difference). Four trials fulfilled the inclusion criteria.Results: a fewer days of stimulation (p < 0.0001), a fewer total amount of r-FSH administered (p < 0.0001) and a higher serum estradiol levels on the day of hCG administration (p < 0.0001) were observed for the r-LH supplementation protocol. However, differences were not observed in number of oocyte retrieved, number of mature oocytes, clinical pregnancy per oocyte retrieval, implantation and miscarriage rates.Conclusions: more randomized controlled trials are necessary before evidence-based recommendations regarding exogenous LH supplementation in ovarian stimulation protocols with FSH and GnRH-agonist for assisted reproduction treatment can be provided.

Mola hidatiforme completa e eclâmpsia: relato de caso

O desenvolvimento de pré-eclâmpsia ou eclâmpsia antes da 20ª semana deve levar à suspeita de mola hidatiforme. Descrevemos um caso de mola hidatiforme completa (MHC) e eclâmpsia concomitante em paciente com 20 anos que apresentava sangramento genital, anemia, tamanho uterino excessivo e cistos de ovário, associados a hipertensão arterial e proteinúria. Os níveis de b-hCG estavam elevados e a função tiroidiana, alterada. A ultra-sonografia mostrou-se compatível com MHC. Após o esvaziamento uterino apresentou cefaléia e alterações visuais, seguidas por convulsões tônico-clônicas que cessaram com sulfato de magnésio hepta-hidratado a 50%. No seguimento pós-molar foi diagnosticado tumor trofoblástico gestacional (TTG) prontamente tratado com quimioterapia. A associação de MHC e eclâmpsia determina esvaziamento uterino imediato e seguimento pós-molar rigoroso, pelo risco aumentado de desenvolvimento de TTG.

Primary Pulmonary Choriocarcinoma After Human Chorionic Gonadotropin Normalization Following Hydatidiform Mole A Report of Two Cases

BACKGROUND: Primary pulmonary choriocarcinoma (PPC) is rare and frequently leads to death.CASES: Two young patients presented with previous molar pregnancy and spontaneous serum human chorionic gonadotropin (hCG) normalization. Patient 1 was referred to our center after partial response to chemotherapy. Pulmonary lobectomy was performed, and hCG rapidly declined. During further chemotherapy, liver metastasis was detected by positron emission tomography. Right hepatectomy was performed, and hCG declined for 28 days, but increased again despite chemotherapy. This patient died from hepatic failure 3 years after diagnosis. Patient 2 presented with persistently high hCG, though the affected organ was not identified. Chemotherapy was unsuccessful. Patient reevaluation showed an isolated pulmonary mass. Pulmonary lobectomy was performed; 2 weeks later, hCG was normal and consolidation with 2 cycles of chemotherapy was administered. The patient has been in remission for 24 months. PPC was confirmed by histo pathology and immunohistochemistry in both cases. Gestational origin of the tumor was confirmed by molecular genetic analysis (polymorphic microsatellite markers).CONCLUSION: The possibility of choriocarcinoma cannot be overlooked in young women with an isolated pulmonary mass. Early diagnosis, prompt chemotherapy, and surgical resection in a specialized center improves the prognosis. (J Reprod Med 2010;55:311-316)

Maternal and perinatal outcomes of first pregnancy after chemotherapy for gestational trophoblastic neoplasia in Brazilian women

Objective. To evaluate maternal and perinatal outcomes of first pregnancy after chemotherapy for gestational trophoblastic neoplasia (GTN) in Brazilian patients.Methods. This study included 252 subsequent pregnancies after chemotherapy for GTN treated between 1960-2005. Correlations of maternal and perinatal outcomes with chemotherapy regimen (single or multiagent) and the time interval between chemotherapy completion and first subsequent pregnancy were investigated.Results. There was a significant increase in adverse maternal outcomes in women who conceived <6 months than 6-12 months (76.2% and 19.6%; p<0.0001; OR=13.12; CI 95%=3.87-44.40) and >12 months (76.2% and 21.7%; P<0.0001; OR=11.56; CI 95%=3.98-33.55) after chemotherapy. Spontaneous abortion frequency was higher <6 months (71.4%) than 6-12 months (17.6%; p<0.0001: OR=11.66; CI 95%=3.55-38.22) and >12 months (9.4%; p<0.0001: OR=23.97: CI 95%=8.21-69.91) after chemotherapy. There was no difference in adverse perinatal outcomes (stillbirth, fetal malformation, and preterm birth) related to the interval after chemotherapy and Subsequent pregnancy. The overall occurrence of adverse maternal and perinatal outcomes did not significantly differ between patients on single or multiagent regimens.Conclusion. Adverse maternal outcomes and spontaneous abortion were more frequent among patients who conceived within 6 months of chemotherapy completion. In these cases, careful prenatal monitoring and hCG level measurement 6 weeks after the completion of any new pregnancy are recommended. (C) 2008 Elsevier B.V. All rights reserved.

Efficacy of the motile sperm organelle morphology examination (MSOME) in predicting pregnancy after intrauterine insemination

Background: Although the motile sperm organelle morphology examination (MSOME) was developed merely as a selection criterion, its application as a method for classifying sperm morphology may represent an improvement in the evaluation of semen quality. The aim of this study was to determine the prognostic value of normal sperm morphology using MSOME with regard to clinical pregnancy (CP) after intrauterine insemination (IUI).Methods: A total of 156 IUI cycles that were performed in 111 couples were prospectively analysed. Each subject received 75 IU of recombinant FSH every second day from the third day of the cycle. Beginning on the 10th day of the cycle, follicular development was monitored by vaginal ultrasound. When one or two follicles measuring at least 17 mm were observed, recombinant hCG was administered, and IUI was performed 12-14 h and 36-40 h after hCG treatment. Prior to the IUI procedure, sperm samples were analysed by MSOME at 8400x magnification using an inverted microscope that was equipped with DIC/Nomarski differential interference contrast optics. A minimum of 200 motile spermatozoa per semen sample were evaluated, and the percentage of normal spermatozoa in each sample was determined.Results: Pregnancy occurred in 34 IUI cycles (CP rate per cycle: 21.8%, per patient: 30.6%). Based on the MSOME criteria, a significantly higher percentage of normal spermatozoa was found in the group of men in which the IUI cycles resulted in pregnancy (2.6+/-3.1%) compared to the group that did not achieve pregnancy (1.2+/-1.7%; P = 0.019). Logistic regression showed that the percentage of normal cells in the MSOME was a determining factor for the likelihood of clinical pregnancy (OR: 1.28; 95% CI: 1.08 to 1.51; P = 0.003). The ROC curve revealed an area under the curve of 0.63 and an optimum cut-off point of 2% of normal sperm morphology. At this cut-off threshold, using the percentage of normal sperm morphology by MSOME to predict pregnancy was 50% sensitive with a 40% positive predictive value and 79% specificity with an 85% negative predictive value. The efficacy of using the percentage of normal sperm morphology by MSOME in predicting pregnancy was 65%.Conclusions: The present findings support the use of high-magnification microscopy both for selecting spermatozoa and as a routine method for analysing semen before performing IUI.

Características das Curvas de Regressão da Gonadotrofina Coriônica Pós-mola Hidatiforme Completa

Objetivos: construir a curva de regressão do b-hCG pós-mola hidatiforme completa (MHC) com remissão espontânea e comparar com a curva de regressão pós-MHC com tumor trofoblástico gestacional (TTG). Análise comparativa da curva de regressão do b-hCG das portadoras de MHC, acompanhadas no Serviço, com a curva de regressão observada por outros autores1-3. Métodos: foi realizada avaliação clínica e laboratorial (dosagem sérica de b-hCG), na admissão e no segmento pós-molar, de todas as pacientes com MHC, atendidas entre 1990 e 1998 no Hospital das Clínicas de Botucatu - Unesp. O resultado da determinação seriada do b-hCG foi analisado em curvas log de regressão. A evolução da curva de regressão do b-hCG foi analisada e comparada em MHC com remissão espontânea e MHC com TTG numa curva log de regressão, com intervalo de confiança de 95%. A curva log de regressão do grupo de remissão espontânea foi comparada com curvas consideradas padrão1,2. Foram construídas curvas log individuais de todas as pacientes e classificadas de acordo com os quatro tipos de curva (I, II, III e IV), propostos para o seguimento pós-molar³. Resultados: 61 pacientes com MHC tiveram seguimento pós-molar completo, 50 (82%) apresentaram remissão espontânea e 11 (18%) desenvolveram TTG. No grupo de pacientes com MHC e remissão espontânea, o tempo para alcançar a normalização dos níveis do b-hCG, após o esvaziamento molar, foi até 20 semanas. As pacientes que desenvolveram TTG apresentaram desvio precoce da curva de regressão normal do b-hCG, 4 a 6 semanas após o esvaziamento molar. Nestas pacientes, a quimioterapia foi introduzida em média na 9ª semana pós-esvaziamento molar. Conclusões: a curva de regressão do b-hCG pós-MHC com remissão espontânea apresentou declínio log exponencial, semelhante ao observado por outros autores1,2, e diferente das MHC com TTG. Foram identificados três tipos de curvas de regressão do b-hCG, semelhantes aos de Goldstein³, I, II e IV, e outros dois tipos diferentes de regressão do b-hCG: V (regressão normal) e VI (regressão anormal).

Malformação arteriovenosa uterina após doença trofoblástica gestacional

OBJETIVO: investigar a presença e resultados de malformações vasculares uterinas (MAVU) após doença trofoblástica gestacional (DTG). MÉTODOS: estudo retrospectivo com inclusão de casos diagnosticados entre 1987 e 2004; 2764 pacientes após DTG foram acompanhadas anualmente com ultra-sonografia transvaginal e Doppler colorido no Centro de Neoplasia Trofoblástica Gestacional da Santa Casa da Misericórdia (Rio de Janeiro, RJ, Brasil). Sete pacientes tiveram diagnóstico final de MAVU baseado em análise ultra-sonográfica - índice de pulsatilidade (IP), índice de resistência (IR) e velocidade sistólica máxima (VSM) - e achados de imagens de ressonância nuclear magnética (RNM). Dosagens negativas de beta-hCG foram decisivas para estabelecer o diagnóstico diferencial com DTG recidivante. RESULTADOS: a incidência de MAVU após DTG foi 0,2% (7/2764). Achados ultra-sonográficos de MAVU: IP médio de 0,44±0,058 (extremos: 0,38-0,52); IR médio de 0,36±0,072 (extremos: 0,29-0,50); VSM média de 64,6±23,99 cm/s (extremos: 37-96). A imagem de RNM revelou útero aumentado, miométrio heterogêneo, espaços vasculares tortuosos e vasos parametriais com ectasia. A apresentação clínica mais comum foi hemorragia transvaginal, presente em 52,7% (4/7) dos casos. Tratamento farmacológico com 150 mg de acetato de medroxiprogesterona foi empregado para controlar a hemorragia, após a estabilização hemodinâmica. Permanecem as pacientes em seguimento, assintomáticas até hoje. Duas pacientes engravidaram com MAVU, com gestações e partos exitosos. CONCLUSÃO: presente sangramento transvaginal em pacientes com beta-hCG negativo e história de DTG, deve-se considerar a possibilidade de MAVU e solicitar avaliação ultra-sonográfica com dopplervelocimetria. O tratamento conservador é a melhor opção na maioria dos casos de MAVU pós-DTG.

Preditores Clínicos e Histopatológicos de Tumor Trofoblástico Gestacional pós-Mola Hidatiforme Completa

Objetivo: definir os preditores clínicos e histopatológicos mais eficientes da evolução da mola hidatiforme completa (MHC) para tumor trofoblástico gestacional (TTG). Métodos: estudo prospectivo clínico e histopatológico de todas as portadoras de MHC, atendidas entre 1990 e 1998 no Hospital das Clínicas de Botucatu -- UNESP. A avaliação clínica pré-esvaziamento molar classificou a gravidez molar em: MHC de alto risco e MHC de baixo risco. Foram analisados os preditores clínicos para TTG, estabelecidos por Goldstein et al.¹ e por outros autores2--10. A avaliação histopatológica incluiu a determinação do diagnóstico de MHC, segundo os critérios de Szulman e Surti11, e o reconhecimento dos fatores de risco para TTG, de Ayhan et al.8. Os preditores clínicos e histopatológicos foram correlacionados com o desenvolvimento de TTG pós-molar. Resultados: em 65 portadoras de MHC, cistos do ovário maiores que 6 cm e tamanho uterino maior que 16 cm foram os preditores clínicos mais eficientes de TTG. A proliferação trofoblástica, a atipia nuclear, a necrose/hemorragia, a maturação trofoblástica e a relação cito/sinciciotrofoblasto não foram preditores significativos para TTG. A correlação entre preditor clínico e histopatológico para o desenvolvimento de TTG não foi possível porque nenhum parâmetro histopatológico foi significativo. Conclusões: mais estudos são necessários para avaliar possíveis preditores de persistência (TTG) e sua aplicação no contexto clínico das MHC. Enquanto isso, a determinação seriada de hCG sérico permanece o único indicador prognóstico seguro para TTG pós-MHC.

Sincronização da ovulação em fêmeas suínas submetidas ao desmame precoce

Verificou-se a eficiência de protocolos para sincronizar a ovulação em porcas desmamadas precocemente. Trinta porcas com média de 4,4± 2,0 partos e estádio de lactação de 14,8± 0,7 dias foram distribuídas em três grupos de 10 animais: 1- nenhum tratamento hormonal; 2- 1000 UI de PMSG, via intramuscular (IM), 48h pós-desmame e 0,25mg de GnRH, IM, 72h após a aplicação do PMSG; 3- 1000 UI de PMSG, IM, 48h pós-desmame e 500 UI de hCG, IM, 72h após o PMSG. O momento da ovulação foi detectado por ultra-sonografia transretal. A taxa de sincronização (ovulação até 48h após aplicação de hCG ou GnRH) dos grupos 2 e 3 (94,7%) foi maior (P<0,01) que no grupo controle (40%). Com o uso dos protocolos de sincronização de ovulação, as fêmeas tratadas apresentaram, em relação ao grupo controle, tendência de maior taxa de prenhez (95% vs. 70%; P<0,10) e similares intervalo do desmame ao estro (96,5± 3,0 vs. 130,2± 31,4h) e número de leitões nascidos vivos por fêmea gestante no primeiro cio pós-desmame (10,9± 0,8 vs. 12,0± 0,9). Dessa maneira, os protocolos de sincronização usados neste estudo foram eficientes em sincronizar a ovulação, e podem viabilizar o uso da inseminação artificial em horários predeterminados.

Elevated progesterone concentrations enhance prostaglandin F-2 alpha synthesis in dairy cows

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effects of postbreeding gonadotropin treatments on conception rates of lactating dairy cows subjected to timed artificial insemination or embryo transfer in a tropical environment

The objective of experiment 1 was to evaluate the effects of treatments with human chorionic gonadotropin (hCG) or GnRH 7 d after induced ovulation on reproductive performance of lactating dairy cows submitted to timed artificial insemination (TAI) or timed embryo transfer (TET). A total of 834 potential breedings were used from 661 lactating Holstein cows (37.3 +/- 0.3 kg of milk/d). Cows had ovulation synchronized and were assigned randomly to receive TAI on d 0 or TET on d 7. Within each group, cows were assigned randomly to receive on d 7 no additional treatment (control; n(TAI) = 156; n(TET) = 126), a 100 mu g i.m. injection of GnRH (n(TAI) = 155; n(TET) = 124), or a 2,500 TU i.m. injection of hCG (ITA = 151; n(TET) = 122). Postbreeding treatment affected the percentages of pregnant cows at TET on d 28 (control: 38.1%; GnRH: 52.4%; hCG: 45.1%) and on d 60 (control: 32.5%; GnRH: 41.1%; hCG: 38.5%), but postbreeding treatment did not affect percentages of pregnant cows at TAT on d 28 (control: 30.1%; GnRH: 32.2%; hCG: 32.4%) or on d 60 (control: 25.6%; GnRH: 27.1%; hCG: 29.8%). The objective of experiment 2 was to evaluate the effect of a treatment with GnRH 7 d after TET on reproductive performance of lactating dairy cows that received a previous GnRH treatment at TET. A total of 285 potential breedings were used from 257 lactating Holstein cows (35.1 +/- 0.8 kg of milk/d). Cows had ovulation synchronized and were assigned for TET on d 7. Immediately after TET, all cows were treated with a 100 mu g i.m. injection of GnRH. on d 14, cows were assigned randomly to receive (G7-14; n = 147) or not (G7; n = 138) an additional injection of GnRH. Pregnancy diagnosis were performed on d 28 and 60. The additional treatment with GnRH on d 14 did not affect the percentages of pregnant cows on d 28 (G7: 48.5%; G7-14: 42.9%) or on d 60 (G7: 39.8%; G7-14: 37.4%). In conclusion, treatment with GnRH or hCG 7 d after induced ovulation increased conception rates in lactating dairy cows submitted to TET, but not in cows submitted to TAI. Moreover, treatment with GnRH 7 d after TET did not enhance reproductive performance of lactating dairy cows that received a previous GnRH treatment at TET.

The efficacy of a single chain recombinant equine luteinizing hormone (reLH) in mares: Induction of ovulation, hormone profiles, and inter-ovulatory intervals

The objectives of this study were to determine the efficacy of recombinant equine luteinizing hormone (reLH) in shortening the time to ovulation in cycling mares and to determine the effects of treatment on endogenous hormones and inter-ovulatory intervals. In study 1, mares of light horse breeds (3-20 years) were treated with either a vehicle, various doses of reLH, or human chorionic gonadotropin (hCG). Cycling mares were examined by palpation and ultrasound per rectum daily or every 12 h from the time of treatment to ovulation. In studies 2 and 3, jugular blood samples were collected daily or every 12 h from the time of treatment to ovulation for analysis of LH, follicle stimulating hormone (FSH), estradiol-17 beta (E-2), and progesterone (P-4) by radioimmunoassays (RIA). Increasing doses of reLH (0.3, 0.6, 0.75, and 0.9 mg) showed increasing effectiveness at inducing ovulation within 48 h of treatment. Treatments with the 0.75 and 0.9 mg doses of reLH resulted in 90% and 80% ovulation rates, which were similar to hCG treatment (85.7%). Except for the early rise in LH after treatment with 0.5, 0.65, and 1.0 mg of reLH, hormone profiles appeared to be similar between control and treated cycles. Inter-ovulatory intervals were similar between control and treatment cycles. In conclusion, reLH is a reliable and effective ovulatory agent that does not significantly alter endogenous hormone profiles or affect inter-ovulatory intervals.(c) 2007 Elsevier B.V. All rights reserved.

Strategies to improve the ovarian response to equine pituitary extract in cyclic mares

Equine pituitary extract (EPE) has been reported to induce heightened follicular development in mares, but the response is inconsistent and lower than results obtained in ruminants undergoing standard superovulatory protocols. Three separate experiments were conducted to improve the ovarian response to EPE by evaluating: (1) effect of increasing the frequency or dose of EPE treatment; (2) use of a potent gonadotropin-releasing hormone agonist (GnRH-a) prior to EPE stimulation (3) administration of EPE twice daily in successively decreasing doses. In the first experiment. 50 mares were randomly assigned to one of four treatment groups. Mares received (1) 25 mg EPE once daily; (2) 50 mg EPE once daily (3) 12.5 mg EPE twice daily; or (4) 25 mg EPE twice daily. All mares began EPE treatment 5 days after detection of ovulation and received a single dose of cloprostenol sodium 7 days postovulation. EPE was discontinued once half of a cohort of follicles reached a diameter of greater than or equal to35 mm and hCG was administered. Mares receiving 50 mg of EPE once daily developed a greater number (P = 0.008) of preovulatory follicles than the remaining groups of EPE-treated mares, and more (P = 0.06) ovulations were detected for mares receiving 25 mg EPE twice daily compared to those receiving either 25 mg EPE once daily and 12.5 mg EPE twice daily. Embryo recovery per mare was greater (P = 0.05) in the mares that received 12.5 mg EPE twice daily than those that received 25 mg EPE once daily. In Experiment 2, 20 randomly selected mares received either 25 mg EPE twice daily beginning 5 days after a spontaneous ovulation. or two doses of a GnRH-a agonist upon detection of a follicle greater than or equal to35 mm and 25 mg EPE twice daily beginning 5 days after ovulation. Twenty-four hours after administration of hCG, oocytes were recovered by transvaginal aspiration from all follicles greater than or equal to35 mm. No differences were observed between groups in the numbers of preovulatory follicles generated (P = 0.54) and oocytes recovered (P = 0.40) per mare. In Experiment 3, 18 mares were randomly assigned to one of two treatment groups. Then, 6-11 days after ovulation, mares were administered a dose of PGF(2gamma) and concomitantly began twice-daily treatments with EPE given in successively declining doses, or a dose of PGF(2alpha), but no EPE treatment. Mares administered EPE developed a higher (P = 0.0004) number of follicles :35 mm, experienced more (P = 0.02) ovulations, and yielded a greater (P = 0.0006) number of embryos than untreated mares. In summary, doubling the dose of EPE generated a greater ovarian response, while increasing the frequency of treatment, but not necessarily the dose. improved embryo collection. Additionally, pretreatment with a GnRH-a prior to ovarian stimulation did not enhance the response to EPE or oocyte recovery rates. (C) 2002 Elsevier B.V. All rights reserved.