Private IVF Research Articles

Apoptosis-inhibitor Aven is downregulated in defective spermatogenesis and a novel estrogen target gene in mammalian testis

To study the expression and localization of Aven in rat and human testis from azoospermic patients with different etiologies and its regulation by estrogens. Experimental study. University research center and private IVF clinic. Six men with obstructive azoospermia, five with hypospermatogenesis, and six with Sertoli cell-only syndrome; male Wistar rats. Testicular biopsies and rat seminiferous tubules (SeT) cultured in the presence or absence of 17β-estradiol (E(2)). Testicular cell localization of Aven protein was analyzed by immunohistochemistry. Expression levels of Aven in testicular biopsies and cultured SeT, in the presence or absence of 17β-estradiol, were determined by quantitative reverse transcription-polymerase chain reaction and Western blot. Aven is expressed in Sertoli cells, spermatocytes, and spermatogonia of both rat and human testis. Aven is underexpressed in the testis of men with nonobstructive azoospermia, and its expression levels correlate with severity of spermatogenic status. Aven expression is regulated by E(2) in rat SeT cultured exvivo. The results suggest that deregulation of the expression of the apoptosis inhibitor Aven may be related to male factor infertility. Moreover, Aven is an estrogen target gene and may be involved in the mechanism of testicular apoptosis control by estrogens.

Patients with endometriosis and patients with poor ovarian reserve have abnormal follicle-stimulating hormone receptor signaling pathways

To study the integrity of the FSH receptor (FSHR) signaling pathway in granulosa-lutein cells at the time of egg retrieval and its relationship with the infertility diagnosis. In vitro assays. University laboratory and private IVF center. Patients undergoing IVF: 35 controls (no ovarian factor, NOF), 28 poor responders (PR), 32 patients with endometriosis (EM), and 22 patients with polycystic ovary syndrome (PCOS). Quantitative reverse transcriptase-polymerase chain reaction (PCR) in granulosa-lutein cells from pooled follicles. Correlation between expression of FSHR and FSH-regulated genes PAPP/Cyp19A1. Positive correlations among FSHR, PAPP, and Cyp19A1 expression levels are observed in NOF and are lost, with different patterns, in poor responder patients and those with endometriosis. Patients with endometriosis are an heterogeneous group including patients with poor ovarian reserve who behave like other poor responders patients (endometriosis-A) and patients with good response to ovarian stimulation (endometriosis-B) who show a specific alteration of the FSHR signaling pathway. These preliminary data suggest that the different signaling pathways activated through the FSHR in normal ovaries (NOF) are disrupted in poor responders and in patients with endometriosis. A better knowledge of the molecular origin of these errors may guide clinicians in the choice of personalized ovulation induction protocols for each typeof ovarian dysfunction.

Detorsion and conservative therapy for twisted adnexa: Our experience

Objective:1) To determine if detorsion of the twisted adnexa is better than traditional adnexectomy to conserve the adnexa and preserve its function. 2) To determine the feasibility of detorsion in conservation of adnexa.Design:Prospective Study from September 2004 to September 2008.Setting:Private IVF and Endoscopy Centre.Patients:22 patients with twisted adnexa (15 non-pregnant and 7 pregnant).Intervention:Surgical intervention and either detorsion of adnexa or adnexectomy.Main Outcome Measures:Ovarian preservation and conservation of ovarian function in 77.2% cases determined by: a) Follicular development on sonography (performed for one year after adnexectomy). b) Subsequent surgery for unrelated cause showing healthy ovaries. c) controlled ovarian hyperstimulation and successful oocyte retrieval subsequently.Results:We could conserve the adenexa in 77.2% cases. Laparoscopic detorsion was performed in 11/15(73.33 %) of non-pregnant women and adnexectomy done in four women 26.66%. Among the seven pregnant women, adnexa could be preserved in 6/7(85.7%) and only one woman required adnexectomy. Laparotomy was required in 2/22(9%) women both of which were in late second trimester of pregnancy. In one case (4.54%) we had recurrence of torsion. 88.23% of the women with conserved adnexa showed preservation of ovarian function.Conclusion:Our study showed that timely diagnosis and intervention could make the difference between ovarian loss and salvage- an outcome of great importance in population of reproductive age females. Laparoscopy with its many benefits proves to be superior to laparotomy.

Sperm DNA fragmentation levels in testicular sperm samples from azoospermic males as assessed by the sperm chromatin dispersion (SCD) test

To analyze sperm DNA fragmentation (SDF) in testicular sperm samples from patients with azoospermia either from spermatogenic failure or from duct obstruction. Several technologies can be applied in the evaluation of SDF, but given the ease and low costs, the sperm chromatin dispersion test (SCD) has emerged as a promising standard. Prospective blind observational cohort study. University-affiliated private IVF setting. Azoospermic patients from couples undergoing intracytoplasmic sperm injection cycles. Testicular sperm extraction (TESE). We determined testicular SDF, and a basic comparison between nonobstructive (n = 22) and obstructive azoospermia (n = 40) was performed. We also correlated SDF with embryo quality and pregnancy outcome. SDF in the testicular sperm of patients with nonobstructive azoospermia was significantly higher, 46.92% (SEM = 4.47), than that of patients with obstructive azoospermia, 35.96% (SEM = 2.63). A moderate relationship between embryo morphology and testicular SDF was detected. Logistic regression analysis of the effect of testicular SDF on pregnancy outcome revealed no significant effect (odds ratio = 1.015). Ours is the first report of SDF analysis in testicular sperm by using SCD in azoospermia. This result suggests that spermatogenesis failure may result in a severe affectation of sperm DNA integrity. The degree of DNA fragmentation using the SCD test is not reflected in pregnancy chances, and the explanation could be that embryos have been selected.

The p53 codon 72 single nucleotide polymorphism lacks a significant effect on implantation rate in fresh in vitro fertilization cycles: an analysis of 1,056 patients

To determine whether the p53 codon 72 single nucleotide polymorphism, a change of the amino acid arginine (Arg) to proline (Pro) resulting from a single nucleotide mutation of guanine (G) to cytosine (C), has a clinically significant effect on implantation rate in fresh IVF cycles. Prospective cohort analysis. University-affiliated private IVF center. One thousand fifty-six female patients undergoing fresh nondonor IVF cycles. Embryo implantation rate. Of the 1,056 patients (2,600 total embryos transferred) undergoing their first IVF cycle, 289 had no implantation events and attempted a second cycle. Of the 289 patients in their second cycle, 72 had no implantation events and attempted a third cycle. The p53 codon 72 single nucleotide polymorphism frequencies in the first cycle (homozygous major allele Arg/Arg [G_G] = 45%, heterozygous allele Arg/Pro [G_C] = 44%, and homozygous minor allele Pro/Pro [C_C] = 11%) did not differ significantly across subsequent IVF cycles. There was no statistically significant difference in embryo implantation rate with respect to the single nucleotide polymorphism. The p53 codon 72 single nucleotide polymorphism lacks a clinically significant effect on embryo implantation rate in patients undergoing fresh nondonor IVF cycles.

Human papilloma virus (HPV) genotyping using HPV DNA chip in a population of women undergoing treatment for infertility

OBJECTIVE: The objective of the present study was to use powerful new molecular techniques (DNA chips) to determine the prevalence and the genotype of HPV infection in a population of women consulting for infertility.DESIGN: Prospective study in a Private IVF.MATERIALS AND METHODS: A prospective study was conducted on women consulting in a period of 4 months for investigation of infertility at the IVF Center Clinique de la DHUYS. One hundred and eleven consecutive women aged from 25 to 45 years (average33.3) underwent a liquid based prep cervical cytological screening combined to systematic HPV detection and genotyping. Cytology and HPV testing were performed on the same sample using a validated cytologic fixative liquid (Qualicyt). The HPV genotyping test used (Greiner bio-one, PapilloCheck) allows the simultaneous detection of 24 different HPV types: 18 high risk (HR) HPV types (16/18/31/33/35/39/45/51/52/53/56/58/59/66/68/70/73/82) and 6 low risk (LR) HPV types (6/11/40/42/43/44-55). The principle of the assay is based on the detection of a E1 fragment gene from human HPV. DNA fragments are amplified by PCR and their products are hybridized to specific DNA probe fixed on the DNA chip. Positive and negative internal controls are available for each sample on the same chip. The 111 infertile women were compared with 550 female controls aged from 25 to 45 years (average32.2) wich where routinely consulting at the same period in a regular OBGYN office of the same geographic area. For this control population, HPV testing was performed only in case of abnormal cytology.RESULTS: No cytologic abnormalities where observed in the infertile women population compared to 3,45% (19/550) of the controls. HPV screening on infertile population showed 2,7% (3/111) HPV HR positives tests (genotypes HPV39 / HPV51 / HPV56). No Low risk HPV where detected. For a french study∗ the prevalence of HR HPV in a sexually active population (20- 44) resulted in 15,2%.CONCLUSIONS: Using a micro array technology for genotyping HPV, we did not notice an increase of cytologic abnormalities in our women infertile population. HPV presence appear less frequently in this type of women population that are involved in a long term monogamous relationship. HPV DNA Chip allows a rapid, sensitive and specific detection of HPV infections. We suggest that this technology can now be a reliable diagnostic tool for routine tests with huge potential diagnostics applications. ∗Boulanger JC et al. Gynecologie Obstetrique & Fertilite, 2004,vol32, n3, p218-22. OBJECTIVE: The objective of the present study was to use powerful new molecular techniques (DNA chips) to determine the prevalence and the genotype of HPV infection in a population of women consulting for infertility. DESIGN: Prospective study in a Private IVF. MATERIALS AND METHODS: A prospective study was conducted on women consulting in a period of 4 months for investigation of infertility at the IVF Center Clinique de la DHUYS. One hundred and eleven consecutive women aged from 25 to 45 years (average33.3) underwent a liquid based prep cervical cytological screening combined to systematic HPV detection and genotyping. Cytology and HPV testing were performed on the same sample using a validated cytologic fixative liquid (Qualicyt). The HPV genotyping test used (Greiner bio-one, PapilloCheck) allows the simultaneous detection of 24 different HPV types: 18 high risk (HR) HPV types (16/18/31/33/35/39/45/51/52/53/56/58/59/66/68/70/73/82) and 6 low risk (LR) HPV types (6/11/40/42/43/44-55). The principle of the assay is based on the detection of a E1 fragment gene from human HPV. DNA fragments are amplified by PCR and their products are hybridized to specific DNA probe fixed on the DNA chip. Positive and negative internal controls are available for each sample on the same chip. The 111 infertile women were compared with 550 female controls aged from 25 to 45 years (average32.2) wich where routinely consulting at the same period in a regular OBGYN office of the same geographic area. For this control population, HPV testing was performed only in case of abnormal cytology. RESULTS: No cytologic abnormalities where observed in the infertile women population compared to 3,45% (19/550) of the controls. HPV screening on infertile population showed 2,7% (3/111) HPV HR positives tests (genotypes HPV39 / HPV51 / HPV56). No Low risk HPV where detected. For a french study∗ the prevalence of HR HPV in a sexually active population (20- 44) resulted in 15,2%. CONCLUSIONS: Using a micro array technology for genotyping HPV, we did not notice an increase of cytologic abnormalities in our women infertile population. HPV presence appear less frequently in this type of women population that are involved in a long term monogamous relationship. HPV DNA Chip allows a rapid, sensitive and specific detection of HPV infections. We suggest that this technology can now be a reliable diagnostic tool for routine tests with huge potential diagnostics applications. ∗Boulanger JC et al. Gynecologie Obstetrique & Fertilite, 2004,vol32, n3, p218-22.

Systemic methotrexate to treat ectopic pregnancy does not affect ovarian reserve

To evaluate whether methotrexate (MTX) compromises ovarian reserve and future reproductive outcome in women undergoing assisted reproductive technology (ART), when it is used as first-line treatment for ectopic pregnancy (EP). Prospective, observational study. University-affiliated private IVF unit. Twenty-five women undergoing IVF-ICSI who were treated with MTX (1 mg/kg IM) for an EP after ART. Evaluation of reproductive outcome and serum anti-Müllerian hormone (AMH) levels. Serum AMH was evaluated before administering MTX and >or=1 week after the resolution of the EP. Reproductive outcome was evaluated by comparing subsequent IVF-ICSI cycles after EP resolution. Serum AMH levels, cycle length, gonadotropin dose required, peak serum E(2) level, oocytes collected, and embryos obtained. Serum AMH levels before MTX were not statistically significantly different from those after treatment (3.7 +/- 0.3 ng/mL vs. 3.9 +/- 0.3 ng/mL). Patients undergoing a subsequent cycle after systemic treatment for EP had similar cycle durations (10.3 vs. 10.8 d), gonadotropin requirements (2,775 vs. 2,630.3 IU), peak E(2) levels (1,884.3 vs. 1,523.6 pg/mL), number of oocytes retrieved (12.1 vs. 10.5), and total number of embryos obtained (7.1 vs. 6.5). Single-dose MTX is a safe first-treatment choice that does not compromise future reproductive outcomes in women who are diagnosed with EP after ART.

GnRH agonist and antagonist protocols for stage I–II endometriosis and endometrioma in in vitro fertilization/intracytoplasmic sperm injection cycles

To investigate the outcomes of intracytoplasmic sperm injection (ICSI) cycles after controlled ovarian hyperstimulation (COH) with GnRH antagonist or GnRH agonist (GnRH-a) in mild-to-moderate endometriosis and endometrioma. Prospective randomize trial. A private IVF center. A total of 246 ICSI cycles in 246 patients were divided into three groups: women with mild-to-moderate endometriosis (n = 98); women who had ovarian surgery for endometrioma (n = 81); women with endometrioma and no history of previous surgery (n = 67). Patients in each group were randomized to COH with either triptrolein or cetrorelix. Clinical parameters, characteristics of COH, and ICSI results were analyzed. Outcomes of COH with both GnRH antagonist and GnRH-a were similar in patients with mild-to-moderate endometriosis. Implantation rates were 15.9% vs. 22.6% and clinical pregnancy rates were 27.5% vs. 39% with GnRH antagonist and GnRH-a protocols, respectively, in patients who had ovarian surgery for endometrioma. Implantation rates were 12.5% vs. 14.8% and clinical pregnancy rates were 20.5% vs. 24.2% with GnRH antagonist and GnRH-a protocols, respectively, in patients with endometrioma and no history of ovarian surgery. Considering the implantation and clinical pregnancy rates, COH with both GnRH antagonist and GnRH-a protocols may be equally effective in patients with mild-to-moderate endometriosis and endometrioma who did and did not undergo ovarian surgery.

Optimization of estradiol supplementation during the luteal phase improves the pregnancy rate in women undergoing in vitro fertilization–embryo transfer cycles

To evaluate the influence of different E2 supplementation doses during the luteal phase on implantation and pregnancy rates in women undergoing intracytoplasmic sperm injection (ICSI) cycles. Prospective, randomized study. A private IVF unit. One hundred sixty-six women younger than 40 years who were undergoing IVF with long protocol controlled ovarian hyperstimulation (COH). A total of 231 cycles were investigated. Group 1 (P only) included 80 cycles, group 2 (P and 2 mg of E2) included 73 cycles, and group 3 (P and 6 mg of E2) included 78 cycles. Supplementation in the luteal phase with different doses of E2 (0, 2, or 6 mg/d). Serum E2 and P levels in the late luteal phase, and implantation rate and pregnancy rate (PR) were documented. The data were analyzed with regard to the entire study population and further stratified according to the E2 dose used. Significantly higher implantation rate and PR were recorded in those who received low dose E2 supplementation compared with no substitution (PR 23.1% vs. 32.8%). The best implantation and pregnancy results were found significantly in the group with high dose E2 supplementation (PR 51.3%). For women treated with a long GnRH analogue protocol for COH, addition of a high dose of E2 to daily P supplementation significantly improved the IVF-embryo transfer results.

Deleterious effect of the presence of hydrosalpinx on implantation and pregnancy rates with in vitro fertilization

To determine the effect of the presence of a unilateral or bilateral hydrosalpinx on the outcome with IVF-ET. Retrospective analysis of clinical and laboratory data. Hospital-based private IVF center. Eight hundred forty-six patients with tubal disease younger than age 40 years undergoing 1,766 stimulation cycles. In 118 cycles, a hydrosalpinx was noted sonographically (group I) whereas, in 1,648 cycles, no such image was documented. Pregnancy and implantation rates. Group I displayed a significantly lower pregnancy rate per transfer than group II (16.84% versus 36.83%) and a lower implantation rate (3.92% versus 11.53%). The presence of hydrosalpinx adversely affects the outcome of IVF.

Clinical assessment of recombinant human follicle-stimulating hormone in stimulating ovarian follicular development before in vitro fertilization.

ObjectiveTo compare the efficacy and the safety of recombinant human FSH (hFSH) with urinary hFSH for stimulating follicular development in women undergoing IVF-ET.DesignMulticenter, prospective, randomized, open, parallel group, clinical study.SettingEight European academic IVF units and one private IVF unit.PatientsInfertile female patients aged 18 to 38 years suffering from tubal disease, mild endometriosis, or unexplained infertility.InterventionsPretreatment with buserelin acetate was followed by recombinant or urinary hFSH treatment started at an initial dose of 225 IU FSH/d. Dose adjustment was allowed after 5 days of FSH. After administration of hCG, a standard IVF-ET procedure was performed.Main Outcome MeasuresFollicular development, oocyte retrieval, fertilized oocytes, duration and dose of FSH, and pregnancy. The hypothesis formulated before the study was that no difference was expected between the two FSH preparations.ResultsSixty patients were treated with recombinant hFSH and 63 with urinary hFSH. The mean number (±SD) of growing follicles (mean diameter >10 mm) was 10.3 ± 4.9 and 11.2 ± 5.2, of follicles (mean diameter > 14 mm) was 7.8 ± 3.6 and 9.2 ± 4.5, of retrieved oocytes was 9.3 ± 5.0 and 10.7 ± 5.3, and of fertilized oocytes was 5.6 ± 3.8 and 6.5 ± 4.3, for recombinant and urinary hFSH, respectively. The duration of FSH treatment was 9.9 ± 2.3 and 9.4 ± 1.8 days and the average total dose was 2270 ± 714 and 2095 ± 591 IU of FSH, for recombinant and urinary hFSH, respectively. Thirteen pregnancies were recorded in the recombinant hFSH group and 11 in the urinary hFSH group. Nine patients delivered 13 live infants in the recombinant hFSH group and eight delivered 13 live infants in the urinary hFSH group. In terms of safety, no difference was recorded between the groups and no anti-FSH antibodies were found in any of the patients.ConclusionsThis clinical study shows that recombinant hFSH is as safe and effective as urinary hFSH in stimulating ovarian follicular development.