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Objective To evaluate the influence on sperm parameters and in vitro fertilization (IVF) outcomes of the administration of 400 mg/day of vitamin E for 3 months to men from infertile couples who are undergoing IVF. Design Double-blind, placebo-controlled, randomized study. Setting Human reproduction unit of a university hospital. Patient(s) A total of 101 couples, 50 in the vitamin E group and 51 in the placebo group, undergoing IVF, among whom 64.4% of cases had an abnormal spermiogram according to World Health Organization (WHO) criteria. Intervention(s) Vitamin E (α-tocopherol), 400 mg daily by mouth for 3 months, with sperm analysis performed immediately before starting the treatment and 3 months later on the day of IVF. Main Outcome Measure(s) WHO sperm parameters and IVF outcomes. Result(s) Although there was a statistically significant increase in progressive motility in the vitamin E group compared with before-treatment values, a similar increase occurred in the placebo group. Normal morphology was even better in the placebo group. Regarding IVF outcomes, better fertilization rates were observed in the placebo group, but the live-birth rate per transfer was statistically significantly higher in the vitamin E group: 17 (41.46%) of 41 versus 9 (20.46%) of 44 in the placebo group. Although the clinical pregnancy rates (both per transfer and per cycle started) and the implantation rate were somewhat higher in the vitamin E group (43.9% and 25%; 36.0% and 22.0%; and 24.7% and 14.1%, respectively), the increase was not statistically significant. Conclusion(s) The effect of vitamin E on classic sperm parameters was not an improvement over placebo. Nonetheless, vitamin E administration was associated with a statistically significantly higher live-birth rate, and there was a trend toward better results in other IVF parameters. Clinical Trial Registration Number Eudra CT 2007-000960-25.

Objective To ascertain whether the oocytes of women who are obese or overweight have a different fatty acid (FA) profile than women with normal weight. Design Prospective case-control study. Setting Two IVF centers. Patient(s) A total of 205 women undergoing IVF and intracytoplasmic sperm injection (ICSI) were included in the study, totaling 922 oocytes. Intervention(s) The unfertilized and the immature oocytes from the women who underwent IVF/ICSI were subjected to FA analysis with capillary gas chromatography. Women were classified according their body mass index (BMI) as normal, overweight, or obese. Germinal vesicle oocytes, metaphase I oocytes, and unfertilized metaphase II oocytes were analyzed separately. Main Outcome Measure(s) Fatty acid profile. Result(s) A very different oocyte FA pattern was observed for each BMI. Women with normal weight had higher levels of saturated FAs, and lower levels of monosaturated FAs. Women who were obese had lower levels of n-3 polyunsaturated FA, and the lowest n-6:n-3 ratios. Regarding specific FAs, docosahexaenoic acid levels were lower in women with normal weight than in those who are overweight, and in women who are overweight than in those who are obese. The opposite occurred with eicosapentaenoic acid, with the highest levels in women who have normal weight followed by those who are overweight and lower levels in those women who were obese. When FA analysis was restricted to a subset of oocytes, many of these differences persisted. Conclusion(s) Our study shows that oocytes from women who are obese or overweight have a different FA composition. This difference in levels could be related to the IVF poor outcome in these women. Therefore, this different composition could suggest that offspring of women who are obese or overweight have an unfavorable milieu even before conception.

OBJECTIVE: To characterize the most relevant changes in the lipidome of endometrial fluid aspirate (EFA) in non-implantative cycles. DESIGN: Lipidomics in a prospective cohort study. SETTINGS: Reproductive unit of a university hospital. PATIENTS: Twenty-nine women undergoing an IVF cycle. Fifteen achieved pregnancy and 14 did not. INTERVENTION: Endometrial fluid aspiration immediately before performing embryo transfer. MAIN OUTCOME MEASURES: Clinical pregnancy rate and lipidomic profiles obtained on an ultra-high performance liquid chromatography coupled to time-of-flight mass spectrometry (UHPLC-ToF-MS)-based analytical platform. RESULTS: The comparative analysis of the lipidomic patterns of endometrial fluid in implantative and non-implantative IVF cycles revealed eight altered metabolites: seven glycerophospholipids and an omega-6 polyunsaturated fatty acid. Then, women with a non-implantative cycle were accurately classified with a support vector machine algorithm including these eight lipid metabolites. The diagnostic performances of the algorithm showed an area under the receiver operating characteristic curve, sensitivity, specificity, and accuracy of 0.893 ± 0.07, 85.7%, 80.0%, and 82.8%, respectively. CONCLUSION: A predictive lipidomic signature linked to the implantative status of the endometrial fluid has been found. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s10815-019-01670-z) contains supplementary material, which is available to authorized users.

Research question What is the influence of the Caesarean section–pregnancy interval (CSPI) on the risk of uterine rupture, and what are the repercussions on IVF pregnancy rates of prolonging it? Study design Systematic searches were performed using PubMed MEDLINE to identify studies published up until July 2017 for articles with the following keywords: ‘interdelivery interval’ and ‘uterine rupture’; ‘interpregnancy interval’ and ‘uterine rupture’; ‘interpregnancy interval’ and ‘cesarean section’; and ‘uterine rupture’ and ‘cesarean section’. The search identified 1609 articles, of which six were included (involving 56,419 women). Four reported significantly higher uterine rupture rates in cases of a short CSPI. Results From the analysis, the uterine rupture rate can be modelled by a formula corresponding to a hyperbolic curve. There is no clear cut-off in uterine rupture in relation to CSPI. The curve showed a sharp decrease in uterine rupture until the 10th month of CSPI (uterine rupture rate 0.7%), then a moderate and steady decrease until the 40th month (uterine rupture rate 0.4%) and afterwards a very mild decrease. From the data it is possible to calculate, according to the age of the woman, the expected reduction in IVF rates and uterine rupture as CSPI increases. Conclusion The risk of uterine rupture in relation to CSPI can be represented by means of a hyperbolic curve. After a 10-month CSPI, the expected uterine rupture rate is close to 0.7%. The impact of prolonging or reducing this interval on IVF pregnancy rates can be easily obtained from the table included in the article. This should be helpful in the decision-making process for both patients and physicians.

[EN] Purpose. Some microbiota patterns have been associated with favorable IVF prognosis and others with pathological conditions. The endometrial fluid aspirate (EFA) contains antibacterial proteins that are enriched in implantative IVF cycles, but the antimicrobial effect of EFA has not been addressed. We aimed to evaluate the antimicrobial activity of the human endometrial fluid during the natural cycle. Methods. EFA was obtained through an embryo transfer catheter in 38 women, aged 18-40 years, with regular cycles attending to a fertility clinic. The antimicrobial activity of EFAs was tested against two strains of Staphylococcus aureus; one strain each of Streptococcus agalactiae, Enterococcus faecalis, Escherichia coli, and Klebsiella pneumoniae; and three yeasts (Candida albicans, Candida glabrata, and Candida krusei). Results. All samples exhibited antibacterial activity against S. aureus. In addition, 32.4% of EFAs were active against one of the other microorganisms assayed, 16.2% against two, and 5.4% against four of them. In contrast, none exhibited antibacterial activity against E. coli or K. pneumoniae. The antimicrobial activity differs considerably between EFA samples, and we failed to observe a cycle-related pattern. Conclusions. EFA presented two antimicrobial activity patterns: (a) one common to all the samples, exhibiting activity against S. aureus and lack of activity against E. coli and K. pneumoniae, and (b) an individualized pattern, showing activity against some of the other microorganisms tested. The intensity of antibacterial activity differs between EFA samples. Our data suggest that the uterine microbiota is controlled by means of endometrial fluid components. This study was partially supported by a Grant for Fertility Innovation (GFI, 2011) from Merck, Darmstadt, Germany. M. Bregón-Villahoz is recipient of a predoctoral grant from the Universidad del País Vasco-Euskal Herriko Unibertsitatea (UPV/EHU) (PIF19/316). The authors thank the technical and human support provided by DNA Bank Service (SGIker) of the University of the Basque Country (UPV/EHU) and European funding (ERDF and ESF). CIC bioGUNE is accredited with the Severo Ochoa Excellence award by the Spanish Ministerio de Economía y Competitividad, MINECO (SEV-2016-0644).

To investigate the influence of vitamin D status on in vitro fertilisation (IVF) results, a meta-analysis of 15 cohort studies of 3711 women undergoing IVF was performed. Women were classified into...

Study question Is there any difference in the protein composition of the endometrial fluid aspirate (EFA) obtained the day of embryo transfer in in vitro fertilization (IVF) cycles achieving and not achieving pregnancy? Summary answer Comparative analysis identified a differential protein expression pattern in 'implantative' and 'non-implantative' IVF cycles. What is known already EFA allows non-invasive characterization of the endometrium, and may contain important information on its receptivity when performing (IVF) cycles. Endometrial side of implantation has usually been studied with endometrial biopsy in an IVF cycle prior to embryo transfer, focusing on 'receptive/non-receptive' endometria and with low-throughput proteomic techniques. Study design, size, duration We have compared the protein expression patterns in EFA from a total of 110 women undergoing IVF, corresponding to 50 implantative and 60 non-implantative IVF cycles. Discovery (38 patients) and Validation (42 patients) sample cohorts were analyzed using a high-throughput differential proteomic approach. Then, the differential expression of glycogen phosphorylase B (PYGB) was validated by western blotting in an additional cohort (30 patients). The study period was 18 months. Participants/materials, setting, methods The population under study consisted of 110 women aged 18-40 years old, undergoing their first or second IVF/ intracytoplasmic sperm injection cycle, with normal uterus and endometrium, and 1-2 good quality embryos, and embryo transfer being performed on Day 3. Endometrial fluid aspiration was performed immediately before the embryo transfer. Samples (80) were initially distributed in two independent cohorts and analyzed by liquid chromatography-mass spectrometry. The first cohort was used for the discovery and the second for the validation of the results. Filter-aided sample preparation was used for the in-solution tryptic digestion of the proteins present in the samples, followed by label-free mass spectrometry analysis. In order to unravel the molecular features of receptivity, the lists of differential proteins were thoroughly analyzed using different bioinformatic tools, including GSEA, IPA and GO analysis. Main results and the role of chance A false discovery rate-based correction of the t-test P-values was carried out in order to strengthen the reliability of the results. Functional analyses denoted the deregulation of important processes governing receptivity, such as antimicrobial response, cell-cell interaction, immune response and inflammatory signaling, among others. Overall eight proteins were commonly deregulated in both studied datasets and brain form glycogen phosphorylase (PYGB) was selected for confirmatory analysis. Limitations, reasons for caution Our results were obtained from patients with normal uterus and endometrium and with good quality embryos, who had fresh Day-3 embryo transfer, in stimulated cycles. Therefore, our observations may not be applicable to poor prognosis cases or non-stimulated cycles. Wider implications of the findings This work provides insights into the molecular features of implantative IVF cycles using non-invasive methods. It reveals that EFA may reflect an increased inflammatory state in non-implantative endometrium. Additionally, it proposes PYGB as a potential biomarker for endometrial receptivity or implantation success. This knowledge opens a new avenue for developing embryo transfer strategies, through the improvement of embryo culture media or modifying endometrial fluid composition to increase pregnancy rates. Study funding/competing interest(s) This study was partially funded by a Grant for Fertility Innovation (GFI, 2011) from Merck (Darmstadt, Germany). Authors declare no competing interests. Trial registration number Not applicable.

The role of acquired thrombophilia has been accepted as an etiology of recurrent miscarriage (RM); however, the contribution of specific inherited thrombophilic genes to this disorder has remained controversial. An increased incidence of RM has been suggested in women with inherited thrombophilia.In this prospective study, assisted women with RM or repeated implant failure (RIF) were subjected to Thromboincode analysis, in order to identify 12 genetic variants for Factor V Leiden, Factor V Hong Kong, Factor V Cambridge, FII, FXIII, FXII, and A1 carriers. Patients included in this study were separated in RM cases (n=43), RIF cases (n=36) and RIF+RM (n=76). As a control group, patients undergoing IVF treatment (n=34) were used and a previously described 249 cases population as a representative sample of Spanish population were selected. Level of statistical significance was p0.05 and groups were compared by Fisher test, except for age that was compared by t-test.Regarding FXIII, higher values were observed in RM (69.76%), RIF (70%) and in RM+RIF (68.42%) group when compared with our control group (52.94%) and general Spanish population (56.5%), but these differences were statistically significant only in RIF group (p=0.043, p=0.01).Concerning our findings, both RM and RIF patients had a very similar panel of thrombophilia polymorphisms, suggesting that, in both, thrombophilia might have an important contribution. High frequency of Val34Leu polymorphism in RM/ RIF presumably speaks in favor of a multifactorial RM genesis, wherean altered thrombophilia status plays a role.

The endogenous cannabinoid system has been characterized in some female reproductive organs but little is known about the expression and localization pattern of cannabinoid-degrading enzymes in relation to the CB1 cannabinoid receptor in human oocytes. In this study, we focus on the investigation of the presence and differential distribution of fatty acid amide hydrolase (FAAH) and monoglyceride lipase (MGLL) in relation to CB1 during the maturation of human oocytes. We used a total of 290 human oocytes not suitable for in vitro fertilization/intracytoplasmic sperm injection (ICSI): germinal-vesicle (GV) and metaphase-I (MI) stages and metaphase-II (MII) oocytes that had not developed into an embryo after ICSI. Cannabinoid-degrading enzymes and the cannabinoid CB1 receptor were present in human oocytes. Specifically, FAAH was detected in the periphery of the oocyte from the GV to MI stage and co-localized with CB1. Later, by the MII stage, FAAH was spread within the oocyte, whereas MGLL immunostaining was homogeneous across the oocyte at all stages of maturation and only overlapped with CB1 at the GV stage. This coordinated redistribution of cannabinoid system proteins suggests a role for this system in the maturation of the female gamete.

Research question What is the frequency of cervical pregnancy in women undergoing assisted reproductive technologies (ART) and what are the risk factors? Design Case-control study of women undergoing assisted reproductive technology (ART) at 25 private assisted reproduction clinics run by the same group in Spain. Two control groups (tubal ectopic pregnancies and intrauterine pregnancies) were established. The main outcome measure was frequency of cervical pregnancy. Demographic, clinical factors and IVF parameters were assessed for their influence on cervical pregnancy risk. Results Thirty-two clinical pregnancies were achieved out of 91,067 ongoing pregnancies, yielding a rate of 3.5/10,000. Cervical pregnancies represented 2.02% of all ectopic pregnancies (32/1582). The main risk factors two or more previous pregnancies (OR 2.68; CI 1.18 to 6.07); two or more previous miscarriages (OR 4.21; CI1.7 to 10.43), one or more previous curettages (OR 3.99, CI 1.67 to 9.56), two or more previous curettages (OR 4.71; CI 1.19 to 18.66) and smoking (OR 2.82 CI 1.14 to 6.94). History of caesarean sections and tubal pregnancy was not associated with an elevated cervical pregnancy risk. Infertility conditions and endometrial thickness were similar across the three groups. The proportion of women from whom fewer than 10 oocytes were retrieved was higher in the clinical pregnancy group than in the IUP group. Conclusions In ART, the main risk factors for cervical ectopic pregnancy are a history of at least two pregnancies, miscarriages, at least one curettage and smoking. IVF parameters do not seem to influence the development of clinical pregnancies. Cervical pregnancies are less common in ART than previously reported, attributable to improvements in ART; a publication bias in early IVF reports cannot be ruled out.

To assess whether there are proteins in endometrial fluid aspirate (EFA) that predict implantation. The population under study consisted of 285 women undergoing embryo transfer (ET). Endometrial fluid aspiration was performed immediately before ET. Results of proteomic analysis of EFA were compared between 33 cases who achieved pregnancy and 33 who did not. Samples were analysed by 2D electrophoresis and mass spectrometry. Blood samples were studied by ELISA Pregnancy rates and maternal complications were compared to those in women refusing aspiration. We found 23 proteins differentially expressed in the EFA in conception cycles: 4 up-regulated proteins and 19 down-regulated (FC = 0.31 0.78) (among others, arginase-1, actin B, PARK-7, cofilin-1, stathmin, annexin-2 and CAPZB). Among the five studied proteins that were differentially expressed in EFA, none was differentially expressed in serum. The aspiration procedure had no impact on pregnancy rate. No maternal complications were reported. We found a very different protein profile in implantative cycles, the majority of proteins being down-regulated. This probably reflects a different endometrial functional status, more favourable to implantation. EFA proteomic analysis could be a useful tool in the planning ET strategies.

The aim of this study was to ascertain the incidence of pelvic inflammatory disease (PID) after intrauterine insemination (IUI). A systematic review was conducted using three different approaches: a search of IUI registries; a search of published meta-analyses; and a search of prospective randomized trials. Search terms were 'IUI', 'complications', 'infection' and 'PID'. Two IUI registers were identified that met the inclusion criteria, totalling 365,874 cycles, with 57 PID cases being reported. The post-IUI PID rate was 0.16/1000 (95% CI 0.2 to 0.3/1000). The frequency was higher in husband sperm cycles (0.21/1000) (28/135,839) than in donor sperm cycles (0.03/1000) (1/33,712) (P < 0.05; OR 6.95). Nineteen meta-analyses were retrieved, which included 156 trials, totalling 43,048 cycles, with no PID case being reported. Seventeen prospective clinical trials published between 2013 and 2014 were identified, totalling 4968 cycles; no PID case was reported. The reported rate of post-IUI clinical PID is low (0.16/1000), about 40% higher than reported in the general population of women during their reproductive life. No antibiotic prophylaxis should be recommended unless there is an associated risk factor.

In this retrospective study, the efficiency of carrying out rescue intrauterine insemination (IUI) in low-responder patients undergoing IVF when no oocytes were retrieved after follicular aspiration and when HCG timing was adequate was analysed. A historical control group was used. Over 13 years, women undergoing IVF with failure to obtain oocytes at follicular aspiration underwent rescue IUI if the following criteria were met: adequate HCG timing; one normal tube; motile sperm count after preparation over 3 million/ml; and ultrasound visualization of one to six follicles over 13 mm. The rescue IUI was carried out 1 h after follicular aspiration. Results were compared with those of a standard IUI population (5394 cycles) in the same period. Confidence intervals were calculated using Poisson 97.5% confidence upper tail limits when no event was observed in the study sample. No pregnancies were achieved among the 54 cases who underwent rescue IUI (confidence interval: 0 to 6.8%). This pregnancy rate was lower than that observed in the general IUI population (17.5%) (relative risk, 19.2). After adjusting for age and endometriosis, the relative risk was 11.7. The rescue IUI is an inefficient procedure. Its efficacy is unlikely to exceed 7% pregnancy rate per IUI.

Aims: To explore whether the transfer of very poor quality (VPQ) embryos is associated with an increase in congenital malformations or perinatal problems. Methods: In this retrospective case-control study, 74 children conceived by in vitro fertilization (IVF) and/or intracytoplasmic sperm injection (ICSI) resulting exclusively from the transfer of VPQ embryos were compared with 1,507 children born after the transfer of top morphological quality (TQ) embryos over the same period of time in the same centers. Results: The prevalence of birth defects in children resulting from VPQ embryos was 1.35% (1/74), similar to the 1.72% (26/1,507) when only TQ embryos were transferred; the rate of chromosomal abnormalities detected was also similar (0.0 vs. 0.4%), as was perinatal mortality. After correcting for multiplicity (higher in the TQ group), the aforementioned parameters remained similar in the two groups. Conclusion: Congenital malformations and perinatal complications do not seem to be more common in children born after transfer of VPQ embryos in IVF/ICSI cycles. Given our preliminary data, which need to be confirmed in much larger studies, when only VPQ embryos are available for transfer in IVF/ICSI cycles, we do not believe that they should be discarded with the intention of avoiding birth defects or perinatal complications.

This study assessed the influence of adding embryos with different embryo quality on pregnancy rate and multiple pregnancy rate (MPR). The study included 1891 IVF transfers performed at two centres with different embryo transfer policies. Pregnancy rate and MPR were analysed following three models and then including embryo quality. A predictive mathematical model and two scatter plots were constructed. The model based on embryo independence was incompatible with the observed data, while both the ground and collaborative models provided excellent fits. The collaborative model, however, predicted multiple pregnancies, especially triplets, more accurately. Transfer of additional embryos, irrespective of embryo quality, always increased pregnancy rate and MPR. When implantation rate was low, there was a marked increase in pregnancy rate but only a relatively small increase in MPR. In contrast, with higher implantation rates, the increase in pregnancy rate was mainly due to the increase in MPR, with the same singleton pregnancy rate. Transfer of additional embryos, irrespective of embryo quality, follows a collaborative pattern and always results in an increase in pregnancy rate and MPR. The scatter plots accurately predicted the influence of the different combinations of number and embryo quality on pregnancy rate and MPR.

Objective To assess whether, in GnRH agonist IVF cycles where there is a risk of ovarian hyperstimulation syndrome (OHSS), the addition of cabergoline to the hydroxyethyl starch (HES) infusion could decrease OHSS incidence and severity. Materials and methods Prospective randomized study. The population under study consisted of women undergoing IVF cycles with GnRH agonist protocols, at risk of OHSS (more than 20 follicles observed larger than 12 mm in diameter and/or estradiol levels of 3000–5000 pg/mL). Women received a slow infusion of 500 mL of 6% HES during follicular aspiration alone or combined with 0.5 mg cabergoline administration for 8 days, starting on the day of hCG administration. Results The rates of OHSS (both early and late) were very similar in the HES alone group (3.19% (3/94)) and in the HES plus cabergoline group (5.68% (5/88)), as were the rates of severe cases of OHSS (1.06% and 2.27%). Pregnancy rates (PR) were also similar in the two groups (ongoing PR per transfer, 47.56% and 47.50%). Comments The co-administration of cabergoline in patients receiving HES due to OHSS risk did not reduce the rate or severity of OHSS in GnRH agonist IVF cycles.

The aim of our study was to ascertain the influence of hCG levels at oocyte pick-up on IVF outcomes, and their relationship with clinical parameters.A prospective study was performed including 473 women undergoing IVF, aged under 40 years. Blood samples to analyze hCG levels were obtained at the time of follicular aspiration, 36 h after the administration of 250 μg of recombinant hCG.Neither the numbers of oocytes obtained or fertilized, nor the pregnancy rate, were correlated with hCG levels. Moreover, hCG values were very similar in women who did and did not become pregnant (123.3 ± 48.7 and 117.5 ± 44.7 mUI/mL). Cases in which no oocytes were recovered after follicular aspiration had similar hCG levels to those in which more than 1 oocyte was obtained. On the other hand, hCG levels were negatively related to body mass index, weight, and age.These data indicate that after the administration of 250 μg of recombinant hCG, hCG levels are not responsible for failure to recover oocytes. Specifically, there was no correlation between plasma hCG levels and the number of oocytes obtained or other markers of IVF outcome. There was, however, an inverse relationship with BMI, body weight and age.

The endogenous opioid system has been characterized in some female reproductive organs, but little is known about the expression of these receptors in human oocytes. This study investigated the presence and differential distribution of the opioid receptors during the maturation of human oocytes. A total of 821 human oocytes from an intracytoplasmic sperm injection (ICSI) programme were studied including 213 at germinal-vesicle (GV) stage and 164 at metaphase-I (MI) stage and 444 failed fertilization metaphase-II (MII) oocytes. Additionally 31 MII oocytes corresponding to cases where ICSI was not attempted and 50 failed fertilization MII oocytes from the IVF programme were included. Western blot analysis revealed the presence of the delta (OPRD1), kappa (OPRK1) and mu (OPRM1) opioid receptors in human oocytes. The OPRK1 and OPRM1 immunostaining patterns changed during the maturation of the oocyte, while the OPRD1 pattern was the same throughout. In particular, OPRD1 were detected in peripheral tissue from the GV to the MII stage. OPRK1 were found peripherally at the GV stage, more internally at MI and homogeneously at MII. Finally, OPRM1 were located peripherally at the GV stage and homogeneously in MI and MII oocytes. Opioids may have a role in oocyte maturation, acting via receptors. The opioid system has been well characterized in the central nervous system, but it is now known that opioids also act in reproductive organs. However, little is known about the presence and function of this system in human oocytes and its role in their maturation. In this study, we investigated the presence and differential distribution of three (delta, kappa and mu) opioid receptors (proteins which bind the opioids) during the maturation of human oocytes. A total of 821 human oocytes (from 253 patients) not suitable for intracytoplasmic sperm injection (ICSI) or which did not develop into an embryo after ICSI were studied. Thus, we have verified the presence of the delta, kappa and mu opioid receptors in human oocytes. The kappa and mu localization changed during the maturation of the oocyte, while the Delta localization was the same throughout. In particular, the delta receptor was detected in the periphery of the oocyte. On the other hand, the kappa receptor was found peripherally at the beginning, more internally during maturation and homogeneously at the end of maturation. Finally, the Mu receptor was located peripherally at the beginning of maturation and homogeneously in the rest of the maturation stages. This finding suggests a possible role for opioids, acting via receptors, in the maturation of the oocyte.

This single-centre, randomized, parallel group, comparative study aimed to identify potential benefits of mid-follicular recombinant human LH (r-HLH) supplementation in women aged 35-39 years undergoing ovarian stimulation for intracytoplasmic sperm injection (ICSI). The main endpoint was the number of metaphase II oocytes retrieved. After pituitary suppression with a gonadotrophin-releasing hormone agonist, ovarian stimulation was initiated with recombinant human FSH (r-HFSH; 300-450 IU/day). On stimulation day 6, patients were randomized to receive r-HFSH alone or r-HFSH + r-HLH (r-HLH 150 IU/day) for the remainder of the stimulation period. Final follicular maturation was triggered with 250 mug of recombinant human chorionic gonadotrophin. After assessing oocyte nuclear maturity, oocyte were fertilized by ICSI and afterwards embryo quality was analyzed. Of the 131 women enrolled, 68 were allocated to r-HFSH alone and 63 to r-HFSH + r-HLH. No significant differences were observed in markers of either oocyte or embryo quality or quantity. However, higher rates of implantation and live birth per started cycle were observed with r-HLH supplementation than with r-HFSH alone. Although additional large studies are required to further investigate these findings, r-HLH supplementation for women aged 35-39 years undergoing ICSI is recommended as it may have a beneficial action on implantation.

Introduction: Couples who have not achieved a pregnancy within one year of unprotected intercourse qualify for a fertility work-up. A semen analysis is part of this work-up and includes the assessment of sperm concentration, motility and morphology. The predictive value of sperm morphology has been

BACKGROUND: The aim was to ascertain whether using ultrasound guidance during intrauterine insemination (IUI) could increase pregnancy rates (PRs). METHODS: The population under study consisted of 73 consecutive couples subjected to IUI in our Human Reproduction Unit, between June and December 2006, with a total of 231 IUI cycles performed. The patients were randomized using a computer-generated random numeric table into two groups: ultrasound-guided IUI group (n = 33) and clinical IUI group (n = 40). RESULTS: The PR was 16.0% per cycle in ultrasound-guided IUI and 16.8% in the control group, no statistically significant differences being observed between the groups. The 95% confidence interval for the difference in PRs of 0.8% was - 8.8 to 10. There were no differences in PR per woman, nor in first-cycle PR. The cumulative PR was also similar in both populations. Although the initial intention was to perform a study involving a larger number of cases, after a first interim analysis, the study was interrupted due to its futility. There were no differences in PR according to the different cervico-uterine angles. CONCLUSIONS: Ultrasound-guided IUI does not produce better results than blind insemination, because the PR per cycle is similar. ClinicalTrials.gov ID NCT00809952.

Objective To describe the expression of cannabinoid receptors CB1 and CB2 and cannabinoid-degrading enzymes fatty acid amide hydrolase (FAAH) and monoglyceride lipase (MGLL) in human granulosa cells and to investigate their differential distribution with respect to CB1 at various stages during the nuclear maturation of the oocyte. Design Analysis of granulosa cells from germinal vesicle (GV), metaphase I (MI), and MII oocytes by quantitative reverse transcriptase—polymerase chain reaction, Western blot, and indirect immunofluorescence assays. Setting Academic research laboratory. Patient(s) Patients from the Human Reproduction Unit of Cruces University Hospital undergoing intracytoplasmic sperm injection. Intervention(s) We analyzed the granulosa cells of 300 oocytes from 53 patients. The oocyte maturation stages were 75 at GV stage, 51 at MI, and 174 at MII. Main Outcome Measure(s) The mRNA and protein expression of CB1, CB2, FAAH, and MGLL and localization in granulosa cells at each oocyte maturation stage. Result(s) CB1, FAAH, and MGLL are present in human granulosa cells during oocyte maturation, but the presence of CB2 receptor is not entirely clear in those cells. CB1 and FAAH were detected in the periphery of the granulosa cells from the GV to the MII oocytes, and they colocalized in some portions of the cell membrane. On the other hand, MGLL immunostaining was more homogeneous across the cell and overlapped with CB1 only weakly. Conclusion(s) The presence of the cannabinoid system in granulosa cells suggests a possible role of this system in the nuclear maturation of the oocyte.

Objective : To compare the results of intrauterine insemination (IUI) when GnRH antagonist was added—to avoid IUI on weekend—with those obtained with the standard IUI protocol. Study design : In an IUI program under ovarian stimulation with gonadotropins when one or more follicles of 15–16 mm were seen, if it was not possible for logistic reasons (weekend) to perform the insemination 72 h later, GnRH antagonist was administered until human chorionic gonadotropin (hCG) administration. The IUI was performed on Monday. We compared the results of this IUI ``weekend-free'' group with our results in standard IUI cycles, where IUI was performed 36–38 h after reaching optimal follicular growth. Results : Both groups were comparable regarding the main demographic parameters, except for higher estradiol levels, due to the prolonging ovarian stimulation. The per cycle pregnancy rate (PR) were very similar in both groups: 15.7% in the weekend-free IUI versus 16.5% in standard IUI. The multiple pregnancy rate and the hyperstimulation rate were also similar. A non-significant trend to higher high-order multiple pregnancy was observed in the weekend-free IUI. Conclusions : In IUI cycles under ovarian suprastimulation with gonadotrophins, the use of GnRH antagonist allows the manipulation of the follicular development in such a way that it is possible to avoid inseminations on the weekends, without apparently reducing the PR.

To ascertain the influence of the duration of the 'interval loading-discharging embryos' (ILDE) on the results of embryo transfer.The population under study consisted of 450 consecutive fresh embryo transfers. ILDE was measured in all transfers. Pregnancy and implantation rates were analyzed.Conceptional cycles had a lower (mean +/- SD) ILDE than non conceptional cycles (53.5 +/- 43.6 s vs 63.7 +/- 49.3). When only easy transfers--defined as those not requiring cervical tenaculum--were considered, similar differences were observed. The following pregnancy rates were obtained according ILDE duration: 38.9% (ILDE30 s), 33.2% (ILDE 31-60), 31.6% (ILDE 61-120) and 19.1% (ILDE120) (P0.05). When only easy transfers were considered, similar results were obtained: 40.0%, 33.3%, 32.0% and 19.4%, respectively. Similar results were observed regarding implantation rates regarding the whole population (21.2%, 15.4%, 15.9% and 9.4%, respectively; P0.01) as well as when only easy transfers were considered.ILDE duration is a prognostic factor of pregnancy rate and of implantation rate in IVF. The longer the ILDE duration, the lower the pregnancy and implantation rates. The decrease in pregnancy and implantation rates is gradual until an ILDE of 120 s, and decreases sharply afterwards. It is recommended to speed up the embryo transfer process, wherever possible. ILDE120 s carries a poor prognosis and should, when possible, be avoided.

To date, there is scarce information on the metabolic and biochemical repercussions of Zn-deficiency in fish. In this work, the effect of dietary Zn-deficiency on the diet utilization and the metabolism of activated oxygen species in rainbow trout (Oncorhynchus mykiss) has been studied. Fish were randomly separated in different lots according to their Zn-starvation and diet intake. In crude extracts of liver, gut and muscle, total and isoenzymatic superoxide dismutase and catalase activities were analysed. Lipid peroxidation was also determined in the same tissues. Western blotting was performed using antibodies against manganese- and copperzinc-containing superoxide dismutase. Lots fed on the Zn-deficient diet and with low intake showed significantly lower weight gain and feed-conversion efficiency indexes than control trouts. However, these parameters returned to control values when trouts were recovered by feeding them a control diet ad libitum. In control trouts, three independent copperzinc superoxide dismutase isozymes were detected in liver, whereas only one isozyme was present in the other lots. However, by Western blotting analysis the presence of a manganese superoxide dismutase was found in liver from all lots except in control trouts. Catalase activity and lipid peroxidation values were mainly detected in liver and gut, respectively, and both parameters increased in all lots with respect to the control group. Our results thus suggest that in rainbow trout an oxidative stress appears to occur as a consequence of Zn-deficiency.

Objective To evaluate, in patients with hydrosalpinges, the effect on in vitro fertilization (IVF) outcome of the insertion by hysteroscopy of an intratubal blocking device, in cases where laparoscopic salpingectomy or laparoscopy was contraindicated. Study design A prospective interventional case series study was conducted in fifteen women with unilateral (N = 6) or bilateral hydrosalpinges (N = 9) submitted for IVF. In all of them, laparoscopic salpingectomy was contraindicated. Hysteroscopic insertion of the Essure intratubal device in a consultation room setting was performed. IVF results were compared with those of women where hydrosalpinx was treated by laparoscopic salpingectomy (48 women, 76 cycles). Results There were no complications during or immediately after the procedure in any of the patients. There were four pregnancies from 16 embryo-transfers with own oocytes, one spontaneous pregnancy after unilateral Essure insertion, and one pregnancy after oocyte donation. In one case the hydrosalpinx grew and pelvic inflammatory disease developed 6 months after the insertion, requiring bilateral adnexectomy. Although not of statistical significance, IVF pregnancy rates were somewhat lower than in the laparoscopic salpingectomy group, which was attributed to the lower ovarian reserve before Essure insertion. Conclusion The hysteroscopic insertion of the Essure intratubal device prior to IVF is a reasonable option in cases where laparoscopic salpingectomy is contraindicated. Larger series are required to assess pregnancy outcome.

To assess the value of the levonorgestrel-releasing intrauterine device (LNG-IUD) in the treatment of pelvic pain in patients with endometriosis in which previous medical and surgical treatments have failed.This prospective study evaluated pelvic pain by means of both a visual analog grading scale and a verbal descriptive grading scale. Pain was evaluated before insertion of the LNG-IUD and afterwards, at 3-month intervals. Adverse effects and reasons for removal were recorded. Me dian follow-up was 22 months (range, 3-36 months). Response rates were expressed as a percentage of the initial cohort in whom the LNG-IUD was inserted, including cases in which the LNG-IUD was extracted.Three months after LNG-IUD insertion, almost 50% of patients reported an improvement in pain symptoms, with this figure rising to 60% after 6 months and almost 70% by the end of follow-up. A trend of better responses was observed when endometriosis was diagnosed10 years before, as well as in stage I-II endometriosis, but these differences were not statistically significant.In endometriosis patients with recurrent pelvic pain in whom previous medical and surgical treatments have failed, LNG-IUD insertion should be considered before radical surgery is performed.

Objective To relate the glutathione peroxidase (GPX) activity level in human seminal plasma with standard semen parameters and spermatozoa fertilization potential in terms of fertilization and pregnancy rates in an IVF program. Design Prospective study. Setting Human Reproduction Unit at Cruces Hospital (Vizcaya, Spain). Patient(s) Three hundred consecutive males from infertile couples participating in the IVF program. Intervention(s) None. Main Outcome Measure(s) Analysis of GPX activity in seminal plasma by spectrophotometry. Result(s) GPX activity in seminal plasma was significantly lower in patients with abnormal sperm as assessed by 1999 and 2010 World Health Organization (WHO) criteria, compared with normozoospermic individuals. There was a more significant decrease in those samples with severe sperm pathologies. GPX values were significantly lower in samples with severe asthenozoospermia, oligozoospermia, and teratozoospermia compared with normal samples. However, there was no correlation between GPX activity in seminal plasma in IVF patients and fertilization rates or pregnancy outcome. Conclusion(s) Although seminal plasma GPX activity was related to semen quality according to WHO parameters, such an association was not found with IVF–intracytoplasmic sperm injection (ICSI) outcome, presumably because of the well-known ability of IVF-ICSI procedures to overcome sperm deficiencies in the fertilization process.

7 páginas., [Background]: The objective of this study is to investigate the influence of male age on human fertility, defined as the birth rate for a given population. [Methods]: Data from the Spanish National Statistics Institute (INE) for the year 2004 from a total of 454,753 newborn infants and sorted by male and female age groups were evaluated. In order to correct the influence of female age-related fertility, a different analysis was performed considering only women under 30 years of age. [Results]: From a demographic point of view, male fertility started to decline at 35–39 years of age. This decline is constant and follows an exponential pattern (slope –0.11 to –0.12). The trend persisted when the data were adjusted for every 1,000 men in the age group, as well as when only women under the age of 30 were considered. Male fertility showed a 21–23% annual decrease starting at the age of 39. [Conclusion]: An exponential decrease in human fertility which is independent of the woman’s age was observed with male aging. This decay is probably due to a downfall in male fecundity, closely related to a decline in sperm quality. However, social or behavioral causes for this trend cannot be excluded.

Endocannabinoid anandamide and cannabinoid receptors have been described in some organs of the female reproductive system, but little is known about the expression of these receptors in human oocytes. The aim of the study was to describe the expression of cannabinoid receptors in human oocytes and to investigate their differential distribution at various stages of meiotic resumption in human oocytes. A total of 750 human oocytes from 214 patients were analysed by Western blot, immunocytochemistry and PCR. For this study, oocytes that were not suitable for intracytoplasmic sperm injection (ICSI) (germinal-vesicle and metaphase-I stages), as well as metaphase-II oocytes that had not developed into an embryo after ICSI were used. Western blot analysis revealed the presence of CB1 and CB2 receptor proteins in human oocytes. CB1 and CB2 receptor immunostaining patterns changed during the various stages of meiotic resumption. Localization of CB1 receptor was peripheral at germinal-vesicle stage, homogeneous over the entire oocyte at metaphase I and peripheral at mature metaphase II. CB2 receptor localization was peripheral at germinal-vesicle and metaphase-I stages but homogeneous over the entire cell at metaphase II. This finding suggests a possible role for endocannabinoids, acting via receptors, in the maturation of female gametes and fertilization. The number of couples with sterility problems attending fertility programmes is rising but treatment is not always successful. Important problems associated with failure to conceive remain unresolved because many physiological aspects of human reproduction are still unknown. Endocannabinoids are endogenous chemical compounds that mimic the action of the main psychoactive component of marijuana, delta-9-tetrahydrocannabinol. An endogenous cannabinoid named anandamide has been found in human follicular fluid. Thus, in order to develop knowledge in this field, in the present study we have described the presence of the cannabinoid receptors CB1 and CB2 (the proteins required to mediate the action of the cannabinoids) in the early stages of meiotic resumption of oocytes (the stages before ovulation) and we could postulate that the endocannabinoids could act in the regulation of maturation of oocytes. Our study, together with other studies, indicates that the endocannabinoid system may play a role in human reproduction.

Objective To compare the pregnancy rates (PRs) in intrauterine insemination (IUI) using recombinant FSH (rec-FSH) or highly purified urinary FSH (HP-FSH). Design Systematic review and metaanalysis. Setting University hospital. Patient(s) None. Intervention(s) Electronic and manual searches. Main Outcome Measure(s) PR, per first cycle PR and per woman PR. Result(s) Six randomized trials (713 women, 1,581 cycles) were identified. In three the same doses of rec-FSH and HP-FSH were used ("equal dose" group), whereas in the other three the ratio HP-FSH:rec-FSH dose was 1.5. The global metaanalysis showed no differences in PRs. The PR per cycle was similar across the 1.5 ratio group (14.51% vs. 14.93%; relative risk [RR], 0.970; 95% confidence interval [CI], 0.687–1.369). However, the metaanalysis of the equal dose group, showed differences in the PR in favor of rec-FSH (16.36% vs. 12.31%; RR, 1.394; 95% CI, 1.004–1.936). Per woman PR analysis showed similar results (41.44% vs. 31.55%; RR, 1.273; 95% CI, 0.987–1.643). Per first cycle PR analysis showed a similar trend, although the difference did not reach significance (RR, 1.434; 95% CI, 0.934–2.203). Conclusion(s) Rec-FSH was associated with higher per cycle PR than HP-FSH, when used at the same dose, whereas the PR were similar when the dose of rec-FSH was 50% lower.

To ascertain if the daily activity in the in vitro fertilization (IVF) laboratory is related to pregnancy rates (PR) and fertilization rates (FR) in an IVF program.A retrospective study was performed to compare the PR and the FR obtained in 845 oocyte retrievals (OR) and 713 embryo transfers (ET), according to the daily workload. Different cutoffs were established:or = 3 OR per day vs.3 OR per day;or = 3 ET per day vs.3 ET per day, and also a cutoff considering the global activity in 3 different categories: optimal (level I), overload (level II) and high overload (level III), both the day of OR and of ET.The PR on the days withor = 3 OR and with3 OR were similar, as were the days withor = 3 ET or3 ET. There were no differences in PR when the activity the day of OR was level I, II or III (24.4%, 25.2% and 28.3%, respectively) or when the activity on the day of ET was level I, II or III (29.6%, 37.3% and 23.7%, respectively).We failed to show any adverse results on our IVF program associated with the daily workload.

Objective The purpose of this study was to compare the pregnancy rate in an artificial insemination donor program in women with minimal endometriosis and in women without endometriosis. Study Design A prospective double-blinded study was conducted in women with azoospermic partners. Results The per-cycle pregnancy rate was 8.6% (9/104 women) in the minimal endometriosis group vs 13.3% (26/196 women) in the control group. The per-woman pregnancy rate was 37.5% (9/24 women) in the minimal endometriosis group and 51.0% (26/51 women) in the control group. Conclusion Pregnancy rates were statistically similar in normal women and in women with minimal endometriosis.

BACKGROUND: Recent reports have suggested that ultrasound (US) guidance during embryo transfer might improve pregnancy rates. METHODS: A prospective randomized (computer-generated random table) trial was performed to compare embryo transfer under abdominal US guidance (n 255 women) with clinical touch embryo transfer (n 260). RESULTS: The clinical pregnancy rate was 26.3% (67/255) in the US-guided transfer group compared with 18.1% (47/260) in the clinical touch transfer group (P < 0.05). The implantation rate was 11.1% (100/903) in the US group compared with 7.5% (66/884) in the clinical touch group (P < 0.05). US-guided transfer was associated with a decrease in the difficulty of the transfers: 97% of transfers were easy in the US-guided group compared with 81% in the clinical touch group (P < 0.05). CONCLUSIONS: US-guided embryo transfer increased pregnancy and implantation rates in IVF cycles, as well as the frequency of easy transfers. It is suggested that the decrease in cervical and uterine trauma can play a role in the increase in pregnancy rates associated with USguided transfer. It is recommended that embryo transfer should be performed under US guidance.

Introduction: The lipid composition of spermatozoa plays an important role for the viability, maturity and functions of those cells. Has been suggested that the proportion of unsaturated fatty acids (UFA) may have an influence over the membrane fluidity and flexibility. The aim of the study was to evaluate the relationship between motility and fatty acid composition of human spermatozoa before and after in vitro capacitation. Materials and Methods: The population under study consisted of 50 consecutive males from infertile couples subjected to FIV/ICSI program in our Human Reproduction Unit from Cruces Hospital (Vizcaya, Spain), between June and September 2007. Patients were divided in two groups according to their sperm motility: group I (n = 25) corresponded to ≥40% of progressively motile sperm and group II (n = 25)