SARS-CoV-2 is not found in human semen during mild COVID-19 acute stage.

The aim of this study was to verify the presence of SARS-CoV-2 in the seminal sample of men during the acute phase of COVID-19. A prospective study was performed with inclusion of twenty-two men diagnosed with COVID-19 through RT-PCR from pharyngeal smear samples and who were in the acute phase of infection. These men were evaluated regarding medical history and physical examination. Furthermore, seminal samples of each men were collected 7, 14 and 21 days after the infection was confirmed. The sample were used for seminal analysis, as well as for the presence of SARS-CoV-2 using RT-PCR technique. In addition, cell culture was performed with subsequent repetition of the analysis of viral presence. None of the semen samples collected was positive for the detection of the virus that causes COVID-19. Most of the men evaluated had a mild condition and the loss of smell was the most frequent symptom. There were no significant changes in seminal parameters within the period of study. Based on our pilot data, patients with a mild form of COVID-19 in the acute stage of the disease are unlikely to have SARS-CoV-2 in semen.

The risk of birth defects with conception by ART.

STUDY QUESTION: What is the association between ART conception and treatment parameters and the risk of birth defects? SUMMARY ANSWER: Compared to naturally conceived singleton infants, the risk of a major nonchromosomal defect among ART singletons conceived with autologous oocytes and fresh embryos without use of ICSI was increased by 18%, with increases of 42% and 30% for use of ICSI with and without male factor diagnosis, respectively. WHAT IS KNOWN ALREADY: Prior studies have indicated that infertility and ART are associated with an increased risk of birth defects but have been limited by small sample size and inadequate statistical power, failure to differentiate results by plurality, differences in birth defect definitions and methods of ascertainment, lack of information on ART treatment parameters or study periods spanning decades resulting in a substantial historical bias as ART techniques have improved. STUDY DESIGN, SIZE, DURATION: This was a population-based cohort study linking ART cycles reported to the Society for Assisted Reproductive Technology Clinic Outcome Reporting System (SART CORS) from 1 January 2004 to 31 December 2015 that resulted in live births from 1 September 2004 to 31 December 2016 in Massachusetts and North Carolina and from 1 September 2004 to 31 December 2015 for Texas and New York: these were large and ethnically diverse States, with birth defect registries utilizing the same case definitions and data collected, and with high numbers of ART births annually. A 10:1 sample of non-ART births were chosen within the same time period as the ART birth. Naturally conceived ART siblings were identified through the mother's information. Non-ART children were classified as being born to women who conceived with ovulation induction (OI)/IUI when there was an indication of infertility treatment on the birth certificate, but the woman did not link to the SART CORS; all others were classified as being naturally conceived. PARTICIPANTS/MATERIALS, SETTING, METHODS: The study population included 135 051 ART children (78 362 singletons and 56 689 twins), 23 647 naturally conceived ART siblings (22 301 singletons and 1346 twins) and 9396 children born to women treated with OI/IUI (6597 singletons and 2799 twins) and 1 067 922 naturally conceived children (1 037 757 singletons and 30 165 twins). All study children were linked to their respective State birth defect registries to identify major defects diagnosed within the first year of life. We classified children with major defects as either chromosomal (i.e. presence of a chromosomal defect with or without any other major defect) or nonchromosomal (i.e. presence of a major defect but having no chromosomal defect), or all major defects (chromosomal and nonchromosomal). Logistic regression models were used to generate adjusted odds ratios (AORs) and 95% CI to evaluate the risk of birth defects due to conception with ART (using autologous oocytes and fresh embryos), and with and without the use of ICSI in the absence or presence of male factor infertility, with naturally conceived children as the reference. Analyses within the ART group were stratified by combinations of oocyte source (autologous, donor) and embryo state (fresh, thawed), with births from autologous oocytes and fresh embryos as the reference. Analyses limited to fresh embryos were stratified by oocyte source (autologous, donor) and the use of ICSI. Triplets and higher-order multiples were excluded. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 21 998 singleton children (1.9%) and 3037 twin children (3.3%) had a major birth defect. Compared to naturally conceived children, ART singletons (conceived from autologous oocytes, fresh embryos without the use of ICSI) had increased risks of a major nonchromosomal birth defect (AOR 1.18, 95% 1.05, 1.32), cardiovascular defects (AOR 1.20, 95% CI 1.03, 1.40), and any birth defect (AOR 1.18, 95% CI 1.09, 1.27). Compared to naturally conceived children, ART singletons conceived (from autologous oocytes, fresh embryos) with the use of ICSI, the risks were increased for a major nonchromosomal birth defect (AOR 1.30, 95% CI 1.16, 1.45 without male factor diagnosis; AOR 1.42, 95% CI 1.28, 1.57 with male factor diagnosis); blastogenesis defects (AOR 1.49, 95% CI 1.08, 2.05 without male factor; AOR 1.56, 95% CI 1.17, 2.08 with male factor); cardiovascular defects (AOR 1.28, 95% CI 1.10,1.48 without male factor; AOR 1.45, 95% CI 1.27, 1.66 with male factor); in addition, the risk for musculoskeletal defects was increased (AOR 1.34, 95% CI 1.01, 1.78 without male factor) and the risk for genitourinary defects in male infants was increased (AOR 1.33, 95% CI 1.08, 1.65 with male factor). Comparisons within ART singleton births conceived from autologous oocytes and fresh embryos indicated that the use of ICSI was associated with increased risks of a major nonchromosomal birth defect (AOR 1.18, 95% CI 1.03, 1.35), blastogenesis defects (AOR 1.65, 95% CI 1.08, 2.51), gastrointestinal defects (AOR 2.21, 95% CI 1.28, 3.82) and any defect (AOR 1.11, 95% CI 1.01, 1.22). Compared to naturally conceived children, ART singleton siblings had increased risks of musculoskeletal defects (AOR 1.32, 95% CI 1.04, 1.67) and any defect (AOR 1.15, 95% CI 1.08, 1.23). ART twins (conceived with autologous oocytes, fresh embryos, without ICSI) were at increased risk of chromosomal defects (AOR 1.89, 95% CI 1.10, 3.24) and ART twin siblings were at increased risk of any defect (AOR 1.26, 95% CI 1.01, 1.57). The 18% increased risk of a major nonchromosomal birth defect in singleton infants conceived with ART without ICSI (∼36% of ART births), the 30% increased risk with ICSI without male factor (∼33% of ART births), and the 42% increased risk with ICSI and male factor (∼31% of ART births) translates into an estimated excess of 386 major birth defects among the 68 908 singleton children born by ART in 2017. LIMITATIONS, REASONS FOR CAUTION: In the SART CORS database, it was not possible to differentiate method of embryo freezing (slow freezing vs vitrification), and data on ICSI was only available in the fresh embryo ART group. In the OI/IUI group, it was not possible to differentiate type of non-ART treatment utilized, and in both the ART and OI/IUI groups, data were unavailable on duration of infertility. WIDER IMPLICATIONS OF THE FINDINGS: The use of ART is associated with increased risks of a major nonchromosomal birth defect, cardiovascular defect and any defect in singleton children, and chromosomal defects in twins; the use of ICSI further increases this risk, the most with male factor infertility. These findings support the judicious use of ICSI only when medically indicated. The relative contribution of ART treatment parameters versus the biology of the subfertile couple to this increased risk remains unclear and warrants further study. STUDY FUNDING/COMPETING INTEREST(S): This project was supported by grant R01 HD084377 from the National Institute of Child Health and Human Development. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of Child Health and Human Development, or the National Institutes of Health, nor any of the State Departments of Health which contributed data. E.W. is a contract vendor for SART; all other authors report no conflicts. TRIAL REGISTRATION NUMBER: N/A.

Risks of nonchromosomal birth defects, small-for-gestational age birthweight, and prematurity with in vitro fertilization: effect of number of embryos transferred and plurality at conception versus at birth.

PURPOSE: Excess embryos transferred (ET) (> plurality at birth) and fetal heartbeats (FHB) at 6 weeks' gestation are associated with reductions in birthweight and gestation, but prior studies have been limited by small sample sizes and limited IVF data. This analysis evaluated associations between excess ET, excess FHB, and adverse perinatal outcomes, including the risk of nonchromosomal birth defects. METHODS: Live births conceived via IVF from Massachusetts, New York, North Carolina, and Texas included 138,435 children born 2004-2013 (Texas), 2004-2016 (Massachusetts and North Carolina), and 2004-2017 (New York) were classified by ET and FHB. Major birth defects were reported by statewide registries within the first year of life. Logistic regression was used to estimate adjusted odds ratios (AORs) and 95% CIs of the risks of a major nonchromosomal birth defect, small-for-gestational age birthweight (SGA), low birthweight (LBW), and preterm birth (≤36 weeks), by excess ET, and excess ET + excess FHB, by plurality at birth (singletons and twins). RESULTS: In singletons with [2 ET, FHB =1] and [≥3 ET, FHB=1], risks [AOR (95% CI)] were increased, respectively, for major nonchromosomal birth defects [1.13 (1.00-1.27) and 1.18 (1.00-1.38)], SGA [1.10 (1.03-1.17) and 1.15 (1.05-1.26)], LBW [1.09 (1.02-1.13) and 1.17 (1.07-1.27)], and preterm birth [1.06 (1.00-1.12) and 1.14 (1.06-1.23)]. With excess ET + excess FHB, risks of all adverse outcomes except major nonchromosomal birth defects increased further for both singletons and twins. CONCLUSION: Excess embryos transferred are associated with increased risks for nonchromosomal birth defects, reduced birthweight, and prematurity in IVF-conceived births.

MFGE8 regulates TGF-ß-induced epithelial mesenchymal transition in endometrial epithelial cells in vitro.

This study investigated the role of milk fat globule-epidermal growth factor-factor 8 (MFGE8) in TGF-ß-induced epithelial-mesenchymal transition (EMT) of endometrial epithelial cells. These were in vitro studies using human endometrial epithelial cells and mouse blastocysts. We investigated the ability of TGF-ß to induce EMT in endometrial epithelial cells (HEC-1A) by assessment of cytological phenotype (by light and atomic force microscopy), changes in expression of the markers of cell adhesion/differentiation E- and N-cadherin, and of the transcription factor Snail (by immunofluorescence and immunoblotting), and competence to support embryo attachment in a mouse blastocyst outgrowth assay. We also studied the effects of E-cadherin expression in cells transfected by retroviral shRNA vectors specifically silencing MFGE8. Results demonstrated that TGF-ß induced EMT as demonstrated by phenotypic cell changes, by a switch of cadherin expression as well as by upregulation of the expression of the mesenchymal markers Snail and Vimentin. Upon MFGE8 knockdown, these processes were interfered with, suggesting that MFGE8 and TGF-ß together may participate in regulation of EMT. This study demonstrated for the first time that endometrial MFGE8 modulates TGF-ß-induced EMT in human endometrium cells.

Predictive factors for ovarian response in a corifollitropin alfa/GnRH antagonist protocol for controlled ovarian stimulation in IVF/ICSI cycles.

BACKGROUND: This secondary analysis aimed to identify predictors of low (<6 oocytes retrieved) and high ovarian response (>18 oocytes retrieved) in IVF patients undergoing controlled ovarian stimulation with corifollitropin alfa in a gonadotropin-releasing hormone (GnRH) antagonist protocol. METHODS: Statistical model building for high and low ovarian response was based on the 150 µg corifollitropin alfa treatment group of the Pursue trial in infertile women aged 35-42 years (n = 694). RESULTS: Multivariable logistic regression models were constructed in a stepwise fashion (P <0.05 for entry). 14.1 % of subjects were high ovarian responders and 23.2 % were low ovarian responders. The regression model for high ovarian response included four independent predictors: higher anti-Müllerian hormone (AMH) and antral follicle count (AFC) increased the risk, and higher follicle-stimulating hormone (FSH) levels and advancing age decreased the risk of high ovarian response. The regression model for low ovarian response also included four independent predictors: advancing age increased the risk, and higher AMH, higher AFC and longer menstrual cycle length decreased the risk of low ovarian response. CONCLUSIONS: AMH, AFC and age predicted both high and low ovarian responses, FSH predicted high ovarian response, and menstrual cycle length predicted low ovarian response in a corifollitropin alfa/GnRH antagonist protocol. TRIAL REGISTRATION NUMBER: NCT01144416 , Protocol P06029.

Impact of serum estradiol levels on the implantation rate of cleavage stage cryopreserved-thawed embryos transferred in programmed cycles with exogenous hormonal replacement.

PURPOSE: To investigate the impact of late follicular phase serum estradiol (E2) levels on implantation and pregnancy outcomes of cleavage stage cryopreserved/thawed embryos transferred in programmed cycles with exogenous hormonal replacement. METHODS: Retrospective cohort analysis of IVF patients with transfer of cryopreserved-thawed day-3 embryos in E2 and progesterone (P4) supplemented cycles (n = 208 cycles). MAIN OUTCOME MEASURES: implantation and pregnancy rates according to late follicular phase serum E2 levels and early secretory phase E2/P4 ratios. RESULTS: Logistic regression performed for embryo implantation and for pregnancy outcome in relation to E2 (day 15), P4 (day 15 and 16), before (crude analysis) and after adjustment (adjusted analysis) for baseline characteristics (including age, BMI, serum basal cycle day 3 FSH levels, embryo quality, endometrial lining thickness) showed no significant association. Similarly, ROC analysis showed no impact of cycle day 16 E2/P4 ratio. CONCLUSIONS: Neither late follicular phase serum E2 nor the early E2/P4 ratio were able to predict implantation or pregnancy outcome of day-3 cryopreserved-thawed embryos transferred in artificially programmed cycles.

Globozoospermia is mainly due to DPY19L2 deletion via non-allelic homologous recombination involving two recombination hotspots.

To date, mutations in two genes, SPATA16 and DPY19L2, have been identified as responsible for a severe teratozoospermia, namely globozoospermia. The two initial descriptions of the DPY19L2 deletion lead to a very different rate of occurrence of this mutation among globospermic patients. In order to better estimate the contribution of DPY19L2 in globozoospermia, we screened a larger cohort including 64 globozoospermic patients. Twenty of the new patients were homozygous for the DPY19L2 deletion, and 7 were compound heterozygous for both this deletion and a point mutation. We also identified four additional mutated patients. The final mutation load in our cohort is 66.7% (36 out of 54). Out of 36 mutated patients, 69.4% are homozygous deleted, 19.4% heterozygous composite and 11.1% showed a homozygous point mutation. The mechanism underlying the deletion is a non-allelic homologous recombination (NAHR) between the flanking low-copy repeats. Here, we characterized a total of nine breakpoints for the DPY19L2 NAHR-driven deletion that clustered in two recombination hotspots, both containing direct repeat elements (AluSq2 in hotspot 1, THE1B in hotspot 2). Globozoospermia can be considered as a new genomic disorder. This study confirms that DPY19L2 is the major gene responsible for globozoospermia and enlarges the spectrum of possible mutations in the gene. This is a major finding and should contribute to the development of an efficient molecular diagnosis strategy for globozoospermia.

Characterization of secreted proteins of 2-cell mouse embryos cultured in vitro to the blastocyst stage with and without protein supplementation.

PURPOSE: To identify the secreted proteins of murine embryos grown in vitro. METHODS: Two-cell mouse embryos (n=432) were randomly allocated to culture to the blastocyst stage in protein-free and in protein-supplemented (3 % BSA) media. Proteins were separated by SDS-PAGE; bands were visualized by coomassie staining, followed by in-gel trypsin digestion and liquid chromatography-tandem mass spectrometry. RT-PCR and confocal microscopy were used to confirm gene/protein expression in blastocysts. RESULTS: Of all individually identified proteins, 34 and 23 were found in embryos cultured without and with BSA, respectively, and 20 were common. Identified proteins having an N-terminal secretory sequence or transmembrane domains located on the extracellular backbone were postulated as secreted proteins. Gene and protein expression for two selected molecules were confirmed. Functional analysis revealed over-represented processes related to lipid metabolism, cyclase activity, and cell adhesion/membrane functions. CONCLUSIONS: This study provided evidence to further characterize secreted proteins by mouse embryos grown from the 2-cell to the blastocyst stage in vitro. Because of homology between murine and human, these results may provide information to be translated to the clinical setting.

The search for biomarkers of human embryo developmental potential in IVF: a comprehensive proteomic approach.

The objective of these studies was to identify differentially expressed peptides/proteins in the culture media of embryos grown during in vitro fertilization (IVF) treatment to establish their value as biomarkers predictive of implantation potential and live birth. Micro-droplets of embryo culture media from IVF patients (conditioned) and control media maintained under identical culture conditions were collected and frozen at -80°C on Days 2-3 of in vitro development prior to analysis. The embryos were transferred on Day 3. The peptides were affinity purified based on their physico-chemical properties and profiled by mass spectrometry for differential expression. The identified proteins were further characterized by western blot and ELISA, and absolute quantification was achieved by multiple reaction monitoring (MRM). We identified up to 14 differentially regulated peptides after capture using paramagnetic beads with different affinities. These differentially expressed peptides were used to generate genetic algorithms (GAs) with a recognition capability of 71-84% for embryo transfer cycles resulting in pregnancy and 75-89% for those with failed implantation. Several peptides were further identified as fragments of Apolipoprotein A-1, which showed consistent and significantly reduced expression in the embryo media samples from embryo transfer cycles resulting in viable pregnancies. Western blot and ELISA, as well as quantitative MRM results, were confirmatory. These results demonstrated that peptide/protein profiles from the culture medium during early human in vitro development can discriminate embryos with highest and lowest implantation competence following uterine transfer. Further prospective studies are needed to establish validated thresholds for clinical application.

A novel model of human implantation: 3D endometrium-like culture system to study attachment of human trophoblast (Jar) cell spheroids.

There is an urgent need to develop optimized experimental models to examine human implantation. These studies aimed to (i) establish a human endometrium-like three-dimensional (3D) culture system, and (ii) examine the attachment of trophoblast-like Jar spheroids to the culture. In the present work, 3D endometrial cultures were constructed with fibrin-agarose as matrix scaffold, and using epithelial and stromal cells from both human primary cultures and established cell lines. An attachment assay between trophoblast cells and the 3D culture was developed. Epithelial cells (cytokeratin(+)) concentrated on top of the matrix forming a monolayer, and stromal cells (vimentin(+)) resided within the matrix, resembling the normal endometrial structure. The capability of primary epithelial cells to form glands spontaneously was observed. Human trophoblast cells (Jar cells) were hCG(+) by immunostaining, allowed to form spheroids, and confirmed to secrete hCG into the medium. Time-dependent experiments demonstrated a high rate of attachment of Jar spheroids to the epithelium, and adhesion was strongly related to the various cell types present in the 3D culture. An architecturally and functionally competent 3D endometrial culture system was established, that coupled with Jar spheroids mimicking trophoblast cells, provides a unique in vitro model for the study of certain aspects of human implantation.

A study of the cost, accuracy, and benefits of 3-dimensional sonography compared with hysterosalpingography in women with uterine abnormalities.

OBJECTIVES: We conducted a prospective blinded study to evaluate the costs, accuracy, risks, and benefits of 3-dimensional (3D) transvaginal sonography compared to hysterosalpingography. METHODS: A total of 101 women aged 26 to 44 years with evidence of uterine anomalies were enrolled. All participants had routine hysterosalpingography as part of their infertility evaluation as well as 3D transvaginal sonography as part of the study. Surgical findings were used as the standard for final diagnosis. RESULTS: A total of 6 normal uteri and 119 uterine anomalies were classified: 30 congenital uterine anomalies (3 arcuate, 1 unicornuate, 4 bicornuate, 2 didelphys, and 20 septate uteri) and 89 acquired anomalies (38 polyps, 30 fibroids, 17 synechiae, and 4 adenomyosis). Congenital anomalies were correctly identified in 30 of 30 cases by 3D sonography but from 10 to 30 of 30 cases by hysterosalpingography. The detection rates for acquired uterine anomalies were lower for both techniques: 44 to 89 of 89 cases for 3D sonography and 22 to 74 of 89 cases for hysterosalpingography. Only 7 of the 20 septi would have been surgically corrected if patients only had hysterosalpingography. On the contrary, 30 of 30 patients with congenital uterine anomalies, 2 of 4 patients with adenomyosis, and all 6 patients with normal uteri were spared from surgery with diagnoses by 3D sonography. No adverse effects were reported after sonography, and only 6 minor ones were reported after hysterosalpingography. CONCLUSIONS: Three-dimensional transvaginal sonography provides visualization and evaluation of the uterine cavity with similar or better accuracy than standard hysterosalpingography in the office setting, with lower cost and morbidity.

Does the degree of hypothalamic-pituitary-ovarian recovery after oral contraceptive pills affect outcomes of IVF/ICSI cycles receiving GnRH-antagonist adjuvant therapy in women over 35 years of age?

PURPOSE: To evaluate if the degree of recovery of serum gonadotropins after oral contraceptive pills (OCP) pretreatment has an impact on ovarian response in GnRH-antagonist IVF cycles in women of advanced maternal age. METHODS: In this retrospective cohort study, we included 98 women 35-42 years undergoing their first IVF cycle receiving gonadotropins and a fixed GnRH-antagonist adjuvant protocol. Data analysis was carried out according to changes in serum FSH, LH and estradiol (E(2)) levels (basal and post-OCP) divided in quartiles, and also according to absolute levels. The main outcomes were peak serum E(2), number of mature oocytes retrieved, length of stimulation, and amount of gonadotropins used. RESULTS: By quartile analysis, patients with the highest levels of serum gonadotropins suppression and also patients with gonadotropin rebound needed larger amounts of LH during the treatment. On the other hand, women with absolute suppression of FSH/LH had increased length of stimulation. CONCLUSIONS: The results of this study provide data that assist in clinical management. Gonadotropin serum levels after OCP treatment provide information for optimization of supplementation with LH in GnRH-antagonist cycles in women over age 35.

Expression of milk fat globule EGF-factor 8 (MFG-E8) mRNA and protein in the human endometrium and its regulation by prolactin.

Milk fat globule epidermal growth-factor 8 (MFG-E8) has not been previously linked to endometrial physiology. We reported on MFG-E8 mRNA up-regulation in the human endometrium during the window of implantation (WOI) using microarrays. Prolactin (PRL) secreted by stromal cells has been suggested to modulate protein expression. The objective of this study was to characterize the endometrial expression of MFG-E8 and its ligand αvß3 integrin during the menstrual cycle and its possible regulation by PRL. MFG-E8 mRNA (real-time RT-PCR) and protein expression (immunohistochemistry and immunoblotting) were analyzed in human endometrial biopsies at different times of the menstrual cycle, as well as in primary endometrial cell cultures. In primary cultures of epithelial cells, MFG-E8 intracellular protein expression was evaluated in absence or presence of PRL (0.2 and 1 µg/ml). The results show that MFG-E8 protein is almost exclusively localized to the epithelium in whole endometrial biopsies. Both MFG-E8 mRNA and protein expression increased in the luteal phase and were highest during the WOI; epithelial protein location of αvß3 integrin also peaked on cycle Day 24. Cultured epithelial cells showed a diffuse staining of MFG-E8 over the cytoplasmic area; however, some cells presented a punctuated staining pattern. PRL treatment of epithelial cells for 72 h in vitro significantly increased MFG-E8 protein intracellular expression. This is the first report on MFG-E8 protein localization to the human endometrial epithelium and its up-regulation during the WOI. The pattern of glandular expression of its ligand αvß3 integrin was remarkably similar. In vitro data support a modulatory role for PRL as a stromal/epithelial paracrine factor controlling MFG-E8.

Assisted reproduction for the validation of gestational age assessment methods.

Despite major achievements in medicine, preterm birth (PTB) remains a leading cause of infant morbidity and mortality, worldwide. Research efforts have been devoted towards a better understanding of the multifactorial aetiology of PTB and its subtypes, with the purpose of prevention and control. The availability of valid and reliable gestational age data is a prerequisite for PTB classification. Pregnancies conceived through assisted reproduction treatments provide an opportunity for the exact determination of gestational age using date of delivery and dates of fertilization or implantation. The purpose of this review article is to evaluate the current evidence for or against the various methods that can be applied to measure gestational age, namely the first day of the last menstrual period, ultrasound before 20 weeks of gestation and post-natal assessments, and to propose the use of assisted reproduction treatments populations for further validation of these methods.

Gonadotrophin ovulation induction and enhancement outcomes: analysis of more than 1400 cycles.

Ovulation induction (OI) or ovulation enhancement (OE) with gonadotrophins can be a reasonable treatment option for patients with a variety of infertility diagnoses. It must be used with extensive monitoring and management given the risk of multiple pregnancy,especially high-order multiples. This retrospective study evaluated per cycle outcomes of a large cohort of 1452 gonadotrophin OI/OE cycles at an academic infertility centre, and the efficacy of specific guidelines in limiting multiple pregnancy. The lowest possible gonadotrophin doses were used and cycle cancellation was recommended if more than three dominant follicles were present, and/or ifserum oestradiol was above 1500 pg/ml. Overall, pregnancy rate (PR) was 12% and live birth rate was 7.7%, with an increasing trend in younger patients (P = 0.0002 and <0.0001, respectively). Multiple clinical PR was 2.6% with 1.9% twins and 0.7% triplets and above.The birthweight of a singleton from a vanishing twin pregnancy (n = 8)was significantly lower than other singletons (2882 g versus 3250 g,P = 0.013). Reducing multiple pregnancies from OI/OE cycles remains an important and challenging goal. In this large cohort, high-order multiple clinical PR was limited to 0.7% per cycle by using specific management strategies while maintaining a reasonable PR.

Successful extracorporeal mature oocyte harvesting after laparoscopic oophorectomy following controlled ovarian hyperstimulation for the purpose of fertility preservation in a patient with borderline ovarian tumor.

PURPOSE: To report the successful extracorporeal recovery of mature oocytes after laparoscopic oophorectomy following ovarian hyperstimulation for the purpose of fertility preservation in a patient with recurrent serous borderline ovarian tumor. METHODS: A 25-year-old nulligravida woman presented with recurrence of a borderline serous adenocarcinoma in the right ovary after been treated conservatively with left oophorectomy for the same. RESULT(S): The patient underwent ovarian stimulation followed by a laparoscopic oophorectomy and ex-vivo retrieval of oocytes. Twenty two oocytes were recovered: fourteen metaphases II, two metaphases I, five prophases I and one degenerate. CONCLUSION(S): Mature oocytes were successfully retrieved ex-vivo from the hyperstimulated ovary recovered via laparoscopy. The procedure can be performed in a quick manner, with standard equipment, without damaging the ovary, the follicles or the oocytes, and without the risk of cancer cell spillage associated with the standard transvaginal oocyte retrieval if there is concern of ovarian surface/peritoneal metastatic disease.

Do GnRH analogues directly affect human endometrial epithelial cell gene expression?

We examined whether Gonadotrophin-releasing hormone (GnRH) analogues [leuprolide acetate (LA) and ganirelix acetate (GA)] modulate gene expression in Ishikawa cells used as surrogate for human endometrial epithelial cells in vitro. The specific aims were: (i) to study the modulatory effect of GnRH analogues by RT-PCR [in the absence and presence of E(2) and P4, and cyclic adenosine monophosphate (cAMP)] on mRNA expression of genes modulated during the window of implantation in GnRH analogues/rFSH-treated assisted reproductive technology cycles including OPTINEURIN (OPTN), CHROMATIN MODIFYING PROTEIN (CHMP1A), PROSAPOSIN (PSAP), IGFBP-5 and SORTING NEXIN 7 (SNX7), and (ii) to analyze the 5'-flanking regions of such genes for the presence of putative steroid-response elements [estrogen-response elements (EREs) and P4-response element (PREs)]. Ishikawa cells were cytokeratin+/vimentin- and expressed ERalpha, ERbeta, PR and GnRH-R proteins. At 6 and 24 h, neither LA nor GA alone had an effect on gene expression. GnRH analogues alone or following E(2) and/or P4 co-incubation for 24 h also had no effect on gene expression, but P4 significantly increased expression of CHMP1A. E(2) + P4 treatment for 4 days, alone or followed by GA, had no effect, but E(2) + P4 treatment followed by LA significantly decreased IGFBP-5 expression. The addition of 8-Br cAMP did not modify gene expression, with the exception of IGFBP-5 that was significantly increased. The GnRH analogues did not modify intracellular cAMP levels. We identified conserved EREs for OPN, CHMP1A, SNX7 and PSAP and PREs for SNX7. We conclude that GnRH analogues appear not to have major direct effects on gene expression of human endometrial epithelial cells in vitro.

Two singleton live births after the transfer of cryopreserved-thawed day-3 embryos following an unstimulated in-vitro oocyte maturation cycle.

The objective was to report two singleton live births after transfer of cryopreserved-thawed day-3 embryos resulting from an unstimulated in-vitro oocyte maturation (IVM) cycle. A 29-year-old female patient with polycystic ovaries (PCO) underwent an unstimulated IVM cycle. A total of 43 prophase-I oocytes were retrieved; 21 oocytes achieved in-vitro maturation to the metaphase-II stage at 36 h post-retrieval and 18 oocytes were fertilized (two pronuclei) after intracytoplasmic sperm injection. Two embryos were transferred in the fresh cycle (no pregnancy) and 15 day-3 embryos (post-oocyte microinjection) were cryopreserved. Subsequently, the patient became pregnant after each of two embryo transfer cycles from cryopreserved-thawed embryos (three and two embryos transferred respectively), with delivery of a single, term, healthy baby after each transfer. It is concluded that healthy live births were documented in a PCO patient undergoing unstimulated IVM followed by transfer of day-3 cryopreserved (slow-freeze)-thawed embryos, adding these methodologies to the armamentarium of assisted reproductive technologies.

A descriptive study of asthma in young adults conceived by IVF.

Although asthma has been previously associated with preterm delivery and low birthweight, evidence supporting a relationship between IVF and asthma remains inconclusive. The purpose of this study was to characterize asthma experiences in the oldest IVF-conceived generation in the USA. A cross-sectional study was conducted among 173 young adults (age: 18-26 years) conceived by conventional IVF between 1981 and 1990 at a major fertility treatment centre. A self-administered questionnaire was used with standard questions adapted from the 2008 Behavioural Risk Factor Surveillance System to assess asthma characteristics. Sixteen percent of participants reported a lifetime diagnosis of asthma; nearly half of those were no longer experiencing asthma symptoms at the time of the survey. The asthma profile of young adults conceived by IVF appeared to be favourable compared with the general population of the USA. Although few statistically significant results were obtained, low birthweight infants and individuals of a multiple gestation tended to be diagnosed at a later stage and were more likely to be current asthmatics seeking healthcare services than normal-weight infants and individuals of a singleton gestation. Further studies using larger samples and more advanced designs are needed to confirm these preliminary findings.

Assessment of Birth Defects and Cancer Risk in Children Conceived via In Vitro Fertilization in the US.

Importance: Children with birth defects have a greater risk of developing cancer, but this association has not yet been evaluated in children conceived with in vitro fertilization (IVF). Objective: To assess whether the association between birth defects and cancer is greater in children conceived via IVF compared with children conceived naturally. Design, Setting, and Participants: This cohort study of live births, birth defects, and cancer from Massachusetts, New York, North Carolina, and Texas included 1 000 639 children born to fertile women and 52 776 children conceived via IVF (using autologous oocytes and fresh embryos) during 2004-2016 in Massachusetts and North Carolina, 2004-2015 in New York, and 2004-2013 in Texas. Children were followed up for an average of 5.7 years (6 008 985 total person-years of exposure). Data analysis was conducted from April 1 to August 31, 2020. Exposures: Conception by IVF for state residents who gave birth to liveborn singletons during the study period. Birth defect diagnoses recorded by statewide registries. Main Outcomes and Measures: Cancer diagnosis as recorded by state cancer registries. Cox proportional hazards regression models were used to estimate hazard ratios (HRs) and 95% CIs for birth defect-cancer associations separately in fertile and IVF groups. Results: A total of 1 000 639 children (51.3% boys; 69.7% White; and 38.3% born between 2009-2012) were in the fertile group and 52 776 were in the IVF group (51.3% boys; 81.3% White; and 39.6% born between 2009-2012). Compared with children without birth defects, cancer risks were higher among children with a major birth defect in the fertile group (hazard ratio [HR], 3.15; 95% CI, 2.40-4.14) and IVF group (HR, 6.90; 95% CI, 3.73-12.74). The HR of cancer among children with a major nonchromosomal defect was 2.07 (95% CI, 1.47-2.91) among children in the fertile group and 4.04 (95% CI, 1.86-8.77) among children in the IVF group. The HR of cancer among children with a chromosomal defect was 15.45 (95% CI, 10.00-23.86) in the fertile group and 38.91 (95% CI, 15.56-97.33) in the IVF group. Conclusions and Relevance: This study found that among children with birth defects, those conceived via IVF were at greater risk of developing cancer compared with children conceived naturally.