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Objective Dual trigger is used to induce final oocyte maturation during the process of controlled ovarian hyperstimulation, yet yielding controversial results. Also, there are yet no data regarding the effect of the dosage of the dual trigger on clinical outcomes. Based on the Patient-Oriented Strategies Encompassing IndividualizeD Oocyte Number (POSEIDON) criteria, this study aimed to determine the clinical difference of a single bolus versus two boluses of gonadotropin-releasing hormone agonist (GnRHa) in POSEIDON group IV patients using dual trigger.Methods We screened a total of 1,256 patients who underwent in vitro fertilization (IVF) cycles who met the POSEIDON group IV criteria. Six hundred and twenty-nine patients received one bolus of GnRHa, and 627 patients were given two boluses. All patients received the same dose of recombinant human chorionic gonadotropin during the dual trigger cycle.Results Metaphase II oocyte retrieval rate, fertilization rate and clinical pregnancy rate did not differ between the two groups. However, a lower percentage of at least one top-quality embryo transfer (34.3% vs. 26.0%, P=0.001) in the two bolus-GnRHa group was noted.Conclusion A double bolus of GnRHa did not show superior clinical results compared to a single bolus of GnRHa in the dual trigger IVF cycle. Therefore, GnRHa doses for use should be decided based on individual clinical situations considering cost-effectiveness and patient compliance, but further investigation will be needed.

Objective Premature ovarian insufficiency (POI) is a clinical disease that is diagnosed by the loss of ovarian function before the age of 40. Despite recent progress in molecular diagnosis, the genetic etiology of POI is not well established. The aim of this study is to reveal pathogenic genetic variants involved in POI. Study design and main outcome measures To reveal pathogenic genetic variants involved in POI, whole exome sequencing was performed in nonconsanguineous family members with POI. Constitutional variants were filtered against population databases and a missense mutation of natriuretic peptide C (NPPC) (c.131A>G, p.Q44R) was selected as a convincing candidate mutation among 14 heterozygous mutant alleles in 13 genes. Results The wild-type NPPC and mutant NPPC (NPPC131A>G) were expressed in HeLa cells, and cells expressing NPPC131A>G secreted unique peptides. The ProP 1.0 Server, a neural network prediction tool, predicted the presence of a cleavage site at the substituted arginine residue (p.Q44R) of NPPC. The molecular weight of predicted cleaved peptides processed from mutant NPPC precursor corresponded to that of the actual mutant peptide. The cGMP synthetic activity of NPR2-expressing cells was significantly decreased by interaction with the mutant NPPC peptide compared with wild-type NPPC. Conclusions The peptide generated by a rare mutation of NPPC might influence paracrine C-type natriuretic peptide (CNP)-mediated preantral follicle development and/or sustain meiotic arrest in oocytes. We therefore suggest that a mutation of the NPPC gene is involved in the pathogenesis of POI.

OBJECTIVE To evaluate the effects of the duration of cryostorage on clinical outcomes after embryo transfer of vitrified blastocysts stored in an open-device slush-nitrogen (SN2 ) system. METHODS A retrospective cohort study was carried out on 1632 autologous vitrified-warmed blastocyst transfer cycles between January 2013 and June 2014. Duration of cryostorage was divided into four groups: Group I: 0-6 months (n=937); Group II: 7-12 months (n=299); Group III: 13-24 months (n=165); and Group IV: ≥25 months (n=231). The effects of the duration of cryostorage on the survival rate (SR), clinical pregnancy rate (CPR), live birth rate (LBR), and neonatal outcomes of vitrified blastocysts stored in an open-device SN2 system were evaluated. RESULTS There were no significant differences between groups in SR, CPR, LBR, and neonatal outcomes after autologous vitrified-warmed blastocyst transfer. Multivariate logistic regression analysis showed no effect on LBR from duration of cryostorage. CONCLUSION Vitrification using SN2 and long-term cryostorage in an open-device system are safe and effective and do not significantly affect clinical outcomes after embryo transfer.

This study aimed to evaluate the effect of hyaluronan-selected/physiological intracytoplasmic sperm injection (PICSI) on fertilization and quality of cleavage-stage embryos in infertile couples with ≤1% of spermatozoa with normal strict morphology (severe teratozoospermia). Seventy-seven couples underwent PICSI between October 2017 and December 2018 (PICSI group), while 75 couples underwent conventional intracytoplasmic sperm injection (ICSI) between January 2016 and September 2017 (ICSI group). Good quality embryos (GQEs) were evaluated based on morphology. Patient and cycle characteristics were comparable between the PICSI and ICSI groups, except for age and anti-Müllerian hormone (AMH) level (38.4 ± 3.9 years vs. 36.3 ± 4.3 years, p = .002 and 2.06 ± 1.99 ng/mL vs. 2.97 ± 3.25 ng/mL, p = .040). The fertilization rate per oocyte inseminated and GQE rate were significantly higher in the PICSI group than in the ICSI group (82.7% vs. 71.7%, p ˂ .001 and 52.8% vs. 34.0%, p ˂ .001). Furthermore, the absence of GQEs was found to be lower in the PICSI group (13.0% vs. 30.7%, p = .008). Multivariate analysis adjusted for age and AMH level identified PICSI as an unfavorable and independent factor for the absence of GQEs (adjusted odds ratio, 0.333; 95% confidence interval, 0.125–0.890). PICSI seems to be superior to ICSI in terms of fertilization and embryo quality in couples with severe teratozoospermia.

Previous studies have reported that the mitochondrial DNA (mtDNA) contents of cumulus cells (CCs) in ovarian follicular fluid are correlated with embryo quality. Quantification of mtDNA CCs has been suggested as a biomarker of embryo viability. The aim of this study was to determine the relationship between mitochondrial DNA (mtDNA)/genomic DNA (gDNA) ratio in CCs and IVF outcomes such as fertilization rates and embryo quality in infertile women. This is an observational study on 144 cumulus-oocyte complexes obtained from 144 patients undergoing IVF-intracytoplasmic sperm injection (ICSI) at a single fertility center. The CCs in ovarian follicular fluid from patients undergoing IVF-ICSI were collected by ovum pick-up. A relative copy number quantification was used to determine mtDNA/gDNA ratio. Quantitative real-time PCR for various markers (β2M and mtMinArc gene) was used to determine average mtDNA/gDNA ratio of CCs. Investigation of the correlation between mtDNA/gDNA ratio in CCs and IVF outcomes showed no statistically significant correlation between the mtDNA/gDNA ratio in CCs and fertilization rates. However, mtDNA/gDNA ratio and embryo quality showed a statistically significant positive correlation. A significantly higher mtDNA/gDNA ratio was observed in the good quality embryo group compared with the poor quality embryo group (P < 0.05). In addition, the mtDNA/gDNA ratio showed negative correlation with the patient's age (correlation coefficient= -0.228, P < 0.05). Results of this study demonstrate a negative correlation of mtDNA/gDNA ratio in CCs with patient's age, and a low copy number of mtDNA in CCs may have adverse effects on embryo quality in IVF cycles. These results suggest that the ratio of mtDNA/gDNA in CCs may serve as a biomarker in predicting IVF outcomes.

The process of selecting a good quality embryo to improve the pregnancy outcomes is very important. The aim of our study was to elaborate the embryo selection process in a single vitrified-warmed blastocyst transfer (VBT) cycle by analyzing pre-vitrified and post-warmed blastocyst morphological factors to improve pregnancy outcomes. In this retrospective cohort study, we performed 329 single VBT cycles. The pre-vitrified and post-warmed morphological factors of all blastocysts were analyzed. Logistic regression analysis was conducted to select the independent morphological factor associated with ongoing pregnancy. The expansion of blastocoel (mid blastocoel; aOR 2.27, 95% CI.0.80–6.42, p = 0.12, expanded blastocoel; aOR 3.15, 95% CI.1.18–8.44, p = 0.02) in a pre-vitrified blastocyst and the grade of inner cell mass (ICM) (grade B; aOR 0.47, 95% CI.0.27–0.83, p = 0.01, grade C; aOR 0.22, 95% CI 0.09–0.56 p

Aim Antioxidants have been studied to be effective in improving embryo qualities from in vitro fertilization. L-carnitine (LC) has been known to reduce reactive oxygen species and enhance adenosine triphosphate production, which contribute to the development of a high-quality embryo. This is the first study to include both mouse and human subjects and aimed to evaluate whether LC supplementation in culture media has any beneficial effect on the development of the embryos, as well as its clinical outcomes. Methods Mouse embryos were used as models in the animal studies for cell immunofluorescent staining evaluation. Inner cell mass and trophectoderm (TE) cells were counted and statistically analyzed between LC and control groups. For human studies, medical records of patients with infertility undergoing in vitro fertilization procedures from January to May 2017 were included and the embryos were divided into two groups at the two pronuclear stage. Statistical analysis was performed to compare the embryo status and clinical outcomes of the two groups. Results In the animal study, the LC group showed significantly higher numbers of cells in the inner cell mass and trophectoderm, indicating better development. In the human studies, there were significantly higher numbers of good-quality embryos on days 2, 3 and 5 in the LC group than in the control. The clinical outcomes, such as implantation, clinical pregnancy and ongoing pregnancy rates, were also higher in the LC group than in the control. Conclusion LC supplementation in culture media improved human embryo quality and eventually achieved better pregnancy outcomes.

To evaluate clinical utility of antral follicle count (AFC) and anti-Müllerian hormone (AMH) in predicting in vitro fertilization (IVF) outcomes among the patients over 40 years old in their first IVF cycles.Total 219 patients aged 40 or older who underwent their first IVF with gonadotropin-releasing hormone antagonist protocol from January 2013 to September 2014 in CHA Gangnam fertility center were retrospectively analyzed. AFC and serum samples were measured prior to IVF treatment. The main outcomes were clinical pregnancy rate and live birth.36 out of 219 patients achieved clinical pregnancy (16.4%) and 27 out of 219 patients delivered (12.3%). The receiver operating characteristic curve analysis to predict clinical pregnancy showed that both age and AFC equally had higher accuracy by area under the curve (AUC = 0.657, P 0.01) than serum AMH (AUC 0.613, P = 0.03). The optimum cut-off value of age was ≤41 and that of AFC was3 to predict clinical pregnancy. For the prediction of live birth, AFC had the highest accuracy (AUC 0.698, P 0.01), followed by age (AUC 0.674, P 0.01) and the number of total retrieved oocytes (AUC 0.620, P = 0.02). The optimum cut-off value of age was ≤41, that of AFC was3 and that of the number of total retrieved oocytes were6. With multivariate regression analysis, age and AMH were significantly correlated with clinical pregnancy (age, odds ratio [OR] 0.53, P 0.01; AMH, OR 1.31, P = 0.04), whereas age and AFC were association with live birth significantly (age, OR 0.41, P 0.01; AFC, OR 1.10, P = 0.02).In patients aged over 40, AFC and AMH were shown to be good biomarkers for the prediction of clinical pregnancy and live birth. Although AMH was positively correlated with clinical pregnancy and had no association with live birth, the predictive value of AFC and AMH were similar for both clinical pregnancy and live birth. To predict the live birth, age ≤41, AFC3 and total retrieved oocytes6 appeared to be meaningful. This study demonstrated the significance of AMH and AFC as predictors of clinical pregnancy and live birth for old aged women at their first IVF cycle with gonadotropin-releasing hormone antagonist protocol.

Supplementation of growth hormone (GH) during controlled ovarian stimulation (COS) has been suggested to improve ovarian response. Despite potential benefits in poor responders, multiple injections of GH during COS are inconvenient. We conducted a randomized controlled study to evaluate the efficacy and safety of sustained-release human GH in poor responders undergoing in vitro fertilization (IVF). This was a single-center, randomized, open-label, parallel study. Infertile women who satisfied the Bologna criteria for poor responders were randomized into GH treatment and control groups. The treatment group received a sustained-release GH (Eutropin Plus® 20 mg) three times before and during COS (mid-luteal, late luteal, and menstrual cycle day 2). The baseline characteristics and IVF outcomes were compared between the two groups. A total of 127 patients were included in the analysis. The mean age was 39.6 years and mean anti-Mullerian hormone level was 0.6 ng/ml. There was no significant difference in the baseline characteristics between GH treatment and control groups. The number of follicles on the human chorionic gonadotropin triggering day (3.1 ± 2.3 vs. 2.4 ± 1.6, P = 0.043) and the proportion of metaphase II oocytes (67.5 vs. 52.3%, P = 0.030) were higher in the GH group than in controls. The percentage of clinical and ongoing pregnancy and miscarriage was not different between the two groups. Supplementation of sustained-release GH before and during COS improved ovarian response, with an increase in mature oocytes in poor responders. Further studies are needed to ensure this benefit in general infertility patients.

Purpose: Subfertility associated with polycystic ovary syndrome (PCOS) mainly originates from oligoovulation/anovulation. Although insulin resistance and androgen excess are known to cause PCOS-associated implantation failure, the consequences of PCOS on endometrial homeostasis and pathophysiology have not been comprehensively understood. In this study, we examined whether the pathophysiologic milieu of PCOS intrinsically affects expression profiles of genes related to insulin signaling and facilitative glucose transporters (GLUTs) in the human endometrium and/or during in vitro decidualization. Study Design: Seven healthy women with regular menstrual cycles and 13 patients with PCOS were recruited for this study. To mimic the hyperandrogenic or hyperinsulinemic milieu in the endometrium of patient with PCOS (PCOSE) in vitro, human endometrial stromal cells (hESCs) were treated with dihydrotestosterone (DHT) or insulin, respectively. Results: In PCOSE, messenger RNA (mRNA) levels of insulin receptor (IR), IR substrate (IRS) 1, and IRS2 were significantly increased. Furthermore, GLUT1 and GLUT12 were aberrantly increased. Chronic exposure to insulin or DHT aberrantly increased IRS1/IRS2 phosphorylation and protein levels of GLUT1 and GLUT12 in hESCs, suggesting that not only hyperinsulinemic but also hyperandrogenic conditions affect insulin signaling and glucose metabolism. The mRNA microarrays demonstrated that DHT dysregulates various gene sets, including cell cycle and glucose metabolism, in hESCs. Furthermore, DHT suppressed the expression of GLUT1 and GLUT12 as well as decidualization markers, IGFBP1 and prolactin, during in vitro decidualization. Conclusions: The hyperandrogenic milieu affects gene expression profiles, including gene sets associated with insulin signaling, cell cycle, glucose metabolism, and/or glucose transport, in human endometrium and during in vitro decidualization.

Basal luteinizing hormone (LH) levels have also been suggested to impact on ovarian responsiveness as well as basal follicular stimulating hormone (FSH) levels. The aim of this study was to compare the in vitro fertilization (IVF) outcomes according to cycle day 3 FSH/LH ratio and to assess the proper stimulation protocol between gonadotropin-releasing hormone (GnRH) agonist and GnRH antagonist protocols. The retrospective cohort study recruited a total of 1211 women having the laboratory values of FSH (

Introduction Miscarriage is the most common placental-related complication of pregnancy. It has been extensively investigated to discover the underlying mechanism(s) by which miscarriage occurs, but in many cases the etiology still remains unclear. The aim of this study was to analyze genome-wide expression profiles of placental villi (PV) from unexplained miscarriage with a pathway-oriented method for identifying underlying mechanism(s) of unexplained miscarriage. Methods We investigated PV of 18 women with unexplained miscarriage and 11 women underwent normal pregnancy. Each PV was obtained through dilatation & evacuation and chorionic villous sampling, respectively. Genome-wide expression profiles of PV were analyzed by Gene Set Enrichment Analysis (GSEA) to find dysregulated signaling pathways in PV of unexplained miscarriage. Results Unsupervised hierarchical clustering showed heterogeneity of expression profiles between PV of normal developing pregnancy and unexplained miscarriage. GSEA, a supervised analysis, with KEGG pathways revealed that several gene sets associated with mitochondrial function including glutathione metabolism and oxidative phosphorylation are dysregulated in PV from unexplained miscarriage. RT-PCR, real-time RT-PCR and/or immunohistochemistry reinforced that expression of genes constituting these gene sets enriched in normal pregnancy and Cu/Zn-superoxide dismutase was down-regulated in PV of unexplained miscarriage. Discussion Structural vulnerability of placental villi for reactive oxygen species (ROS), which is caused by systemic down-regulation of mitochondrial pathways involved in mitochondrial redox balance and functions, aggravates oxidative stress with increased ROS production in PV of unexplained miscarriage. Conclusion Systemic vulnerability for ROS in PV could be a major cause of unexplained miscarriage.

To compare the IVF outcomes of vitrification-thawed blastocyst transfer cycles utilizing different endometrial preparation methods.We retrospectively assessed IVF outcomes in 611 patients (648 cycles) who underwent blastocyst frozen embryo transfer (FET) between January 2007 and December 2009. All embryos had been cryopreserved by a vitrification method following a previous IVF cycle. Patients were prepared for transfer by using either the natural cycle (n = 310/Group 1), the natural cycle with ovulation induction employing human chorionic gonadotropin (n = 134/Group 2), or a hormonally manipulated artificial cycle with estrogen and progesterone supplementation (n = 204/Group 3).Multivariate logistic regression analysis showed a significant difference in clinical pregnancy rate between Groups 3 (30.4%) and 1 (41.9%) (odds ratio [OR], 0.567; 95% confidence interval [CI], 0.379-0.847, P = 0.006) whereas the difference between Groups 2 and 1 was not significant (41.8% vs. 41.9%; OR, 0.683; 95% CI, 0.435-1.073; P = 0.098). Other significant variables affecting clinical pregnancy rate were the number of embryos transferred, the grade of transferred embryos, and maximal endometrial thickness.The results showed that, using vitrification-thawed blastocyst transfer, employment of natural cycles with or without hCG treatment was associated with better outcomes than was the use of hormonally manipulated cycles.

The present study aims to evaluate whether multiplex ligation-dependent probe amplification (MLPA) technique with subtelomeric probes is to be an alternative method of routine G-banding chromosome analysis from pregnancy loss. A review of 5 years (from 2005 to 2009) of karyotype for products of conception (POCs) was carried out. From June 2010 to June 2012, MLPA was performed in parallel with karyotype analysis on 347 miscarriages. Karyotyped miscarriages served as controls in this blinded study. Abnormal results were confirmed by fluorescence in situ hybridization. A review of 5 years of karyotype results for POCs indicated that 11.46 % of cases failed to karyotyping. In the study periods, MLPA results were successfully obtained from all cases including 51 (14.7 %) culture failed cases, chromosomal abnormalities were detected in 27 (52.9 %) of cases which failed to grow or could not be cultivated. It took 3 weeks by conventional karyotyping, but it required at least 24 h and at most a week by MLPA from tissue sampling to final reporting. 47 cases showed discordant results between karyotyping and MLPA because of maternal cell contamination, polyploidy, mosaicism, or balanced translocation. MLPA technique is relatively low cost, less labor intensive and reduces waiting time with high accuracy compared with conventional cytogenetic analysis. Therefore, MLPA can be the first approach for chromosome analysis from pregnancy loss.

Objective To evaluate whether endometrial and subendometrial blood flow parameters measured using three-dimensional power Doppler ultrasound (3D PD-US) can predict pregnancy after IUI. Design Prospective clinical study. Setting Infertility center in a referral hospital. Patient(s) One hundred six women who underwent ovulation induction and IUI. Intervention(s) A color Doppler ultrasound and a 3D PD-US examination were performed on the day of IUI. Main Outcome Measure(s) Pulsatility index (PI), resistance index (RI), and systolic/diastolic (S/D) ratio of uterine artery, and vascularization index (VI), flow index (FI), and vascularization flow index (VFI) of the endometrium as well as those of subendometrial region. These measurements were analyzed in relation to IUI outcome (pregnant vs. nonpregnant). Result(s) The pregnant group had higher endometrium VI, FI, and VFI scores than the nonpregnant group. In contrast, the subendometrial region VI, FI, and VFI scores did not differ between the groups, nor did the uterine artery PI, RI, and S/D. Pregnancies did not occur when endometrial blood flow had not been detected. Conclusion(s) Three-dimensional PD-US was useful for evaluating endometrial and subendometrial neovascularization in IUI cycles. Endometrial blood flow parameters may be useful predictors for pregnancy.

OBJECTIVE: The Rotterdam definition for polycystic ovary syndrome (PCOS) states two out of three criteria must be met. However the clinical significance and management of isolated polycystic ovary (PCO) is still unclear. The aim of our study is to investigate the clinical significance of isolated PCO in assisted reproduction by comparing clinical and laboratory aspects of patients with morphologically normal ovaries (Nov), isolated PCO, and polycystic ovary syndrome (PCOS). DESIGN: This is a retrospective medical record review study. MATERIALS AND METHODS: Medical records of 444 patients that visited our infertility clinic between 2011 and 2013 were retrospectively reviewed. Patients in the three groups, isolated PCO, PCOS, and Nov, were subdivided into subgroups according to age, previous history of fertility or infertility. Cumulative embryo score (CES) and the number of good quality embryos were used to assess embryo quality. RESULTS: The number of oocytes retrieved was significantly higher in PCO group compared to Nov group (12.94 11.1 vs. 8.03 7.4, respectively, p