The composition dynamics of transposable elements in human blastocysts.

Transposable elements (TEs) are mobile DNA sequences that can replicate themselves and play significant roles in embryo development and chromosomal structure remodeling. In this study, we investigated the variation of TEs in blastocysts with different parental genetic backgrounds. We analyzed the proportions of 1137 TEs subfamilies from six classes at the DNA level using Bowtie2 and PopoolationTE2 in 196 blastocysts with abnormal parental chromosomal diseases. Our findings revealed that the parental karyotype was the dominant factor influencing TEs frequencies. Out of the 1116 subfamilies, different frequencies were observed in blastocysts with varying parental karyotypes. The development stage of blastocysts was the second most crucial factor influencing TEs proportions. A total of 614 subfamilies exhibited different proportions at distinct blastocyst stages. Notably, subfamily members belonging to the Alu family showed a high proportion at stage 6, while those from the LINE class exhibited a high proportion at stage 3 and a low proportion at stage 6. Moreover, the proportions of some TEs subfamilies also varied depending on blastocyst karyotype, inner cell mass status, and outer trophectoderm status. We found that 48 subfamilies displayed different proportions between balanced and unbalanced blastocysts. Additionally, 19 subfamilies demonstrated varying proportions among different inner cell mass scores, and 43 subfamilies exhibited different proportions among outer trophectoderm scores. This study suggests that the composition of TEs subfamilies may be influenced by various factors and undergoes dynamic modulation during embryo development.

The effects of different endometrial preparation regimens on pregnancy outcomes in frozen-thawed embryo transfer cycles: a prospective randomized controlled study.

OBJECTIVE: An increasing number of research have emerged to compare the pregnancy outcomes between the natural cycle and the hormone replacement therapy (HRT) cycle in preparing the endometrium for frozen-thawed embryo transfer (FET), but the results are controversial. This prospective randomized controlled study was hence designed to obtain more solid evidence. MATERIALS AND METHODS: In this study, patients with regular menstrual cycle length (21-35 days) who underwent FET between January 2010 to December 2017 were recruited for this study. Upon further filtering with the selection criteria of patients being, a total of 405 patients were recruited and randomized. Finally, analysis was performed on 384 patients: 178 belonged to the natural cycle group whereas the remaining 206 were in the HRT group. The primary outcome was live birth rate, while the secondary outcomes were implantation rate, clinical pregnancy rate, early miscarriage rate, late miscarriage rate, multiple birth rate and low birth weight rate. RESULTS: The live birth rate (37.6% vs 30.1%, p = 0.119) of natural cycle group were higher than those of the hormone replacement therapy group, although the difference was not significant. The secondary outcomes were not found to differ significantly between the two groups. Nonetheless, the endometrium was found to be thicker in the natural cycle group (10.75 mm) than the HRT group (9.00 mm) (p < 0.001). CONCLUSION: No significant differences were observed between the pregnancy outcomes of the natural cycle group and the HRT group which comprised of patients with regular menstrual cycle length.

Increased ammonium in culture medium may promote cellular apoptosis and negatively affect pluripotency of human blastocysts.

PURPOSE: To determine the association between ammonium concentration in culture medium and blastocyst development and to assess the influence of increased ammonium concentration on the expression of Bax, Bcl-2 and Oct4. METHODS: A total of 254 cleavage-stage embryos were individually cultured in 30µL G2-plus medium on Day 3, and then culture media samples were collected on Day 5 for ammonium concentration determination immediately after evaluating the embryos morphology. Poor-quality blastocysts (combined score of CC) were used for gene expression analysis. The blastocyst formation rate, good-quality blastocyst rate and relative expression levels of Bax, Bcl-2 and Oct4 were analyzed. RESULTS: Based on receiver operating characteristic curve, the cutoff value of ammonium concentration produced by embryos was 16.07 µmol/L (AUC = 0.722, 95% CI 0.637-0.807; P = 0.000), so all embryos were assigned to two groups according to the cutoff value: normal group (< 16.07 µmol/L) and increased group (≥ 16.07 µmol/L). There was a significant difference in blastocyst formation rate (80.5% vs 59.0%, P < 0.01) between normal group and increased group, as well as for good-quality blastocyst rate (21.0% vs 3.4%, P < 0.01). A significantly higher expression level of Bax (P < 0.05) and considerably lower expression level of Oct4 (P < 0.01) were observed in increased group compared to normal group. CONCLUSION: Our data demonstrated for the first time that increased ammonium concentration in culture medium may promote cellular apoptosis and negatively affect pluripotency of human blastocyst.

Machine Learning Electrocardiogram for Mobile Cardiac Pattern Extraction.

BACKGROUND: Internet-of-things technologies are reshaping healthcare applications. We take a special interest in long-term, out-of-clinic, electrocardiogram (ECG)-based heart health management and propose a machine learning framework to extract crucial patterns from noisy mobile ECG signals. METHODS: A three-stage hybrid machine learning framework is proposed for estimating heart-disease-related ECG QRS duration. First, raw heartbeats are recognized from the mobile ECG using a support vector machine (SVM). Then, the QRS boundaries are located using a novel pattern recognition approach, multiview dynamic time warping (MV-DTW). To enhance robustness with motion artifacts in the signal, the MV-DTW path distance is also used to quantize heartbeat-specific distortion conditions. Finally, a regression model is trained to transform the mobile ECG QRS duration into the commonly used standard chest ECG QRS durations. RESULTS: With the proposed framework, the performance of ECG QRS duration estimation is very encouraging, and the correlation coefficient, mean error/standard deviation, mean absolute error, and root mean absolute error are 91.2%, 0.4 ± 2.6, 1.7, and 2.6 ms, respectively, compared with the traditional chest ECG-based measurements. CONCLUSIONS: Promising experimental results are demonstrated to indicate the effectiveness of the framework. This study will greatly advance machine-learning-enabled ECG data mining towards smart medical decision support.

Comparative clinical outcome following individualized follitropin delta dosing in Chinese women undergoing ovarian stimulation for in vitro fertilization /intracytoplasmic sperm injection.

BACKGROUND: To compare the efficacy and safety of follitropin delta in its individualized fixed-dose regimen with follitropin alfa in a conventional adjustable dosing regimen in Chinese women.  METHODS: This was a subgroup analysis of the randomized, multi-center, assessor-blind, non-inferiority trial (GRAPE) including 759 Chinese women (aged 20-40 years) recruited in 16 reproductive medicine clinics in China. Women were randomized in a 1:1 ratio to be treated with either follitropin delta dose based on anti-Müllerian hormone (AMH) and body weight or conventional dosing with follitropin alfa following a gonadotropin-releasing hormone (GnRH) antagonist protocol. The primary outcome was ongoing pregnancy rate assessed 10-11 weeks after embryo transfer in the fresh cycle (non-inferiority margin -10.0%). RESULTS: 378 in the follitropin delta group and 381 in the follitropin alfa group were randomized and exposed. Non-inferiority was confirmed with respect to ongoing pregnancy with rates of 31.0% vs. 25.7% for follitropin delta compared to follitropin alfa, estimated mean difference of 5.1% (95% confidence interval (CI) -1.3% to 11.5%). The clinical pregnancy rate (35.4% vs. 31.5%, P = 0.239) and live birth rate (31.0% vs. 25.5%, P = 0.101) were comparable between the follitropin delta group and the follitropin alfa group. Overall, the individualized follitropin delta treatment resulted in fewer oocytes retrieved compared to follitropin alfa treatment (10.3 ± 6.2 vs. 12.5 ± 7.5, P < 0.001), which was mainly due to fewer oocytes (10.5 ± 6.4 vs. 13.9 ± 7.8) in women with AMH ≥ 15 pmol/L. Accordingly there was a lower incidence of early ovarian hyper-stimulation syndrome (OHSS) and/or preventive interventions (6.1% vs. 11.0%, P = 0.013). A daily follitropin delta dose of 10.2 µg (95% CI: 9.3-11.2 µg) was estimated to provide the same number of oocytes retrieved as a starting dose of 150 IU/d of follitropin alfa. CONCLUSION: Follitropin delta in its individualized fixed-dose regimen showed similar efficacy and improved safety compared with follitropin alfa in a conventional adjustable dosing regimen in Chinese women. CLINICAL TRIAL REGISTRATION NUMBER: NCT03296527.

Long-term GnRH agonist pretreatment before frozen embryo transfer improves pregnancy outcomes in women with adenomyosis.

RESEARCH QUESTION: Do frozen embryo transfer (FET) cycles following long-term gonadotrophin-releasing hormone agonist (GnRHa) pretreatment have better pregnancy outcomes than fresh embryo transfer cycles with long or ultra-long GnRHa protocol in these patients? DESIGN: This study included 537 women with adenomyosis divided into three groups: (Group A) FET cycles following long-term GnRHa pretreatment (192 patients); (Group B) fresh embryo transfer cycles with the ultra-long GnRHa protocol (241 patients); (Group C) fresh embryo transfer cycles with the long GnRHa protocol (104 patients). RESULTS: The total gonadotrophin dose and stimulation duration were significantly lower in Group A than in Groups B and C. The implantation and live birth rates were significantly higher in Group A than in Groups B and C. In the long-term GnRHa pretreatment and FET treatment of Group A, implantation (odds ratio [OR] 1.729, 95% confidence interval [CI] 1.073-2.788, P = 0.025), clinical pregnancy (OR 1.665, 95% CI 1.032-2.686, P = 0.037) and live birth rates (OR 1.694, 95% CI 1.045-2.746, P = 0.033) increased and miscarriage rate (OR 0.203, 95% CI 0.078-0.530, P = 0.001) decreased when compared with Group C. Comparison of Groups A and B showed that with the long-term GnRHa pretreatment, FET was a protective factor for live birth rate (OR 1.350, 95% CI 1.017-1.792, P = 0.038). CONCLUSION: FET following long-term GnRHa pretreatment has a better IVF/intracytoplasmic sperm injection outcome, and a potential benefit in terms of a lower gonadotrophin dose, and a shorter stimulation duration than fresh embryo transfer combined with a long or ultra-long GnRHa protocol.

The relationship of total progressive motile sperm count with the outcome of IUI? An analysis of 5171 cycles.

Background: The role of motile sperm count in intrauterine insemination (IUI) success rate is controversial. This retrospective cohort study performed among unselected infertile couples undergoing IUI was to explore the association between the total progressive motile sperm count (TPMSC) and the live birth rate (LBR) following IUI.Methods: The total cohort of 5363 cycles, 2666 infertile couples between January 2015 and December 2018 and finally 5171 cycles, 2647 couples were included for analysis in Sun Yat-sen memorial hospital of Sun Yat-sen University. The primary outcome was LBR per cycle. And the secondary outcome measure was clinical pregnancy rate (CPR) per cycle.Results: From the receiver operating characteristic (ROC) analysis of female age predicting live birth, female age cutoff was defined as 28 years. With a female age of ≤28 years, the CPRs were 11.5%, 14.9%, 16.1%, and 15.8% in quartile groups of pre-wash TPMSC, respectively. For the LBRs the values were 9.4%, 12.9%, 14.4%, and 11.3%, and there were also no significant differences in quartile groups of pre-wash TPMSC with ≤24 million (M), [24M-50M], [50M-97M], >97M. No statistically significant differences in the CPRs (p = .051) and LBRs (p = .088) were also observed in the quartiles groups of post-wash TPMSC. With a female age of >28 years, the CPR in couples with post-wash TPMSC ≤22.32 M was significantly lower than with post-wash TPMSC >81.0 M (p = .007). There was an obvious trend in which CPRs and LBRs increased with the post-wash TPMSC during the <81 M interval in women >28 years.Conclusions: The optimal female age cutoff for live birth was 28 years in IUI cycles. Pre-wash and post-wash TPMSC were not significantly associated with CPR and LBR per cycle. When female age >28 years, there was a better outcome with post-wash TPMSC >22.32 million.

Microinjection manipulation decreases the expression of GABA-A receptor signaling pathway genes in mouse embryos derived using intracytoplasmic sperm injection.

BACKGROUND: The GABA-A receptor signaling pathway regulates proliferation, differentiation, apoptosis, and responses to overt DNA damage during embryonic development. METHODS: To analyze the gene expression after intracytoplasmic sperm injection (ICSI) and in in vivo mouse embryos, the global pattern of gene expression dataset, GSE23009, was obtained from the Gene Expression Omnibus database. Genes with differential expression were identified using the R software package, and RT-qPCR was performed to confirm the microarray results. RESULTS: Mouse blastocysts derived from ICSI fertilization had decreased expression of GABA-A receptor signaling pathway genes. However, the mechanisms underlying these changes were not elucidated. The gene expression of the GABA-A pathway was not significantly different between blastocysts obtained from IVF and in vivo fertilization. However, microinjection after IVF significantly reduced the expression of the GABA-A pathway gene to levels similar to those in the ICSI group. CONCLUSION: Based on our results, decreased gene expression is a result of the microinjection manipulation performed during ICSI.

Evaluation of chromosomal abnormalities from preimplantation genetic testing to the reproductive outcomes: a comparison between three different structural rearrangements based on next-generation sequencing.

PURPOSE: The aim of this study was to determine factors affecting the chromosome imbalance in blastocysts and reproductive outcomes by a comparison between the reciprocal translocation (REC), inversion (INV), and Robertsonian translocation (ROB) carriers. METHODS: Couples with one partner carrying translocation or inversion underwent preimplantation genetic testing for chromosomal structural rearrangement (PGT-SR) cycles, including 215 PGT-SR cycles performed in subsequent 164 frozen-thawed embryo transfer cycles and 61 prenatal diagnoses of fetuses and 59 normal live birth babies. A total of 899 samples were processed by whole-genome amplification followed by next-generation sequencing (NGS). Karyotype and chromosome microarray analyses were used to confirm the PGT results from the amniotic fluid samples. RESULTS: A total of 843 blastocysts from 124 REC, 21 INV, and 35 ROB carriers were diagnosed by PGT-SR. The percentage of unbalanced blastocysts was significantly higher in REC than in INV and ROB carriers (64.31% vs. 28.05% vs. 37.02%). Stratification analysis of female carrier age and gonadotropin doses showed no significant increase in unbalanced chromosomal abnormalities in the three groups. Also, the different breakpoints in chromosomal arms did not affect the rate of unbalanced chromosomes in the embryos. Logistic regression indicated blastocyst quality as a statistically significant risk factor associated with unbalanced chromosomal abnormalities from translocation carriers (P < 0.001). The source of abnormalities in the three groups showed significant differences such that the abnormalities in REC mostly originated from parental translocation but the abnormalities in INV were mainly de novo variations. 164 blastocysts were transferred, and there were no significant differences in the clinical pregnancy rate and miscarriage rate. A total of 59 healthy babies were born, and there were no significant differences in the gender ratio and birth height, except the birth weight of boys between INV and ROB groups (P = 0.02). The results of amniocentesis revealed that more fetuses have normal chromosomal karyotypes than balanced carriers, particularly in the REC group. CONCLUSIONS: Reciprocal translocation carriers have more risk of unbalanced rearrangement, but embryonic chromosome abnormalities of inversion carriers come mainly from de novo variations. This is the first study specifically comparing three different PGT-SRs using the NGS method and evaluating their reproductive outcomes. Our findings will provide the reciprocal translocation, inversion, and Robertsonian translocation carrier couples with more accurate genetic counseling on the reproductive risk of chromosomal imbalance.

Does an FSH surge at the time of hCG trigger improve IVF/ICSI outcomes? A randomized, double-blinded, placebo-controlled study.

STUDY QUESTION: Does an artificially induced FSH surge at the time of hCG trigger improve IVF/ICSI outcomes? SUMMARY ANSWER: An additional FSH bolus administered at the time of hCG trigger has no effect on clinical pregnancy rate, embryo quality, fertilization rate, implantation rate and live birth rate in women undergoing the long GnRH agonist (GnRHa) protocol for IVF/ICSI. WHAT IS KNOWN ALREADY: Normal ovulation is preceded by a surge in both LH and FSH. Few randomized clinical trials have specifically investigated the role of the FSH surge. Some studies indicated that FSH given at hCG ovulation trigger boosts fertilization rate and even prevents ovarian hyperstimulation syndrome (OHSS). STUDY DESIGN, SIZE, DURATION: This was a randomized, double-blinded, placebo-controlled trial conducted at a single IVF center, from June 2012 to November 2013. A sample size calculation indicated that 347 women per group would be adequate. A total of 732 women undergoing IVF/ICSI were randomized, using electronically randomized tables, to the intervention or placebo groups. Participants and clinical doctors were blinded to the treatment allocation. PARTICIPANTS/MATERIALS, SETTING, METHODS: Patients aged ≤42 years who were treated with IVF/ICSI owing to tubal factor, male factor, unexplained, endometriosis and multiple factors were enrolled in this trial. Subjects all received a standard long GnRHa protocol for IVF/ICSI and hCG 6000-10 000 IU to trigger oocyte maturation. A total of 364 and 368 patients were randomized to receive a urinary FSH (uFSH) bolus (6 ampules, 450 IU) and placebo, respectively, at the time of the hCG trigger. The primary outcome measure was clinical pregnancy rate. The secondary outcome measures were FSH level on the day of oocyte retrieval, number of oocytes retrieved, good-quality embryo rate, live birth rate and rate of OHSS. MAIN RESULTS AND THE ROLE OF CHANCE: There were no significant differences in the baseline demographic characteristics between the two study groups. There were also no significant differences between groups in cycle characteristics, such as the mean number of stimulation days, total gonadotrophin dose and peak estradiol. The clinical pregnancy rate was 51.6% in the placebo group and 52.7% in the FSH co-trigger group, with an absolute rate difference of 1.1% (95% CI -6.1% to 8.3%). The number of oocytes retrieved was 10.47 ± 4.52 and 10.74 ± 5.01 (P = 0.44), the rate of good-quality embryos was 37% and 33.9% (P = 0.093) and the implantation rate was 35% and 36% (P = 0.7) in the placebo group and the FSH co-trigger group, respectively. LIMITATIONS, REASONS FOR CAUTION: This was a single-center study, which may limit its effectiveness. The use of uFSH is a limitation, as this is not the same as the natural FSH. We did not collect follicular fluid for further study of molecular changes after the use of uFSH as a co-trigger. WIDER IMPLICATIONS OF THE FINDINGS: Based on previous data and our results, an additional FSH bolus administered at the time of hCG trigger has no benefit on clinical pregnancy rates in women undergoing the long GnRHa protocol in IVF/ICSI: a single hCG trigger is sufficient. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Key Research and Development Program of China (2016YFC1000205); Sun Yat-Sen University Clinical Research 5010 Program (2016004); the Science and Technology Project of Guangdong Province (2016A020216011 and 2017A020213028); and Science Technology Research Project of Guangdong Province (S2011010004662). There are no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: The trial was registered in the Chinese Clinical Trial Registry (ChiCTR-TRC-12002246). TRIAL REGISTRATION DATE: 20 May 2012. DATE OF FIRST PATIENT'S ENROLMENT: 10 June 2012.

Rapamycin preserves the primordial follicle pool during cisplatin treatment in vitro and in vivo.

Rapamycin has been proven to effectively inhibit the activation of primordial follicles while cisplatin-induced the loss of primordial follicles due to the over-activation of the primordial follicle stockpile. Whether rapamycin could inhibit the loss of primordial follicles induced by cisplatin is still unknown. The ovaries of neonatal Sprague Dawley rats were cultured in vitro in different doses of rapamycin (0.08, 0.16, and 0.32 µg/ml) and cisplatin (0.1, 0.4, and 0.8 µg/ml). The immature BALB/c mice were administered cisplatin with or without rapamycin by intraperitoneal injection. Ovaries were collected to analyze the histomorphology, the messenger RNA (mRNA) expression of anti-Mullerian hormone (AMH), growth differentiation factor 9 (GDF9), and bone morphogenetic protein 15 (BMP15) and the expression of key proteins of mammalian target of rapamycin (mTOR) pathway. Growing follicle counts of ovaries cultured in vitro in the R0.16 and R0.32 groups were decreased and the ratio of growing to primordial follicles was also decreased in a dose-dependent manner. In the C0.8 group, growing follicles were decreased compared with the other groups while the ratio was substantially increased in the C0.4 and C0.8 group. Co-treatment attenuated primordial follicle loss and reduced the upregulated ratio induced by cisplatin. Ovarian follicle dynamics in vivo was consistent with the in vitro results. Primordial follicles counts were statistically increased and the ratio was reduced in the rapamycin group compared with the control group. Primordial follicle counts were dramatically reduced in the cisplatin group whereas co-treatment with rapamycin slightly recovered its counts. There was no obvious difference in the number of growing follicles between the cisplatin group and other groups. The ratio was significantly increased in cisplatin-treated mice whereas decreased in the co-treatment group. The apoptosis rate of antral follicles in cisplatin-treated mice was higher than the other groups while the apoptosis rate was decreased in the co-treatment group in vivo. Compared with the control and rapamycin group, the mRNA expression of AMH, GDF9, and BMP15 were downregulated in the cisplatin group. The co-treatment group recovered the mRNA expression of BMP15. In addition, the expression of key protein of mTOR pathway rpS6 and its phosphorylated forms were increased in the cisplatin-treated group while co-treatment decreased their expression. Rapamycin attenuated the loss of primordial follicles induced by cisplatin through the inhibitory effect of rapamycin on the mTOR pathway. These results suggest that rapamycin may be an effective drug for the protection of ovarian function during chemotherapy.

A whole-genome sequencing-based novel preimplantation genetic testing method for de novo mutations combined with chromosomal balanced translocations.

PURPOSE: To explore a new preimplantation genetic testing (PGT) method for de novo mutations (DNMs) combined with chromosomal balanced translocations by whole-genome sequencing (WGS) using the MGISEQ-2000 sequencer. METHODS: Two families, one with maternal Olmsted syndrome caused by DNM (c.1246C>T) in TRPV3 and a paternal Robertsonian translocation and one with paternal Marfan syndrome caused by DNM (c.4952_4955delAATG) in FBN1 and a maternal reciprocal translocation, underwent PGT for monogenetic disease (PGT-M), chromosomal aneuploidy, and structural rearrangement. WGS of embryos and family members were performed. Bioinformatics analysis based on gradient sequencing depth was performed, and parent-embryo haplotyping was conducted for DNM diagnosis. Sanger sequencing, karyotyping, and chromosomal microarray analysis were performed using an amniotic fluid sample to confirm the PGT results. RESULTS: After 1 PGT cycle, WGS of 2 embryos from the Olmsted syndrome family revealed euploid embryos without DNMs; after 2 cycles, the 11 embryos from the Marfan syndrome family showed only 1 normal embryo without DNM, copy number variations (CNVs), or aneuploidy. Moreover, 1 blastocyst from the Marfan syndrome family was transferred back to the uterus; the amniocentesis test results were confirmed by PGT and a healthy infant was born. CONCLUSIONS: WGS based on parent-embryo haplotypes was an effective strategy for PGT of DNMs combined with a chromosomal balanced translocation. Our results indicate this is a reliable and effective diagnostic method that is useful for clinical application in PGT of patients with DNMs.

Design Implementation and Evaluation of a Mobile Continuous Blood Oxygen Saturation Monitoring System.

OBJECTIVE: In this study, we built a mobile continuous Blood Oxygen Saturation (SpO2) monitor, and for the first time, explored key design principles towards daily applications. METHODS: We firstly built a customized wearable computer that can sense two-channel photoplethysmogram (PPG) signals, and transmit the signals wirelessly to smartphone. Afterwards, we explored many SpO2 model building principles, focusing on linear/nonlinear models, different PPG parameter calculation methods, and different finger types. Moreover, we further compared PPG sensor placement principles by comparing different hand configurations and different finger configurations. Finally, a dataset collected from eleven human subjects was used to evaluate the mobile health monitor and explore all of the above design principles. RESULTS: The experimental results show that the root mean square error of the SpO2 estimation is only 1.8, indicating the effectiveness of the system. CONCLUSION: These results indicate the effectiveness of the customized mobile SpO2 monitor and the selected design principles. SIGNIFICANCE: This research is expected to facilitate the continuous SpO2 monitoring of patients with clinical indications.

VIP activates primordial follicles of rat through ERK-mTOR pathway in tissue culture.

In vitro activation of primordial follicles is becoming more essential in assisted reproductive technologies. Vasoactive intestinal peptide (VIP) is one of the members of the neurotrophin family which has demonstrated to have an impact on follicle development in recent years. This study aims to investigate the effect of VIP on the activation of primordial follicles in neonatal rat in an in vitro culture system and to determine the relevant molecular mechanism of their activation. Ovaries of 4-day-old rats were examined for the expression of VIP receptors and were cultured in mediums containing VIP with or without inhibitors of the ERK-mTOR signalling pathway. They were then collected for histological analysis or measurement of the molecular expression of this pathway. The receptors of VIP were found in granular cells and oocytes of primordial and early-growing follicles in neonatal ovary. The ratio of growing follicle increased in the presence VIP at different concentrations, with the highest level of increase being observed in the 10-7 mol/L VIP-treated group. The ratio of PCNA-positive granular cells was also increased, while that of the apoptotic oocytes were decreased, and protein analysis showed increased phosphorylation of ERK1/2, mTOR and RPS6 in the VIP-treated group. However, the effect of VIP on the activation of primordial follicle became insignificant with the addition of MEK inhibitor (U0126) or mTORC1 inhibitor (rapamycin). This study indicated that VIP could activate neonatal rat primordial follicle through the ERK-mTOR signalling pathway, suggesting a strategy for in vitro primordial follicle recruitment.

Live Births after in vitro Fertilization with Fertility-Sparing Surgery for Borderline Ovarian Tumors: A Case Series and Literature Review.

This is a case series reporting live births following in vitro fertilization (IVF) after fertility-sparing surgery in patients with borderline ovarian tumors (BOTs). Seventeen patients, in whom fertility-sparing surgery for BOTs followed by IVF was performed from July 1, 2011 to September 30, 2017, in our IVF center, were included. Here we retrospectively present their BOT characteristics and clinical outcomes of assisted reproduction technology (ART) and then discuss the literature on management of patients with BOTs after fertility-sparing surgery. The study sample of 17 patients started IVF-assisted reproduction within a median of 24 months after fertility-sparing surgery. Eight patients achieved live births after IVF followed by fresh or frozen-thawed embryo transfer, and one is currently pregnant after the second IVF treatment. Four of these 17 women experienced 6 recurrences of BOTs after IVF and delivery in a median of 29 months of follow-up. The literature reports a pooled estimate of 63.5% for pregnancy and 21.7% for recurrence of BOTs in patients with the same condition. Pregnancy outcomes of IVF in these women are usually better than those of generally infertile women. Moreover, our review of the literature suggests that ART has no negative impact on the prognosis of BOTs.

Novel mutation in the ZP1 gene and clinical implications.

PURPOSE: Empty follicle syndrome (EFS) is a complex reproductive disorder characterized by the repeated failure to aspirate oocytes from mature ovarian follicles during in vitro fertilization (IVF). In addition to some cases caused by iatrogenic problems and known genetic factors, there are still many unexplained aspects of EFS. Here, we aimed to assess the clinical and genetic characteristics of two EFS patients. METHODS: We have characterized two primary infertility patients with EFS in a nonconsanguineous family from China. Both the patients presented similar clinical phenotypes, that is a few granulosa cells but no oocytes could be retrieved during repeated cycles with normal follicular development, E2 levels, and bioavailable hCG plasma levels. Abnormal oocytes were obtained once or twice between multiple IVF cycles. We performed Sanger sequencing of the LHCGR and ZP1~ZP4 genes in the patients, and further bioinformatics analysis was performed to identify pathogenic elements in the genes. RESULTS: A novel mutation, c.181C>T (p.Arg61Cys), and a known mutation, c.1169_1176delTTTTCCCA (p.Ile390Thrfs*16), in the ZP1 gene were both identified in patient 2, but no mutations were identified in patient 1. The novel mutation inherited from her mother was absent in the control cohort and the ExAc database. The arginine residue is conserved at this position, and its replacement by cysteine was predicted to be deleterious. In another allele, a paternal frameshift mutation was predicted to introduce premature stop codons, resulting in the deletion of 234 amino acids from the C-terminus of the ZP1 protein. CONCLUSIONS: Our findings presented compound heterozygous mutations in ZP1 associated with EFS and abnormal oocytes and provided further new evidence for the genetic basis of EFS and support for the genetic diagnosis of infertile individuals.

Downregulation of serum long noncoding RNA GAS5 may contribute to insulin resistance in PCOS patients.

Polycystic ovary syndrome (PCOS) is a common endocrine disease that affects reproductive-aged women and mostly characterized by insulin resistance (IR). The underlying mechanism remains unknown. Long noncoding RNAs (lncRNAs) have been demonstrated to be involved in various levels of biological regulation process of cell development, metabolism, and differentiation. This study aims to investigate the relationship between IR and differential expression of lncRNA Growth-arrest specific transcript 5 (GAS5) in patients' serum with and without PCOS. A total of 76 cases of serum was collected from non-PCOS and PCOS patients with and without IR to measure interleukin-18 (IL-18) and GAS5 expression, which were correlated with IR status. The IL-18 concentration in serums was significantly increased in PCOS patients with IR. GAS5 expression was decreased in serums in PCOS patients with IR. Result of correlation analysis shows that there is a negative association between GAS5 expression and homeostasis model of assessment for insulin resistance (HOMA-IR). GAS5 was yielded the ROC curve (AUC). Our study implied that elevated IL-18 expression and downregulation of GAS5 in serums might contribute to IR in PCOS patients.

Brief co-incubation of gametes benefits the outcomes of newborns.

PURPOSE: The objective of this study was to determine whether ammonium accumulates in IVF media during fertility process and whether the brief co-incubation of gametes (bIVF) benefited the outcomes of newborns. METHODS: Ammonium levels in IVF media during gamete co-incubation were measured and the effects of bIVF on neonatal outcomes were evaluated retrospectively in this study. RESULTS: A total of 609 live newborns cycles were included in this study. The results showed that ammonium levels in the conventional IVF (cIVF) media was significantly increased than that in bIVF and control media (27.32 ± 5.60 vs 20.71 ± 3.89, P = 0.03; 27.32 ± 5.60 vs 19.46 ± 1.31, P = 0.01, respectively). In the cIVF group, the mean gestational age was significantly lower (37.36 ± 2.29 vs. 37.74 ± 1.94 weeks, P = 0.031) and the incidence of preterm birth (< 37 weeks) was higher than that in the bIVF group (25.80 vs. 17.63%, P = 0.015). Singleton cycles and twin cycles were then analyzed respectively. The gestational age and birth weight of the singleton cycles were similar between the two groups. However, of the twin cycles, the gestational age was significantly decreased and the rate of preterm birth was increased significantly in the cIVF group (35.76 ± 2.31 vs. 36.48 ± 1.73, P = 0.013; 53.33 vs. 31.52%, P = 0.002, respectively). CONCLUSIONS: There is an ammonium accumulation in IVF media during co-incubation of gametes. And bIVF reduces the risk of preterm birth (< 37 weeks), especially with regard to preterm birth of the twin cycles, and seems to be a safe alternative method for improving the neonatal outcomes compared with cIVF.

Genome-wide screening differential long non-coding RNAs expression profiles discloses its roles involved in OHSS development.

OBJECTIVE: To screen differentially expressed lncRNAs involved in OHSS. OHSS is defined as ovarian hyperstimulation syndrome. It is characterized as enlarged ovary and increased vascular permeability. DESIGN: Case-control study. SETTING: University-affiliated hospital. PATIENT(S): Patients with OHSS high risk (n = 30) and low risk (n = 30) were included in this study. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): LncRNAs from women with OHSS high risk and low risk were used for high-throughput sequencing profiling. The eight most differentially expressed lncRNAs in granulosa cells were validated by semi-quantitative reverse transcription-polymerase chain reaction analysis. RESULT(S): A total of 23,815 lncRNAs were detected and 482 were differentially expressed (fold-change ≥2; p < 0.05, FDR value < 0.001), of which 205 were upregulated and 277 were downregulated. Lnc-SEC16B.1-6, lnc-SNURF-13, lnc-LGR6-6, and lnc-H2AFY2-2 were up-regulated, while lnc-BRD2-2, lnc-HSPA6-2, and lnc-CLIC6-5 were downregulated significantly in granulosa cells. These results were confirmed by qRT-PCR. KEGG pathways and Gene Ontology enrichment analysis revealed that several biological processes were significantly associated. Meanwhile, the lncRNA/miRNA interaction network was established according to ceRNA network model. CONCLUSION(S): Comprehensive expression screening identified eight novel lncRNAs associated with risk factors of OHSS process. Although it is unclear how these altered lncRNAs regulate the process of OHSS, our findings suggest these lncRNAs may be novel players in OHSS development.

A novel mutation in the TUBB8 gene is associated with complete cleavage failure in fertilized eggs.

PURPOSE: Cleavage of the zygote during human reproduction is a key event of early embryonic development. The genetic events associated with idiopathic embryonic cleavage failure are not certain. Mutations in the tubulin beta 8 class VIII (TUBB8) gene have been reported to be associated with oocyte maturation, fertilization, and developmental arrest. Here, we aimed to assess the clinical and genetic characteristics of complete cleavage failure in fertilized eggs. METHODS: We have characterized a patient with a 9-year history of primary infertility in a non-consanguineous family from China. The patient presented complete cleavage failure in all two-pronuclear (2PN) fertilized oocytes after 2 cycles of in vitro fertilization (IVF). We performed Sanger sequencing of the TUBB8 gene in the patient, and further bioinformatics analysis to identify pathogenesis of gene. RESULTS: A novel homozygous mutation, c.322G > A (p.Glu108Lys), was detected, and this change was absent from 179 control subjects. Glutamic acid is highly conserved at this position, and replacement by lysine was predicted to be repelled by the α-tubulin positive region, disrupting the α-ß tubulin interaction. CONCLUSIONS: Our findings presented a homozygous mutation of TUBB8 associated with complete cleavage failure in fertilized eggs and provided new data for the genotype-phenotype of TUBB8-related diseases.