Reproductive outcomes of cesarean scar pregnancies treated with uterine artery embolization combined with curettage.

OBJECTIVE: The aim of this study was to review the reproductive outcomes of women with a cesarean scar pregnancy (CSP) treated with dilation and curettage (D&C) after uterine artery embolization (UAE). MATERIALS AND METHODS: This was a retrospective study to review women who received UAE followed by D&C for CSP between January 2010 and December 2019 at the Changhua Christian Hospital, Changhua in Taiwan. Data were collected from both electronic and paper medical records. Patients were contact via phone call to follow up reproductive outcomes between January 2021 and March 2021. These subsequent reproductive outcomes (including pregnancy rate, secondary infertility rate, miscarriage rate, live birth rate, and recurrent CSP rate) were recorded and analyzed. RESULTS: A total of 53 cases of women who received UAE followed by D&C for CSP were identified. The women's average age was 34.8 ± 5.1 years. The mean gestational age at diagnosis was 6.2 ± 1.1 weeks. The mean level for human chorionic gonadotropin was 23,407.7 ± 29,105.5 mIU/ml. The average of blood loss during D&C was 19.2 ± 43.6 ml. The average hospitalization time after D&C was 3.5 ± 1.1 days. Of the 53 cases, 10 patients were lost to follow-up and 43 patients agreed to follow-up on reproductive outcomes in 2021. Twenty-three patients who desired to conceive were analyzed. Nineteen out of these 23 women (82.6%) succeeded in conceiving again and gave birth to 15 healthy babies (78.9%). Only one woman (1/19, 5.3%) experienced recurrence of CSP. The average time interval between previous CSP treatment and subsequent conception was 10.4 ± 6.7 months. CONCLUSION: UAE combined with curettage treatment in CSP patients results in a positive rate of subsequent pregnancy outcomes. This minimally invasive procedure may be considered as one of the treatment options for CSP, as it enables preservation of fertility after treatment.

Automatic Sleep-Arousal Detection with Single-Lead EEG Using Stacking Ensemble Learning.

Poor-quality sleep substantially diminishes the overall quality of life. It has been shown that sleep arousal serves as a good indicator for scoring sleep quality. However, patients are conventionally asked to perform overnight polysomnography tests to collect their physiological data, which are used for the manual judging of sleep arousals. Even worse, not only is this process time-consuming and cumbersome, the judgment of sleep-arousal events is subjective and differs widely from expert to expert. Therefore, this work focuses on designing an automatic sleep-arousal detector that necessitates only a single-lead electroencephalogram signal. Based on the stacking ensemble learning framework, the automatic sleep-arousal detector adopts a meta-classifier that stacks four sub-models: one-dimensional convolutional neural networks, recurrent neural networks, merged convolutional and recurrent networks, and random forest classifiers. This meta-classifier exploits both advantages from deep learning networks and conventional machine learning algorithms to enhance its performance. The embedded information for discriminating the sleep-arousals is extracted from waveform sequences, spectrum characteristics, and expert-defined statistics in single-lead EEG signals. Its effectiveness is evaluated using an open-accessed database, which comprises polysomnograms of 994 individuals, provided by PhysioNet. The improvement of the stacking ensemble learning over a single sub-model was up to 9.29%, 7.79%, 11.03%, 8.61% and 9.04%, respectively, in terms of specificity, sensitivity, precision, accuracy, and area under the receiver operating characteristic curve.

Microfluidic channel integrated with a lattice lightsheet microscopic system for continuous cell imaging.

In this study, a continuous cell-imaging system with subcellular resolution was developed by integrating a microfluidic platform with lattice lightsheet microscopy (LLSM). To reduce aberrations of the lightsheet propagating into the device, a microfluidic channel sealed with a water refractive index-matched thin film was fabricated. When the lightsheet emerged from the water-immersed objectives and penetrated through the water refractive-matched thin film into the microfluidic channel at an incident angle, less light scattering and fewer aberrations were found. Suspended cells flowed across the lattice lightsheet, and an imaging system with the image plane perpendicular to the lightsheet was used to sequentially acquire cell images. By applying a thinner lattice lightsheet, higher-resolution, higher-contrast images were obtained. Furthermore, three-dimensional cell images could be achieved by reconstructing sequential two-dimensional cell images.

Hypertension Risk in Young Women With Polycystic Ovary Syndrome: A Nationwide Population-Based Cohort Study.

Objective: A number of publications have assessed the prevalence of hypertension in polycystic ovary syndrome (PCOS) patients with inconclusive results. Since in general populations the occurrence of hypertension is related to age and comorbidities, we investigated the incidence rate and hazard ratios (HRs) of hypertension between healthy subjects and young women with PCOS as well as comorbidities. Methods: We conducted a population-based retrospective cohort study by using the National Health Insurance Research Database in Taiwan. The cohort included women with the diagnosis of PCOS between 2000 and 2012. Those without PCOS were selected as the control group at a ratio of 4:1 by an age-matched systematic random-sampling method. Cox proportional hazard regression analysis was used to determine the effects of PCOS on the risks of developing hypertension. Stratification analysis was performed to elucidate the interaction among PCOS and the comorbidities, which affect the incidence of hypertension. Results: The PCOS cohort consisted of 20,652 patients with PCOS and the comparison cohort consisted of 82,608 matched patients without PCOS. There was no difference in the distribution of age between the PCOS cohort and the comparison cohort (29.1 ± 6.8 vs. 29.0 ± 6.5, p = 0.32). The incidence rates of hypertension were 7.85 and 4.23 per 1,000 person-years in the PCOS and comparison groups, respectively. A statistically significant higher risk of hypertension was found in the PCOS cohort (adjusted HR = 1.62, 95% confidence interval = 1.48-1.76) than in the comparison cohort. After a joint analysis of comorbidities, the adjusted HR of hypertension was 9.44 (95% confidence interval = 7.27-12.24) for PCOS patients with comorbidities of diabetes mellitus (DM) and hyperlipidemia compared with women with neither PCOS nor DM and hyperlipidemia. Conclusion: The risk of developing hypertension in young women with PCOS was higher than in controls in this cohort study. The comorbidities of DM and hyperlipidemia could interact with PCOS patients and further increase the risk of hypertension. An earlier screening for hypertension and comorbidities in patients with PCOS may be warranted, even in young women.

Association between the Genetic Variants of Glutathione Peroxidase 4 and Severity of Endometriosis.

It has been reported that oxidative and nitrative stress might be the pathogenesis of endometriosis. This prospective case-control study attempted to check the connection between single nucleotide polymorphism (SNP) of three antioxidant enzymes (glutathione peroxidase 4 (GPX4), thioredoxin 2 (TXN2), thioredoxin reductase 1 (TXNRD1)) and endometriosis. We recruited 90 patients with histology-approved endometriosis as the case group and 130 age-matched women for an annual pap smear examination as the control group. The stage of endometriosis was evaluated with revised ASRM score. Both groups were genotyped in the peripheral leukocytes for the SNP of GPX4 (rs713041), TXN2 (rs4821494) and TXNRD1 (rs1128446) by PCR-based methods. An X2 test was used to analysis of the difference of allele frequency and SNP distribution between two groups. The results revealed GPX4 (rs713041) has a significantly different distribution between two groups (C:T = 116 (44.6%):144 (55.4%) in control and C:T = 104 (57.8%): 76 (42.2%) in endometriosis groups, p = 0.007). The SNP in TXN2 (rs4821494) also showed a difference in allele frequency (G:T = 180 (69.2%):80 (30.8%) in control and G:T = 141 (78.3%):39 (21.6%) in endometriosis group, p = 0.030). In addition, the SNP GPX4 (rs713041) was associated with the severity of the endometriosis. Women who have advanced stage endometriosis were different from mild endometriosis in genetic variants of GPX4 gene (p = 0.001). In conclusion, the relationship between endometriosis and SNP of antioxidant enzymes, GPX4 and TXN2, was confirmed by the present study. According to the result, we suggested that the GPX4 might contribute to the pathogenesis of endometriosis.

Risk of Psoriasis in Patients with Polycystic Ovary Syndrome: A National Population-Based Cohort Study.

Both polycystic ovary syndrome (PCOS) and psoriasis are associated with insulin resistance and metabolic syndrome. Nonetheless, the incidence of psoriasis in patients with PCOS is unclear. We used the Longitudinal Health Insurance Research Database (LHID) in Taiwan from 2000 to 2012 to perform a retrospective population-based cohort study to elucidate the occurrence of psoriasis in PCOS patients. Patients with PCOS without psoriasis in the index year (the year of PCOS diagnosis) were recruited as the PCOS group. Those without PCOS nor psoriasis (control group) were selected using propensity score matching at a ratio of 4:1. Hazard ratios (HRs) were obtained using the Cox proportional hazards regression model. In total, 4707 and 18,828 patients were included in the PCOS and control groups, respectively. The incidence rates of psoriasis in the control and PCOS groups were 0.34 and 0.70 per 1000 person-years, respectively. The risk of psoriasis was higher in the PCOS group by an HR of 2.07 (95% confidence interval [CI] = 1.25-3.43) compared with the control group. In conclusion, the incidence of psoriasis in the PCOS group was higher than that in the control group. Further studies should be conducted to investigate the mechanism underlying the association, and to benefit the long-term management of patients with PCOS.

Interleukin-3 Polymorphism is Associated with Miscarriage of Fresh in Vitro Fertilization Cycles.

The aim of this study was to examine the association between interleukin (IL) genes polymorphisms and in vitro fertilization (IVF) outcome. A prospective cohort analysis was performed at a Women's Hospital IVF centre of 1015 female patients undergoing fresh non-donor IVF cycles. The effects of the following six single nucleotide polymorphisms (SNPs) in five IL genes on IVF outcomes were explored: IL-1α (rs1800587 C/T), IL-3 (rs40401 C/T), IL-6 (rs1800795 C/G), IL-15 (rs3806798 A/T), IL-18 (rs187238 C/G) and IL-18 (rs1946518 G/T). The main outcome measures included clinical pregnancy, embryo implantation, abortion and live birth rates. There were no statistically significant differences in clinical pregnancy, embryo implantation and live birth rates in the analysis of 1015 patients attempting their first cycle of IVF. Infertile women with IL-3 homozygous major genotype had a higher abortion rate than those with heterozygous and homozygous minor genotype (16.5% vs. 7.9%, P = 0.025). In conclusion, our results indicated that the IL-3 rs40401 polymorphism is associated with increased risk of abortion of IVF patients. Future studies with inclusion of other ethnic populations must be conducted to confirm the findings of this study.

Anti-Müllerian Hormone Gene Polymorphism is Associated with Clinical Pregnancy of Fresh IVF Cycles.

The aim of this study was to examine the effects of single-nucleotide polymorphisms (SNPs) in the anti-Müllerian hormone (AMH) and AMH type II receptor (AMHRII) genes on in vitro fertilization (IVF) outcomes. In this prospective cohort study, we genotyped the AMH 146 T > G, AMHRII -482 A > G and AMHRII IVS1 +149 T > A variants in 635 women undergoing their first cycle of controlled ovarian stimulation for IVF. DNA was extracted from the peripheral blood of all participants, and the SNPs were genotyped by real-time polymerase chain reaction. The distributions, frequencies of genes, and correlation with clinical pregnancy of IVF were analyzed. The AMH 146 T > G G/G genotype in women was associated with a lower clinical pregnancy rate (T/T: 55.0%, T/G: 51.8%, G/G: 40.0%; p < 0.05). Women with the AMH 146 T > G GG genotype were half as likely to have a clinical pregnancy compared with women with TT genotypes (OR = 0.55, 95% CI: 0.34⁻0.88, p = 0.014). With multivariate analysis, the AMH 146 T > G GG genotype remains as a significant independent factor to predict clinical pregnancy (p = 0.014). No significant difference was found between AMHRII polymorphisms and clinical pregnancy outcomes of IVF. In conclusion, our results show that AMH 146 T > G seems to be a susceptibility biomarker capable of predicting IVF pregnancy outcomes. Further studies should focus on the mechanism of these associations and the inclusion of other ethnic populations to confirm the findings of this study.

Performance of preimplantation genetic testing for aneuploidy in IVF cycles for patients with advanced maternal age, repeat implantation failure, and idiopathic recurrent miscarriage.

OBJECTIVE: The primary objective of this study was to investigate whether preimplantation genetic testing for aneuploidy (PGT-A) of blastocysts through array comparative genomic hybridization (aCGH) improves live birth rates (LBR) in IVF cycles for patients with high prevalence of aneuploidy. MATERIALS AND METHODS: This study included 1389 blastocysts with aCGH results derived from 296 PGT-A cycles in IVF patients with advanced maternal age (AMA) (n = 87, group A), those with repeated implantation failure (RIF) (n = 82, group B), those with recurrent miscarriage (RM) (n = 82, group C), and oocyte donors (OD) (n = 45, young age, as a control group). Another 61 AMA patients without PGT-A procedures were used as a control group for group A. Vitrification was performed after blastocyst biopsy, and thawed euploid embryos were transferred in a nonstimulated cycle. RESULTS: For the AMA group, a significant increase in LBRs was found in the PGT-A group compared with the non-PGT-A group (54.1% vs. 32.8%, p = 0.018). Consistent LBRs (54.1%, 51.6%, 55.9%, and 57.1%, respectively, in group A, B, C, and young age group) were obtained for all the indications. CONCLUSIONS: LBRs can be improved using PGT-A of blastocysts with aCGH in IVF cycles for patients with a high rate of aneuploidy, especially for patients with AMA.

The role of glucocorticoids in ovarian development of sleep deprived rats.

OBJECTIVE: Sleep deprivation (SD) adversely affects female reproductive function. In this study, we investigated the role of glucocorticoids in ovarian development in sleep deprived rats. MATERIALS AND METHODS: Female rats were subjected to SD for 1-4 days. Concentrations of serum estradiol and corticosterone were assessed. Betamethasone (BET) and/or recombinant human follicle-stimulating hormone (FSH) were administered to 21-day-old female rats for 2 days to evaluate ovarian status for follicular development. Intact preantral follicles were mechanically dissected from the rat's ovaries and cultured for 72 h with or without FSH in the presence or absence of BET to evaluate follicular development. RESULTS: SD led to a significant difference in serum estradiol concentrations between the sham and SD groups, and corticosterone concentrations were significantly elevated in groups with more than 2 days of SD (P < 0.05). FSH stimulated ovarian growth in immature rats, whereas BET inhibited ovarian development caused by the FSH treatment. Treatment of the preantral follicles with FSH induced an increase in both follicle size and follicular cell number, while follicular cell differentiation was accompanied by enhanced inhibin-α and connexin 43 expression. Inhibition of FSH-stimulated follicular growth through BET treatment exhibited a dose-dependent reciprocal trend; as the BET dose increased (0.001-1 µg/mL), preantral follicular growth decreased. This decrease was associated with a decrease in follicular cell numbers and suppression of a proliferating cell nuclear antigen, inhibin-α, and connexin 43 expression. CONCLUSION: The findings suggest that the adverse effects of SD may inhibit follicular development during ovarian hyperstimulation by corticosterone elevation in rat.

Effect of morphokinetics and morphological dynamics of cleavage stage on embryo developmental potential: A time-lapse study.

OBJECTIVE: Using a non-invasive method to select the most competent embryo is essential in in vitro fertilization (IVF). Since the beginning of clinical application of time-lapse technology, several studies have proposed models using the time-lapse imaging system for predicting the IVF outcome. This study used both morphokinetic and morphological dynamic parameters to select embryos with the highest developmental potential. MATERIALS AND METHODS: A total of 23 intracytoplasmic sperm injection treatment cycles with 138 fertilized oocytes were included in this study. All embryos were cultured to the blastocyst stage, and embryo development was recorded every 10 min by using a time-lapse imaging system. Morphokinetic parameters and eight major abnormal division behaviors were studied to determine their effects on blastocyst formation. The most influential variables were used in hierarchical classification for blastocyst formation prediction. RESULTS: Several parameters were significantly related to the developmental potential. Embryos with the timing of pronuclear fading (tPNF) of >26.4 h post insemination (hpi), the timing of division to two cells (t2) of >29.1 hpi, and the timing of division to four cells (t4) of >41.3 hpi showed the lowest blastocyst formation rate. The abnormal division behaviors of fragmentation >50%, direct cleavage, reverse cleavage, and delayed division or developmental arrest were found to be detrimental to blastocyst formation. On the basis of these results, we propose a hierarchical model classification, in which embryos are classified into groups A-D according to their developmental potential. The blastocyst formation rates of groups A, B, C, and D were 80.0%, 77.8%, 53.7%, and 22.2% (p < 0.001). The good blastocyst rates of groups A, B, C, and D were 60.0%, 44.4%, 14.6%, and 11.1% (p = 0.007). CONCLUSION: We propose a hierarchical classification system for blastocyst formation prediction, which provides information for embryo selection by using a time-lapse imaging system.

B-Cell Lymphoma 6 (BCL6) Is a Host Restriction Factor That Can Suppress HBV Gene Expression and Modulate Immune Responses.

Hepatitis B virus (HBV) infection causes acute and chronic liver inflammation. Recent studies have demonstrated that some viral antigens can suppress host innate and adaptive immunity, and thus lead to HBV liver persistency. However, the cellular factors that can help host cells to clear HBV during acute infection remain largely unknown. Here, we used HBV-cleared and HBV-persistent mouse models to seek for cellular factors that might participate in HBV clearance. HBV replicon DNA was delivered into the mouse liver by hydrodynamic injection. RNA-Seq analysis was conducted to identify immune-related genes that were differentially expressed in HBV-persistent and HBV-cleared mouse models. A cellular factor, B cell lymphoma 6 (BCL6), was found to be significantly upregulated in the liver of HBV-cleared mice upon HBV clearance. Co-expression of BCL6 and a persistent HBV clone rendered the clone largely cleared, implicating an important role of BCL6 in controlling HBV clearance. Mechanistic studies demonstrated that BCL6 functioned as a repressor, binding to and suppressing the activities of the four HBV promoters. Correspondingly, BCL6 expression significantly reduced the levels of HBV viral RNA, DNA, and proteins. BCL6 expression could be stimulated by inflammatory cytokines such as TNF-α; the BCL6 in turn synergized TNF-α signaling to produce large amounts of CXCL9 and CXCL10, leading to increased infiltrating immune cells and elevated cytokine levels in the liver. Thus, positive feedback loops on BCL6 expression and immune responses could be produced. Together, our results demonstrate that BCL6 is a novel host restriction factor that exerts both anti-HBV and immunomodulatory activities. Induction of BCL6 in the liver may ultimately assist host immune responses to clear HBV.

Endoperoxide formation by an α-ketoglutarate-dependent mononuclear non-haem iron enzyme.

Many peroxy-containing secondary metabolites have been isolated and shown to provide beneficial effects to human health. Yet, the mechanisms of most endoperoxide biosyntheses are not well understood. Although endoperoxides have been suggested as key reaction intermediates in several cases, the only well-characterized endoperoxide biosynthetic enzyme is prostaglandin H synthase, a haem-containing enzyme. Fumitremorgin B endoperoxidase (FtmOx1) from Aspergillus fumigatus is the first reported α-ketoglutarate-dependent mononuclear non-haem iron enzyme that can catalyse an endoperoxide formation reaction. To elucidate the mechanistic details for this unique chemical transformation, we report the X-ray crystal structures of FtmOx1 and the binary complexes it forms with either the co-substrate (α-ketoglutarate) or the substrate (fumitremorgin B). Uniquely, after α-ketoglutarate has bound to the mononuclear iron centre in a bidentate fashion, the remaining open site for oxygen binding and activation is shielded from the substrate or the solvent by a tyrosine residue (Y224). Upon replacing Y224 with alanine or phenylalanine, the FtmOx1 catalysis diverts from endoperoxide formation to the more commonly observed hydroxylation. Subsequent characterizations by a combination of stopped-flow optical absorption spectroscopy and freeze-quench electron paramagnetic resonance spectroscopy support the presence of transient radical species in FtmOx1 catalysis. Our results help to unravel the novel mechanism for this endoperoxide formation reaction.

The influence of female age on the cumulative live-birth rate of fresh cycles and subsequent frozen cycles using vitrified blastocysts in hyper-responders.

OBJECTIVE: The aim of this research was to study the influence of female age on the cumulative live-birth rate of fresh and subsequent frozen cycles using vitrified blastocysts of the same cohort in hyper-responders. MATERIALS AND METHODS: This was a retrospective study of 1137 infertile women undergoing their first in vitro fertilization treatment between 2006 and 2013. The main outcome measure was cumulative live births among the fresh and all vitrified blastocyst transfers combined after the same stimulation cycle. The results were also analyzed according to age (i.e., <35 years, 35-39 years, and ≥ 40 years). RESULTS: The mean number of retrieved oocytes was 19.9 ± 8.5 oocytes. The cumulative pregnancy rate was 89.2% and the cumulative live-birth rate was 73.3%. The cumulative live-birth rate declined from 73.9% for women younger than 35 years old to 67.3% for women 35-39 years old to 57.9% for women 40 years or older. CONCLUSION: Combined fresh and vitrified blastocyst transfer cycles can result in a high cumulative live-birth rate. The cumulative live-birth rates among older women are lower than the rates among younger women when autologous oocytes are used.

Diversity of Aquatic Pseudomonas Species and Their Activity against the Fish Pathogenic Oomycete Saprolegnia.

Emerging fungal and oomycete pathogens are increasingly threatening animals and plants globally. Amongst oomycetes, Saprolegnia species adversely affect wild and cultivated populations of amphibians and fish, leading to substantial reductions in biodiversity and food productivity. With the ban of several chemical control measures, new sustainable methods are needed to mitigate Saprolegnia infections in aquaculture. Here, PhyloChip-based community analyses showed that the Pseudomonadales, particularly Pseudomonas species, represent one of the largest bacterial orders associated with salmon eggs from a commercial hatchery. Among the Pseudomonas species isolated from salmon eggs, significantly more biosurfactant producers were retrieved from healthy salmon eggs than from Saprolegnia-infected eggs. Subsequent in vivo activity bioassays showed that Pseudomonas isolate H6 significantly reduced salmon egg mortality caused by Saprolegnia diclina. Live colony mass spectrometry showed that strain H6 produces a viscosin-like lipopeptide surfactant. This biosurfactant inhibited growth of Saprolegnia in vitro, but no significant protection of salmon eggs against Saprolegniosis was observed. These results indicate that live inocula of aquatic Pseudomonas strains, instead of their bioactive compound, can provide new (micro)biological and sustainable means to mitigate oomycete diseases in aquaculture.

Transforming growth factor-ß1 up-regulates connexin43 expression in human granulosa cells.

STUDY QUESTION: Does transforming growth factor-ß1 (TGF-ß1) up-regulate connexin43 (Cx43) to promote cell-cell communication in human granulosa cells? SUMMARY ANSWER: TGF-ß1 up-regulates Cx43 and increases gap junction intercellular communication activities (GJIC) in human granulosa cells, and this effect occurs via the activin receptor-like kinase (ALK)5-mediated Sma- and Mad-related protein (SMAD)2/3-SMAD4-dependent pathway. WHAT IS KNOWN ALREADY: TGF-ß1 and its receptors are expressed in human granulosa cells, and follicular fluid contains TGF-ß1 protein. In human granulosa cells, Cx43 gap junctions play an important role in the development of follicles and oocytes. STUDY DESIGN, SIZE, DURATION: This is an experimental study which was performed over a 1-year period. PARTICIPANTS/MATERIALS, SETTING, METHODS: Immortalized human granulosa cells (SVOG cells) and primary human granulosa-lutein cells obtained from women undergoing IVF in an academic research center were used as the study models. Cx43 mRNA and protein expression levels were examined after exposure of SVOG cells to recombinant human TGF-ß1. An activin/TGF-ß type I receptor inhibitor, SB431542, and small interfering RNAs targeting ALK4, ALK5, SMAD2, SMAD3 and SMAD4 were used to verify the specificity of the effects and to investigate the molecular mechanisms. Real-time-quantitative PCR and western blot analysis were used to detect the specific mRNA and protein levels, respectively. GJIC between SVOG cells were evaluated using a scrape loading and dye transfer assay. Results were analyzed by one-way analysis of variance. MAIN RESULTS AND THE ROLE OF CHANCE: TGF-ß1 treatment increased phosphorylation of SMAD2/3 (P < 0.0001) and up-regulated Cx43 mRNA and protein levels (P < 0.001) in SVOG cells and these stimulatory effects were abolished by the TGF-ß type I receptor inhibitor SB431542. In addition, the up-regulatory effect of TGF-ß1 on Cx43 expression (mRNA and protein) was confirmed in primary cultures of human granulosa-lutein cells (P < 0.05). The small interfering RNA-mediated knockdown of ALK5, but not ALK4, abolished the TGF-ß1-induced phosphorylation of SMAD2/3 and the up-regulation of Cx43. Furthermore, knockdown of SMAD2/3 or the common SMAD, SMAD4, abolished the stimulatory effects of TGF-ß1 on Cx43 expression in SVOG cells. The TGF-ß1-induced up-regulation of Cx43 contributed to the increase of GJIC between SVOG cells (P < 0.001). LIMITATIONS, REASONS FOR CAUTION: The results of this study were generated from in vitro system and may not reflect the intra-ovarian microenvironment in vivo. WIDER IMPLICATIONS OF THE FINDINGS: Our studies represent the first comprehensive research of molecular mechanisms of TGF-ß1 in the regulation of Cx43 expression and GJIC in human granulosa cells and demonstrate that TGF-ß1 may play a crucial role in the local modulation of cell-cell communication. Deepening our understanding of the molecular determinants will offer important insights into ovarian physiology and lead to the development of potential therapeutic methods for fertility regulation. STUDY FUNDING/COMPETING INTERESTS: This research was supported by an operating grant from the Canadian Institutes of Health Research to P.C.K.L. There are no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: NA.

Preimplantation genetic screening of blastocysts by multiplex qPCR followed by fresh embryo transfer: validation and verification.

BACKGROUND: Aneuploidy is an important etiology of implantation failure and quantitative real-time polymerase chain reaction (qPCR) seems a promising preimplantation genetic screening (PGS) technology to detect aneuploidies. This verification study aimed at verifying the impact on reproductive outcomes in in vitro fertilization (IVF) cycles using fresh embryo transfer (FET) in which the embryos were selected by blastocyst biopsy with qPCR-based PGS in our settings. RESULTS: A total of 13 infertile couples with more than once failed in vitro fertilization were enrolled during July to October of 2014. PGS was conducted by qPCR with selectively amplified markers to detect common aneuploidies (chromosomes 13, 18, 21, X, and Y). The design of the qPCR molecular markers adopted the locked nucleic acid (LNA) strategy. The blastocyst biopsy was performed on Day 5/6 and the PGS was done on the same day, which enabled FET. A total of 72 blastocysts were biopsied. Successful diagnoses were established in all embryos and the rate of successful diagnosis was 100 %. The aneuploidy rate was 38.9 % (28/72). 28 embryos were transferred. The clinical pregnancy rate was 61.5 % (8/13) per cycle. Early first trimester abortion was encountered in 1 and the ongoing pregnancy rate was 53.8 % (7/13) per cycle. CONCLUSION: This study verified the favorable outcome of adopting PGS with qPCR + FET in our own setting. Expanding the repertoire of aneuploidies being investigated (from a limited set to all 24 chromosomes) is underway and a randomized study by comparing qPCR and other PGS technologies is warranted.

Sleep deprivation enhances peripheral serotonin secretion to regulate the large follicle steroidogenesis of rats.

OBJECTIVE: Sleep deprivation (SD) leads to the disturbance of the estrous cycle. Serotonin, the levels of which increase with SD, has been shown to inhibit luteinizing hormone production and the receptor has been found in the follicles. In this study, the serotonin effect on preovulatory follicular steroidogenesis is investigated and the underlying mechanisms are elucidated. MATERIALS AND METHODS: Female rats were subjected to SD for a time span of 1-4 days using the dish-over-water-method with a Rechtschaffen apparatus. Serum estradiol and serotonin concentrations were assessed; thereafter, they were evaluated with the effect of serotonin on the estradiol production and steroidogenic acute regulatory (StAR) protein expression in a serum-free culture system. Preovulatory follicles were dissected mechanically from the ovaries of 21-day-old rats, which induced follicle growth, and cultured for 24 hours with or without recombinant human follicle-stimulating hormone (FSH) in the presence or absence of serotonin. RESULTS: SD, led to a significant decrease in serum estradiol concentrations, while serotonin concentrations were significantly elevated (all p < 0.05). Follicles were cultured with a constant dose of FSH (50 mIU/mL) and increasing doses of serotonin, estradiol production was reduced by 20%. The inhibitory effect of serotonin was concentration dependent. The addition of serotonin (0.1 µg/mL) decreased FSH-induced estradiol production and attenuated FSH-stimulated follicular StAR protein expression. The inhibitory effects of serotonin could be reduced by the serotonin receptor antagonist ketanserin. CONCLUSION: These findings suggest that decreased serum estradiol concentrations in SD rats may be the result of serotonin-related inhibition of estradiol production and decreased large follicle expression of StAR protein.

MS/MS networking guided analysis of molecule and gene cluster families.

The ability to correlate the production of specialized metabolites to the genetic capacity of the organism that produces such molecules has become an invaluable tool in aiding the discovery of biotechnologically applicable molecules. Here, we accomplish this task by matching molecular families with gene cluster families, making these correlations to 60 microbes at one time instead of connecting one molecule to one organism at a time, such as how it is traditionally done. We can correlate these families through the use of nanospray desorption electrospray ionization MS/MS, an ambient pressure MS technique, in conjunction with MS/MS networking and peptidogenomics. We matched the molecular families of peptide natural products produced by 42 bacilli and 18 pseudomonads through the generation of amino acid sequence tags from MS/MS data of specific clusters found in the MS/MS network. These sequence tags were then linked to biosynthetic gene clusters in publicly accessible genomes, providing us with the ability to link particular molecules with the genes that produced them. As an example of its use, this approach was applied to two unsequenced Pseudoalteromonas species, leading to the discovery of the gene cluster for a molecular family, the bromoalterochromides, in the previously sequenced strain P. piscicida JCM 20779(T). The approach itself is not limited to 60 related strains, because spectral networking can be readily adopted to look at molecular family-gene cluster families of hundreds or more diverse organisms in one single MS/MS network.