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Is it possible to develop a quantitative method for detecting parental DNA contamination in conventional IVF using preimplantation genetic testing for aneuploidy (PGT-A)?In this study, a quantification method was established for the parental contamination test (qPCT), which ensured more reliable results, and then verified its effectiveness for vitrified conventional IVF embryos. A total of 120 surplus vitrified blastocysts from patients who underwent prior routine IVF cycles were available for study.The results of the prospective clinical study of qPCT-PGT-A showed that the maternal contamination rate was 0.83% (1/120) and that the risk of paternal contamination was negligible. The 24 frozen embryo transfer cycles resulted in 16 clinical pregnancies, including 13 live births, one late inevitable miscarriage and two ongoing pregnancies.The risk of PGT in embryos with potential parental contamination is relatively low, and PGT-A is applicable for vitrified conventional IVF embryos.

Background：There is an urgent clinical need for preimplantation genetic testing (PGT) in frozen conventional human in vitro fertilization (IVF) embryos. However, it is difficult to detect parental contamination from sperm and cumulus cells in frozen conventional human IVF embryos during PGT with current methods. We established a quantification method for the parental contamination test, named qPCT, which ensures the results more reliable and substantially reduces the risk of PGT in frozen conventional IVF embryos.Methords：34 patients who had experienced repeated implantation failure or abortion due to chromosomal abnormalities after embryo transfer in prior routine IVF cycles. A total of 120 surplus frozen blastocysts from them were available for the study. Parental DNA contamination in the biopsied TE cells was detected via qPCT assay during PGT. To analyze the ploidy status of blastocysts, the amplified DNA of TE samples was sequenced using a NextSeq 550 sequencer with a single-ended read length of 55 bp..Result(s): A new quantification method for parental contamination testing (qPCT) in single-cell whole-genome amplification (WGA) products based on allelic ratio analysis was established and validated in an artificial model by comparing 22 results obtained before and after adding different numbers of sperm and cumulus cells to biopsied TE cells. The results of the prospective clinical study of qPCT-PGT-A showed that the maternal contamination rate was 0.83% (1/120) and the risk of paternal contamination was negligible. The euploidy rate in these blastocysts was 47.50% (57/120), and 21 frozen embryo transfer (FET) cycles resulted in ten ongoing clinical pregnancies and four healthy births.Conclusion(s): The evidence we provide in the study shows a low risk of PGT in embryos simultaneously with adhered sperm and cumulus cells. The qPCT assay can be used to detect the risk of potential contamination and ensure the accuracy of PGT results, thereby improving the clinical outcome of IVF.

ObjectiveTo establish a method to assess risks associated with preimplantation genetic testing (PGT) in embryos simultaneously with adhered sperm and cumulus cells.DesignA prospective pilot study.SettingUniversity teaching hospital.Patient(s)120 frozen blastocysts that could be biopsied from 34 patients who had experienced repeated implantation failure or abortion due to chromosomal abnormalities after embryos transfer in prior routine IVF cycles.Intervention(s)Chromosome screening and parental DNA contamination testing was performed in the surplus frozen IVF blastocysts from 34 patients.Main Outcome Measure(s)Parental DNA contamination rate and euploidy rate in biopsied blastocysts.Result(s)A new quantification method for parental contamination testing (qPCT) in single-cell whole-genome amplification (WGA) products based on allelic ratio analysis was established and validated in an artificial model by comparing 22 results obtained before and after adding different numbers of sperm and cumulus cells to biopsied TE cells. The results of the prospective clinical study of qPCT-PGT-A showed that the maternal contamination rate was 0.83% (1/120) and the risk of paternal contamination was negligible. The euploidy rate in these blastocysts was 47.50% (57/120), and 21 frozen embryo transfer (FET) cycles resulted in ten ongoing clinical pregnancies and four healthy births.Conclusion(s)The evidence we provide in the study shows a low risk of PGT in embryos simultaneously with adhered sperm and cumulus cells. The qPCT assay can be used to detect the risk of potential contamination and ensure the accuracy of PGT results, thereby improving the clinical outcome of IVF.CapsuleDuring PGT for human frozen conventional IVF embryos, paternal source pollution is negligible, while maternal pollution can not be ignored. qPCT method can effectively detect the parent DNA contamination in WGA products of biopsied TE cells.

Emerging studies have demonstrated the promising clinical value of circulating tumor cells (CTCs) for diagnosis, disease assessment, treatment monitoring and prognosis in epithelial ovarian cancer. However, the clinical application of CTC remains restricted due to diverse detection techniques with variable sensitivity and specificity and a lack of common standards.We enrolled 160 patients with epithelial ovarian cancer as the experimental group, and 90 patients including 50 patients with benign ovarian tumor and 40 healthy females as the control group. We enriched CTCs with immunomagnetic beads targeting two epithelial cell surface antigens (EpCAM and MUC1), and used multiple reverse transcription-polymerase chain reaction (RT-PCR) detecting three markers (EpCAM, MUC1 and WT1) for quantification. And then we used a binary logistic regression analysis and focused on EpCAM, MUC1 and WT1 to establish an optimized CTC detection model.The sensitivity and specificity of the optimized model is 79.4% and 92.2%, respectively. The specificity of the CTC detection model is significantly higher than CA125 (92.2%Our study presents an optimized detection model for CTCs, which consists of the expression levels of three markers (EpCAM, MUC1 and WT1). In comparison with CA125, our model has high specificity and demonstrates better diagnostic values, especially for early-stage ovarian cancer. Detection of CTC

Objective: DUSP6 is a negative regulator of the ERK signaling pathway and plays an important role in chemotherapy-resistance. Previously we showed that DUSP6 is overexpressed in ovarian cancer side population (SP) cells that possess cancer stem cell-like properties and are quiescent and chemotherapy-resistant. Here, we explore the effects of DUSP6 on chemotherapy-resistance by examining its regulation of the ERK signaling pathway and G0/G1 cell cycle arrest. Methods: mRNA and protein expression of DUSP6 and G0/G1 cell cycle checkpoint regulating proteins (CyclinD1, CyclinD3 and CyclinE2) was evaluated among ovarian cancer cell lines and tissue samples. Ovarian cancer cells were transiently transfected to overexpress DUSP6. After treatment with cisplatin, cell viability was measured by the MTS assay at 48 hours and the half maximal inhibitory concentration (IC50) for each cell line was calculated. Subcellular localization and cell cycle analysis were determined by using immunofluorescence and FACS, respectively. Results: SKOV3 and OVCAR8 SP cells were shown to express higher levels of DUSP6 and lower levels of CyclinD3 compared with non-SP (NSP) cells (P

BackgroundHigh grade serous ovarian cancer (HGSOC) is the most common subtype of ovarian cancer. Although platinum-based chemotherapy has been the cornerstone for HGSOC treatment, nearly 25% of patients would have less than 6 months of interval since the last platinum chemotherapy, referred to as platinum-resistance. Currently, no precise tools to predict platinum resistance have been developed yet.MethodsNinety-nine HGSOC patients, who have finished cytoreductive surgery and platinum-based chemotherapy in Peking University Third Hospital from 2018 to 2019, were enrolled. Whole-genome sequencing (WGS) and whole-exome sequencing (WES) were performed on the collected tumor tissue samples to establish a platinum-resistance predictor in a discovery cohort of 57 patients, and further validated in another 42 HGSOC patients.ResultsA high prevalence of alterations in DNA damage repair (DDR) pathway, including BRCA1/2, was identified both in the platinum-sensitive and resistant HGSOC patients. Compared with the resistant subgroup, there was a trend of higher prevalence of homologous recombination deficiency (HRD) in the platinum-sensitive subgroup (78.95% vs. 47.37%, p=0.0646). Based on the HRD score, microhomology insertions and deletions (MHID), copy number changes load, duplication load of 1–100 kb, single nucleotide variants load, and eight other mutational signatures, a combined predictor of platinum-resistance, named as DRDscore, was established. DRDscore outperformed in predicting the platinum-sensitivity than the previously reported biomarkers with a predictive accuracy of 0.860 at a threshold of 0.7584. The predictive performance of DRDscore was validated in an independent cohort of 42 HGSOC patients with a sensitivity of 90.9%.ConclusionsA multi-genomic signature-based analysis enabled the prediction of initial platinum resistance in advanced HGSOC patients, which may serve as a novel assessment of platinum resistance, provide therapeutic guidance, and merit further validation.

Copper- and nitrogen-codoped reduced graphene oxide material (Cu/N-rGO) was prepared with a hydrothermal method. Its versatile catalytic performances were demonstrated toward the oxidative degradation of rhodamine B (RhB) and oxygen reduction reaction (ORR). The Cu and N codoping of graphene enhanced not only its activation ability toward H2O2, but also its electrocatalytic ability for ORR. It was observed that the use of 3%Cu/N-rGO together with 40 mmol&middot, L&minus, 1 H2O2 and 4 mmol&middot, 1 Na2CO3 could remove more than 94% of the added RhB (30 mg&middot, 1) in 20 min through a catalytic Fenton-like degradation. Quenching experiments and electron paramagnetic resonance (EPR) measurements indicated that the main reactive species generated in the catalytic oxidation process were surface-bound &bull, OH. The modified graphene also showed good electrocatalytic activity for ORR reaction in alkaline media through a four-electron mechanism. On the electrode of Cu/N-rGO, the ORR reaction exhibited an onset potential of &minus, 0.1 V and a half-wave potential of &minus, 0.248 V, which were correspondingly close to those on a Pt/C electrode. In comparison with a Pt/C electrode, the 3%Cu/N-rGO electrode showed much greater tolerance to methanol. Such outstanding catalytic properties are attributed to the abundant active sites and the synergism between Cu and N in Cu/N-rGO.

To investigate the correlation of circulating long non-coding RNA nuclear-enriched abundant transcript 1 (lncRNA NEAT1) expression with disease risk, severity, prognosis and inflammatory cytokine levels in sepsis patients.152 sepsis patients and 150 health controls (HCs) were enrolled in this study. Plasma and serum samples were obtained from sepsis patients and HCs, and lncRNA NEAT1 expression in plasma was determined by quantitative polymerase chain reaction, while levels of inflammatory cytokines in serum were detected by enzyme linked immune sorbent assay.LncRNA NEAT1 expression was remarkably higher in sepsis patients than in HCs (P 0.001). Receiver operating characteristic (ROC) curve disclosed a good predictive value of lncRNA NEAT1 expression for sepsis risk with area under curve (AUC) of 0.730 (95% CI: 0.740-0.861). Subsequent multivariate logistic regression analysis demonstrated that lncRNA NEAT1 expression was independently associated with higher sepsis risk (P 0.001). In sepsis patients, lncRNA NEAT1 expression was also observed to be positively correlated with Acute Physiology and Chronic Health Evaluation (APACHE) II score (P 0.001), serum tumor necrosis factor-α (P 0.001), interleukin (IL)-1β (P = 0.043), IL-6 (P = 0.001) and IL-8 (P = 0.038), while negatively correlated with IL-10 (P 0.001). In addition, lncRNA NEAT1 expression was increased in non-survivors compared to survivors (P = 0.006), and ROC curve revealed a good prognostic value of lncRNA NEAT1 for non-survivor risk with AUC 0.641 (95% CI: 0.536-0.746).Circulating lncRNA NEAT1 correlates with increased disease risk, elevated severity and unfavorable prognosis as well as higher expression of pro-inflammatory cytokines in sepsis patients.

The novel thiacalix[4]-1,3-2,4-aza-biscrown 3 in 1,3-alternate conformation, was facilely prepared by “1 + 2” cyclocondensation of p-tert-butylthiacalix[4]arene tetrahydrazide derivative 2 with o-phthalaldehyde in yield of 78%. The liquid–liquid extraction experiment showed that compound 3 was excellent receptor for zwitterionic α-amino acids and soft cations Ag+ and Hg2+. The extraction percentage of methionine was as high as 78%.

To evaluate the incidence, predisposing factors, early diagnosis and treatment options of heterotopic pregnancy (HP) following in vitro fertilization and embryo transfer (IVF-ET) procedure. A retrospective review study was performed to identify the HP cases after IVF-ET at the Reproductive Centre in Guangdong Women and Children’s Hospital in China between the years of 2002–2007. Twelve out of 1,476 pregnancies (0.81%) were diagnosed for HP, of which nine patients elected for exploratory salpingectomy, two patients received selective fetal reduction by embryo aspiration under ultrasound guidance, and one patient opted for expectant treatment. Postoperatively, four intrauterine pregnancies were continued to develop until term while two were delivered at 35 weeks of gestation. The achievement ratio of continuous pregnancy was 66.7% (6/9). The incidence of HP is increasing due to the widespread use of assisted reproductive technology. An early transvaginal sonography performed by experienced radiologist/radiographer is considered to be essential and beneficial in establishing early diagnosis of HP. Both salpingectomy and selective fetal reduction by embryo aspiration can be administered as one of the effective therapies for HP with the optimal outcome of intrauterine pregnancy.