Parental methylome reprogramming in human uniparental blastocysts reveals germline memory transition.

Uniparental embryos derived from only the mother (gynogenetic [GG]) or the father (androgenetic [AG]) are unique models for studying genomic imprinting and parental contributions to embryonic development. Human parthenogenetic embryos can be obtained following artificial activation of unfertilized oocytes, but the production of AG embryos by injection of two sperm into one denucleated oocyte leads to an extra centriole, resulting in multipolar spindles, abnormal cell division, and developmental defects. Here, we improved androgenote production by transferring the male pronucleus from one zygote into another haploid androgenote to prevent extra centrioles and successfully generated human diploid AG embryos capable of developing into blastocysts with an identifiable inner cell mass (ICM) and trophectoderm (TE). The GG embryos were also generated. The zygotic genome was successfully activated in both the AG and GG embryos. DNA methylome analysis showed that the GG blastocysts partially retain the oocyte transcription-dependent methylation pattern, whereas the AG blastocyst methylome showed more extensive demethylation. The methylation states of most known imprinted differentially methylated regions (DMRs) were recapitulated in the AG and GG blastocysts. Novel candidate imprinted DMRs were also identified. The production of uniparental human embryos followed by transcriptome and methylome analysis is valuable for identifying parental contributions and epigenome memory transitions during early human development.

Publisher Correction: Chromatin analysis in human early development reveals epigenetic transition during ZGA.

In this Letter, the 'Open chromatin' label in Fig. 4a should have been centred above the first three columns, and the black horizontal line underneath the label should have been removed. In addition, there should have been a vertical black line between the last two sets of panels for consistency. Minor changes have also been made to Fig. 1 and to the legend of Fig. 3. These errrors have been corrected online, and see Supplementary Information to the accompanying Amendment for the original Fig. 4.

Chromatin analysis in human early development reveals epigenetic transition during ZGA.

Upon fertilization, drastic chromatin reorganization occurs during preimplantation development 1 . However, the global chromatin landscape and its molecular dynamics in this period remain largely unexplored in humans. Here we investigate chromatin states in human preimplantation development using an improved assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) 2 . We find widespread accessible chromatin regions in early human embryos that overlap extensively with putative cis-regulatory sequences and transposable elements. Integrative analyses show both conservation and divergence in regulatory circuitry between human and mouse early development, and between human pluripotency in vivo and human embryonic stem cells. In addition, we find widespread open chromatin regions before zygotic genome activation (ZGA). The accessible chromatin loci are readily found at CpG-rich promoters. Unexpectedly, many others reside in distal regions that overlap with DNA hypomethylated domains in human oocytes and are enriched for transcription factor-binding sites. A large portion of these regions then become inaccessible after ZGA in a transcription-dependent manner. Notably, such extensive chromatin reorganization during ZGA is conserved in mice and correlates with the reprogramming of the non-canonical histone mark H3K4me3, which is uniquely linked to genome silencing3-5. Taken together, these data not only reveal a conserved principle that underlies the chromatin transition during mammalian ZGA, but also help to advance our understanding of epigenetic reprogramming during human early development and in vitro fertilization.

Resolution enhancement via guided filtering for spatial-frequency multiplexing single-shot high-speed imaging.

The frequency recognition algorithm for multiple exposures (FRAME) is a progressive single-shot high-speed videography technique that employs the spatial-frequency multiplexing concept to provide high temporal and spatial resolution. However, the inherent crosstalk from the zero-frequency component to the carrier-frequency component leads to resolution degradation and artifacts. To improve recovered frames' quality, we propose a FRAME reconstruction method using guided filters for a removal of the zero-frequency component, which can minimize the artifacts while enhance spatial resolution. A total variation (TV) denoising operation is involved to remove artifacts further to achieve optimized performances. Simulations and experiments were conducted to demonstrate the robust and efficient post-processing capability of the proposed method. With a two-frame experimental system, the results of a USAF 1951 resolution target reveal a 1.8-fold improvement in spatial resolution from 16 lp/mm to 28.5 lp/mm. For complex dynamic scenarios, the wide field of high-speed fuel spray was shot and the proposed method can resolve two droplets with a 30 µm distance which outperforms the traditional method.

Effective protection: the embryonic development and clinical outcomes of emergency vitrification of 1246 oocytes and Day 0-Day 5 embryos in a natural disaster.

STUDY QUESTION: Can emergency vitrification protect embryos and oocytes during natural disasters or other events that prevent normal practice to achieve satisfactory embryonic development and clinical outcomes at a later time? SUMMARY ANSWER: Emergency vitrification of oocytes and Day 0-Day 5 (D0-D5) embryos during disasters is a safe and effective protective measure. WHAT IS KNOWN ALREADY: When some destructive events such as floods, earthquakes, tsunamis, and other accidents occur, emergency vitrification in embryo laboratories to protect human embryos, oocytes, and sperm is one of the important measures of an IVF emergency plan. However, there are few detailed reports on emergency vitrification in a state of disaster, especially about oocytes and D0 zygotes. Therefore, the effectiveness and safety of emergency vitrification of oocytes and D0-D5 embryos in disaster states are still unclear. STUDY DESIGN, SIZE, DURATION: A retrospective study was made in the Reproductive Medicine Center of the First Affiliated Hospital of Zhengzhou University from January 2018 to November 2022. The record rainstorms in Zhengzhou, China, caused severe flooding, traffic disruptions, and power outages. From 17:30, 20 July 2021 to 17:30, 21 July 2021, 1246 oocytes and D0-D5 embryos of 155 patients were vitrified whilst the laboratory had only an emergency power supply. PARTICIPANTS/MATERIALS, SETTING, METHODS: As of 21 December 2021, 1149 emergency vitrified oocytes and D0-D5 embryos of 124 patients underwent frozen-thawed embryo transfer (FET). They were divided into the following four groups according to the days of embryo culture in vitro: oocyte group, Day 0-Day 1 (D0-D1) group, Day 2-Day 3 (D2-D3) group, and Day 4-Day 5 (D4-D5) group. Control groups for each were selected from fresh cycle patients who underwent IVF/ICSI from January 2018 to October 2021. Control and emergency vitrification patients were matched on criteria that included age, fertilization method, days of embryonic development, and number and grade of transferred embryos. A total of 493 control patients were randomly selected from the eligible patients and matched with the emergency vitrification groups in a ratio of 4:1. The results of assisted reproduction and follow-up of pregnancy were analyzed. The embryonic development, clinical outcomes, and birth outcomes in each group were statistically analyzed. MAIN RESULTS AND THE ROLE OF CHANCE: A significant difference was observed in fertilization rate (81% versus 72%, P = 0.022) between the oocyte group and the control group. Significant differences were also observed in the monozygotic twin pregnancy rate (10% versus 0%, P = 0.038) and ectopic pregnancy rate (5% versus 0%, P = 0.039) between the D0-D1 group and the control group. No significant differences (P > 0.05) were observed between vitrified oocytes/D0-D1 embryos/D2-D3 embryos and the control group on the number of high-quality embryos (3.17 ± 3.00 versus 3.84 ± 3.01, P = 0.346; 5.04 ± 3.66 versus 4.56 ± 2.87, P = 0.346; 4.85 ± 5.36 versus 5.04 ± 4.64, P = 0.839), the number of usable blastocysts (1.22 ± 1.78 versus 1.21 ± 2.03, P = 0.981; 2.16 ± 2.26 versus 1.55 ± 2.08, P = 0.090; 2.82 ± 3.23 versus 2.58 ± 3.32, P = 0.706), clinical pregnancy rate (56% versus 57%, P = 0.915; 55% versus 55%, P = 1.000; 40% versus 50%, P = 0.488), miscarriage rate (30% versus 15%, P = 0.496; 5% versus 11%, P = 0.678; 17% versus 20%, P = 1.000), and live birth rate (39% versus 49%, P = 0.460; 53% versus 50%, P = 0.772; 33% versus 40%, P = 0.635). No significant differences (P > 0.05) were observed between the D4-D5 group and the control group on clinical pregnancy rate (40% versus 55%, P = 0.645), miscarriage rate (0% versus 18%, P = 1.000), and live birth rate (40% versus 45%, P = 1.000). LIMITATIONS, REASONS FOR CAUTION: The retrospective study design is a limitation. The timing and extent of natural disasters are unpredictable, so the sample size of vitrified oocytes, zygotes, and embryos is beyond experimental control. WIDER IMPLICATIONS OF THE FINDINGS: This study is the first study analyzing embryonic development, clinical outcomes, and birth outcomes of large samples of oocytes, D0 zygotes, and D1-D5 embryos after emergency vitrification under the disaster conditions. The results show that emergency vitrification is a safe and effective protective measure applicable to oocytes and D0-D5 embryos. The embryology laboratories need to be equipped with an emergency uninterrupted power supply capable of delivering for 6-8 h at full load. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Natural Science Foundation of China (grant 81871206). The authors declare that they have no conflicts of interest. All authors have completed the ICMJE Disclosure form. TRIAL REGISTRATION NUMBER: N/A.

The preimplantation genetic testing for monogenic disorders strategy for blocking the transmission of hereditary cancers through haplotype linkage analysis by karyomapping.

PURPOSE: Providing feasible preimplantation genetic testing strategies for monogenic disorders (PGT-M) for prevention and control of genetic cancers. METHODS: Inclusion of families with a specific pathogenic mutation or a clear family history of genetic cancers. Identification of the distribution of hereditary cancer-related mutations in families through genetic testing. After a series of assisted reproductive measures such as down-regulation, stimulation, egg retrieval, and in vitro fertilization, a biopsy of trophectoderm cells from a blastocyst was performed for single-cell level whole-genome amplification (WGA). Then, the detection of chromosomal aneuploidies was performed by karyomapping. Construction of a haplotype-based linkage analysis to determine whether the embryo carries the mutation. Meanwhile, we performed CNV testing. Finally, embryos can be selected for transfer, and the results will be verified in 18-22 weeks after pregnancy. RESULTS: Six couples with a total of 7 cycles were included in our study. Except for cycle 1 of case 5 which did not result in a transferable embryo, the remaining 6 cycles produced transferable embryos and had a successful pregnancy. Four couples have had amniotic fluid tests to confirm that the fetus does not carry the mutation, while 1 couple was not tested due to insufficient pregnancy weeks. And the remaining couples had to induce labor due to fetal megacystis during pregnancy. CONCLUSION: Our strategy has been proven to be feasible. It can effectively prevent transmission of hereditary cancer-related mutations to offspring during the prenatal stage.

Identification of carrier status of Xp22.31 microdeletions associated with X-linked ichthyosis at the single-cell level using haplotype linkage analysis by karyomapping.

PURPOSE: Currently, owing to the limitations of high-throughput sequencing depth and the allele dropout caused by the whole-genome amplification, detection of chromosomal variants in embryos with CNVs <5 Mb is unsatisfactory at the single-cell level using only conventional sequencing methods. Therefore, here we aimed to use a strategy of preimplantation genetic testing for monogenic (PGT-M) to compensate for the shortcomings of conventional sequencing methods. The purpose of this study is to report the effectiveness of haplotype linkage analysis by karyomapping for preimplantation diagnosis microdeletion diseases. METHODS: Six couples carrying chromosomal microdeletions associated with X-linked ichthyosis were recruited, and all couples entered the PGT process. Multiple displacement amplification (MDA) method was used to amplify the whole-genome DNA of trophectoderm cells. Then karyomapping based on single nucleotide polymorphism (SNP) was used for haplotype linkage analysis to detect alleles carrying microdeletions, and CNVs of embryos were identified to determine euploid identity. Amniotic fluid tests were performed in the second trimester to verify the PGT-M results. RESULTS: All couples were tested for chromosomal microdeletions, with deletion fragments ranging in size from 1.60 to 1.73 Mb, and one partner in each couple did not carry the microdeletion. Three couples successfully underwent PGT-M assisted conception and obtained healthy live births. CONCLUSION: This study shows that haplotype linkage analysis by karyomapping could effectively detect the carrier status of embryos with microdeletions at the single-cell level. This approach may be applied to the preimplantation diagnosis of various chromosomal microvariation diseases.

Adverse maternal and neonatal outcomes of preimplantation genetic testing with trophectoderm biopsy: a retrospective cohort study of 3373 intracytoplasmic sperm injection single frozen-thawed blastocyst transfer cycles.

PURPOSE: To investigate whether trophectoderm biopsy increases the risk of adverse maternal and neonatal outcomes in intracytoplasmic sperm injection (ICSI) single frozen-thawed blastocyst transfer cycles. METHODS: This respective cohort study enrolled 3373 ICSI single frozen-thawed blastocyst transfer cycles with and without trophectoderm biopsy. Statistical methods including univariate logistic regression analysis, multivariate logistic regression analysis, and stratified analyses were performed to explore the impact of trophectoderm biopsy on adverse maternal and neonatal outcomes. RESULTS: The rates of adverse maternal and neonatal outcomes were comparable between the two groups. Univariate analysis showed that the live birth rate (45.15% vs. 40.75%; P = 0.010) in the biopsied group was statistically higher than that in the unbiopsied group, and the rates of miscarriage (15.40% vs. 20.00%; P = 0.011) and birth defects (0.58% vs. 2.16%; P = 0.007) were statistically lower in the biopsied group. After adjusting for confounding factors, the rates of miscarriage (aOR = 0.74; 95% CI = 0.57-0.96; P = 0.022) and birth defects (aOR = 0.24, 95% CI = 0.08-0.70, P = 0.009) in the biopsied group were significantly lower than those in the unbiopsied group. Stratified analyses showed that the birth defects rate after biopsy was significantly reduced in the subgroups of age < 35 years old, BMI ≥ 24 kg/m2, artificial cycle with downregulation, poor-quality blastocysts, and Day 5 poor-quality blastocysts. CONCLUSION: Preimplantation genetic testing (PGT) with trophectoderm biopsy does not increase the risk of adverse maternal and neonatal outcomes in ICSI single frozen-thawed blastocyst transfer cycles, and PGT can effectively reduce the rates of miscarriage and birth defects.

The Molecular Basis of Catalysis by SDR Family Members Ketoacyl-ACP Reductase FabG and Enoyl-ACP Reductase FabI in Type-II Fatty Acid Biosynthesis.

The short-chain dehydrogenase/reductase (SDR) superfamily members acyl-ACP reductases FabG and FabI are indispensable core enzymatic modules and catalytic orientation controllers in type-II fatty acid biosynthesis. Herein, we report their distinct substrate allosteric recognition and enantioselective reduction mechanisms. FabG achieves allosteric regulation of ACP and NADPH through ACP binding across two adjacent FabG monomers, while FabI follows an irreversible compulsory order of substrate binding in that NADH binding must precede that of ACP on a discrete FabI monomer. Moreover, FabG and FabI utilize a backdoor residue Phe187 or a "rheostat" α8 helix for acyl chain length selection, and their corresponding triad residues Ser142 or Tyr145 recognize the keto- or enoyl-acyl substrates, respectively, facilitating initiation of nucleophilic attack by NAD(P)H. The other two triad residues (Tyr and Lys) mediate subsequent proton transfer and (R)-3-hydroxyacyl- or saturated acyl-ACP production.

miR-188-3p-targeted regulation of ATG7 affects cell autophagy in patients with nonobstructive azoospermia.

BACKGROUND: Nonobstructive azoospermia (NOA) is one of the most difficult forms of male infertility to treat, and its pathogenesis is still unclear. miRNAs can regulate autophagy by affecting their target gene expression. Our previous study found that miR-188-3p expression in NOA patients was low. There are potential binding sites between the autophagy gene ATG7 and miR-188-3p. This study aimed to verify the binding site between miR-188-3p and ATG7 and whether miR-188-3p affects autophagy and participates in NOA by regulating ATG7 to influence the autophagy marker genes LC3 and Beclin-1. METHODS: Testicular tissue from 16 NOA patients and 16 patients with normal spermatogenesis and 5 cases in each group of pathological sections were collected. High-throughput sequencing was performed to detect mRNA expression differences. Quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting, immunohistochemical staining and immunofluorescence were used to detect protein localization and expression. Autophagosome changes were detected by electron microscopy. The targeting relationship between miR-188-3p and ATG7 was confirmed by a luciferase assay. RESULTS: ATG7 protein was localized in the cytoplasm of spermatogenic cells at all levels, and the ATG7 gene (p = 0.019) and protein (p = 0.000) were more highly expressed in the NOA group. ATG7 expression after overexpression/inhibition of miR-188-3p was significantly lower (p = 0.029)/higher (p = 0.021) than in the control group. After overexpression of miR-188-3p, the ATG7 3'UTR-WT luciferase activity was impeded (p = 0.004), while the ATG7 3'UTR-MUT luciferase activity showed no significant difference (p = 0.46). LC3 (p = 0.023) and Beclin-1 (p = 0.041) expression in the NOA group was significantly higher. LC3 and Beclin-1 gene expression after miR-188-3p overexpression/inhibition was significantly lower (p = 0.010 and 0.024, respectively) and higher (p = 0.024 and 0.049, respectively). LC3 punctate aggregation in the cytoplasm decreased after overexpression of miR-188-3p, while the LC3 punctate aggregation in the miR-188-3p inhibitor group was higher. The number of autophagosomes in the miR-188-3p mimic group was lower than the number of autophagosomes in the mimic NC group. CONCLUSIONS: LC3 and Beclin-1 were more highly expressed in NOA testes and negatively correlated with the expression of miR-188-3p, suggesting that miR-188-3p may be involved in the process of autophagy in NOA. miR-188-3p may regulate its target gene ATG7 to participate in autophagy anDual luciferase experiment d affect the development of NOA.

A novel monogenic preimplantation genetic testing strategy for sporadic polycystic kidney caused by de novo PKD1 mutation.

Autosomal dominant hereditary polycystic kidney disease (ADPKD) is the most common inherited kidney disease that causes end-stage renal disease and kidney failure. Preimplantation genetic testing for monogenic (PGT-M) can effectively prevent the transmission of genetic diseases from parents to the offspring before pregnancy. However, PGT-M currently adopts the single nucleotide polymorphism (SNP) linkage analysis for embryo's pathogenic gene carrying status and linkage analysis requires proband of the family. Here we report a new PGT-M strategy using single sperm SNP linkage analysis for male patient with sporadic ADPKD caused by de novo PKD1 mutation. We recruited five couples with male patient with ADPKD caused by de novo PKD1 mutation, and 39 embryos from six PGT-M cycles were detected. The five couples had at least one embryo that does not carry the PKD1 mutation. Within these five couples, the accuracy of carrier status of embryos was confirmed by amniotic fluid gene detection of two couples and two couples successfully delivered healthy fetuses. Therefore, the new PGT-M strategy of using single sperm SNP linkage analysis was proved to be feasible and effective for male patient with ADPKD caused by de novo PKD1 mutation.

Dynamic changes in clinical and CT characteristics of COVID-19 cases with different exposure histories: a retrospective study.

BACKGROUND: To assess the dynamic changes in clinical and CT characteristics of COVID-19 patients with different epidemiology histories. METHODS: Fifty-three discharged COVID-19 patients were enrolled at Beijing YouAn Hospital, Capital Medical University, between January 21 and March 10, 2020. Spearman correlation analysis was performed between CT scores and laboratory indicators. Patients were divided into the Wuhan group (lived in or with travel to Wuhan, numbering 30 cases) and non-Wuhan group (close contacts or unknown exposure, totaling 23 cases). The CT and laboratory findings were compared between and within groups during the clinical process. RESULTS: Fever (88.7%), cough (64.2%), fatigue (34%), and abnormal laboratory indicators, including lymphopenia, reduced albumin, albumin/globulin (A/G), and elevated C-reactive protein (CRP), were mainly observed. Subpleural ground-glass opacities (86.8%) were usually detected at admission. The CT scores were highly correlated with lymphocytes, CRP, albumin, and A/G at initial and follow-ups (all p < 0.05). Four days after admission, most patients (66.7% Wuhan, 47.8% non-Wuhan) showed progression, and the CT scores of Wuhan significantly increased (p = 0.015). Eight days after admission, the vast majority of patients (69.2% Wuhan, 100% non-Wuhan, p = 0.006) presented improvement, and the CT scores of non-Wuhan were significantly lower than Wuhan (p = 0.006). Pneumonia was completely absorbed in most patients 2-4 weeks after discharge. CONCLUSIONS: CT plays a crucial role in the early diagnosis and monitoring of changes in COVID-19. Lymphocytes, CRP, albumin, and A/G are expected to predict disease severity and prognosis. Viral pathogenicity in non-endemic areas may be weaker than core-infected areas. In most patients, lung lesions can disappear around 4 weeks after discharge.

Improved clinical outcomes of preimplantation genetic testing for aneuploidy using MALBAC-NGS compared with MDA-SNP array.

BACKGROUND: To assess whether preimplantation genetic testing for aneuploidy with next generation sequencing (NGS) outweighs single nucleotide polymorphism (SNP) array in improving clinical outcomes. METHODS: A retrospective analysis of the clinical outcomes of patients who underwent PGT-A treatment in a single center from January 2013 to December 2017.A total of 1418 couples who underwent PGT-A treatment were enrolled, of which 805 couples used NGS for PGT-A, while the remaining 613 couples used SNP array for PGT-A. Clinical pregnancy rate, miscarriage rate and healthy baby rate were compared between the MALBAC-NGS-PGT-A and MDA-SNP-PGT-A groups. RESULTS: After testing karyotypes of 5771 biopsied blastocysts, 32.2% (1861/5771) were identified as chromosomally normal, while 67.8% were chromosomally abnormal. In terms of clinical outcomes, women in the MALBAC-NGS-PGT-A group had a significantly higher clinical pregnancy rate (50.5% vs 41.7%, p = 0.002) and healthy baby rate (39.6% vs 31.4%, p = 0.003), and a lower miscarriage rate (15.5% vs 22.8%, p = 0.036). CONCLUSION: This is the largest study reporting the extensive application of NGS-based PGT-A, whilst comparing the clinical outcomes of MALBAC-NGS-PGT-A and MDA-SNP-PGT-A. The results provide greater evidence supporting the wider use of NGS in PGT-A, not only for its lower cost but also for its improved clinical outcomes compared to SNP-based PGT-A.

Mapping allele with resolved carrier status of Robertsonian and reciprocal translocation in human preimplantation embryos.

Reciprocal translocations (RecT) and Robertsonian translocations (RobT) are among the most common chromosomal abnormalities that cause infertility and birth defects. Preimplantation genetic testing for aneuploidy using comprehensive chromosome screening for in vitro fertilization enables embryo selection with balanced chromosomal ploidy; however, it is normally unable to determine whether an embryo is a translocation carrier. Here we report a method named "Mapping Allele with Resolved Carrier Status" (MaReCs), which enables chromosomal ploidy screening and resolution of the translocation carrier status of the same embryo. We performed MaReCs on 108 embryos, of which 96 were from 13 RecT carriers and 12 were from three RobT carriers. Thirteen of the sixteen patients had at least one diploid embryo. We have confirmed the accuracy of our carrier status determination in amniotic fluid karyotyping of seven cases as well as in the live birth we have thus far. Therefore, MaReCs accurately enables the selection of translocation-free embryos from patients carrying chromosomal translocations. We expect MaReCs will help reduce the propagation of RecT/RobT in the human population.

Live births following preimplantation genetic testing for dynamic mutation diseases by karyomapping: a report of three cases.

PURPOSE: The preimplantation genetic testing for monogenic defects (PGT-M) is a beneficial strategy for the patients suffering from a Mendelian disease, which could protect their offspring from inheriting the disease. The purpose of this study is to report the effectiveness of PGT-M based on karyomapping for three cases of dynamic mutation diseases with trinucleotide repeat expansion. METHODS: PGT-M was carried out on three couples, whose family members were diagnosed with Huntington's disease or spinocerebellar ataxias 2 or 12. The whole genome amplification was obtained using the multiple displacement amplification (MDA) method. Then, karyomapping was performed to detect the allele that is carrying the trinucleotide repeat expansion using single nucleotide polymorphism (SNP) linkage analyses, and the copy number variations (CNVs) of the embryos were also identified. Prenatal diagnosis was performed to validate the accuracy of PGT-M. RESULTS: PGT-M was successfully performed on the three couples, and they accepted the transfers of euploid blastocysts without the relevant pathogenic allele. The clinical pregnancies were acquired and the prenatal diagnosis of the three families confirmed the effectiveness of karyomapping. The three born babies were healthy and free of the pathogenic alleles HTT, ATXN2, or PPP2R2B corresponding to Huntington's disease, spinocerebellar ataxias 2 or 12, respectively. CONCLUSION: This study shows that karyomapping is a highly powerful and efficient approach for dynamic mutation detection in preimplantation embryos. In this work, we first report the birth of healthy babies that are free of the pathogenic gene for dynamic mutation diseases in patients receiving PGT-M by karyomapping.

Quantitative Association Between Serum/Dietary Magnesium and Cardiovascular Disease/Coronary Heart Disease Risk: A Dose-Response Meta-analysis of Prospective Cohort Studies.

BACKGROUND: The quantitative association between serum/dietary magnesium and cardiovascular disease (CVD) remains unclear. We conducted a dose-response meta-analysis to evaluate the quantitative association between serum/dietary magnesium and CVD, including coronary heart disease (CHD). METHODS: PubMed, China National Knowledge Infrastructure, and Web of Science were searched for publications. STATA 12.0 was used to analyze data. We used the random-effects model to reduce heterogeneity. RESULTS: Eighteen prospective cohort studies with 544,581 participants and 22,658 CVD cases were included. The follow-up duration was 1-28 years. The pooled relative risk (RR) of CVD for the relatively normal versus lowest serum and dietary magnesium level was 0.64 {[95% confidence interval (CI): 0.51-0.80] and 0.90 [95% CI: 0.84-0.96]}. The pooled RR of CHD for the relatively normal versus lowest serum and dietary magnesium level was 0.70 (95% CI: 0.57-0.85) and 0.86 (95% CI: 0.77-0.94). We noted a significant association between increasing serum magnesium levels (per 0.1-mg/dL increase) and risk of CVD (RR: 0.93, 95% CI: 0.88-0.97) and CHD (RR: 0.90, 95% CI: 0.84-0.96) and between dietary magnesium levels (per 100-mg/d increase) and risk of CVD (RR: 0.90, 95% CI: 0.83-0.96) and CHD (RR: 0.92, 95% CI: 0.82-0.98). Serum/dietary Mg level comparisons presented a 7%-10% decrease in CVD/CHD risk. The dose-response meta-analyses showed linear relationships between serum magnesium and CVD (Pnonlinearity = 0.833) or CHD (Pnonlinearity = 0.193) and dietary magnesium and CVD (Pnonlinearity = 0.463) or CHD (Pnonlinearity = 0.440). CONCLUSIONS: Increasing dietary magnesium or serum magnesium level is linearly and inversely associated with the risk of total CVD and CHD events.

Analysis of Related Causes for No Embryos Transferred and Corresponding Coping Measures in Assisted Reproductive Technology.

OBJECTIVE: To analyze the related causes for no embryos transferred in assisted reproductive technology (ART) in order to provide corresponding coping measures for infertile couples. STUDY DESIGN: The data of 607 couples who underwent ART and had no embryos transferred in our reproductive center between January 2010 and January 2014 were retrospectively analyzed. RESULTS: The cycles of no embryos transferred accounted for 3.99% (607/15,224) of total cycles. Of those, complete fertilization failure, oocyte retrieval failure, and complete abnormal fertilization accounted for 28.3% (172/607), 25.7% (156/607) and 22.24% (135/607), respectively. The incidence of complete abnormal fertilization was higher in IVF than in ICSI (p<0.05). In both IVF and ICSI cycles, the incidences of no embryos transferred were higher in the patients retrieving ≤3 oocytes than in the patients retrieving >3 oocytes (p<0.05). In IVF cycles the incidences of no embryos transferred were higher in the patients with primary infertility than in those with secondary infertility (p<0.05). CONCLUSION: The main causes of no embryos transferred are complete fertilization failure, oocyte retrieval failure, and complete abnormal fertilization. Retrieving adequate number of mature oocytes is the key to success of ART. Patients who experienced complete abnormal fertilization in IVF or the patients with primary infertility who experienced complete fertilization failure or normal fertilization without cleavage should receive ICSI in the next treatment.

Effects of Vitrification on Outcomes of In Vivo-Mature, In Vitro-Mature and Immature Human Oocytes.

BACKGROUND/AIMS: To observe the effects of vitrification on spindle, zona pellucida, embryonic aneuploidy and DNA injury in in vivo-maruted, in vitro-mature and immature human oocytes. METHODS: Between January 2009 and February 2015, 223 immature oocytes from 450 infertile patients, and 31 in vivo-mature oocytes from 3 infertile couples were collected. Of the 223 immature oocytes, 113 were used for in vitro culture before vitrification. Some oocytes were randomly divided into in vivo-mature group (group A, n = 15), in vitro-mature group (group B, n = 88) and immature group (group C, n = 85), and then the oocytes with spindle in these three groups after freezing-thawing were selected to use for Polscope imaging, embryonic aneuploidy screening and embryo development evaluation. Other oocytes were randomly divided into group A (n = 16), group B (n = 25) and group C (n = 25) for detecting DNA injury. RESULTS: After thawing, spindle occurrence rate, spindle Retardance value, and cleavage rate were significantly higher in groups A and B than in group C (all P < 0.05), but there were no statistical differences in fertility rate, high-quality embryo rate, blastulation rate and aneuploidy rate amongst the three groups (all P > 0.05). Zona pellucida density (ZPD) was significantly lower in group A than in groups B and C both before and after vitrification (all P < 0.05). ZPD was significantly higher after thawing than before vitrification (all P < 0.05), but zona pellucida thickness (ZPT) was not significantly changed in all the three groups (all P > 0.05). Rate of comet cells was significantly lower in group A than in groups B and C (all P < 0.01). Comet tail was significantly longer in group C than in groups B and A (all P < 0.05). CONCLUSION: In vivo- and in vitro-mature human oocytes are more suitable to vitrification than immature human oocytes. Spindle Retardance value has more predictive value for embryonic development potential than ZPD and ZPT.

Role of PAFAH1B1 in human spermatogenesis, fertilization and early embryonic development.

Spermatogenesis, fertilization and subsequent embryonic development are complex processes that require tight regulation. The PAFAH1B1 gene plays important roles in these reproductive events in mice, but its expression and roles in human reproduction have not been investigated. Expression analysis of testicular tissue by reverse transcription quantitative PCR and immunohistochemistry revealed varied expression levels among samples of different spermatogenic abilities (as assessed by the Johnsen score), with protein expression restricted to spermatogonia, spermatocytes and spermatids. Immunofluorescence on spermatozoa showed expression over the acrosome and midpiece regions of ejaculated samples, whereas a high proportion of percutaneous epididymal sperm aspiration-derived spermatozoa showed expression restricted to the midpiece. Analysis for PAFAH1B1 mRNA also revealed different expression levels among unfertilized oocytes, zygotes, cleavage stage embryos and blastocysts, with protein localized at the membrane level in oocytes and zygotes, and gradually distributing within the cytoplasm of cleavage stage embryos and blastocysts. Interestingly, microinjection of PAFAH1B1 siRNA into zygotes significantly (P = 0.024) increased fragmentation formation rates in subsequent embryonic development stages. Altogether, these are the first results to support a role for PAFAH1B1 in human spermatogenesis and early embryonic development.

Effects of group culture on the development of discarded human embryos and the construction of human embryonic stem cell lines.

PURPOSE: To explore the effect of group culture on the developmental potential of discarded embryos in in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) cycles and establish the human embryonic stem cell lines for future research. METHORDS: Fresh discarded embryos were collected from the IVF/ICSI-ET program in the reproductive medical center of the first affiliated hospital of Zhengzhou university in this study. All zygotes were individually cultured from Day 1 to Day 3. On Day 3, discard embryos were then cultured in group of 1-4 embryos per droplet (30 µl/droplets) with a constant culture medium until Day 5 or 6. Mechanical method was used to isolate the inner cell mass (ICM) of blastocyst from the embryo. Then we inoculated the ICM on feeder layer. After identification of those cells, the human embryonic stem cell lines (hESCs) were established. RESULTS: In this study, we collected 1,223 fresh discarded embryos and they were sequential cultured to the blastocysts (18.07 %, 221/1,223), in which good quality blastocysts were 61(4.98 %, 61/1,223). There was no significant difference in the patients. The embryos from 1PN, 2PN, 3PN were sequential cultured to the blastocyst s(39.31 %,92/234;12.87 %,64/497;13.21 %,65/492),in which good quality blastocysts was 13.6 %(32/92),2.61 %(13/64), 3.04 %(15/65).1PN embryo's blastulation rate and quality embryo formation rate was significantly higher than the 2PN and 3PN embryos' (P <0.05). Three embryos group cultivation has the highest blastulation rate and quality embryo formation rate (P <0.05). In total, we successfully established 4 hESCs lines. CONCLUSION: The group culture of human discard embryos can improve the blastulation rate and blastocyst quality to some extent. Three embryos group cultivate is the better culture number. Human discard embryos are good source for establishment of hESCs.