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STUDY QUESTION Is there an efficient and cost-effective detection platform for different genetic conditions about embryos? SUMMARY ANSWER A multifunctional haplotyping-based preimplantation genetic testing platform was provided for detecting different genetic conditions. WHAT IS KNOWN ALREADY Genetic disease and chromosomal rearrangement have been known to significantly impact fertility and development. Therefore, preimplantation genetic testing for aneuploidy (PGT-A), monogenic disorders (PGT-M) and structural rearrangements (PGT-SR), a part of ART, has been presented together to minimize the fetal genetic risk and increase pregnancy rate. For patients or their families who are suffering from chromosome abnormality, monogenic disease, unexplained repeated spontaneous abortion or implantation failure, after accepting genetic counseling, they may be suggested to accept detection from more than one PGT platforms about the embryos to avoid some genetic diseases. However, PGT platforms work through different workflows. The high costliness, lack of material and long-time operation of combined PGT platforms limit their application. STUDY DESIGN, SIZE, DURATION All 188 embryonic samples from 43 families were tested with HaploPGT platform, and most of their genetic abnormalities had been determined by different conventional PGT methods beforehand. Among them, there were 12 families only carrying structural rearrangements (115 embryos) in which 9 families accepted implantation and 5 families had normal labor ART outcomes, 7 families only carrying monogenic diseases (26 embryos) and 3 families carrying both structural rearrangements and monogenic diseases (26 embryos). Twelve monopronucleated zygotes (1PN) samples and 9 suspected triploid samples were collected from 21 families. PARTICIPANTS/MATERIALS, SETTINGS, METHODS Here, we raised a comprehensive PGT method called HaploPGT, combining reduced representation genome sequencing, read-count analysis, B allele frequency and haplotyping analysis, to simultaneously detect different genetic disorders in one single test. MAIN RESULTS AND THE ROLE OF CHANCE With 80 million reads (80M) genomic data, the proportion of windows (1 million base pairs (Mb)) containing two or more informative single nucleotide polymorphism (SNP) sites was 97.81%, meanwhile the genotyping error rate stabilized at a low level (2.19%). Furthermore, the informative SNPs were equally distributed across the genome, and whole-genomic haplotyping was established. Therefore, 80M was chosen to balance the cost and accuracy in HaploPGT. HaploPGT was able to identify abnormal embryos with triploid, global and partial loss of heterozygosity, and even to distinguish parental origin of copy number variation in mosaic and non-mosaic embryos. Besides, by retrospectively analyzing 188 embryonic samples from 43 families, HaploPGT revealed 100% concordance with the available results obtained from reference methods, including PGT-A, PGT-M, PGT-SR and PGT-HLA. LIMITATIONS, REASON FOR CAUTION Despite the numerous benefits HaploPGT could bring, it still required additional family members to deduce the parental haplotype for identifying balanced translocation and monogenic mutation in tested embryos. In terms of PGT-SR, the additional family member could be a reference embryo with unbalanced translocation. For PGT-M, a proband was normally required. In both cases, genomic information from grandparents or parental siblings might help for haplotyping theoretically. Another restriction was that haploid, and diploid resulting from the duplication of a haploid, could not be told apart by HaploPGT, but it was able to recognize partial loss of heterozygosity in the embryonic genome. In addition, it should be noted that the location of rearrangement breakpoints and the situation of mutation sites were complicated, which meant that partial genetic disorders might not be completely detected. WIDER IMPLICATIONS OF THE FINDINGS HaploPGT is an efficient and cost-effective detection platform with high clinical value for detecting genetic status. This platform could promote the application of PGT in ART, to increase pregnancy rate and decrease the birth of children with genetic diseases. STUDY FUNDING/COMPETING INTEREST(S) This study was supported by grants from the National Natural Science Foundation of China (81873478, to L.H.), National Key R&D Program of China (2018YFC1003100, to L.H.), the Natural Science Foundation of Hunan Province (Grant 2022JJ30414, to P.X.), Hunan Provincial Grant for Innovative Province Construction (2019SK4012) and the Scientific Research Foundation of Reproductive and Genetic Hospital of China International Trust & Investment Corporation (CITIC)-Xiangya (YNXM-201910). Haplotyping analysis has been licensed to Basecare Co., Ltd. L.K., Y.M., K.K., D.Z., N.L., J.Z. and R.D. are Basecare Co., Ltd employees. The other authors declare no competing interests. TRIAL REGISTRATION NUMBER N/A.

BACKGROUND: This study aimed to explore the imaging characteristics, diversity and changing trend in CT scans of pediatric patients infected with Delta-variant strain by studying imaging features of children infected with Delta and comparing the results to those of children with original COVID-19. METHODS: A retrospective, comparative analysis of initial chest CT manifestations between 63 pediatric patients infected with Delta variant in 2021 and 23 pediatric patients with COVID-19 in 2020 was conducted. Corresponding imaging features were analyzed. In addition, the changing trend in imaging features of COVID-19 Delta-variant cases were explored by evaluating the initial and follow-up CT scans. RESULTS: Among 63 children with Delta-variant COVID-19 in 2021, 34 (53.9%) showed positive chest CT presentation; and their CT score (1.10 ± 1.41) was significantly lower than that in 2020 (2.56 ± 3.5) (P = 0.0073). Lesion distribution: lung lesions of Delta cases appear mainly in the lower lungs on both sides. Most children had single lobe involvement (18 cases, 52.9%), 14 (41.2%) in the right lung alone, and 14 (41.2%) in both lungs. A majority of Delta cases displayed initially ground glass (23 cases, 67.6%) and nodular shadows (13 cases, 38.2%) in the first CT scan, with few extrapulmonary manifestations. The 34 children with abnormal chest CT for the first time have a total of 92 chest CT examinations. These children showed a statistically significant difference between the 0-3 day group and the 4-7 day group (P = 0.0392) and a significant difference between the 4-7 day group and the more than 8 days group (P = 0.0003). CONCLUSIONS: The early manifestations of COVID-19 in children with abnormal imaging are mostly small subpleural nodular ground glass opacity. The changes on the Delta-variant COVID-19 chest CT were milder than the original strain. The lesions reached a peak on CT in 4-7 days and quickly improved and absorbed after a week. Dynamic CT re-examination can achieve a good prognosis.

The Hippo pathway is an evolutionarily conserved developmental pathway that controls organ size by integrating diverse regulatory inputs, including actomyosin-mediated cytoskeletal tension. Despite established connections between the actomyosin cytoskeleton and the Hippo pathway, the upstream regulation of actomyosin in the Hippo pathway is less defined. Here, we identify the phosphoinositide-3-phosphatase Myotubularin (Mtm) as a novel upstream regulator of actomyosin that functions synergistically with the Hippo pathway during growth control. Mechanistically, Mtm regulates membrane phospholipid PI(3)P dynamics, which, in turn, modulates actomyosin activity through Rab11-mediated vesicular trafficking. We reveal PI(3)P dynamics as a novel mode of upstream regulation of actomyosin and establish Rab11-mediated vesicular trafficking as a functional link between membrane lipid dynamics and actomyosin activation in the context of growth control. Our study also shows that MTMR2, the human counterpart of Drosophila Mtm, has conserved functions in regulating actomyosin activity and tissue growth, providing new insights into the molecular basis of MTMR2-related peripheral nerve myelination and human disorders.

Summary Multiple morphological abnormalities of the sperm flagella (MMAF)-induced asthenoteratozoospermia is a common cause of male infertility. Previous studies have identified several MMAF-associated genes, highlighting the condition’s genetic heterogeneity. To further define the genetic causes underlying MMAF, we performed whole-exome sequencing in a cohort of 643 Chinese MMAF-affected men. Bi-allelic DNAH10 variants were identified in five individuals with MMAF from four unrelated families. These variants were either rare or absent in public population genome databases and were predicted to be deleterious by multiple bioinformatics tools. Morphological and ultrastructural analyses of the spermatozoa obtained from men harboring bi-allelic DNAH10 variants revealed striking flagellar defects with the absence of inner dynein arms (IDAs). DNAH10 encodes an axonemal IDA heavy chain component that is predominantly expressed in the testes. Immunostaining analysis indicated that DNAH10 localized to the entire sperm flagellum of control spermatozoa. In contrast, spermatozoa from the men harboring bi-allelic DNAH10 variants exhibited an absence or markedly reduced staining intensity of DNAH10 and other IDA components, including DNAH2 and DNAH6. Furthermore, the phenotypes were recapitulated in mouse models lacking Dnah10 or expressing a disease-associated variant, confirming the involvement of DNAH10 in human MMAF. Altogether, our findings in humans and mice demonstrate that DNAH10 is essential for sperm flagellar assembly and that deleterious bi-allelic DNAH10 variants can cause male infertility with MMAF. These findings will provide guidance for genetic counseling and insights into the diagnosis of MMAF-associated asthenoteratozoospermia.

Leukocyte recruitment is a critical step in the pathogenesis of inflammatory and immunological responses. Cell adhesion molecules (CAMs) are involved in controlling cell movements and the recruitment process, and the integrin family of CAMs plays a key role. During cell movement, integrin function is dynamically and precisely regulated. However, this balance might be broken under pathological conditions. Thus, the functional regulation and molecular mechanisms of integrins related to diseases are often a focus of research. Integrin β2 is one of the most commonly expressed integrins in leukocytes that mediate leukocyte adhesion and migration, and it plays an important role in immune responses and inflammation. In this review, we focus on specific functions of integrin β2 in leukocyte recruitment, the conformational changes and signal transduction of integrin β2 activation, the similarities between murine and human factors, and how new insights into these processes can inform future therapies for inflammation and immune diseases.

BackgroundThe risk of developing gestational diabetes mellitus (GDM) is higher in women undergoing assisted reproductive treatment than in women conceiving spontaneously.ObjectivesTo determine whether the GDM risk after day-3 embryo transfer differs from the GDM risk after day-5 blastocyst transfer.MethodsProspective observational study in women becoming pregnant after first fresh embryo or blastocyst transfer.ResultsA total of 1579 women got pregnant and had live birth; 1300 women got day-3 embryo transfer only, whereas 279 women received at least 1 blastocyst. Of 1579 women, 252 developed GDM. Age, body mass index, baseline estradiol, baseline high-density lipoprotein, and progesterone on the day of human chorionic gonadotropin injection were not different in women receiving day-3 embryos only vs women receiving at least 1 blastocyst. The number and quality of retrieved oocytes were not different in women receiving day-3 embryo transfer from those receiving blastocysts. Our study confirmed already established GDM risk factors such as age and body mass index, baseline estradiol, and high-density lipoprotein, as well as progesterone after ovarian stimulation. We furthermore demonstrate that the GDM incidence in women receiving day-5 blastocyst transfer was significantly higher than those who received day-3 embryo transfer (21.15% vs 14.85%; P = 0.009). Considering confounding factors, we likewise saw that blastocyst transfer was an independent procedure-related GDM risk factor [P = 0.009, Exp (B): 1.56, 95% CI: 1.12-2.18].ConclusionBlastocyst transfer after in vitro fertilization/intracytoplasmic sperm injection increases the risk of developing GDM.

Objective To investigate the effects of non-Robertsonian translocation with chromosome fusion (N-RBCF) on preimplantation embryos. Design Case series. Setting University-affiliated center. Patient(s) Twelve couples with N-RBCF. Intervention(s) Assisted reproduction with preimplantation genetic testing in chromosomal structural rearrangement (PGT-SR). Main Outcome Measure(s) Normal embryo rate, unbalanced translocation rate. Result(s) PGT was performed in 12 N-RBCF carriers, of whom 4 carried Y-autosome fusions and 8 autosomal fusions. A total of 12 (63.2%) of 19 blastocysts exhibited normal/balanced embryos, and only one (5.3%) embryo exhibited unbalanced translocations among Y-autosome fusion cases. In contrast to these findings, the percentage of normal/balanced blastocysts in 8 autosomal N-RBCF cases was 28.2% (11/39), whereas the unbalanced translocation rate was 53.8%. Furthermore, the percentage of normal/balanced embryos in the Robertsonian translocation group was significantly higher than that of the 8 autosomal N-RBCF (48.7% vs. 28.2%) cases. The rates of abnormality from chromosomal fusion in the 8 autosomal N-RBCF cases were significantly higher than those noted in the Robertsonian translocation (53.8% vs. 31.4%) subjects. The results of the stratified analysis according to the carrier’s sex demonstrated that the rates of unbalanced translocation were significantly higher in the male autosomal N-RBCF subjects than those from the corresponding Robertsonian translocation (55% vs. 19.7%) cases. Conclusion(s) A low number of unbalanced translocations was identified in blastocysts from N-RBCF subjects who carried the Y fusion. The risk of unbalanced translocation in autosomal N-RBCF was higher than that of the Robertsonian translocation, notably in male carriers.

PURPOSE: To verify the diagnostic power of vertical and horizontal thickness profiles of the corneal sublayers generated by ultra-high resolution optical coherence tomography (UHROCT) in subclinical and suspected keratoconus. METHODS: In this cross-sectional study, 25 eyes with confirmed keratoconus, 63 eyes with suspected keratoconus, 15 eyes with subclinical keratoconus, and 42 normal eyes were investigated. Vertical and horizontal thickness profiles of the corneal epithelium, Bowman's layer, and stroma were measured by UHR-OCT. Diagnostic indices included ratios of thickness distribution and multimeric discriminant functions calculated by multiple logistic regression based on them. Receiver operating characteristic curves were used to verify the predictive accuracy by the area under the curve (AUC). RESULTS: Function consisting of two indices (vertical maximum ectasia index of epithelium and horizontal maximum ectasia index of Bowman's layer) performed well to discriminate subclinical keratoconus (AUC = 0.967) and suspected keratoconus (AUC = 0.932) from normal. In addition, when four indices were combined, the diagnostic power for subclinical keratoconus (AUC = 0.984) and suspected keratoconus (AUC = 0.971) was further increased. However, both binary and quaternary functions could not adequately discriminate suspected from subclinical keratoconus. CONCLUSIONS: UHR-OCT–generated thickness indices from the vertical and horizontal thickness profiles of the corneal epithelium and Bowman's layer showed an evident diagnostic efficacy in discriminating suspected and subclinical keratoconus from normal eyes. The early changes in keratoconus might prefer thickness distribution in corneal sublayers rather than corneal thickness or topography. [ J Refract Surg . 2021;37(7):438–445.]

Background Previous studies suggested that non-invasive preimplantation genetic testing (niPGT) for intracytoplasmic sperm injection (ICSI) blastocysts can be used to identify chromosomal ploidy and chromosomal abnormalities. Here, we report the feasibility and performance of niPGT for conventional in vitro fertilization (IVF) blastocysts. Methods This was a prospective observational study. In the preclinical stage, whole genome amplification and NGS were performed using the sperm spent culture medium (SCM). Then, trophectoderm (TE) biopsies and corresponding SCM derived from 27 conventional IVF monopronuclear embryos were collected. In the clinical stage, samples from 25 conventional IVF cycles and 37 ICSI cycles from April 2020–August 2021 were collected for performance evaluation. Results Preclinically, we confirmed failed sperm DNA amplification under the current amplification system. Subsequent niPGT from the 27 monopronuclear blastocysts showed 69.2% concordance with PGT results of corresponding TE biopsies. In the clinical stage, no paternal contamination was observed in any of the 161 SCM samples from conventional IVF. While maternal contamination was observed in 29.8% (48/161) SCM samples, only 2.5% (4/161) samples had a contamination ratio ≥ 50%. Compared with that of TE biopsy, the performances of NiPGT from 161 conventional IVF embryos and 122 ICSI embryos were not significantly different (P > 0.05), with ploidy concordance rates of 75% and 74.6% for IVF and ICSI methods, respectively. Finally, evaluation of the euploid probability of embryos with different types of niPGT results showed prediction probabilities of 82.8%, 77.8%, 62.5%, 50.0%, 40.9% and 18.4% for euploidy, sex-chromosome mosaics only, low-level mosaics, multiple abnormal chromosomes, high-level mosaics and aneuploidy, respectively. Conclusions Our research results preliminarily confirm that the niPGT approach using SCM from conventional IVF has comparable performance with ICSI and might broadening the application scope of niPGT.

IntroductionMorphological evaluation is used to select embryos for in vitro fertilisation. However, it does not fully reflect the implantation potential. Preimplantation genetic testing for aneuploidies (PGT-A) can detect embryonic aneuploidy, but biopsy procedure is invasive. Currently, a non-invasive PGT (ni-PGT) approach using spent medium is being evaluated. However, the clinical benefit of ni-PGT has not been clearly demonstrated. A multicentre randomised trial is needed to verify whether ni-PGT can be an new effective tool for evaluating embryos.Methods and analysisOverall, 1148 couples aged 35~42 (women) receiving in vitro fertilization–intracytoplasmic sperm injection are planned to be enrolled. Couples will be digitally randomised to (1) ni-PGT and (2) conventional morphology groups at a 1:1 treatment ratio. The primary outcome will be the ongoing pregnancy rate related to the first transfer cycle within 6 months after oocyte retrieval.Ethics and disseminationThe study protocol is approved by the Ethics Committee of Peking University Third Hospital and the participating hospitals. The results will be disseminated through international conferences and scientific journals.Trial registration numberNCT04339166.

As a treasure among Chinese national musical instruments, guzheng is an important part of traditional Chinese music. As the art of national music goes to the world, the art of guzheng has been widely promoted. As the best form to carry forward the art of guzheng, the teaching of guzheng majors in colleges is significant in teaching and continuously improves guzheng art accomplishment. Oral teaching and step-by-step music theory and technique teaching are typical ways of teaching musical instrument performance in colleges. However, under big data, Chinese education is undergoing a profound change, and the combination of big data and education has become a new contemporary education method. This work studies the guzheng professional education in colleges under big data. First, this work aims at the existing outstanding issues of guzheng teaching in colleges and studies the challenges and optimization paths of guzheng professional education in colleges under big data. Second, this work proposes a multiscale residual attention fusion network (MSRAFNET) to evaluate the teaching quality of guzheng majors in colleges in the era of big data. The feature extraction of the network model is mainly completed by the residual module, which is composed of several multiscale residual learning units. Adding an attention mechanism to the multiscale residual learning unit can enhance the feature extraction of key information by the network and reduce the interference of redundant information, which is more conducive to the learning of data features. It adopts the design of GAP and Dropout to reduce spatial parameters in network training, and the effect of antioverfitting is better. Third, this work systematically evaluates the optimization path of Guzheng education and MSRAFNET, and the systematic experiments verify the superiority of the designed method.

Nanoscale small extracellular vesicles (sEVs, exosomes) in tears allow us to investigate the multisignatures of diseases. However, the translations of tear sEVs for biomarker discovery and clinical diagnostics are practically limited by low recovery, long processing time, and small sample volume. Here, we report an incorporated tear-exosomes analysis

Epilepsy is a neurological disease caused by ab-normal neural electrical discharges. Electroencephalography (EEG) is a powerful tool to measure the brain electrical activity and has been widely used for seizure detection. Manual EEG analysis is labor-intensive and time-consuming. Automatic seizure detection is urgently demanded for long-time seizure monitoring. Many methods have been proposed for automatic seizure detection based on EEG signals. However, most of the existing methods are patient-specific with limited generaliz-ability. Few studies investigate inter-patient seizure detection, which remains challenging. The aim of the present study is therefore to develop advanced algorithms for efficient inter-patient seizure detection using EEG. To this end, dynamic brain network is employed to capture the spatiotemporal dynamics of the connectivity among brain regions. A novel graph neural network referred to as graph isomorphic network is proposed for effective local-global spatiotemporal feature extraction and seizure classification. The proposed method is evaluated with the CHB-MIT open dataset with a ten-fold cross-validation. The results reveal excellent performance for the proposed method, with accuracy, sensitivity, and specificity of 96.2%, 95.4%, and 97.0% respectively, significantly higher than the results reported in the literature. Our results provide useful information for inter-patient seizure detection, particularly for long-time ambulatory seizure monitoring.

This study aimed to report the three-dimensional reconstruction of the foramen ovale (FO) based on computed tomography angiography and describe its shape and related angles. A retrospective analysis of 199 adult patients who were hospitalised at the Department of Neurosurgery, The Second Affiliated Hospital of Bengbu Medical College, Bengbu, China, from January to December 2020 was conducted. The original DICOM files of patients' computed tomography scans were processed by 3D Slicer software to reconstruct the three-dimensional skull. The morphological characteristics of the FO on both sides were analysed. Their size, related angles and volumes, and the differences between the two sides and gender were compared. A total of 398 FO from 199 patients were studied. The most frequent shape of the FO was oval, accounting for 54.27%. The mean lengths of the right and the left sides were 5.40±1.51 and 5.10±1.18mm, respectively. The mean width on the right and left sides was 3.23±1.16 and 3.33±1.19 mm, respectively. The FO is most commonly oval in shape. Clinicians may use the anatomical characteristics regarding the size and shape of the FO for diagnosis and treatment. Key Words: Foramen ovale, Computed tomographic angiography, 3-Dimensional anatomy.

Coronavirus disease 2019 (COVID-19) is a highly contagious respiratory disease that threatens global health. During the pandemic period of COVID-19, the task for prevention in the general ward of cardiovascular surgery is fairly arduous. The present study intends to summarize our experience with infection control, including ward setting, admission procedures, personnel management, health education, and so on, to provide references for clinical management.

Objective To measure free and total 25-hydroxyvitamin D [25(OH)D] immediately before embryo transfer and analyze its association with early pregnancy outcome parameters such as biochemical pregnancy, implantation rate, and clinical pregnancy rates in women undergoing fresh embryo transfer after their first ovarian hyperstimulation. Design Prospective cohort study. Setting Academically affiliated private fertility center. Patient(s) A total of 2,569 women undergoing fresh embryo transfer after ovarian hyperstimulation. Interventions(s) None. Main Outcome Measure(s) The study end points were biochemical pregnancy rate, implantation rate, clinical pregnancy rate, ectopic pregnancy rate, early miscarriages, and ongoing pregnancy rate. Free and total 25(OH)D concentrations were measured 1 day before embryo transfer. Result(s) Total 25(OH)D correlated with free 25(OH)D. Total and free 25(OH)D serum concentrations were similar in those patients reaching and not reaching the study outcomes (biochemical pregnancy rate, implantation rate, clinical pregnancy rate, ectopic pregnancy rate, early miscarriages, and ongoing pregnancy rate). There was likewise no statistical difference when analyzing the frequency of all study outcomes in quintiles of either total or free 25(OH)D. In addition, the study population was divided into three groups according to the total vitamin D status based on clinical practice guideline. All outcomes were similar in women with adequate, insufficient, and deficient total 25(OH)D. Multiple linear regression analysis considering confounding likewise indicated no association of free or total vitamin D with any of the study outcomes. Conclusion(s) Neither free nor total 25(OH)D concentration at embryo transfer was associated with successful embryo implantation in women undergoing fresh transfer after ovarian hyperstimulation.

Chromobox 4 (CBX4) is a core component of polycomb-repressive complex 1 with important roles in cancer biology and tissue homeostasis. Aberrant expression of CBX4 has been implicated in several human malignancies. However, its role and underlying mechanisms in the tumorigenesis of lung adenocarcinoma (LUAD) have not been defined in vivo. Here, we found that expression of CBX4 was frequently up-regulated in human LUAD samples and correlated with poor patient survival. Importantly, genetic ablation of CBX4 greatly dampened lung tumor formation and improved survival in the KrasG12D/P53L/L (KP) autochthonous mouse model of LUAD. In addition, CBX4 depletion significantly inhibited proliferation and anchorage-independent growth of KP mouse embryonic fibroblasts. Moreover, ectopic CBX4 expression clearly promoted proliferation and anchorage-independent growth in both human and mouse LUAD cells, whereas silencing of CBX4 exerted opposite effects. Mechanistically, CBX4 promoted growth of LUAD cells through activation of the Wnt/β-catenin pathway. Furthermore, expression levels of CBX4 were positively correlated with β-catenin in human LUAD samples. In conclusion, our data suggest that CBX4 plays an oncogenic role via the Wnt/β-catenin pathway and could serve as a potential therapeutic target in LUAD.

Background: PCSK9 (proprotein convertase subtilisin/kexin 9), mainly secreted by the liver and released into the blood, elevates plasma low-density lipoprotein cholesterol by degrading low-density lipoprotein receptor. Pleiotropic effects of PCSK9 beyond lipid metabolism have been shown. However, the direct effects of PCSK9 on platelet activation and thrombosis, and the underlying mechanisms, as well, still remain unclear. Methods: We detected the direct effects of PCSK9 on agonist-induced platelet aggregation, dense granule ATP release, integrin αIIbβ3 activation, α-granule release, spreading, and clot retraction. These studies were complemented by in vivo analysis of FeCl 3 -injured mouse mesenteric arteriole thrombosis. We also investigated the underlying mechanisms. Using the myocardial infarction (MI) model, we explored the effects of PCSK9 on microvascular obstruction and infarct expansion post-MI. Results: PCSK9 directly enhances agonist-induced platelet aggregation, dense granule ATP release, integrin αIIbβ3 activation, P-selectin release from α-granules, spreading, and clot retraction. In line, PCSK9 enhances in vivo thrombosis in a FeCl 3 -injured mesenteric arteriole thrombosis mouse model, whereas PCSK9 inhibitor evolocumab ameliorates its enhancing effects. Mechanism studies revealed that PCSK9 binds to platelet CD36 and thus activates Src kinase and MAPK (mitogen-activated protein kinase)–extracellular signal-regulated kinase 5 and c-Jun N-terminal kinase, increases the generation of reactive oxygen species, and activates the p38MAPK/cytosolic phospholipase A2/cyclooxygenase-1/thromboxane A 2 signaling pathways downstream of CD36 to enhance platelet activation, as well. Using CD36 knockout mice, we showed that the enhancing effects of PCSK9 on platelet activation are CD36 dependent. It is important to note that aspirin consistently abolishes the enhancing effects of PCSK9 on platelet activation and in vivo thrombosis. Last, we showed that PCSK9 activating platelet CD36 aggravates microvascular obstruction and promotes MI expansion post-MI. Conclusions: PCSK9 in plasma directly enhances platelet activation and in vivo thrombosis, and MI expansion post-MI, as well, by binding to platelet CD36 and thus activating the downstream signaling pathways. PCSK9 inhibitors or aspirin abolish the enhancing effects of PCSK9, supporting the use of aspirin in patients with high plasma PCSK9 levels in addition to PCSK9 inhibitors to prevent thrombotic complications.

Purpose The purpose of this study is to summarize the clinical outcomes of apparently balanced chromosome rearrangement (ABCR) carriers in preimplantation genetic testing (PGT) cycles by next-generation sequencing following microdissecting junction region (MicroSeq) to distinguish non-carrier embryos from balanced carriers. Methods A retrospective study of 762 ABCR carrier couples who requested PGT for structural rearrangements combined with MicroSeq at the Reproductive and Genetic Hospital of CITIC-Xiangya was conducted between October 2014 and October 2019. Results Trophectoderm biopsy was performed in 4122 blastocysts derived from 917 PGT-SR cycles and 3781 blastocysts were detected. Among the 3781 blastocysts diagnosed, 1433 (37.9%, 1433/3781) were balanced, of which 739 blastocysts were carriers (51.57%, 739/1433) and 694 blastocysts were normal (48.43%, 694/1433). Approximately 26.39% of cycles had both carrier and normal embryo transfer, and the average number of biopsied blastocysts was 6.7. In the cumulative 223 biopsied cycles with normal embryo transfer, all couples chose to transfer the normal embryos. In the 225 cycles with only carrier embryos, the couples chose to transfer the carrier embryos in 169/225 (75.11%) cycles. A total of 732 frozen embryo transfer cycles were performed, resulting in 502 clinical pregnancies. Cumulatively, 326 babies were born; all of these babies were healthy and free of any developmental issues. Conclusion Our study provides the first evaluation of the clinical outcomes of a large sample with ABCR carrier couples undergoing the MicroSeq-PGT technique and reveals its powerful ability to distinguish between carrier and non-carrier balanced embryos.

Esophageal squamous cell carcinoma (ESCC) is a worldwide malignancy with high mortality rates and poor prognosis due to the lack of effective biomarkers for early detection. Exosomes have been extensively explored as attractive biomarkers for cancer diagnosis and treatment. However, little is known about exosome metabolomics and their roles in ESCC. Here, we performed a targeted metabolomic analysis of plasma exosomes and identified 196 metabolites, mainly including lipid fatty acids, benzene, amino acids, organic acids, carbohydrates and fatty acyls. We systematically compared metabolome patterns of exosomes via machine learning from patients with recrudescence and patients without recrudescence and demonstrated a marker set consisting of 3'-UMP, palmitoleic acid, palmitaldehyde, and isobutyl decanoate for predicting ESCC recurrence with an AUC of 98%. These metabolome signatures of exosomes retained a high absolute fold change value at all ESCC stages and were very likely associated with cancer metabolism, which could be potentially applied as novel biomarkers for diagnosis and prognosis of ESCC.

Primary hyperparathyroidism (PHPT) is mainly caused by parathyroid adenoma, which produces excess parathyroid hormones. Its pathogenic mechanisms have not yet been fully understood. To investigate the mechanism in the pathogenesis of PHPT, the transcriptome and genome-wide DNA methylation profiles of parathyroid adenoma were analyzed. The candidate genes that may be involved in the PHPT were verified via qRT-PCR, immunohistochemistry, western blot, and methylation-specific PCR. A total of 1650 differentially expressed genes and 2373 differentially methylated regions were identified. After the integration of its transcriptome and DNA methylation data, IL6, SYP, GNA01, and pro-opiomelanocortin (POMC) were the candidate genes that demonstrated a similar pattern between their mRNA expression and DNA methylation status. Of the 4 candidate genes, POMC, a pro-peptide which is processed to a range of bioactive peptide products like ACTH, was further confirmed to be expressed at low levels at both the mRNA and protein levels, which may be due to POMC promoter hypermethylation. Hypermethylation of the POMC promoter may contribute to its low expression, which may be involved in the pathogenesis of PHPT.

Neutrophils are the most abundant leukocyte in human blood. They are critical for fighting infections and are involved in inflammatory diseases. Mitochondria are indispensable for eukaryotic cells, as they control the biochemical processes of respiration and energy production. Mitochondria in neutrophils have been underestimated since glycolysis is a major metabolic pathway for fuel production in neutrophils. However, several studies have shown that mitochondria are greatly involved in multiple neutrophil functions as well as neutrophil-related diseases. In this review, we focus on how mitochondrial components, metabolism, and related genes regulate neutrophil functions and relevant diseases.

Neutrophils are the most abundant leukocytes in humans and are critical for innate immunity and inflammation. Integrins are critical for neutrophil functions, especially for their recruitment to sites of inflammation or infections. Integrin conformational changes during activation have been heavily investigated but are still not fully understood. Many regulators, such as talin, Rap1-interacting adaptor molecule (RIAM), Rap1, and kindlin, are critical for integrin activation and might be potential targets for integrin-regulating drugs in treating inflammatory diseases. In this review, we outline integrin activation regulators in neutrophils with a focus on the above critical regulators, as well as newly discovered modulators that are involved in integrin activation.

In this study, mRNA expression of gastric cancer tissue and clinical data of patients in TCGA-STAD dataset were used, together with the hypoxia-related gene sets in the MsigDB database, to screen hypoxia-related differentially expressed genes (DEGs) in GC. Thereafter, univariate and multivariate Cox regression analyses were carried out on hypoxia-related DEGs. The optimal feature genes related to prognosis were obtained to construct a prognostic risk assessment model. According to the model, the riskScore of GC patients was measured, and GC samples were assigned into high- and low-risk groups in accordance with the median riskScore. Based on the Kaplan-Meier curve and Receiver operating characteristic curve, validity of the prognostic risk assessment model was measured. Gene set enrichment analysis was performed on the two risk groups through Gene set enrichment analysis software. The results revealed that in the high-risk group, 9 signaling pathways were remarkably activated in several terms, like focal adhesion, extracellular matrix receptor interaction, Cell adhesion molecules cams, Cytokine-cytokine receptor interaction, TGF-beta signaling pathway, NOD-like receptor signaling pathway, JAK-STAT signaling pathway, Toll-like receptor signaling pathway and MAPK signaling pathway. In combination with riskScore and clinical factors, univariate and multivariate Cox regression analyses verified the independence of the model. Meanwhile, a nomogram was constructed to predict the 1-, 3- and 5-year survival of GC patients. The calibration curve indicated that the survival status predicted by the nomogram fitted better with actual survival status. On the whole, the prognostic risk model of GC on the basis of hypoxia-related genes demonstrated good predictive ability. It can provide more powerful technical support for clinicians to make prognostic determination and therapeutic plans.

Introduction Intrahepatic cholestasis of pregnancy (ICP) is associated with an increased risk of adverse pregnancy outcomes in mothers and infants. The aim was to evaluate the sensitivity and specificity of biochemical detection of ICP by ROC curve and to determine the threshold of more reliable experimental indicators. Materials and Methods 305 patients and 305 healthy pregnant women were enrolled in the study. Results The average levels of TBA, ALT, and AST in the ICP group were much higher than those in the control group (P Conclusions This study did not find any single specificity and sensitivity markers that could be used to reliably diagnose ICP. In the future, we will pay more attention to the correlation between sensitive biochemical indicators and adverse pregnancy outcomes.

Eukaryotic genomes encode thousands of small and large non-coding RNAs (ncRNAs). However, the expression, functions and evolution of these ncRNAs are still largely unknown. In this study, we have updated deepBase to version 3.0 (deepBase v3.0, http://rna.sysu.edu.cn/deepbase3/index.html), an increasingly popular and openly licensed resource that facilitates integrative and interactive display and analysis of the expression, evolution, and functions of various ncRNAs by deeply mining thousands of high-throughput sequencing data from tissue, tumor and exosome samples. We updated deepBase v3.0 to provide the most comprehensive expression atlas of small RNAs and lncRNAs by integrating ∼67 620 data from 80 normal tissues and ∼50 cancer tissues. The extracellular patterns of various ncRNAs were profiled to explore their applications for discovery of noninvasive biomarkers. Moreover, we constructed survival maps of tRNA-derived RNA Fragments (tRFs), miRNAs, snoRNAs and lncRNAs by analyzing >45 000 cancer sample data and corresponding clinical information. We also developed interactive webs to analyze the differential expression and biological functions of various ncRNAs in ∼50 types of cancers. This update is expected to provide a variety of new modules and graphic visualizations to facilitate analyses and explorations of the functions and mechanisms of various types of ncRNAs.

The tumor microenvironment is known to play an important role in uveal melanoma. Reliable prognostic signatures are needed to aid high risk patients and improve prognosis. Uveal melanoma tissues from three public datasets were analyzed. RNA sequence data of uveal melanoma and corresponding clinical features were obtained from The Cancer Genome Atlas database. Immune and stromal scores were calculated by applying the “ESTIMATE” algorithm. The samples were divided into high and low immune or stromal score groups. We constructed prognostic models by using the ‘lasso’ package and tested them for 500 iterations. The cell signature was validated in another GSE44295 and GSE84976 datasets. We found that the median survival time of the low immune/stromal score group is longer than that of the high-score group. Thirteen immune cells and one stromal cell were concerned significant in predicting poor overall survival rate. Finally, a four-cell model was identified. Further validation revealed that the low-risk group has a significantly better survival than the high-risk group in another two datasets (P < 0.05). Moreover, the high-risk group is more sensitive to immunotherapy and chemotherapy. Summarizing, the proposed immune cells signature is a promising biomarker for estimating overall survival in uveal melanoma.

PURPOSE: To elucidate the genetic cause of intellectual deficiency and/or congenital malformations in two parental reciprocal translocation carriers and provide appropriate strategies of assisted reproductive therapy (ART). MATERIALS AND METHODS: Two similar couples having a child with global developmental delay/intellectual disability symptoms attended the Reproductive and Genetic Hospital of CITIC-Xiangya (Changsha, China) in 2017 and 2019, respectively, in order to determine the cause(s) of the conditions affecting their child and to seek ART to have a healthy baby. Both of the healthy couples were not of consanguineous marriage, denied exposure to toxicants, and had no adverse life history. This study was approved by the Institutional Ethics Committee of the Reproductive & Genetic Hospital of CITIC-Xiangya, and written informed consent was obtained from the parents. Genetic diagnoses were performed by karyotype analysis, breakpoint mapping analysis of chromosomal translocation(s), single-nucleotide polymorphism (SNP) microarray analysis, and whole-exome sequencing (WES) for the two children and different appropriate reproductive strategies were performed in the two families. RESULTS: Karyotype analysis revealed that both patients carried parental reciprocal translocations [46,XY,t(7;16)(p13;q24)pat and 46,XY,t(13;17)(q12.3;p11.2)pat, respectively]. Follow-up breakpoint mapping analysis showed no interruption of associated genes, and SNP microarray analysis identified no significant copy number variations (CNVs) in the two patients. Moreover, WES results revealed that patients 1 and 2 harbored candidate compound heterozygous mutations of MCOLN1 [c.195G>C (p.K65N) and c.1061G>A (p.W354*)] and MCPH1 [c.877A>G (p.S293G) and c.1869_1870delAT (p.C624*)], respectively, that were inherited from their parents and not previously reported. Furthermore, the parents of patient 1 obtained 10 embryos during ART cycle, and an embryo of normal karyotype and non-carrier of observed MCOLN1 mutations according to preimplantation genetic testing for structural rearrangement and monogenic defect was successfully transferred, resulting in the birth of a healthy boy. The parents of patient 2 chose to undergo ART with donor sperm to reduce the risk of recurrence. CONCLUSIONS: Systematic genetic diagnosis of two carriers of inherited chromosomal translocations accompanied by clinical phenotypes revealed their cause of disease, which was critical for genetic counseling and further ART for these families. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s10815-020-01986-1) contains supplementary material, which is available to authorized users.

To investigate changes in choriocapillaris vascular density (CCD) and choroidal thickness (CT) after femtosecond laser in situ keratomileusis (FS-LASIK) in myopic patients.Prospective and observational self-control study. We enrolled 41 myopic patients (41 right eyes), without other ophthalmic or systemic diseases who were eligible and about to FS-LASIK surgery. The CCD was measured in the central 1-mm-diameter circle region of the macular fovea (CCD1) and the 1- to 3-mm ring-like region of the fovea (CCD1-3). The CT was measured at the fovea and at distances of 0.5 and 2.5 mm for the following: nasal, temporal, superior, and inferior to the fovea. Both were measured by optical coherence tomography angiography (OCTA) preoperatively as the baseline and at 1 day, 1 week, 1 month, and 3 months postoperatively.CCD1 and CCD1-3 were significantly lower postoperatively compared with baseline. When the trough was reached, CCD1 declined by 1.51%, and CCD1-3 declined by 1.00% at 1 day postoperatively. However, then they increased with time. CCD1 recovered to baseline at 1 month postoperatively, while CCD1-3 was still lower at the end of follow-up. In addition, the CT was significantly thicker postoperatively and recovered to the baseline at 3 months postoperatively. There was no obvious correlation between CCD, CT, and other parameters.The CCD decreased while the CT increased following FS-LASIK in myopic patients. However, these changes may be a temporary stress response rather than real irreversible injury to the choroid.

Platelet activation plays a pivotal role in physiological hemostasis and pathological thrombosis causing heart attack and stroke. Previous studies conclude that simultaneous activation of Gi and G12/13 signaling pathways is sufficient to cause platelet aggregation. However, using Gq knockout mice and Gq-specific inhibitors, we here demonstrated that platelet aggregation downstream of coactivation of Gi and G12/13 depends on agonist concentrations; coactivation of Gi and G12/13 pathways only induces platelet aggregation under higher agonist concentrations. We confirmed Gi and G12/13 pathway activation by showing cAMP (cyclic adenosine monophosphate) decrease and RhoA activation in platelets stimulated at both low and high agonist concentrations. Interestingly, we found that though Akt and PAK (p21-activated kinase) translocate to the platelet membrane upon both low and high agonist stimulation, membrane-translocated Akt and PAK only phosphorylate at high agonist concentrations, correlating well with platelet aggregation downstream of concomitant Gi and G12/13 pathway activation. PAK inhibitor abolishes Akt phosphorylation, inhibits platelet aggregation in vitro and arterial thrombus formation in vivo. We propose that the PAK-PI3K/Akt pathway mediates platelet aggregation downstream of Gi and G12/13, and PAK may represent a potential antiplatelet and antithrombotic target.

Background and Aims: Chemotherapeutic enteritis is a major dose-limiting adverse reaction to chemotherapy, and few effective drugs are currently available in clinic. Recently the mechanism of intestinal ischemic injury has emerged in the research spotlight due to its notable influence on chemotherapeutic enteritis, but the pathological mechanism of ischemic injury remains unclear. Methods: Irinotecan (CPT-11) was used to establish chemotherapeutic enteritis animal model. Western blotting, gelatin zymography, immunohistochemistry (IHC), intestinal blood flow measurement were used to detect the pathogenesis of ischemia-hypoxia injury. In clinic, the effects of ozone treatment and coagulation examination in patients with chemotherapeutic enteritis were recorded. Results: CPT-11 increased levels of tissue factor (TF) both in the blood and in intestines in mice. Interestingly, the elevation of TF in the blood displayed "double-peak", which was consistent with the intestinal mucosal "double-strike" injury trend. CPT-11 decreased the intestinal blood flow. Moreover, we found that ozone therapy could relieve chemotherapeutic enteritis in mice. We also found that ozone autotransfusion therapy could effectively attenuate chemotherapeutic enteritis and the blood hypercoagulability in cancer patients. Ozone therapy could inhibit TF expression induced by CPT-11 and effectively ameliorate the intestinal mucosal injury in mice. Further research showed that ozone could activate AMPK/SOCS3 to inhibit TLR-4/p38/TF signaling pathway. Conclusions: For the first time, we proposed that TF-induced thrombotic intestinal ischemic injury is a core trigger pathological mechanism of chemotherapeutic enteritis, and provided a new treatment strategy, ozone therapy, to suppress TF expression and treat chemotherapeutic enteritis. Funding Statement: This work was supported by grants from Major Science and Technology Innovation Project of Shandong Province (2018CXGC1220), Major Project of "Science and Technology Innovation Fund" of Nanjing Medical University (2017NJMUCX004), the Natural Science Foundation of Jiangsu province (BK20161033), the Natural Science Foundation of Jiangsu province (BK20181363). Declaration of Interests: The authors declare no conflicts of interest. Ethics Approval Statement: All participants provided written informed consent for participation in the study. This study was approved (2017S053) by the internal review board of the Ethics Committee of Shandong Provincial Qianfoshan Hospital. Animal care and husbandry were carried out in strict accordance with the recommendations of the Guide for the Care and Use of Laboratory Animals (Ministry of Science and Technology of China, 2006) and the Nanjing Medical University Animal Care and Use Committee (NMJU-ACUC). Our study protocol was approved by the NMJU-ACUC, and efforts were made to minimize animal suffering and to reduce the number of mice used.

The tear is a biological fluid that has the diagnostic potential for ocular diseases. Extracellular vesicles (EVs), widly detected in various biofluids including tears, are nanoparticles released by living cells and considered as promising detection sources for noninvasive liquid biopsy. Understanding the roles of tears and tear-EVs in ocular diseases such as dry eye can facilitate the studies of clinical diagnosis, which usually entails detecting such liquid objects with a rapid and effective method. In this study, we used a mass spectrometry-based strategy to analyze peptidome/proteome profiles of tears and EVs for rapid dry eye diagnosis. Nanosized EVs were isolated from tears of both healthy control (HC) individuals and dry eye syndrome (DES) patients, and the tear compositions were further analyzed by tracking their fingerprints with matrix-assisted laser desorption ionization/time-of-flight mass spectrometry. The fingerprints of tear-EVs could be observed in a dose-dependent manner and tears, allowing for comparison of the discriminant peaks between tears and EVs. By analyzing these peaks, the fingerprints of both tear and tear-EVs were showed to have the capability of distinguishing patients with DES from HC donors and providing an efficient way for screening potential DES biomarkers. The proposed tear and EV fingerprinting approach is expected to be a potential tool in the rapid diagnosis of ocular diseases and in-depth research on pathogenesis. Data are available via ProteomeXchange with identifier PXD020217.

Inflammatory bowel disease (IBD) involves chronic inflammation, loss of epithelial integrity, and gastrointestinal microbiota dysbiosis. Effective therapies for IBD have not been established. Accordingly, in this study, we evaluated the effects of inorganic nitrate, a potent nitric oxide (NO) donor and microbiota regulator, in a mouse model of dextran sodium sulfate (DSS)-induced colitis. Mice were pretreated with NaNO3 (2 mM) in their drinking water for 5 days, and NaCl was used as a control. Feces were collected for microbiota analyses. The results showed that oral administration of dietary nitrate could maintained colon consistency, improved colon length, maintained body weight, decreased apoptosis in colon epithelial cells, and ameliorated inflammatory cell infiltration in both the colon and peripheral blood. Microbiota profiling revealed that nitrate regulated dysbiosis. Analysis of the top bacteria at the genus level showed that Bacteroidales_S24-7_group_unidentified, Lactobacillus, Bacteroides, and Prevotellaceae_UCG-001 decreased in the DSS group compared with that in the normal group, whereas Lactobacillus, Ruminococcaceae_UCG-014, and Prevotellaceae_UCG-001 were increased in the DSS + NaNO3 group compared with that in the DSS group. The enriched bacteria in the nitrate group included Gordonibacter, Ureaplasama, and Lachnospiraceae_UCG-006. Moreover, microbiota analysis revealed that nitrate could partially decrease the enriched metabolic pathways (p53 signaling pathway and colorectal cancer pathway) compared with that in the DSS and DSS + NaCl groups. Overall, these findings indicated that nitrate could ameliorate DSS-induced colitis by decreasing inflammation, reducing apoptosis, and regulating the microbiota by activation of the NO3−/NO2−/NO pathway. Nitrate might be a potential treatment for colitis patients in the future clinical application.

Charcoal-stripped fetal bovine serum (CS-FBS) is frequently used in studies on hormone-responsive cancers to provide hormone-free cell culture conditions. CS-FBS may influence the growth of cancer cells; however, the underlying mechanisms remain unclear. In this study, we aimed to clarify the effects of CS-FBS on distinct subtypes of breast cancer cells. We found that the crucial oncoprotein c-Myc was significantly inhibited in estrogen receptor alpha (ER-α)-positive breast cancer cells when cultured in CS-FBS-supplemented medium, but it was not suppressed in ER-α-negative cells. The addition of 17β-estradiol (E2) to CS-FBS-supplemented medium rescued the CS-FBS-induced inhibition of c-Myc, while treatment with 5α-dihydrotestosterone (DHT) suppressed c-Myc expression. Our data demonstrated that CS-FBS may impede the growth of ER-α-positive breast cancer cells via c-Myc inhibition, and this was possibly due to the removal of estrogen. These results highlighted that the core drivers of c-Myc expression were subtype-specific depending on the distinct cell context and special caution should be exercised when using CS-FBS in studies of hormone-responsive cancer cells.

STUDY QUESTION Is the mitochondrial DNA (mtDNA) copy number of cumulus granulosa cells (CGCs) related to the maturation of oocyte cytoplasm? SUMMARY ANSWER Compared with the mtDNA copy number of CGCs from germinal vesicles (GV), CGCs from Metaphase I (MI) oocytes appear to have a lower mtDNA copy number. WHAT IS KNOWN ALREADY The growth and development of CGCs and oocyte are synchronised. The interaction between CGCs and the oocyte provides the appropriate balance of energy, which is necessary for mammalian oocyte development. Moreover, in the oocyte–cumulus complex (OCC), mature oocytes with higher mtDNA copy numbers tend to have corresponding CGCs with higher mtDNA copy numbers. STUDY DESIGN, SIZE, DURATION This is a prospective study of 302 OCCs obtained from 70 women undergoing in vitro fertilisation with intracytoplasmic sperm injection (ICSI) at the Reproductive and Genetic Hospital of CITIC-Xiangya, between 24 February 2018 and 21 December 2019. The CGCs were divided into three groups (GV, MI and MII stages) based on the maturation status of their corresponding oocyte. The sample sizes (n = 302) of CGCs in the three stages were 63 (CGCGV), 70 (CGCMI) and 169 (CGCMII), respectively. Some of the samples (n = 257) was used to quantify the mtDNA copy number, while the rest (n = 45) were used to analyse the expression level of mitochondrial genes. Furthermore, we retrieved 82 immature oocytes from among the 257 OCCs used for mtDNA copy numbers, including 36 GV oocytes and 46 MI oocytes, for analysis of oocyte mtDNA. PARTICIPANTS/MATERIALS, SETTING, METHODS We selected genes with high consistency of real-time PCR results to accurately measure the mtDNA copy number by testing the efficacy and the reproducibility in whole genome amplification (WGA) samples from a human embryonic stem cell line. The CGCs of each oocyte were individually isolated. The mtDNA copy number and gene expression of the CGCs were assessed using real-time PCR techniques. Mitochondrial DNA copy number of the corresponding immature oocytes was also evaluated. MAIN RESULTS AND THE ROLE OF CHANCE MT-ND1, MT-CO1 and β-globin genes were chosen for the assessment of mtDNA content, and mRNA expressions of MT-ND1, MT-CO1, PGC-1α and TFAM were also measured. The genome of 257 CGCs and 82 immature oocytes were amplified according to the multiple displacement amplification (MDA) protocol, and RNA was extracted from 45 CGCs. Compared with CGCGV, CGCMI had a significantly lower mtDNA copy number. In the MT-ND1 assay, the CGCGV: CGCMI was [270 ± 302]: [134 ± 201], P = 0.015. In the MT-CO1 assay, CGCGV: CGCMI was [205 ± 228]: [92 ± 112], P = 0.026. There was no statistical difference in mtDNA between CGCGV and CGCMII. In the MT-ND1 assay, CGCGV: CGCMII was [270 ± 302]: [175 ± 223], P = 0.074. In the MT-CO1 assay, CGCGV: CGCMII was [205 ± 228]: [119 ± 192], P = 0.077. No statistical difference of mtDNA copy number was observed between CGCMI and CGCMII. In the MT-ND1 assay, CGCMI: CGCMII was [134 ± 201]: [175 ± 223], P = 0.422. In the MT-CO1 assay, CGCMI: CGCMII was [92 ± 112]: [119 ± 192], P = 0.478. To verify the reliability of the above results, we further analysed the mtDNA copy number of CGCs of 14 patients with GV, MI and MII oocytes, and the results showed that the mtDNA copy number of CGCMI may be lower. The mtDNA copy number of CGCGV and CGCMI was statistically different in the MT-ND1 assay where CGCGV: CGCMI was [249 ± 173]: [118 ± 113], P = 0.016, but in the MT-CO1 assay, CGCGV: CGCMI was [208 ± 199]: [83 ± 98], P = 0.109. There was no significant difference in mtDNA between CGCGV and CGCMII. In the MT-ND1 assay, CGCGV: CGCMII was [249 ± 173]: [185 ± 200], P = 0.096. In the MT-CO1 assay, CGCGV: CGCMII was [208 ± 199]: [114 ± 139], P = 0.096. There was also no significant difference in mtDNA between CGCMI and CGCMII. In the MT-ND1 assay, CGCMI: CGCMII was [118 ± 113]: [185 ± 200], P = 0.198. In the MT-CO1 assay, CGCMI: CGCMII was [83 ± 98]: [114 ± 139], P = 0.470. Moreover, there were no statistical differences in the expression levels of MT-ND1, MT-CO1, PGC-1α and TFAM between CGCGV, CGCMI and CGCMII (P > 0.05). LARGE SCALE DATA N/A LIMITATIONS, REASONS FOR CAUTION Due to the ethical issues, the study did not quantify the mtDNA content of MII oocytes. Thus, whether the change in mtDNA copy number in CGCs is related to the different developmental stages of oocytes has not been further confirmed. Moreover, the sample size was relatively small. WIDER IMPLICATIONS OF THE FINDINGS The mtDNA copy number of CGCs decreases from the GV phase to the MI phase and stays steady from the MI to MII stage. At different stages of oocyte maturation, the mtDNA of CGCs may undergo self-degradation and replication to meet the energy requirements of the corresponding oocyte and the maturation of the oocyte cytoplasm. STUDY FUNDING/COMPETING INTEREST(S) Funding was provided by the National Key R&D Program of China (Grant 2018YFC1003100, to L.H.), the science and technology major project of the Ministry of Science and Technology of Hunan Province, China (grant 2017SK1030, to G.L.), the National Natural Science Foundation of China (grant 81873478, to L.H.), and Merck Serono China Research Fund for Fertility Experts (to L.H.). There is no conflict of interest.

Alterations of gut microbiota have been implicated in multiple diseases including cancer. However, the gut microbiota spectrum in lung cancer remains largely unknown. Here we profiled the gut microbiota composition in a discovery cohort containing 42 early-stage lung cancer patients and 65 healthy individuals through the 16S ribosomal RNA (rRNA) gene sequencing analysis. We found that lung cancer patients displayed a significant shift of microbiota composition in contrast to the healthy populations. To identify the optimal microbiota signature for noninvasive diagnosis purpose, we took advantage of Support-Vector Machine (SVM) and found that the predictive model with 13 operational taxonomic unit (OTU)-based biomarkers achieved a high accuracy in lung cancer prediction (area under curve, AUC = 97.6%). This signature performed reasonably well in the validation cohort (AUC = 76.4%), which contained 34 lung cancer patients and 40 healthy individuals. To facilitate potential clinical practice, we further constructed a 'patient discrimination index' (PDI), which largely retained the prediction efficiency in both the discovery cohort (AUC = 92.4%) and the validation cohort (AUC = 67.7%). Together, our study uncovered the microbiota spectrum of lung cancer patients and established the specific gut microbial signature for the potential prediction of the early-stage lung cancer.

Since December 2019, there has been an outbreak of a novel coronavirus (COVID-19) infection in Wuhan, China. Meanwhile, the outbreak also drew attention and concern from the World Health Organization (WHO). COVID-19 is another human infectious disease caused by coronavirus. The transmission of COVID-19 is potent and the infection rate is fast. Since there is no specific drug for COVID-19, the treatment is mainly symptomatic supportive therapy. In addition, it should be pointed out that patients with severe illness need more aggressive treatment and meticulous care. Recently, accurate RNA detection has been decisive for the diagnosis of COVID-19. The development of highly sensitive RT-PCR has facilitated epidemiological studies that provide insight into the prevalence, seasonality, clinical manifestations and course of COVID-19 infection. In this review, we summarize the epidemiology and characteristics of COVID-19.

Clinical observation of the association between cancer aggressiveness and embryonic development stage implies the importance of developmental signals in cancer initiation and therapeutic resistance. However, the dynamic gene expression during organogenesis and the master oncofetal drivers are still unclear, which impeded the efficient elimination of poor prognostic tumors, including human hepatocellular carcinoma (HCC). In this study, human embryonic stem cells were induced to differentiate into adult hepatocytes along hepatic lineages to mimic liver development in vitro. Combining transcriptomic data from liver cancer patients with the hepatocyte differentiation model, the active genes derived from different hepatic developmental stages and the tumor tissues were selected. Bioinformatic analysis followed by experimental assays was used to validate the tumor subtype-specific oncofetal signatures and potential therapeutic values. Hierarchical clustering analysis revealed the existence of two subtypes of liver cancer with different oncofetal properties. The gene signatures and their clinical significance were further validated in an independent clinical cohort and The Cancer Genome Atlas database. Upstream activator analysis and functional screening further identified E2F1 and SMAD3 as master transcriptional regulators. Small-molecule inhibitors specifically targeting the oncofetal drivers extensively down-regulated subtype-specific developmental signaling and inhibited tumorigenicity. Liver cancer cells and primary HCC tumors with different oncofetal properties also showed selective vulnerability to their specific inhibitors. Further precise targeting of the tumor initiating steps and driving events according to subtype-specific biomarkers might eliminate tumor progression and provide novel therapeutic strategy.

Human embryonic stem cells (hESCs) have unlimited proliferation capacity and can differentiate into most types of somatic cells. We previously described that the overexpression of FLI1 as well as the activation of protein kinase C (FLI1-PKC) could rapidly and efficiently differentiate hESCs into endothelial cells (ECs). However, the relationship between vascular endothelial growth factor (VEGF) and PKC in hESC-EC differentiation is debated, and the roles of different PKC isoforms in hESC-EC differentiation remain unknown. In this study, after 2 days of induction, we found that the overexpression of FLI1 and the addition of VEGF-A to hESCs (FLI1-VEGF) could generate 19.6% ± 5.4% ECs. The induction efficiency of ECs was reduced from 72.3% ± 7.6% to 37.9% ± 4.9% following the addition VEGF-A siRNA to the FLI1-PKC system. However, the induction of ECs was nearly completely abrogated following the addition of the pan-PKC inhibitor to the FLI1-VEGF, FLI1-PKC, or other systems. The above results suggested that VEGF can partially replace PKC, but PKC plays a more critical role downstream of VEGF during hESC-EC induction. To further investigate which PKC isoform was mainly involved in converting hESCs to ECs, siRNAs were used to knock down nine PKC isoforms in the FLI1-PKC system. Only the knockdown of PKC-ɛ or PKC-η significantly decreased the induction efficiency of ECs to 51.1% ± 5.8% or 50.3% ± 5.1%, respectively. PKC-ɛ/η siRNA could suppress EC induction in other differentiation systems. Moreover, RNA-seq and quantitative polymerase chain reaction analysis also showed that only the expression of PKC-ɛ and PKC-η was robustly upregulated during hESC-EC induction. In summary, our results suggested that VEGF promoted the differentiation of ECs derived from hESCs, which mainly depended on PKC, specifically the PKC-ɛ and PKC-η pathways.

Background Mechanical thrombectomy (MT) is an effective treatment for large-vessel occlusion in acute ischemic stroke, however, only some revascularized patients have a good prognosis. For stroke patients undergoing MT, predicting the risk of unfavorable outcomes and adjusting the treatment strategies accordingly can greatly improve prognosis. Therefore, we aimed to develop and validate a nomogram that can predict 3-month unfavorable outcomes for individual stroke patient treated with MT. Methods We analyzed 258 patients with acute ischemic stroke who underwent MT from January 2018 to February 2021. The primary outcome was a 3-month unfavorable outcome, assessed using the modified Rankin Scale (mRS), 3–6. A nomogram was generated based on a multivariable logistic model. We used the area under the receiver-operating characteristic curve to evaluate the discriminative performance and used the calibration curve and Spiegelhalter’s Z-test to assess the calibration performance of the risk prediction model. Results In our visual nomogram, gender (odds ratio [OR], 3.40; 95%CI, 1.54–7.54), collateral circulation (OR, 0.46; 95%CI, 0.28–0.76), postoperative mTICI (OR, 0.06; 95%CI, 0.01–0.50), stroke-associated pneumonia (OR, 5.76; 95%CI, 2.79–11.87), preoperative Na (OR, 0.82; 95%CI, 0.72–0.92) and creatinine (OR, 1.02; 95%CI, 1.01–1.03) remained independent predictors of 3-month unfavorable outcomes in stroke patients treated with MT. The area under the nomogram curve was 0.8791 with good calibration performance (P = 0.873 for the Spiegelhalter’s Z-test). Conclusions A novel nomogram consisting of gender, collateral circulation, postoperative mTICI, stroke-associated pneumonia, preoperative Na and creatinine can predict the 3-month unfavorable outcomes in stroke patients treated with MT.

Objective To demonstrate the applicability and adaptability of uterine fibroid symptoms and quality of life (UFS-QoL) in assessing the efficacy of treatment in Chinese populations. Methods This is a secondary analysis of a prospective cohort study involving 20 Chinese hospitals and 2,411 Chinese women with fibroids. Patients completed UFS-QoL and short form-36 (SF-36) at pre-surgery, 6-month and 12-month post-treatments. Internal consistency of the quality of life assessed by the UFS-QoL questionnaire using Cronbach’s α coefficient (α). Principal axis factor analysis with orthogonal rotation was established to investigate relationships between items and subscales. Concurrent validity refers to the Spearman's correlation estimate of the correlation between UFS-QoL and SF-36. Using effect size and standardized response mean, the ability to detect change was evaluated by comparing pre- and post-6-month and post-12-month treatment scores. Results Exploratory factor analysis yielded six subscales (concern, activities, energy/mood, control, self-consciousness, and sexual function) with eigenvalues > 1 in UFS-QoL. A 63.61% total variance was explained by the test items. Ceiling effects of self-consciousness and sexual functioning subscales from UFS-QoL were > 15%. UFS-QoL showed a positive and moderate correlation with SF-36 to establish good concurrent validity. And showed good consistency reliability (Cronbach α > 0.7 in all subscales), ability to detect change after treatment. This excluded self-consciousness (α = 0.56), which demonstrated the lowest effect size (0.38) and standardized response means (0.38) 6- and 12-months post-treatment. Conclusions Symptom severity, activity, and mood subscales of the Chinese UFS-QoL were valid and reliable. However, the self-consciousness domain needs further investigation on cultural adaptation, such as cognitive debriefing for how Chinese interpret these questions.

To compare the diagnostic power of strip meniscometry (SM), Schirmer test (ST), and tear meniscus (TM) in mild dry eye disease (DED) and to evaluate the association with DED-related parameters.Forty left eyes with mild DED and 40 left eyes of control participants were investigated. All participants underwent a comprehensive ocular surface examination, including the Ocular Surface Disease Index (OSDI), fluorescein tear film break-up time (FTBUT), ocular surface staining grades, meiboscores, and tear film volume examinations, including SM, ST, tear meniscus height (TMH), and tear meniscus cross-sectional area (TMA) measurements, respectively, by optical coherence tomography (OCT) and Keratograph 5M (K5M). The correlation between these parameters was evaluated, and the receiver operating characteristic (ROC) curve was used to verify the diagnostic power by the area under the curve (AUC).All tear film volume examinations significantly correlated with DED parameters. Among them, the most relevant factor to OSDI scores and FTBUT was SM. In addition, SM (AUC = 0.992), TMH-OCT (AUC = 0.978), and TMA-OCT (AUC = 0.960) showed better diagnostic power than ST (AUC = 0.650) in DED, in which the cutoff value of SM was 3.5 mm (sensitivity, 97.5%; specificity, 95.0%).Compared with ST, SM and TM parameters obtained by OCT were more relevant to ocular surface parameters and can provide a more valuable approach to discriminate mild DED from control participants.This study made a comprehensive comparison of the existing tear volume detection methods and provided a basis for the clinical selection of appropriate detection methods and the diagnosis of mild DED.

Background The development of morphine tolerance is a clinical challenge for managing severe pain. Studies have shown that neuroinflammation is a critical aspect for the development of analgesic tolerance. We found that AMPK-autophagy activation could suppress neuroinflammation and improve morphine tolerance via the upregulation of suppressor of cytokine signaling 3 (SOCS3) by inhibiting the processing and maturation of microRNA-30a-5p. Methods CD-1 mice were utilized for the tail-flick test to evaluate morphine tolerance. The microglial cell line BV-2 was utilized to investigate the mechanism of AMPK-autophagy-mediated posttranscriptional regulation of SOCS3. Proinflammatory cytokines were measured by western blotting and real-time PCR. The levels of SOCS3 and miRNA-processing enzymes were evaluated by western blotting, real-time PCR and immunofluorescence staining. Results Based on experimental verification, miRNA-30a-5p could negatively regulate SOCS3. The AMPK activators AICAR, resveratrol and metformin downregulated miRNA-30a-5p. We found that AMPK activators specifically inhibited the processing and maturation of miRNA-30a-5p in microglia by degrading DICER and AGO2 via autophagy. Furthermore, a miRNA-30a-5p inhibitor significantly improved morphine tolerance via upregulation of SCOS3 in mice. It markedly increased the level of SOCS3 in the spinal cord of mice and subsequently inhibited morphine-induced phosphorylation of NF-κB p65. In addition, a miRNA-30a-5p inhibitor decreased the levels of IL-1β and TNF-α caused by morphine in microglia. Conclusion AMPK-autophagy activation suppresses neuroinflammation and improves morphine tolerance via the upregulation of SOCS3 by inhibiting miRNA-30a-5p.

Background Severe acute pancreatitis (SAP) is the most common gastrointestinal disease and is associated with unpredictable seizures and high mortality rates. Despite improvements in the treatment of acute pancreatitis, the timely and accurate diagnosis of SAP remains highly challenging. Previous research has shown that extracellular vesicles (EVs) in the plasma have significant potential for the diagnosis of SAP since the pancreas can release EVs that carry pathological information into the peripheral blood in the very early stages of the disease. However, we know very little about the metabolites of EVs that might play a role in the diagnosis of SAP. Methods Here, we performed quantitative metabolomic analyses to investigate the metabolite profiles of EVs isolated from SAP plasma. We also determined the metabolic differences of EVs when compared between healthy controls, patients with SAP, and those with mild acute pancreatitis (MAP). Results A total of 313 metabolites were detected, mainly including organic acids, amino acids, fatty acids, and bile acids. The results showed that the metabolic composition of EVs derived from SAP and MAP was significantly different from those derived from healthy controls and identified specific differences between EVs derived from patients with SAP and MAP. On this basis, we identified four biomarkers from plasma EVs for SAP detection, including eicosatrienoic acid (C20:3), thiamine triphosphate, 2-Acetylfuran, and cis-Citral. The area under the curve (AUC) was greater than 0.95 for both discovery (n = 30) and validation (n = 70) sets. Conclusions Our data indicate that metabolic profiling analysis of plasma EVs and the screening of potential biomarkers are of significant potential for improving the early diagnosis and severity differentiation of acute pancreatitis. Graphical abstract

Background Evidence referring to the trade-offs between the benefits and risks of single embryo transfer (SET) versus double embryo transfer (DET) following assisted reproduction technology are insufficient, especially for those women with a defined embryo quality or advanced age. Methods A systematic review and meta-analysis was conducted according to PRISMA guidelines. PubMed, EMBASE, Cochrane Library and ClinicalTrials.gov were searched based on established search strategy from inception through February 2021. Pre-specified primary outcomes were live birth rate (LBR) and multiple pregnancy rate (MPR). Odds ratio (OR) with 95% confidence interval (CI) were pooled by a random-effects model using R version 4.1.0. Results Eighty-five studies (14 randomized controlled trials and 71 observational studies) were eligible. Compared with DET, SET decreased the probability of a live birth (OR = 0.78, 95% CI: 0.71–0.85, P < 0.001, n = 62), and lowered the rate of multiple pregnancy (0.05, 0.04–0.06, P < 0.001, n = 45). In the sub-analyses of age stratification, both the differences of LBR (0.87, 0.54–1.40, P = 0.565, n = 4) and MPR (0.34, 0.06–2.03, P = 0.236, n = 3) between SET and DET groups became insignificant in patients aged ≥40 years. No significant difference in LBR for single GQE versus two embryos of mixed quality [GQE + PQE (non-good quality embryo)] (0.99, 0.77–1.27, P = 0.915, n = 8), nor any difference of MPR in single PQE versus two PQEs (0.23, 0.04–1.49, P = 0.123, n = 6). Moreover, women who conceived through SET were associated with lower risks of poor outcomes, including cesarean section (0.64, 0.43-0.94), antepartum haemorrhage (0.35, 0.15-0.82), preterm birth (0.25, 0.21-0.30), low birth weight (0.20, 0.16-0.25), Apgar1 Conclusions In women aged

PURPOSE: To evaluate the cytogenetic risk of assisted reproductive technology (ART) by comparing the incidence of de novo chromosomal abnormalities between fetuses conceived via in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) and natural conception. MATERIALS AND METHODS: Prenatal invasive diagnostic testing (amniocentesis and cytogenetic analysis) was performed on 1496 fetuses conceived via IVF/ICSI (IVF/ICSI group) and 1396 fetuses from natural conception (NC group). The incidence of de novo chromosomal abnormalities (including aneuploidy and chromosomal structure abnormalities) was used to evaluate the cytogenetic risk of ART. For statistical analysis, χ(2)-test was used for binary dependent variable. The significance level was P 0.05). The incidence of abnormal karyotypes was also not significantly different between the IVF/ICSI and NC groups in different maternal ages, including ≥ 35 years group (7.55% vs. 9.60%, χ(2) = 1.40, P > 0.05) and 0.05). Moreover, there was no difference in the proportion of aneuploid and structural abnormalities in detected karyotypes between the IVF/ICSI and NC groups. Logistic regression analysis showed no significant association between the method of pregnancy and de novo chromosomal abnormalities (odds ratio (OR) 1.03; 95% CI 0.71–1.50; P = 0.86) after adjusting for other confounding factors. CONCLUSION(S). Fetuses conceived via IVF/ICSI had a slight but not statistically significant increase in de novo abnormal karyotypes compared to those in naturally conceived fetuses. Our findings indicate no significant association between de novo fetal chromosomal abnormalities and the pregnancy method in high-risk pregnancies in the second trimester. For these pregnancies with a high risk but with a normal karyotype, further genetic testing is required for diagnosis. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10815-022-02500-5.

In recent years, a rapid development of polymeric hydrogel-based sensors has been witnessed. However, conventional hydrogels often exhibit poor mechanical properties. Additionally, the use of these sensors at temperatures0 °C is limited due to the freezing of the water molecules in the hydrogel matrix. In this study, graphene oxide/poly(acrylamide

Purpose. This study aimed to evaluate the medium-term fit of soft contact lenses (SCLs) and its effects on dry eye by using ultra-high-resolution optical coherence tomography (UHR-OCT) and digital slit-lamp biomicroscopy (DSLB) and identify factors indicating a better contact lens fit. Methods. A total of 14 participants (7 men, 7 women; mean age, 23.1 years) were recruited. Temporal lag and conjunctival coverage were imaged by UHR-OCT. DSLB was used to assess lens decentration and blink-induced movement. In addition, dry eye indices, including ocular surface disease index (OSDI), tear break-up time (TBUT), and Schirmer I test scores, were measured at baseline and 1 day, 1 week, 1 month, 3 months, and 6 months after wearing. Results. Good repeatability was found in the assessments of lens fit characteristics by two researchers and in the assessments by one researcher at different follow-up times. There were no significant differences in lens fit characteristics, TBUT and Schirmer I test scores at each checkpoint (P >0.05). However, the OSDI was significantly higher than baseline after wearing (P

Objective To investigate how serum extracellular vesicles (EVs)-carried miRNA-181b-5p affected the proliferation, cell cycle and apoptosis of acute lymphoblastic leukemia (ALL) cells. Methods Differentially expressed miRNAs related to ALL were screened by bioinformatics analysis, and the localization of target miRNA was searched by its expression. qRT-PCR was adopted to confirm the expression of miRNA-181b-5p. Flow cytometry and fluorescence microscopy were applied to evaluate EVs internalization. MTT assay was employed to verify the proliferation of ALL cells. Cell cycle and apoptosis were analyzed by flow cytometry. Transwell assay was applied to evaluate migration and invasion abilities. Results High expression of miRNA-181b-5p was proved in ALL cell lines, and miRNA-181b-5p enriched in the exosomes and vesicles of blood cells. In the meantime, it was found that EVs carrying miRNA-181b-5p could be internalized by ALL cells and thus the expression of miRNA-181b-5p was up-regulated. Cell function assays showed that the proliferation, migration, invasion abilities of ALL cell lines were promoted in miRNA-181b-5p mimic group or the group co-culturing ALL-derived EVs and BALL-1 cell lines. The percentage of cells in G0/G1 phase was reduced and cell apoptosis was also inhibited. Conclusion miRNA-181b-5p carried by EVs in peripheral blood of ALL patients can enter ALL cells and thus promote the malignancy of ALL cells.