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The aluminum alloy components on automobile often experience high strain rate deformation during accidents. The dynamic deformation behavior and constitutive model are crucial for analysis of vehicle body collisions and failures. In this work, the 7A75 aluminum alloy plate was heat treated according to standard body manufacturing procedures. Quasi-static and dynamic tensile experiments were conducted on the aged 7A75 alloy in a wide strain rate range from 0.001 s−1 to 500 s−1. The dynamic deformation behavior of the alloy was revealed, and the constitutive models were established. It was found that the 7A75 alloy has strong strain rate sensitivity, and its mechanical properties can be significantly improved with the increase of strain rate. As the strain rate increases, the necking instability strain shifts towards higher values, which leads to a larger uniform deformation ability and fracture elongation. This behavior results in a better energy absorption capacity at high strain rates, thus providing a good collision protection property. The constitutive models, including Johnson-Cook (J-C) and Zerilli-Armstrong (Z-A) types, were established and compared. Note that the conventional J-C and Z-A models exhibit severe prediction distortion. Therefore, a modified J-C model incorporating strain rate compensation was introduced, which decrease the errors by 149% and achieves accurate predictions of both dynamic and static deformation behaviors over a wide range of strain rates.

Abstract Background Antimicrobial resistance (AMR) is a major threat to children’s health, particularly in respiratory infections. Accurate identification of pathogens and AMR is crucial for targeted antibiotic treatment. Metagenomic next-generation sequencing (mNGS) shows promise in directly detecting microorganisms and resistance genes in clinical samples. However, the accuracy of AMR prediction through mNGS testing needs further investigation for practical clinical decision-making. Methods We aimed to evaluate the performance of mNGS in predicting AMR for severe pneumonia in pediatric patients. We conducted a retrospective analysis at a tertiary hospital from May 2022 to May 2023. Simultaneous mNGS and culture were performed on bronchoalveolar lavage fluid samples obtained from pediatric patients with severe pneumonia. By comparing the results of mNGS detection of microorganisms and antibiotic resistance genes with those of culture, sensitivity, specificity, positive predictive value, and negative predictive value were calculated. Results mNGS detected bacterial in 71.7% cases (86/120), significantly higher than culture (58/120, 48.3%). Compared to culture, mNGS demonstrated a sensitivity of 96.6% and a specificity of 51.6% in detecting pathogenic microorganisms. Phenotypic susceptibility testing (PST) of 19 antibiotics revealed significant variations in antibiotics resistance rates among different bacteria. Sensitivity prediction of mNGS for carbapenem resistance was higher than penicillins and cephalosporin (67.74% vs. 28.57%, 46.15%), while specificity showed no significant difference (85.71%, 75.00%, 75.00%). mNGS also showed a high sensitivity of 94.74% in predicting carbapenem resistance in Acinetobacter baumannii. Conclusions mNGS exhibits variable predictive performance among different pathogens and antibiotics, indicating its potential as a supplementary tool to conventional PST. However, mNGS currently cannot replace conventional PST.

Hereditary fructose intolerance (HFI) is an autosomal recessive metabolic disease associated with a mutation in the aldolase B gene on chromosome 9q31. In this study, we generated a human-induced pluripotent stem cell (hiPSC) line, FDCHi015-A, from peripheral blood mononuclear cells (PBMCs) of a patient carrying the compound heterozygous mutations c.360_364delCAAA and c.1013C > T in exons 4 and 9 of the ALDOB gene, respectively. The iPSCs with the confirmed patient-specific mutation demonstrate pluripotency markers expression, a normal karyotype, and the ability to differentiate into derivatives of three germ layers.

ObjectiveFructose-1,6-bisphosphatase deficiency (FBP1D) is a rare inborn error due to mutations in the FBP1 gene. The genetic spectrum of FBP1D in China is unknown, also nonspecific manifestations confuse disease diagnosis. We systematically estimated the FBP1D prevalence in Chinese and explored genotype-phenotype association.MethodsWe collected 101 FBP1 variants from our cohort and public resources, and manually curated pathogenicity of these variants. Ninety-seven pathogenic or likely pathogenic variants were used in our cohort to estimate Chinese FBP1D prevalence by three methods: 1) carrier frequency, 2) permutation and combination, 3) Bayesian framework. Allele frequencies (AFs) of these variants in our cohort, China Metabolic Analytics Project (ChinaMAP) and gnomAD were compared to reveal the different hotspots in Chinese and other populations. Clinical and genetic information of 122 FBP1D patients from our cohort and published literature were collected to analyze the genotype-phenotypes association. Phenotypes of 68 hereditary fructose intolerance (HFI) patients from our previous study were used to compare the phenotypic differences between these two fructose metabolism diseases.ResultsThe estimated Chinese FBP1D prevalence was 1/1,310,034. In the Chinese population, c.490G>A and c.355G>A had significantly higher AFs than in the non-Finland European population, and c.841G>A had significantly lower AF value than in the South Asian population (all p values < 0.05). The genotype-phenotype association analyses showed that patients carrying homozygous c.841G>A were more likely to present increased urinary glycerol, carrying two CNVs (especially homozygous exon1 deletion) were often with hepatic steatosis, carrying compound heterozygous variants were usually with lethargy, and carrying homozygous variants were usually with ketosis and hepatic steatosis (all p values < 0.05). By comparing to phenotypes of HFI patients, FBP1D patients were more likely to present hypoglycemia, metabolic acidosis, and seizures (all p-value < 0.05).ConclusionThe prevalence of FBP1D in the Chinese population is extremely low. Genetic sequencing could effectively help to diagnose FBP1D.

Neurodevelopmental disorder with or without autistic features and/or structural brain abnormalities (NEDASB) is a rare autosomal dominant disorder caused by a heterozygous mutation in the NOVA2 gene on chromosome 19q13. Here, we describe the generation and characterization of an iPSC line derived from the peripheral blood of a 7-year-old patient carrying a novel heterozygous mutation in NOVA2 (c.625 del). The iPSCs with the confirmed patient-specific mutation were demonstrated by pluripotency markers, a normal karyotype, and the ability to differentiate into three germ layers. This NOVA2-mutant iPSC line could facilitate disease modeling and therapy development studies for NEDASB.

Abstract Background In China, ~1,072,100 small for gestational age (SGA) births occur annually. These SGA newborns are a high-risk population of developmental delay. Our study aimed to evaluate the genetic profile of SGA newborns in the newborn intensive care unit (NICU) and establish a prognosis prediction model by combining clinical and genetic factors. Methods A cohort of 723 SGA and 1317 appropriate for gestational age (AGA) newborns were recruited between June 2018 and June 2020. Clinical exome sequencing was performed for each newborn. The gene-based rare-variant collapsing analyses and the gene burden test were applied to identify the risk genes for SGA and SGA with poor prognosis. The Gradient Boosting Machine framework was used to generate two models to predict the prognosis of SGA. The performance of two models were validated with an independent cohort of 115 SGA newborns without genetic diagnosis from July 2020 to April 2022. All newborns in this study were recruited through the China Neonatal Genomes Project (CNGP) and were hospitalized in NICU, Children’s Hospital of Fudan University, Shanghai, China. Results Among the 723 SGA newborns, 88(12.2%) received genetic diagnosis, including 42(47.7%) with monogenic diseases and 46(52.3%) with chromosomal abnormalities. SGA with genetic diagnosis showed higher rates in severe SGA(54.5% vs. 41.9%, P=0.0025) than SGA without genetic diagnosis. SGA with chromosomal abnormalities showed higher incidences of physical and neurodevelopmental delay compared to those with monogenic diseases (45.7% vs. 19.0%, P=0.012). We filtered out 3 genes (ITGB4, TXNRD2, RRM2B) as potential causative genes for SGA and 1 gene (ADIPOQ) as potential causative gene for SGA with poor prognosis. The model integrating clinical and genetic factors demonstrated a higher area under the receiver operating characteristic curve (AUC) over the model based solely on clinical factors in both the SGA-model generation dataset (AUC=0.9[95% confidence interval 0.84–0.96] vs. AUC=0.74 [0.64–0.84]; P=0.00196) and the independent SGA-validation dataset (AUC=0.76 [0.6–0.93] vs. AUC=0.53[0.29–0.76]; P=0.0117). Conclusion SGA newborns in NICU presented with roughly equal proportions of monogenic and chromosomal abnormalities. Chromosomal disorders were associated with poorer prognosis. The rare-variant collapsing analyses studies have the ability to identify potential causative factors associated with growth and development. The SGA prognosis prediction model integrating genetic and clinical factors outperformed that relying solely on clinical factors. The application of genetic sequencing in hospitalized SGA newborns may improve early genetic diagnosis and prognosis prediction.

α-L-Rhamnosidases with desirable activity and thermostability profiles could be used for the biocatalytic production of the flavonoid glucoside isoquercetin from natural rutin for functional food. Herein, to improve the catalytic activity of GH78 α-L-rhamnosidase BtRha78A from Bacteroides thetaiotaomicron VPI-5482, a list of residues located at the conserved general acid motif were selected for targeted mutagenesis by the sequence alignment of BtRha78A with homologous α-L-rhamnosidases. Ala-scanning mutagenesis and site-directed mutagenesis based on sequence alignment were performed, and the relative activity on rutin was evaluated. Furthermore, the reaction time curves and enzyme kinetics of better mutants were determined. The results indicate that the conversion rates of mutants V338A, V338I, S340A, and G341A were increased by 21.3%, 20.1%, 13.2%, and 1.6%, respectively, compared with the wild type when using whole-cell biotransformation. Moreover, the catalytic efficiency kcat/KM value of mutant V338A was 1.3-fold higher than that of the wild type. The best mutant, V338A, was employed for the enzymatic preparation of isoquercetin via the biotransformation of rutin at a concentration of 2 mM, and 1.80 g of isoquercetin was obtained. The identification of the best mutant V338A lays the foundation for the efficient preparation of isoquercetin via the biotransformation of rutin, which in turn provides theoretical guidance for its large-scale production.

Background: Neurodevelopmental disorders (NDDs) are a group of diseases that severely affect the physical and mental health of children. The PPP2R5D gene encodes B56δ, the regulatory subunit of protein phosphatase 2A (PP2A). NDDs related to the PPP2R5D gene have recently been defined as Houge–Janssens syndrome 1. Methods: Clinical/whole exome sequencing was performed on approximately 3000 patients with NDDs from 2017 to 2023. In vitro experiments were performed to assess the impairment of variants to protein expression and the assembly of PP2A holoenzyme. The genetic information and phenotypes of the reported patients, as well as patients in this study, were summarized, and the genotype–phenotype relationship was analyzed. The probability of pathogenic missense variants in PPP2R5D was predicted using AlphaMissense (AM), and the relationship between certain phenotype and 3D protein structural features were analyzed. Results: Thirteen new patients carrying twelve PPP2R5D gene variants were detected, including five novel missense variants and one novel frameshift variant. In vitro experiments revealed that the frameshift variant p.H463Mfs*3 resulted in a ~50 kDa truncated protein with lower expression level. Except for E420K and T536R, other missense variants impaired holoenzyme assembly. Furthermore, we found that pathogenic/likely pathogenic (P/LP) variants that have been reported so far were all missense variants and clustered in three conserved regions, and the likelihood of P/LP mutations located in these conserved regions was extremely high. In addition, the macrocephaly phenotype was related to negatively charged residues involved in substrate recruitment. Conclusions: We reported thirteen new patients with PPP2R5D gene variants and expanded the PPP2R5D variant spectrum. We confirmed the pathogenicity of novel variants through in vitro experiments. Our findings in genotype–phenotype relationship provide inspiration for genetic counseling and interpretation of variants. We also provide directions for further research on the mechanism of macrocephaly phenotype.

Objective: We aimed to investigate the clinical and genetic risk factors associated with neonatal severe unconjugated hyperbilirubinemia.Methods: This was a retrospective, 1:1 matched, case–control study. We included 614 neonates diagnosed with severe unconjugated hyperbilirubinemia (serum total bilirubin level ≥425 μmol/L or serum total bilirubin concentration that met exchange transfusion criteria) from the China Neonatal Genomes Project in Children’s Hospital of Fudan University. Clinical exome sequencing data were analyzed using a data analysis pipeline of Children’s Hospital of Fudan University. The factors associated with severe unconjugated hyperbilirubinemia were assessed using univariable and multivariable logistic regression analyses. Interaction analyses were examined between clinical and genetic risk factors.Results: ABO/Rh incompatibility hemolysis (odds ratio [OR] 3.36, 95% confidence interval [CI] 2.32–4.86), extravascular hemorrhage (OR 2.95, 95% CI 2.24–3.89), weight loss (OR 5.46, 95% CI 2.88–10.36), exclusive breastmilk feeding (OR 3.56, 95% CI 2.71–4.68), and the homozygous mutant of UGT1A1 211G>A (OR 2.35, 95% CI 1.54–3.59) were all identified as factors significantly associated with severe unconjugated hyperbilirubinemia. The presence of UGT1A1 211G>A mildly increased the risk of severe unconjugated hyperbilirubinemia caused by ABO/Rh incompatibility hemolysis (OR 3.98, 95% CI 2.19–7.23), although the effect is not statistically significant.Conclusion: ABO/Rh incompatibility hemolysis, extravascular hemorrhage, weight loss, exclusive breastmilk feeding, and the homozygous mutant of UGT1A1 211G>A were found to be risk factors for severe unconjugated hyperbilirubinemia. Clinical factors remain the most crucial and preventable determinants in managing severe unconjugated hyperbilirubinemia, with a minimal genetic contribution. The establishment of preconception care practices and the reinforcement of screening for the aforementioned risk factors are essential steps for preventing severe unconjugated hyperbilirubinemia.

Abstract The Mulibrey (Muscle–liver–brain–eye) nanism caused by loss-of-function variants in TRIM37 gene is an autosomal recessive disorder characterized by severe growth failure and constrictive pericarditis. These patients also suffer from severe respiratory infections, co-incident with an increased mortality rate. Here, we revealed that TRIM37 variants were associated with recurrent infection. Trim37 FIN major (a representative variant of Mulibrey nanism patients) and Trim37 knockout mice were susceptible to influenza virus infection. These mice showed defects in follicular helper T (TFH) cell development and antibody production. The effects of Trim37 on TFH cell differentiation relied on its E3 ligase activity catalyzing the K27/29-linked polyubiquitination of Bcl6 and its MATH domain-mediated interactions with Bcl6, thereby protecting Bcl6 from proteasome-mediated degradation. Collectively, these findings highlight the importance of the Trim37-Bcl6 axis in controlling the development of TFH cells and the production of high-affinity antibodies, and further unveil the immunologic mechanism underlying recurrent respiratory infection in Mulibrey nanism.

Abstract. Background:. Significant brain volume deviation is an essential phenotype in children with neurodevelopmental delay (NDD), but its genetic basis has not been fully characterized. This study attempted to analyze the genetic factors associated with significant whole-brain deviation volume (WBDV). Methods:. We established a reference curve based on 4222 subjects ranging in age from the first postnatal day to 18 years. We recruited only NDD patients without acquired etiologies or positive genetic results. Cranial magnetic resonance imaging (MRI) and clinical exome sequencing (2742 genes) data were acquired. A genetic burden test was performed, and the results were compared between patients with and without significant WBDV. Literature review analyses and BrainSpan analysis based on the human brain developmental transcriptome were performed to detect the potential role of genetic risk factors in human brain development. Results:. We recruited a total of 253 NDD patients. Among them, 26 had significantly decreased WBDV (+2 SDs). NDD patients with significant WBDV had higher rates of motor development delay (49.8% [106/213] vs. 75.0% [30/40], P = 0.003) than patients without significant WBDV. Genetic burden analyses found 30 genes with an increased allele frequency of rare variants in patients with significant WBDV. Analyses of the literature further demonstrated that these genes were not randomly identified: burden genes were more related to the brain development than background genes (P = 1.656e–9). In seven human brain regions related to motor development, we observed burden genes had higher expression before 37-week gestational age than postnatal stages. Functional analyses found that burden genes were enriched in embryonic brain development, with positive regulation of synaptic growth at the neuromuscular junction, positive regulation of deoxyribonucleic acid templated transcription, and response to hormone, and these genes were shown to be expressed in neural progenitors. Based on single cell sequencing analyses, we found TUBB2B gene had elevated expression levels in neural progenitor cells, interneuron, and excitatory neuron and SOX15 had high expression in interneuron and excitatory neuron. Conclusion:. Idiopathic NDD patients with significant brain volume changes detected by MRI had an increased prevalence of motor development delay, which could be explained by the genetic differences characterized herein.

Abstract Animal models play crucial roles in the rapid development of vaccines/drugs for the prevention and therapy of COVID-19, but current models have some deficits when studying the pathogenesis of SARS-CoV-2 on some special tissues or organs. Here, we generated a human ACE2 and SARS-CoV-2 NF/F knockin mouse line that constitutively expresses human ACE2 and specifically expresses SARS-CoV-2 N gene induced by Cre-recombinase. By crossing with Cre transgenic lines allowing for lung-specific and constitutive expression, we generated lung-specific (Sftpc-hACE2-NF/F) and constitutive SARS-CoV-2 N (EIIa-hACE2-NF/F) expressing mice. Upon intranasal infection with a SARS-CoV-2 GFP/ΔN strain which can only replicate in SARS-CoV-2 N expressed cells, we demonstrated that both the Sftpc-hACE2-NF/F and EIIa-hACE2-NF/F mice support viral replication. Consistent with our design, viral replication was limited to the lung tissues in Sftpc-hACE2-NF/F mice, while the EIIa-hACE2-NF/F mice developed infections in multiple tissues. Furthermore, our model supports different SARS-CoV-2 variants infection, and it can be successfully used to evaluate the effects of therapeutic monoclonal antibodies (Ab1F11) and antiviral drugs (Molnupiravir). Finally, to test the effect of SARS-CoV-2 infection on male reproduction, we generated Sertoli cell-specific SARS-CoV-2 N expressed mice by crossing with AMH-Cre transgenic line. We found that SARS-CoV-2 GFP/ΔN strain could infect Sertoli cells, led to spermatogenic defects due to the destruction of blood-testis barrier. Overall, combining with different tissue-specific Cre transgenic lines, the human ACE2 and SARS-CoV-2 NF/F line enables us to evaluate antivirals in vivo and study the pathogenesis of SARS-CoV-2 on some special tissues or organs.

Abstract Background Blau syndrome is a rare autoinflammatory disease caused by autosomal dominant mutations in the CARD15/NOD2 gene. Vascular involvement is a rare phenotype in Blau syndrome patients. In this study, we aimed to describe a 20-year- old Chinese girl with Blau syndrome complicated by renal arteritis. In addition, we summarized a literature review of published cases of vascular involvement in patients with Blau syndrome. Case presentation We describe a 20-year-old girl who was initially misdiagnosed with juvenile idiopathic arthritis (JIA) almost 15 years prior. In October 2019, she developed renal arteritis at the age of 17 years and was eventually diagnosed with Blau syndrome. A de-novo M513T mutation was found in her gene testing. A review of the literature on patients with Blau syndrome and vasculitis showed that a total of 18 cases were reported in the past 40 years. The vast majority of them were predominantly involved medium and large vessel arteritis. Of the 18 patients included in our literature review, 14 patients had aorto-arteritis, and 4 of them had renal artery involvement. Two patients presented with renal artery stenosis, 1with a sinus of Valsalva aneurysm, and 1 with retinal vasculitis. Conclusion A detailed medical history inquiry and a careful physical examination are helpful for the early identification of Blau syndrome, especially for infant onset refractory JIA. Medium-and large-vessel arteritis is a rare clinical manifestation in Blau syndrome patients. Careful examination of the peripheral pulse and measurement of blood pressure at every regular visit may be helpful in the early identification of Blau syndrome-arteritis. Early diagnosis and appropriate treatment may prevent or delay the occurrence of severe symptoms in patients to improve the patient’s quality of life.

Major Histocompatibility Complex Class II (MHC II) deficiency is a rare primary immunodeficiency disorder (PID) with autosomal recessive inheritance pattern. The outcome is almost fatal owing to delayed diagnosis and lacking of effective therapy. Therefore, prompt diagnosis, timely and effective treatment are critical. Here, we report a 117-day-old boy with diarrhea, cough, cyanosis and tachypnea who was failed to be cured by empiric antimicrobial therapy initially and progressed to severe pneumonia and respiratory failure. The patient was admitted to the pediatric intensive care unit (PICU) immediately and underwent a series of tests. Blood examination revealed elevated levels of inflammatory markers and cytomegalovirus DNA. Imaging findings showed signs of severe infection of lungs. Finally, the diagnosis was obtained mainly through next-generation sequencing (NGS). We found out what pathogenic microorganism he was infected via repeated conventional detection methods and metagenomic next-generation sequencing (mNGS) of sputum and bronchoalveolar lavage fluid (BALF). And his whole exome sequencing (WES) examination suggested that CIITA gene was heterozygous mutation, a kind of MHC II deficiency diseases. After aggressive respiratory support and repeated adjustment of antimicrobial regimens, the patient was weaned from ventilator on the 56th day of admission and transferred to the immunology ward on the 60th day. The patient was successful discharged after hospitalizing for 91 days, taking antimicrobials orally to prevent infections post-discharge and waiting for stem cell transplantation. This case highlights the potential importance of NGS in providing better diagnostic testing for unexplained infection and illness. Furthermore, pathogens would be identified more accurately if conventional detection techniques were combined with mNGS.

Although ultra-small nanoclusters (USNCs, < 2 nm) have immense application capabilities in biomedicine, the investigation on body-wide organ responses towards USNCs is scant. Here, applying a novel strategy of single-cell mass cytometry combined with Nano Genome Atlas of multi-tissues, we systematically evaluate the interactions between the host and calcium phosphate (CaP) USNCs at the organism level. Combining single-cell mass cytometry, and magnetic luminex assay results, we identify dynamic immune responses to CaP USNCs at the single cell resolution. The innate immune is initially activated and followed by adaptive immune activation, as evidenced by dynamic immune cells proportions. Furthermore, using Nano Genome Atlas of multi-tissues, we uncover CaP USNCs induce stronger activation of the immune responses in the cartilage and subchondral bone among the five local tissues while promote metabolic activities in the liver and kidney. Moreover, based on the immunological response profiles, histological evaluation of major organs and local tissue, and a body-wide transcriptomics, we demonstrate that CaP USNCs are not more hazardous than the Food and Drug Administration-approved CaP nanoparticles after 14 days of injection. Our findings provide valuable information on the future clinical applications of USNCs and introduce an innovative strategy to decipher the whole body response to implants.

Abstract Stem cell-based tissue regeneration strategies are promising treatments for severe endometrial injuries. However, there are few appropriate seed cells for regenerating a full-thickness endometrium, which mainly consists of epithelia and stroma. Müllerian ducts in female embryonic development develop into endometrial epithelia and stroma. Hence, we first generated human pluripotent stem cells (hPSC)-derived Müllerian duct-like cells (MDLCs) using a defined and effective protocol. The MDLCs are bi-potent, can gradually differentiate into endometrial epithelial and stromal cells, and reconstitute full-thickness endometrium in vitro and in vivo. Furthermore, MDLCs showed the in situ repair capabilities of reconstructing endometrial structure and recovering pregnancy function in full-thickness endometrial injury rats, and their differentiation fate was revealed by single-cell RNA sequencing (scRNA-seq). Our study provides a strategy for hPSC differentiation into endometrial lineages and an alternative seed cell for injured endometrial regeneration.

Abstract Pre-eclampsia (PE), associated with placental malperfusion, is the primary reason for maternal and perinatal mortality and morbidity that can cause vascular endothelial injury and multi-organ injury. Despite considerable research efforts, no pharmaceutical has been shown to stop disease progression. If women precisely diagnosed with PE can achieve treatment at early gestation, the maternal and fetal outcomes can be maximally optimized by expectant management. Current diagnostic approaches applying maternal characteristics or biophysical markers, including blood test, urine analysis and biophysical profile, possess limitations in the precise diagnosis of PE. Biochemical factor research associated with PE development has generated ambitious diagnostic targets based on PE pathogenesis and dissecting molecular phenotypes. This review focuses on current developments in biochemical prediction of PE and the corresponding interventions to ameliorate disease progression, aiming to provide references for clinical diagnoses and treatments.

Abstract Our understanding of full-thickness endometrial regeneration after injury is limited by an incomplete molecular characterization of the cell populations responsible for the organ functions. To help fill this knowledge gap, we characterized 10,551 cells of full-thickness normal human uterine from two menstrual phases (proliferative and secretory phase) using unbiased single cell RNA-sequencing. We dissected cell heterogeneity of main cell types (epithelial, stromal, endothelial, and immune cells) of the full thickness uterine tissues, cell population architectures of human uterus cells across the menstrual cycle. We identified an SFRP4+ stromal cell subpopulation that was highly enriched in the regenerative stage of the human endometria during the menstrual cycle, and the SFRP4+ stromal cells could significantly enhance the proliferation of human endometrial epithelial organoid in vitro, and promote the regeneration of endometrial epithelial glands and full-thickness endometrial injury through IGF1 signaling pathway in vivo. Our cell atlas of full-thickness uterine tissues revealed the cellular heterogeneities, cell population architectures, and their cell–cell communications during the monthly regeneration of the human endometria, which provide insight into the biology of human endometrial regeneration and the development of regenerative medicine treatments against endometrial damage and intrauterine adhesion.

Abstract Background Hereditary fructose intolerance (HFI) caused by aldolase B reduction or deficiency that results in fructose metabolism disorder. The disease prevalence in the Chinese population is unknown, which impedes the formulation of HFI screening and diagnosis strategies. Materials and methods By searching a local cohort (Chinese Children’s Rare Disease Genetic Testing Clinical Collaboration System, CCGT) and public databases (ClinVar and Human Gene Mutation Database) and reviewing HFI-related literature, we manually curated ALDOB pathogenic or likely pathogenic (P/LP) variants according to ACMG guidelines. Allele frequency (AF) information from the local database CCGT and the public databases HuaBiao and gnomAD for ALDOB P/LP variants was used to estimate and the HFI prevalence in the Chinese population and other populations by the Bayesian framework. We collected the genotype and clinical characteristics of HFI patients from the CCGT database and published literature to study genotype–phenotype relationships. Result In total, 81 variants of ALDOB were curated as P/LP. The estimated Chinese HFI prevalence was approximately 1/504,678, which was much lower than that for non-Finland European (1/23,147), Finnish in Finland (1/55,539), admixed American (1/132,801) and Ashkenazi Jewish (1/263,150) populations. By analyzing the genetic characteristics of ALDOB in the Chinese population, two variants (A338V, A338G) had significantly higher AFs in the Chinese population than in the non-Finland European population from gnomAD (all P values

A human induced pluripotent stem cell line (iPSC), FDCHi010-A, was derived from the peripheral blood of a 3-year-old patient with the c.2062C > T (p.R688*) mutation in the AHDC1 gene. The established cell line displayed a typical human embryonic stem cell-like morphology, exhibited a normal euploid karyotype, and fully expressed pluripotency markers. In addition, it retained the ability to differentiate to three germ layers. This cell line with a specific mutation may provide a useful tool for studying the pathogenesis and drug therapy screening of Xia-Gibbs syndrome caused by the AHDC1 gene.

BackgroundTumor necrosis factor (TNF) is an inflammatory cytokine that can coordinate tissue homeostasis by co-regulating the production of cytokines, cell survival, or death. It widely expresses in various tumor tissues and correlates with the malignant clinical features of patients. As an important inflammatory factor, the role of TNFα is involved in all steps of tumorigenesis and development, including cell transformation, survival, proliferation, invasion and metastasis. Recent research has showed that long non-coding RNAs (lncRNAs), defined as RNA transcripts >200 nucleotides that do not encode a protein, influence numerous cellular processes. However, little is known about the genomic profile of TNF pathway related-lncRNAs in GBM. This study investigated the molecular mechanism of TNF related-lncRNAs and their immune characteristics in glioblastoma multiforme (GBM) patients.MethodsTo identify TNF associations in GBM patients, we performed bioinformatics analysis of public databases - The Cancer Genome Atlas (TCGA) and the Chinese Glioma Genome Atlas (CGGA). The ConsensusClusterPlus, CIBERSORT, Estimate, GSVA and TIDE and first-order bias correlation and so on approaches were conducted to comprehensively characterize and compare differences among TNF-related subtypes.ResultsBased on the comprehensive analysis of TNF-related lncRNAs expression profiles, we constructed six TNF-related lncRNAs (C1RL-AS1, LINC00968, MIR155HG, CPB2-AS1, LINC00906, and WDR11-AS1) risk signature to determine the role of TNF-related lncRNAs in GBM. This signature could divide GBM patients into subtypes with distinct clinical and immune characteristics and prognoses. We identified three molecular subtypes (C1, C2, and C3), with C2 showing the best prognosis; otherwise, C3 showing the worst prognosis. Moreover, we assessed the prognostic value, immune infiltration, immune checkpoints, chemokines cytokines and enrichment analysis of this signature in GBM. The TNF-related lncRNA signature was tightly associated with the regulation of tumor immune therapy and could serve as an independent prognostic biomarker in GBM.ConclusionThis analysis provides a comprehensive understanding of the role of TNF-related characters, which may improve the clinical outcome of GBM patients.

Neurodevelopmental disorders (NDDs) have heterogeneity in both clinical characteristics and genetic factors. EBF3 is a recently discovered gene associated with a syndromic form of NDDs characterized by hypotonia, ataxia and facial features. In this study, we report twelve unrelated individuals with EBF3 variants using next-generation sequencing. Five missense variants (four novel variants and one known variant) and seven copy number variations (CNVs) of EBF3 gene were identified. All of these patients exhibited developmental delay/intellectual disability. Ataxia was observed in 33% (6/9) of the patients, and abnormal muscle tone was observed in 55% (6/11) of the patients. Aberrant MRI reports were noted in 64% (7/11) of the patients. Four novel missense variants were all located in the DNA-binding domain. The pathogenicity of these variants was validated by in vitro experiments. We found that the subcellular protein localization of the R152C and F211L mutants was changed, and the distribution pattern of the R163G mutant was changed from even to granular. Luciferase assay results showed that the four EBF3 mutants' transcriptional activities were all significantly decreased (p

Abstract Background Cystic fibrosis (CF) is a common, life-threatening genetic disease in Caucasians but rarely reported in Chinese population. The prevalence and population-specific genetic spectrum of CF in China needs to be systematically estimated and compared with Caucasians. Materials and methods We reviewed 30,951 exome-sequencing samples, including 20,909 pediatric patient samples and 10,042 parent samples, from Chinese Children's Rare Disease Genetic Testing Clinical Collaboration System (CCGT). After the in-lab filtration process, 477 candidate variants of CFTR gene were left and 53 variants were manually curated as pathogenic/likely-pathogenic (P/LP). These P/LP variants were adopted to estimate CF prevalence in three methods: the carrier frequency method, the permutation-combinations method and the Bayesian framework method. Allele frequencies of the 477 CFTR variants were compared with non-Finland European (NFE) and East Asian (EAS) from gnomAD database. To investigate the haplotype structure difference of CFTR, another 2067 whole-genome-sequencing samples from CCGT and 195 NFE from 1000 genome project were analyzed by Shapeit4 software. Result With the 53 manually curated P/LP variants in CFTR gene, we excluded individuals identified or suspected with CF and their parents in our cohorts and estimated the Chinese CF prevalence is approximately 1/128,434. Only 21 (39.6%) of the 53 variants were included in Caucasian specific CF screening panels, resulting in significantly under-estimation of CF prevalence in our children cohort (1/143,171 vs. 1/1,387,395, P = 5e−24) and parent’s cohort (1/110,127 vs. 1/872,437, P = 7e−10). The allele frequencies of six pathogenic variants (G970D, D979A, M469V, G622D, L88X, 1898+5G->T) were significantly higher in our cohorts compared with gnomAD-NFE population (all P-value

Although aging is an increasingly severe healthy, economic, and social global problem, it is far from well-modeling aging due to the aging process’s complexity. To promote the aging modeling, here we did the quantitative measurement based on aging blood transcriptome. Specifically, the aging blood transcriptome landscape was constructed through ensemble modeling in a cohort of 505 people, and 1138 age-related genes were identified. To assess the aging rate in the linear dimension of aging, we constructed a simplified linear aging clock, which distinguished fast-aging and slow-aging populations and showed the differences in the composition of immune cells. Meanwhile, the non-linear dimension of aging revealed the transcriptome fluctuations with a crest around the age of 40 and showed that this crest came earlier and was more vigorous in the fast-aging population. Moreover, the aging clock was applied to evaluate the rejuvenation effect of molecules in vitro, such as Nicotinamide Mononucleotide (NMN) and Metformin. In sum, this study developed a de novo aging clock to evaluate age-dependent precise medicine by revealing its fluctuation nature based on comprehensively mining the aging blood transcriptome, promoting the development of personal aging monitoring and anti-aging therapies.

Neurodevelopmental disorders (NDDs) are a group of high-incidence rare diseases with genetic heterogeneity. PPP2R1A, the regulatory subunit of protein phosphatase 2A, is a recently discovered gene associated with NDDs. Whole/clinical exome sequencing was performed in five patients with a family with NDDs. In vitro experiments were performed to evaluate the mutants’ expression and interactions with the complex. The genotype–phenotype correlations of reported cases as well as our patients with PPP2R1A variants were reviewed. We reported five unrelated individuals with PPP2R1A variants, including two novel missense variants and one frameshift variant. The protein expression of the Arg498Leu variant was less than that of the wild-type protein, the frameshift variant Asn282Argfs*14 was not decreased but truncated, and these two variants impaired the interactions with endogenous PPP25RD and PPP2CA. Furthermore, we found that pathogenic variants clustered in HEAT repeats V, VI and VII, and patients with the Met180Val/Thr variants had macrocephaly, severe ID and hypotonia, but no epilepsy, whereas those with Arg258 amino acid changes had microcephaly, while a few had epilepsy or feeding problems. In this study, we reported five NDD patients with PPP2R1A gene variants and expanded PPP2R1A pathogenic variant spectrum. The genotype and phenotype association findings provide reminders regarding the prognostication and evidence for genetic counseling.

Abstract Background Fabry disease (FD) remains poorly recognized, especially in children in China. Considering the diversity and nonspecific clinical manifestations accompanying with life-threatening aspect of this disease, methods to improve effective screening and management of the suspects are needed. This study aims to explore how it can be done effectively from a multidisciplinary perspective for children with FD at a tertiary children’s hospital in China. Methods A multidisciplinary team (MDT) of pediatric FD experts was launched at Children’s Hospital of Fudan University. Children with high-risk characteristics were referred by the MDT screening team using the dried blood spot (DBS) triple-test (α-galactosidase A, globotriaosylsphingosine, GLA gene). For newborns who were undergoing genetic testing in the hospital, the GLA gene was listed as a routine analysis gene. Evaluation, family screening, and genetic counselling were implemented after screening by the MDT management team. Results Before the establishment of the MDT, no case was diagnosed with FD in the hospital. However, twelve months following the MDT program's implementation, thirty-five children with high-risk profiles were referred for screening by DBS triple-test, with a yield of diagnosis of 14.3% (5/35). These 5 diagnosed children were referred due to a high-risk profile of pain accompanied by dermatological angiokeratoma and hypohidrosis (n = 2), pain accompanied by abnormal liver function (n = 1), pain only (n = 1), and unexplained renal tubular dysfunction (n = 1). Two neonates were detected early with GLA mutations in the hospital, with a yield of detection of 0.14% (2/1420). Furthermore, another 3 children diagnosed with FD were referred from other hospitals. Family screening of these 10 diagnosed children indicated that 9 boys inherited it from their mothers and 1 girl inherited it from her father. Four of them started to receive enzyme replacement therapy. Conclusion Screening and management of children with FD is effective based on a defined screening protocol and a multidisciplinary approach. We should pay more attention to the high-risk profiles of pain, angiokeratoma, decreased sweating, and unexplained chronic kidney disease in children.

Abstract Background Enzyme-based host depletion significantly improves the sensitivity of clinical metagenomics. Recent studies found that real-time adaptive sequencing of DNA molecules was achieved using a nanopore sequencing machine, which enabled effective enrichment of microbial sequences. However, few studies have compared the enzyme-based host depletion and nanopore adaptive sequencing for microbial enrichment efficiency. Results To compare the host depletion and microbial enrichment efficiency of enzyme-based and adaptive sequencing methods, the present study collected clinical samples from eight children with respiratory tract infections. The same respiratory samples were subjected to standard methods, adaptive sequencing methods, enzyme-based host depletion methods, and the combination of adaptive sequencing and enzyme-based host depletion methods. We compared the host depletion efficiency, microbial enrichment efficiency, and pathogenic microorganisms detected between the four methods. We found that adaptive sequencing, enzyme-based host depletion and the combined methods significantly enriched the microbial sequences and significantly increased the diversity of microorganisms (p value

Abstract Background Pathogenic variants of PAX2 cause autosomal-dominant PAX2-related disorder, which includes variable phenotypes ranging from renal coloboma syndrome (RCS), congenital anomalies of the kidney and urinary tract (CAKUT) to nephrosis. Phenotypic variability makes it difficult to define the phenotypic spectrum associated with genotype. Methods We collected the phenotypes in patients enrolled in the China national multicenter registry who were diagnosed with pathogenic variant in PAX2 and reviewed all published cases with PAX2-related disorders. We conducted a phenotype-based cluster analysis by variant types and molecular modeling of the structural impact of missense variants. Results Twenty different PAX2 pathogenic variants were identified in 32 individuals (27 families) with a diagnosis of RCS (9), CAKUT (11) and nephrosis (12) from the Chinese cohort. Individuals with abnormal kidney structure (RCS or CAKUT group) tended to have likely/presumed gene disruptive (LGD) variants (Fisher test, p

Craniosynostosis is a premature fusion of cranial sutures, resulting in abnormally shaped skull and brain development disorder. The description of craniosynostosis in patients with BCL11B mutations is rare. Here, we firstly report a 25-month-old Chinese boy with a novel frameshift variant in BCL11B gene. The patient was identified c.2346_2361del by whole-exome sequencing and was confirmed to be de novo by parental Sanger sequencing. This patient presented clinical phenotype of craniosynostosis as well as global developmental delay. He had a small mouth, thin upper lip, arched eyebrows, a long philtrum, midfacial hypoplasia and craniosynostosis. Brain MRI showed brain extracerebral interval and myelination changes, and brain CT with 3D reconstruction showed multi-craniosynostosis. Our study expands the clinical phenotypes of patients with BCL11B gene mutation, and our findings may help guide clinical treatment and family genetic counseling.

The sperm head-to-tail coupling apparatus ensures sperm head-tail integrity, but mechanistic insights remain limited. Here the authors demonstrate that CENTLEIN links and controls the interaction between SUN5 and PMFBP1, indicating that its impairments might be associated with acephalic spermatozoa syndrome.

Abstract Background Hyaline fibromatosis syndrome is a rare autosomal recessive disorder with ANTXR2 mutations characterised by the accumulation of hyaline substances in tissues. We present a case with the severe form—infantile systemic hyalinosis (ISH)—with long survival and review the literature. Methods and results Trio‐exome sequencing revealed compound heterozygous mutations, including a novel 4.41 kb deletion on 4q21.21 and the previously reported c.1294C > T mutation, in the ANTXR2 gene. He was diagnosed with ISH and treated symptomatically. After follow‐ups until 4 years of age, his recurrent respiratory infections and diarrhoea improved after one severe diarrhoea attack treated with intravenous gamma globulin. He is now awaiting surgical excision of gingival hypertrophy and joint contractures. Conclusion The novel gross deletion in ANTXR2 enriches the genetic mutation spectrum of hyaline fibromatosis syndrome. The manifestation of decreased foetal movement, acute‐infection attack or intravenous gamma globulin treatment may be associated with hyaline fibromatosis syndrome. A review of 116 reported cases reveals that missense mutations in the vWA domain are associated with joint symptoms, respiratory tract infection and diarrhoea, while frameshift mutations are associated with facial deformities and speech delays. We have enriched the current knowledge of the clinical manifestations and genetic mutation spectrum of HFS.

BackgroundGalloway–Mowat syndrome type 3 (GAMOS3) is an extremely rare and severe autosomal-recessive disease characterized by early-onset nephrotic syndrome (NS), microcephaly and neurological impairment. Reported GAMOS cases have gradually increased since pathogenic OSGEP variants were identified as the aetiology in 2017.MethodsUsing whole-exome sequencing and a data analysis process established by Children's Hospital of Fudan University, the clinical and molecular features of 3 infants with OSGEP mutations were summarized. Literature regarding the clinical features of GAMOS3 caused by OSGEP variants was reviewed.ResultsThirty-seven individuals (3 from this study) from 34 families were included. Twenty-two different OSGEP variants were identified. The c.740G>A (p.Arg247Gln) variant in OSGEP was detected in 15 families (44%), all from Asia. Most affected individuals (including patients I and II in this study) showed a typical phenotype, including microcephaly (92%) with brain anomalies (97%), developmental delay (81%), congenital NS (54%), and craniofacial (94%) and skeletal dysmorphism (84%). Renal manifestations varied from proteinuria (94%, median onset = 1.5 months) to NS (83%) and end-stage renal disease (48%, 11 months) during follow-up. Patients with congenital NS had a lower survival probability (median survival time = 3 months) than those without congenital NS (78 months) (P < 0.01, log-rank test).ConclusionGAMOS3 is a progressive renal-neurological syndrome with a poor prognosis, especially with congenital NS. Microcephaly with dysmorphic features are vital clues to further evaluate renal impairment and brain anomalies. Timely molecular diagnosis is crucial for clinical decision-making, appropriate treatment and genetic counselling.

Abstract Background DKC1, the dyskerin encoding gene, functions in telomerase activity and telomere maintenance. DKC1 mutations cause a multisystem disease, dyskeratosis congenita (DC), which is associated with immunodeficiency and bone marrow failure. Methods In this research, we reported a novel intronic mutation of DKC1 causing dyskerin functional loss in a Chinese family. Whole exome sequence (WES) of the proband and validation by sanger sequencing help us identify a pathogenic DKC1 mutation. Minigene splicing assays were performed to evaluate functional change of DKC1. Results A pathogenic DKC1 intronic mutation(c.84 + 7A > G) was identified in the proband, which was inherited from heterozygous mother and not reported before. We detected the novel transcript with a 7 bp intron retention through minigene splicing assay. The newly spliced transcript is so short that would be degraded by nonsense‐mediated mRNA decay in vitro and we infer that the novel DKC1 mutation would influences normal physiological function of dyskerin. Conclusions Our study identified a novel intronic mutation, which expands the spectrum of pathogenic DKC1 gene mutations and can be used in molecular diagnosis. The mutant allele was transmitted to the next generation with high frequency in the family and causes still birth or early death.

Endometrium undergoes repeated repair and regeneration during the menstrual cycle. Previous attempts using gene expression data to define the menstrual cycle failed to come to an agreement. Here we used single-cell RNA sequencing data of C57BL/6J mice uteri to construct a novel integrated cell atlas of mice uteri from the regenerative endometrium to the maturational endometrium at the single-cell level, providing a more accurate cytological-based elucidation for the changes that occurred in the endometrium during the estrus cycle. Based on the expression levels of proliferating cell nuclear antigen, differentially expressed genes, and gene ontology terms, we delineated in detail the transitions of epithelial cells, stromal cells, and immune cells that happened during the estrus cycle. The transcription factors that shaped the differentiation of the mononuclear phagocyte system had been proposed, being Mafb, Irf7, and Nr4a1. The amounts and functions of immune cells varied sharply in two stages, especially NK cells and macrophages. We also found putative uterus tissue-resident macrophages and identified potential endometrial mesenchymal stem cells (high expression of Cd34, Pdgfrb, Aldh1a2) in vivo. The cell atlas of mice uteri presented here would improve our understanding of the transitions that occurred in the endometrium from the regenerative endometrium to the maturational endometrium. With the assistance of a normal cell atlas as a reference, we may identify morphologically unaffected abnormalities in future clinical practice. Cautions would be needed when adopting our conclusions, for the limited number of mice that participated in this study may affect the strength of our conclusions.

BackgroundKCNQ2-related disorder is typically characterized as neonatal onset seizure and epileptic encephalopathy. The relationship between its phenotype and genotype is still elusive. This study aims to provide clinical features, management, and prognosis of patients with novel candidate variants of the KCNQ2 gene.MethodsWe enrolled patients with novel variants in the KCNQ2 gene from the China Neonatal Genomes Project between January 2018 and January 2021. All patients underwent next-generation sequencing tests and genetic data were analyzed by an in-house pipeline. The pathogenicity of variants was classified according to the guideline of the American College of Medical Genetics. Each case was evaluated by two geneticists back to back. Patients' information was acquired from clinical records.ResultsA total of 30 unrelated patients with novel variants in the KCNQ2 gene were identified, including 19 patients with single-nucleotide variants (SNVs) and 11 patients with copy number variants (CNVs). For the 19 SNVs, 12 missense variants and 7 truncating variants were identified. Of them, 36.8% (7/19) of the KCNQ2 variants were located in C-terminal regions, 15.7% (3/19) in segment S2, and 15.7% (3/19) in segment S4. Among them, 18 of 19 patients experienced seizures in the early neonatal period. However, one patient presented neurodevelopmental delay (NDD) as initial phenotype when he was 2 months old, and he had severe NDD when he was 3 years old. This patient did not present seizure but had abnormal electrographic background activity and brain imaging. Moreover, for the 11 patients with CNVs, 20q13.3 deletions involving EEF1A2, KCNQ2, and CHRNA4 genes were detected. All of them presented neonatal-onset seizures, responded to antiepileptic drugs, and had normal neurological development.ConclusionIn this study, patients with novel KCNQ2 variants have variable phenotypes, whereas patients with 20q13.3 deletion involving EEF1A2, KCNQ2, and CHRNA4 genes tend to have normal neurological development.

Developmental and epileptic encephalopathy Type 31 (DEE31) is a severe early infantile epileptic encephalopathy with intellectual and movement disability caused by heterozygous mutations in the Dynamin 1 gene, which encodes DNM1, a GTPase involved in synaptic vesicle fission for receptor-mediated endocytosis on the presynaptic plasma membrane. A human induced pluripotent stem cell (iPSC) line, termed as FDCHI007-A, was generated using non-integrating episomal vector technique from peripheral blood mononuclear cells (PBMCs) of a 3-year-old female DDE31 patient, who had a heterozygous (c.545C > A P. Ala 182Asp) mutation in the DNM1 gene. FDCHI007-A offers a useful cell resource to investigate pathogenic mechanisms in DEE31, as well as a cell-based model for drug development for DEE31.

The skin is the largest organ of the human body. Skin injuries, especially full-thickness injuries, are a major treatment challenge in clinical practice. Therefore, wound dressing materials with therapeutic effects have great practical significance in healthcare. This study used photocrosslinkable gelatin methacryloyl (GelMA) and sulfhydrylated chitosan (CS-SH) to design a double-crosslinked hydrogel for wound dressing. When crosslinked together, the resulting hydrogels showed a highly porous inner structure, and enhanced mechanical properties and moisture retention capacity. The compression modulus of the GelMA/CS-SH hydrogel (GCH) reached up to about 40 kPa and was much higher than that of pure GelMA hydrogel, and the compression modulus was increased with the amount of CS-SH. In vitro study showed no cytotoxicity of obtained hydrogels. Interestingly, a higher concentration of CS-SH slightly promoted the proliferation of cells. Moreover, the double-crosslinked hydrogel exhibited antibacterial properties because of the presence of chitosan. In vivo study based on rats showed that full-thickness skin defects healed on the 15th day. Histological results indicate that the hydrogel accelerated the repair of hair follicles and encouraged the orderly growth of collagen fibers in the wound. Furthermore, better blood vessel formation and a higher expression of VEGFR were observed in the hydrogel group when compared with the untreated control group. Based on our findings, GCH could be a promising candidate for full-thickness wound dressing.

The outcomes of in vitro fertilization (IVF) for endometriotic women are significantly worse than for patients without ovarian endometriosis (OEM), as shown by fewer retrieved oocytes. However, the exact pathophysiological mechanism is still unknown. Thus, we conducted a prospective study that analyzed mRNA and lncRNA transcriptome between granulosa cells (GCs) from patients with fewer retrieved oocytes due to OEM and GCs from controls with male factor (MF) infertility using an RNA sequencing approach. We found a group of significantly differentially expressed genes (DEGs), including NR5A2, MAP3K5, PGRMC2, PRKAR2A, DEPTOR, ITGAV, KPNB1, GPC6, EIF3A, and SMC5, which were validated to be upregulated and negatively correlated with retrieved oocyte numbers in GCs of patients with OEM, while DUSP1 demonstrated the opposite. The molecular functions of these DEGs were mainly enriched in pathways involving mitogen-activated protein kinase (MAPK) signaling, Wnt signaling, steroid hormone response, apoptosis, and cell junction. Furthermore, we performed lncRNA analysis and identified a group of differentially expressed known/novel lncRNAs that were co-expressed with the validated DEGs and correlated with retrieved oocyte numbers. Co-expression networks were constructed between the DEGs and known/novel lncRNAs. These distinctive molecular signatures uncovered in this study are involved in the pathological regulation of ovarian reserve dysfunction in OEM patients.

X-linked MAGT1 deficiency with increased susceptibility to EBV-infection and N-linked glycosylation defect (XMEN) disease is a primary immunodeficiency caused by loss-of-function variants in the MAGT1 gene. Only two patients from one family have been diagnosed with XMEN in China. In this study, we retrospectively analyzed the genetic, clinical, and immunological characteristics of six pediatric patients in a Chinese cohort. Medical records were retrieved, immunological phenotypes were assessed, and infectious microbes in patients were detected. Six male patients (mean age, 6.3 years) from five unrelated families were genetically diagnosed as XMEN. Five patients presented with a major complaint of elevated liver enzymes, while one patient was referred for recurrent fever, cough and skin rash. Five patients developed EBV viremia, and one patient developed non-Hodgkin’s lymphoma. Histopathological findings from liver biopsy tissues showed variable hepatic steatosis, fibrosis, inflammatory infiltration, and glycogenosis. Immune phenotypes included CD4 T-cell lymphopenia, elevated B cells, inverted CD4/CD8 ratios, and elevated αβDNTs. No pathogenic microbes other than EBV were identified in these patients. This study reports the clinical and molecular features of Chinese patients with XMEN. For patients with transaminase elevation, chronic EBV infection and EBV-associated lymphoproliferative disease, the possibility of XMEN should be considered in addition to isolated liver diseases.

Abstract Background Variants in the SLC25A1 gene are associated with a severe neurometabolic disease, D-2- and L-2-hydroxyglutaric aciduria (D/L-2-HGA). A report in 2014 presented the first account of congenital myasthenic syndrome (CMS) with mild intellectual disability (ID) caused by SLC25A1. To date, only two missense variants in SLC25A1 have been linked to CMS. Case presentations A Chinese boy presented fatigable muscular weakness, myasthenic crisis, epilepsy and developmental delay along with mild elevation of urinary 2-ketoglutarate (2-KG) and lactic acid levels. He showed a partial response to pyridostigmine. Genetic analysis using trio whole-exome sequencing (WES), Sanger sequencing, and cosegregation analyses revealed two novel pathogenic variants of SLC25A1 (c.628C > T, p.R210X; c.145G > A, p.V49M). Conclusions We report a boy who carries novel compound heterozygous variants of SLC25A1 and presents a phenotype intermediate between CMS and D/L-2-HGA. This case expands the range of known phenotypes and genotypes associated with SLC25A1.

Summary: Articular cartilage injury and degeneration causing pain and loss of quality-of-life has become a serious problem for increasingly aged populations. Given the poor self-renewal of adult human chondrocytes, alternative functional cell sources are needed. Direct reprogramming by small molecules potentially offers an oncogene-free and cost-effective approach to generate chondrocytes, but has yet to be investigated. Here, we directly reprogrammed mouse embryonic fibroblasts into PRG4+ chondrocytes using a 3D system with a chemical cocktail, VCRTc (valproic acid, CHIR98014, Repsox, TTNPB, and celecoxib). Using single-cell transcriptomics, we revealed the inhibition of fibroblast features and activation of chondrogenesis pathways in early reprograming, and the intermediate cellular process resembling cartilage development. The in vivo implantation of chemical-induced chondrocytes at defective articular surfaces promoted defect healing and rescued 63.4% of mechanical function loss. Our approach directly converts fibroblasts into functional cartilaginous cells, and also provides insights into potential pharmacological strategies for future cartilage regeneration. : In this article, Ouyang and colleagues develop a chemical-driven direct reprogramming of mouse embryonic fibroblasts into PRG4+ chondrocytes, with a screen-defined cocktail VCRTc. Their induced chondrocytes promote in vivo cartilage functional regeneration. The dynamical cellular phenotypes were analyzed by single-cell RNA sequencing, illustrating the inhibition of fibroblast features and activation of chondrogenesis pathways in early reprogramming. Keywords: chemical reprogramming, single-cell analysis, cartilage regeneration, drug discovery

Abstract Background Intellectual disability (ID) constitutes the most common group of neurodevelopmental disorders. Exome sequencing has enabled the discovery of genetic mutations responsible for a wide range of ID disorders. Case presentation In this study, we reported on two male siblings, aged 4 and 2 years, with motor and mental developmental delays and mild dysmorphic facial features. To identify the genetic causes of these symptoms, we employed trio-whole exome sequencing for the proband. We found a novel hemizygous missense variant in the PAK3 gene (c.1112G > A, p.Cys371Tyr), which encodes the p21-activated kinase 3, in the proband, which inherited from mother. The younger brother also has the hemizygous variant, which was confirmed by Sanger sequencing. The variant is located in the kinase domain and was regarded as a likely pathogenic variant in this family. Conclusion We diagnosed two male siblings with developmental delays as having a PAK3 likely pathogenic variant. This finding expands the list of PAK3 gene mutations associated with neurodevelopmental disorders and provides further details on its clinical features.

To investigate the effect of Lentinan (LNT) on lung adenocarcinoma (LUAD) cell stemness and its mechanism. In this study, we founded that LNT significantly reduce the cell proliferation, activity, migration, invasion, and stemness of LUAD cells, and promote their apoptosis compared with the control group in vitro. Moreover, LNT significantly inhibited the volume and weight of tumors of nude mice in vivo. At the same time, LNT can significantly up-regulate miR-216a-5p levels and reduce the protein expression of phospho-JAK2 (Y1007/1008) and phospho-STAT3 (Tyr705), thereby inhibiting the JAK2/STAT3 signaling pathway. Interfering with miR-216a-5p expression and activating the JAK2/STAT3 signaling pathway can significantly reverse LNT inhibitory effects on LUAD. Collectively, LNT can inhibit the JAK2/STAT3 signaling pathway by up-regulating miR-216a-5p, reducing stemness, and promoting LUAD cells apoptosis, then slow down LUAD occurrence and development, providing concepts and experimental foundation treating patients with LUAD.

MEGDEL syndrome and SATB2-associated syndrome (SAS) are both rare congenital disorders with poor prognoses caused by gene mutations. We present the case of a 2-day-old girl with an unexplained abnormal liver function, feeding problem, and dystonia. Using next-generation sequencing, we identified two novel mutations in SERAC1 and a mutation in SATB2. Now, she is 15 months old and has the characteristics of SAS, such as downslanting palpebral fissures and delayed primary dentition. Besides the typical phenotypes of MEGDEL syndrome, such as hypertonia, failure to thrive, deafness, and motor regression, she has progressive cholestasis and is prone to high serum lactate after rehabilitation training and hypoglycemia with low ketone under starving conditions. These phenotypes substantially differ from the transient liver function abnormalities and hypoglycemia reported in the literature.

Neonatal metabolic acidosis (NMA) is a common problem, particularly in critically ill patients in neonatal intensive care units (NICUs). Complex etiologies and atypical clinical signs make diagnosis difficult; thus, it is crucial to investigate the underlying causes of NMA rapidly and provide disorder-specific therapies. Our study aims to provide an overview of the genetic causes of NMA in patients from NICUs. We performed next-generation sequencing (NGS) on neonates with NMA from January 2016 to December 2019. Clinical features, genetic diagnoses, and their effects on clinical interventions were collected for analysis. In the 354 enrolled patients, 131 (37%) received genetic diagnoses; 95 (72.5%) of them were autosomal recessively inherited diseases. Two hundred and fifteen variants spanning 57 genes were classified as pathogenic (P) or likely pathogenic (LP) in 131 patients. The leading cause was metabolic disorders due to 35 genes found in 89 patients (68%). The other 42 NMA patients (32%) with 22 genes had malformations and renal, neuromuscular, and immune-hematological disorders. Seven genes (MMUT, MMACHC, CHD7, NPHS1, OTC, IVD, and PHOX2B) were noted in more than four patients, accounting for 48.9% (64/131) of the identified P/LP variants. Forty-six diagnosed patients with uncorrected NMA died or gave up. In conclusion, 37% of neonates with metabolic acidosis had genetic disorders. Next-generation sequencing should be considered when investigating the etiology of NMA in NICUs. Based on early molecular diagnoses, valuable treatment options can be provided for some genetic diseases to achieve better outcomes.

Next-generation sequencing (NGS) has been used to detect severe combined immunodeficiency (SCID) in patients, and some patients with DNA cross-link repair 1C (DCLRE1C) variants have been identified. Moreover, some compound variants, such as copy number variants (CNV) and single nucleotide variants (SNV), have been reported. The purpose of this study was to expand the genetic data related to patients with SCID carrying the compound DCLRE1C variant. Whole-exome sequencing (WES) was performed for genetic analysis, and variants were verified by performing Sanger sequencing or quantitative PCR. Moreover, we searched PubMed and summarized the data of the reported variants. Four SCID patients with DCLRE1C variants were identified in this study. WES revealed a homozygous deletion in the DCLRE1C gene from exons 1–5 in patient 1, exons 1–3 deletion and a novel rare variant (c.92T>C, p.L31P) in patient 2, exons 1–3 deletion and a novel rare variant (c.328C>G, p.L110V) in patient 3, and exons 1–4 deletion and a novel frameshift variant (c.449dup, p.His151Alafs*20) in patient 4. Based on literature review, exons 1–3 was recognized as a hotspot region for deletion variation. Moreover, we found that compound variations (CNV + SNV) accounted for approximately 7% variations in all variants. When patients are screened for T-cell receptor excision circles (TRECs), NGS can be used to expand genetic testing. Deletion of the DCLRE1C gene should not be ignored when a variant has been found in patients with SCID.

BackgroundAn early and accurate evaluation of the risk of bronchopulmonary dysplasia (BPD) in premature infants is pivotal in implementing preventive strategies. The risk prediction models nowadays for BPD risk that included only clinical factors but without genetic factors are either too complex without practicability or provide poor-to-moderate discrimination. We aim to identify the role of genetic factors in BPD risk prediction early and accurately.MethodsExome sequencing was performed in a cohort of 245 premature infants (gestational age

Angelman syndrome (AS) is a neurodevelopmental genetic disorder, but there has been limited analysis of a large cohort of Chinese children with Angelman syndrome. This study aims to assess the phenotype and genotype of Chinese children with Angelman syndrome. We retrospectively analyzed data through a detailed online survey combined with an on-site study. Furthermore, phenotype analysis stratified by deletion and non-deletion groups was carried out. The responses of family members of 695 individuals with AS revealed that 577 patients (83.02%) had maternal deletions, 65 patients (9.35%) carried UBE3A mutations, 31 (4.46%) patients had UPD15pat (one patient with UPD15pat constituted by a mosaic), 10 patients (1.44%) had imprinting defects and 12 (1.58%) patients only showed abnormal methylation without further detection. We identified 50 different pathogenic variants in this cohort, although 18 of these variants were unreported. Recurrent variant c.2507_2510del (p.K836Rfs*4) was found in 7 patients. In the deletion group, patients were diagnosed at an earlier age, had a more severe clinical phenotype, a higher rate of epilepsy with more multiple seizure types, and more frequently combined medication. Strabismus and sleep disturbances were both common in deletion and non-deletion groups. The top three resources invested in caring for AS children are: daily involvement in patient care, rehabilitation cost, and anti-epileptic treatment. Our study showed the genetic composition of Chinese children with 83.02% of maternal deletions, and the mutation spectrum for UBE3A variants was expanded. Developmental outcomes are associated with genotype, and this was confirmed by deletion patients having a worse clinical phenotype and complex epilepsy.