Gene and drug therapies are being developed to al eviate vision loss in patients with Stargardt’s disease and age-related macular degeneration （AMD）. To evaluate the therapeutic effects of these treatments, organic...Gene and drug therapies are being developed to al eviate vision loss in patients with Stargardt’s disease and age-related macular degeneration （AMD）. To evaluate the therapeutic effects of these treatments, organic sol-vents are routinely used to extract and quantify bisretinoid lipofuscin constituents, such as N-retinylidene-N-retinyl-ethanolamine （A2E） and all-trans-retinal dimer （ATR-dimer）. By high-performance liquid chromatography （HPLC）, we found that A2E and ATR-dimer were both altered by tetrahydrofuran （THF） and chloroform, but were stable in dimethyl sulfoxide （DMSO） or methanol （MeOH）. In addition, cyclohexane and ethanol （EtOH） did not alter ATR-dimer, whereas an alteration of A2E occurred in EtOH. On the basis of these findings, we designed processes II-IV, generated by modifications of process I, a routine method to measure bisretinoid compounds in vivo. Extra amounts of either ATR-dimer or A2E in mouse eyecups were released by processes II-IV versus process I. Efforts to clarify the effects of organic solvents on lipofuscin pigments are important because such studies can guide the handling of these fluoro-phores in related experiments.展开更多
Objective: This study aims to clarify the effect of the active components puerarin and tetrandrine on the chondrogenic differentiation of bone marrow mesenchymal stem cells(BMSCs).Methods: Using network pharmacology, ...Objective: This study aims to clarify the effect of the active components puerarin and tetrandrine on the chondrogenic differentiation of bone marrow mesenchymal stem cells(BMSCs).Methods: Using network pharmacology, protein targets of puerarin and tetrandrine were predicted, and a database of cartilage formation targets was established. The protein target information related to disease was then collected, and the drug-targeting network was constructed by analyzing the protein–protein interactions. Genes related to chondrogenesis induced by puerarin and tetrandrine and chondroblast differentiation signaling pathways were searched. Finally, potential drug-and disease-related genes,as well as proteins, were screened and verified using real-time RT-PCR and western blotting.Results: Network pharmacological studies have shown that puerarin and tetrandrine are involved in BMSCs cartilage differentiation. The experimental results showed that puerarin and tetrandrine could regulate the expression of cartilage differentiation-related genes and proteins. Puerarin increased the protein expression of COL2 A1, COL10 A1, MMP13, and SOX-9,as well as the gene expression of Col2 a1, Mmp13, Tgfb1, and Sox-9. Tetrandrine increased the protein expression of COL2 A1,COL10 A1, MMP13, and SOX-9, as well as the gene expression of Col10 a1, Tgfb1, Sox-9, and Acan. The combination of puerarin and tetrandrine increased the protein expression of COL2 A1, COL10 A1, MMP13, and SOX-9 and the gene expression of Col2 a1,Col10 a1, Sox-9, and Acan.Conclusions: Puerarin, tetrandrine, and their combination can promote the proliferation of BMSCs and induce their differentiation into chondrocytes, and they are thus expected to be inducers of chondrogenic differentiation. These results suggest that puerarin and tetrandrine have potential therapeutic effects on osteoarthritis.展开更多
Background Hemodynamically significant patent ductus arteriosus(hsPDA)is associated with increased comorbidities in neonates.Early evaluation of hsPDA risk is critical to implement individualized intervention.The aim ...Background Hemodynamically significant patent ductus arteriosus(hsPDA)is associated with increased comorbidities in neonates.Early evaluation of hsPDA risk is critical to implement individualized intervention.The aim of the study was to provide a powerful reference for the early identification of high-risk hsPDA population and early treatment decisions.Methods We enrolled infants who were diagnosed with PDA and performed exome sequencing.The collapsing analyses were used to find the risk gene set(RGS)of hsPDA for model construction.The credibility of RGS was proven by RNA sequencing.Multivariate logistic regression was performed to establish models combining clinical and genetic features.The models were evaluated by area under the receiver operating curve(AUC)and decision curve analysis(DCA).Results In this retrospective cohort study of 2199 PDA patients,549(25.0%)infants were diagnosed with hsPDA.The model[all clinical characteristics selected by least absolute shrinkage and selection operator regression(all CCs)]based on six clinical variables was acquired within three days of life,including gestational age(GA),respiratory distress syndrome(RDS),the lowest platelet count,invasive mechanical ventilation,and positive inotropic and vasoactive drugs.It has an AUC of 0.790[95%confidence interval(CI)=0.749–0.832],while the simplified model(basic clinical characteristic model)including GA and RDS has an AUC of 0.753(95%CI=0.706–0.799).There was a certain consistency between RGS and differentially expressed genes of the ductus arteriosus in mice.The AUC of the models was improved by RGS,and the improvement was significant(all CCs vs.all CCs+RGS:0.790 vs.0.817,P<0.001).DCA demonstrated that all models were clinically useful.Conclusions Models based on clinical factors were developed to accurately stratify the risk of hsPDA in the first three days of life.Genetic features might further improve the model performance.展开更多
Background Maple syrup urine disease(MSUD)is an autosomal recessive inherited disorder that affects the degradation of branched-chain amino acids and is associated with acute and chronic brain dysfunction.This study p...Background Maple syrup urine disease(MSUD)is an autosomal recessive inherited disorder that affects the degradation of branched-chain amino acids and is associated with acute and chronic brain dysfunction.This study presents 11 new patients with MSUD and describes the clinical characteristics and gene mutations reported in Chinese individuals.Methods During 2011–2018,11 pedaitric patients with MSUD from 11 Chinese families were analyzed based on clinical characteristics and mass spectrometry,with confirmation via gene sequencing.Novel mutations affecting protein function were predicted with Mutation-Taster,PolyPhen-2,CADD and SIFT software.3D models of the mutated proteins were generated by using the SWISS-MODEL online server,and the models were visualized in PyMOL.The characteristics and gene mutations in patients with MSUD were analyzed retrospectively.Results Seventeen mutations in the BCKDHA,BCKDHB and DBT genes were found,8 of which are novel:c.55C>/T,c.349C>T,c.565C>T,c.808G>A,c.859C>G,and c.1270dupC in BCKDHA;c.275-2A>G in BCKDHB;and c.1291C>T in DBT.Eight patients died.Two patients had severe mental retardation and were physically handicapped.One patient with the intermediate type had relatively good prognosis,with mild psychomotor retardation and adiposity.Four mothers underwent amniocentesis for prenatal diagnosis during their second pregnancy;two fetuses were wild type,and two were carriers of one heterozygous mutation.Conclusions Eight novel mutations were associated with MSUD in Chinese patients.Prenatal diagnosis was successfully performed by genetic analysis.Mutations in the BCKDHB gene were found in the majority of Chinese patients with MSUD.展开更多
Background During next generation sequencing(NGS)data interpretation in critically ill newborns,there is a potential for recognizing and reporting secondary findings(SFs).Early awareness of SFs may provide clues for d...Background During next generation sequencing(NGS)data interpretation in critically ill newborns,there is a potential for recognizing and reporting secondary findings(SFs).Early awareness of SFs may provide clues for disease prevention.In this study,we assessed the frequency of SFs in the China Neonatal Genomes Project(CNGP)participants.Methods A total of 2020 clinical exome sequencing(CES)datasets were screened for variants from a list of 59 genes recommended by the American College of Medical Genetics and Genomics(ACMG)for secondary findings reporting v2.0(ACMG SF v2.0).Identified variants were classified according to the evidence-based guidelines reached by a joint consensus of the ACMG and the Association for Molecular Pathology(AMP).Results Among the 2020 CES datasets,we identified 23 ACMG-reportable genes in 61 individuals,resulting in an overall frequency of SFs at 3.02%.A total of 53 unique variants were identified,including 35 pathogenic and 18 likely pathogenic variants.The common disease categories of SFs associated were cardiovascular and cancer disease.The SF results affected the medical management and follow-up strategy in 49(80.3%)patients.Conclusions We presented the frequency of SFs and their impact on clinical management strategies in CNGP participants.Our study demonstrated that SFs have important practical value in disease prevention and intervention at an early stage.展开更多
Background Infantile hydrocephalus(IHC)is commonly related to other central nervous system diseases,which may have adverse effects on prognosis.The causes of IHC are heterogeneous,and the genetic etiologies are not fu...Background Infantile hydrocephalus(IHC)is commonly related to other central nervous system diseases,which may have adverse effects on prognosis.The causes of IHC are heterogeneous,and the genetic etiologies are not fully understood.This study aimed to analyze the genetic etiologies of an IHC cohort.Methods The data for 110 IHC patients who had received exome sequencing at the Clinical Genetic Center of the Children's Hospital of Fudan University between 2016 and 2019 were reviewed and analyzed retrospectively.An exome-wide association analysis(EWAS)was performed within this cohort using IHC as the study phenotype.Results Of the 110 IHC patients,a pathogenic or likely pathogenic variant was identified in 16(15%)patients,spanning 13 genes.The genes were mainly associated with metabolic disorders,brain abnormalities,and genetic syndromes.IHC patients who had unclear clinical etiology were more likely to possess a genetic etiology.Based on previous studies and on our EWAS results,ZEB1,SBF2,and GNAI2 were over-represented among IHC patients and might alfect the signaling pathways involved in IHC formation.Conclusions Our study showed heterogeneous genetic etiologies in an IHC cohort.It is essential to perform genetic testing on IHC patients who have unclear clinical etiology,and genes associated with metabolic disorders,brain abnormalities,and genetic syndromes should he noted.In addition,when aiming to discover IHC susceptibility genes,genes that might influence the signaling pathways involved in IHC formation should be prioritized.展开更多
基金Project supported by the National Natural Science Foundation of China(Nos.21202146 and 81271018)the Fundamental Research Funds for the Central Universities+1 种基金the Zhejiang Scientific Research Foundation for the Returned Overseas Chinese Scholars(No.J20120556)the Zhejiang Key Laboratory Fund of China(No.2011E10006)
文摘Gene and drug therapies are being developed to al eviate vision loss in patients with Stargardt’s disease and age-related macular degeneration （AMD）. To evaluate the therapeutic effects of these treatments, organic sol-vents are routinely used to extract and quantify bisretinoid lipofuscin constituents, such as N-retinylidene-N-retinyl-ethanolamine （A2E） and all-trans-retinal dimer （ATR-dimer）. By high-performance liquid chromatography （HPLC）, we found that A2E and ATR-dimer were both altered by tetrahydrofuran （THF） and chloroform, but were stable in dimethyl sulfoxide （DMSO） or methanol （MeOH）. In addition, cyclohexane and ethanol （EtOH） did not alter ATR-dimer, whereas an alteration of A2E occurred in EtOH. On the basis of these findings, we designed processes II-IV, generated by modifications of process I, a routine method to measure bisretinoid compounds in vivo. Extra amounts of either ATR-dimer or A2E in mouse eyecups were released by processes II-IV versus process I. Efforts to clarify the effects of organic solvents on lipofuscin pigments are important because such studies can guide the handling of these fluoro-phores in related experiments.
文摘Objective: This study aims to clarify the effect of the active components puerarin and tetrandrine on the chondrogenic differentiation of bone marrow mesenchymal stem cells(BMSCs).Methods: Using network pharmacology, protein targets of puerarin and tetrandrine were predicted, and a database of cartilage formation targets was established. The protein target information related to disease was then collected, and the drug-targeting network was constructed by analyzing the protein–protein interactions. Genes related to chondrogenesis induced by puerarin and tetrandrine and chondroblast differentiation signaling pathways were searched. Finally, potential drug-and disease-related genes,as well as proteins, were screened and verified using real-time RT-PCR and western blotting.Results: Network pharmacological studies have shown that puerarin and tetrandrine are involved in BMSCs cartilage differentiation. The experimental results showed that puerarin and tetrandrine could regulate the expression of cartilage differentiation-related genes and proteins. Puerarin increased the protein expression of COL2 A1, COL10 A1, MMP13, and SOX-9,as well as the gene expression of Col2 a1, Mmp13, Tgfb1, and Sox-9. Tetrandrine increased the protein expression of COL2 A1,COL10 A1, MMP13, and SOX-9, as well as the gene expression of Col10 a1, Tgfb1, Sox-9, and Acan. The combination of puerarin and tetrandrine increased the protein expression of COL2 A1, COL10 A1, MMP13, and SOX-9 and the gene expression of Col2 a1,Col10 a1, Sox-9, and Acan.Conclusions: Puerarin, tetrandrine, and their combination can promote the proliferation of BMSCs and induce their differentiation into chondrocytes, and they are thus expected to be inducers of chondrogenic differentiation. These results suggest that puerarin and tetrandrine have potential therapeutic effects on osteoarthritis.
基金funded by the Cohort Project of Specialized Disease,Clinical Research Center,Children's Hospital of Fudan University(2020ZBDL14)National Key R&D Program of China(2022ZD0116003).
文摘Background Hemodynamically significant patent ductus arteriosus(hsPDA)is associated with increased comorbidities in neonates.Early evaluation of hsPDA risk is critical to implement individualized intervention.The aim of the study was to provide a powerful reference for the early identification of high-risk hsPDA population and early treatment decisions.Methods We enrolled infants who were diagnosed with PDA and performed exome sequencing.The collapsing analyses were used to find the risk gene set(RGS)of hsPDA for model construction.The credibility of RGS was proven by RNA sequencing.Multivariate logistic regression was performed to establish models combining clinical and genetic features.The models were evaluated by area under the receiver operating curve(AUC)and decision curve analysis(DCA).Results In this retrospective cohort study of 2199 PDA patients,549(25.0%)infants were diagnosed with hsPDA.The model[all clinical characteristics selected by least absolute shrinkage and selection operator regression(all CCs)]based on six clinical variables was acquired within three days of life,including gestational age(GA),respiratory distress syndrome(RDS),the lowest platelet count,invasive mechanical ventilation,and positive inotropic and vasoactive drugs.It has an AUC of 0.790[95%confidence interval(CI)=0.749–0.832],while the simplified model(basic clinical characteristic model)including GA and RDS has an AUC of 0.753(95%CI=0.706–0.799).There was a certain consistency between RGS and differentially expressed genes of the ductus arteriosus in mice.The AUC of the models was improved by RGS,and the improvement was significant(all CCs vs.all CCs+RGS:0.790 vs.0.817,P<0.001).DCA demonstrated that all models were clinically useful.Conclusions Models based on clinical factors were developed to accurately stratify the risk of hsPDA in the first three days of life.Genetic features might further improve the model performance.
基金This work was supported by Grants from the National Key Research and Development Program of China(2016YFC0905100).
文摘Background Maple syrup urine disease(MSUD)is an autosomal recessive inherited disorder that affects the degradation of branched-chain amino acids and is associated with acute and chronic brain dysfunction.This study presents 11 new patients with MSUD and describes the clinical characteristics and gene mutations reported in Chinese individuals.Methods During 2011–2018,11 pedaitric patients with MSUD from 11 Chinese families were analyzed based on clinical characteristics and mass spectrometry,with confirmation via gene sequencing.Novel mutations affecting protein function were predicted with Mutation-Taster,PolyPhen-2,CADD and SIFT software.3D models of the mutated proteins were generated by using the SWISS-MODEL online server,and the models were visualized in PyMOL.The characteristics and gene mutations in patients with MSUD were analyzed retrospectively.Results Seventeen mutations in the BCKDHA,BCKDHB and DBT genes were found,8 of which are novel:c.55C>/T,c.349C>T,c.565C>T,c.808G>A,c.859C>G,and c.1270dupC in BCKDHA;c.275-2A>G in BCKDHB;and c.1291C>T in DBT.Eight patients died.Two patients had severe mental retardation and were physically handicapped.One patient with the intermediate type had relatively good prognosis,with mild psychomotor retardation and adiposity.Four mothers underwent amniocentesis for prenatal diagnosis during their second pregnancy;two fetuses were wild type,and two were carriers of one heterozygous mutation.Conclusions Eight novel mutations were associated with MSUD in Chinese patients.Prenatal diagnosis was successfully performed by genetic analysis.Mutations in the BCKDHB gene were found in the majority of Chinese patients with MSUD.
基金Shanghai Municipal Science and Technology Major Project(Grant No.20Z11900600)Clinical Research Plan of Shanghai Hospital Development Center(SHDC2020CR6028-002).
文摘Background During next generation sequencing(NGS)data interpretation in critically ill newborns,there is a potential for recognizing and reporting secondary findings(SFs).Early awareness of SFs may provide clues for disease prevention.In this study,we assessed the frequency of SFs in the China Neonatal Genomes Project(CNGP)participants.Methods A total of 2020 clinical exome sequencing(CES)datasets were screened for variants from a list of 59 genes recommended by the American College of Medical Genetics and Genomics(ACMG)for secondary findings reporting v2.0(ACMG SF v2.0).Identified variants were classified according to the evidence-based guidelines reached by a joint consensus of the ACMG and the Association for Molecular Pathology(AMP).Results Among the 2020 CES datasets,we identified 23 ACMG-reportable genes in 61 individuals,resulting in an overall frequency of SFs at 3.02%.A total of 53 unique variants were identified,including 35 pathogenic and 18 likely pathogenic variants.The common disease categories of SFs associated were cardiovascular and cancer disease.The SF results affected the medical management and follow-up strategy in 49(80.3%)patients.Conclusions We presented the frequency of SFs and their impact on clinical management strategies in CNGP participants.Our study demonstrated that SFs have important practical value in disease prevention and intervention at an early stage.
基金supported by the National Natural Science Foundation of China(81741034)the Shanghai Municipal Commission of Health and Family Planning(GDEK201701)+1 种基金the Shanghai Municipal Science and Technology Major Project(2018SHZDZX01 and 2018SHZDZX05)the Shanghai Shen Kang Hospital Development Center(SHDC12017110).
文摘Background Infantile hydrocephalus(IHC)is commonly related to other central nervous system diseases,which may have adverse effects on prognosis.The causes of IHC are heterogeneous,and the genetic etiologies are not fully understood.This study aimed to analyze the genetic etiologies of an IHC cohort.Methods The data for 110 IHC patients who had received exome sequencing at the Clinical Genetic Center of the Children's Hospital of Fudan University between 2016 and 2019 were reviewed and analyzed retrospectively.An exome-wide association analysis(EWAS)was performed within this cohort using IHC as the study phenotype.Results Of the 110 IHC patients,a pathogenic or likely pathogenic variant was identified in 16(15%)patients,spanning 13 genes.The genes were mainly associated with metabolic disorders,brain abnormalities,and genetic syndromes.IHC patients who had unclear clinical etiology were more likely to possess a genetic etiology.Based on previous studies and on our EWAS results,ZEB1,SBF2,and GNAI2 were over-represented among IHC patients and might alfect the signaling pathways involved in IHC formation.Conclusions Our study showed heterogeneous genetic etiologies in an IHC cohort.It is essential to perform genetic testing on IHC patients who have unclear clinical etiology,and genes associated with metabolic disorders,brain abnormalities,and genetic syndromes should he noted.In addition,when aiming to discover IHC susceptibility genes,genes that might influence the signaling pathways involved in IHC formation should be prioritized.
