Purpose The present study aimed to evaluate the genetic diagnostic yield and accuracy of exome sequencing in Chinese patients with split hand–foot malformation(SHFM),a severe heterogeneous congenital anomaly characte...Purpose The present study aimed to evaluate the genetic diagnostic yield and accuracy of exome sequencing in Chinese patients with split hand–foot malformation(SHFM),a severe heterogeneous congenital anomaly characterized by hypodevelopment of the central ray of the hands and feet.Methods A cohort of seven families and five sporadic patients with SHFM was investigated.Genomic DNA was prepared from the peripheral blood of affected as well as unaffected individuals.Whole exome sequencing(WES)was performed to identify the pathogenic mutations.Array-based comparative genomic hybridization(aCGH),CytoScan,quantitative polymerase chain reaction(qPCR),and Sanger sequencing were performed to validate the findings of WES.WES data of an additional cohort of 24 patients with non-SHFM congenital hand anomalies were analyzed as the control.Results Pathogenic variants of TP63,c.G956A p.R319H,and c.T602A:p.L201H,were identified in two families by WES.In the remaining patients,copy number analysis of the WES data by XHMM software identified pathogenic 10q 24 duplication in five individuals from three families,which was further validated via CytoScan and qPCR;however,WES could not detect duplication in 10q24 in an additional cohort of 24 individuals with non-SHFM congenital hand anomaly.Importantly,qPCR analysis of the 10q24 region copy number revealed a definite consistency with WES data in all individuals.Genotype–phenotype analysis did not present any unique feature that could differentiate between the families with TP63 mutation and 10q24 duplication.Conclusions Our study demonstrated that WES is an accurate and sensitive method to detect the pathogenic 10q24 duplication.Collectively,with TP63 mutation,a single WES testing could yield a diagnosis rate of about 40%(5/12)for the SHFM patients,at least in our cohort.As the genotype–phenotype correlation remains unclear,WES could be used as a cost-effective method for the genetic diagnosis of SHFM.展开更多
Tendinopathy is a common musculoskeletal disorder which results in chronic pain and reduced performance.The therapeutic effect of stem cell derived-small extracellular vesicles(sEVs)for tendinopathy has been validated...Tendinopathy is a common musculoskeletal disorder which results in chronic pain and reduced performance.The therapeutic effect of stem cell derived-small extracellular vesicles(sEVs)for tendinopathy has been validated in recent years.However,whether large extracellular vesicles(lEVs),another subset of extracellular vesicles,possesses the ability for the improvement of tendinopathy remains unknown.Here,we showed that lEVs secreted from iPSC-derived MSCs(iMSC-lEVs)significantly mitigated pain derived from tendinopathy in rats.Immuno-histochemical analysis showed that iMSC-lEVs regulated the heterogeneity of infiltrated macrophages and several inflammatory cytokines in rat tendon tissue.Meanwhile,in vitro experiments revealed that the M1 pro-inflammatory macrophages were repolarized towards M2 anti-inflammatory macrophages by iMSC-lEVs,and this effect was mediated by regulating p38 MAPK pathway.Moreover,liquid chromatography-tandem mass spectrometry analysis identified 2208 proteins encapsulated in iMSC-lEVs,including 134 new-found proteins beyond current Vesiclepedia database.By bioinformatics and Western blot analyses,we showed that DUSP2 and DUSP3,the negative regulator of p38 phosphorylation,were enriched in iMSC-lEVs and could be transported to macrophages.Further,the immunomodulatory effect of iMSC-lEVs on macrophages was validated in explant tendon tissue from tendinopathy patients.Taken together,our results demonstrate that iMSC-lEVs could reduce inflammation in tendinopathy by regulating macrophage heterogeneity,which is mediated via the p38 MAPK pathway by delivery of DUSP2 and DUSP3,and might be a promising candidate for tendinopathy therapy.展开更多
基金We would like to thank the families for their participation in this study.We would also like to thank our funding sources:This study was supported by National Science Funding of China No.81772115 and 81571930 to BW.
文摘Purpose The present study aimed to evaluate the genetic diagnostic yield and accuracy of exome sequencing in Chinese patients with split hand–foot malformation(SHFM),a severe heterogeneous congenital anomaly characterized by hypodevelopment of the central ray of the hands and feet.Methods A cohort of seven families and five sporadic patients with SHFM was investigated.Genomic DNA was prepared from the peripheral blood of affected as well as unaffected individuals.Whole exome sequencing(WES)was performed to identify the pathogenic mutations.Array-based comparative genomic hybridization(aCGH),CytoScan,quantitative polymerase chain reaction(qPCR),and Sanger sequencing were performed to validate the findings of WES.WES data of an additional cohort of 24 patients with non-SHFM congenital hand anomalies were analyzed as the control.Results Pathogenic variants of TP63,c.G956A p.R319H,and c.T602A:p.L201H,were identified in two families by WES.In the remaining patients,copy number analysis of the WES data by XHMM software identified pathogenic 10q 24 duplication in five individuals from three families,which was further validated via CytoScan and qPCR;however,WES could not detect duplication in 10q24 in an additional cohort of 24 individuals with non-SHFM congenital hand anomaly.Importantly,qPCR analysis of the 10q24 region copy number revealed a definite consistency with WES data in all individuals.Genotype–phenotype analysis did not present any unique feature that could differentiate between the families with TP63 mutation and 10q24 duplication.Conclusions Our study demonstrated that WES is an accurate and sensitive method to detect the pathogenic 10q24 duplication.Collectively,with TP63 mutation,a single WES testing could yield a diagnosis rate of about 40%(5/12)for the SHFM patients,at least in our cohort.As the genotype–phenotype correlation remains unclear,WES could be used as a cost-effective method for the genetic diagnosis of SHFM.
基金National Natural Science Foundation of China(Grant No.81870972,82072550)Science and Technology Commission of Shanghai Municipality(Grant No.21DZ2201300).
文摘Tendinopathy is a common musculoskeletal disorder which results in chronic pain and reduced performance.The therapeutic effect of stem cell derived-small extracellular vesicles(sEVs)for tendinopathy has been validated in recent years.However,whether large extracellular vesicles(lEVs),another subset of extracellular vesicles,possesses the ability for the improvement of tendinopathy remains unknown.Here,we showed that lEVs secreted from iPSC-derived MSCs(iMSC-lEVs)significantly mitigated pain derived from tendinopathy in rats.Immuno-histochemical analysis showed that iMSC-lEVs regulated the heterogeneity of infiltrated macrophages and several inflammatory cytokines in rat tendon tissue.Meanwhile,in vitro experiments revealed that the M1 pro-inflammatory macrophages were repolarized towards M2 anti-inflammatory macrophages by iMSC-lEVs,and this effect was mediated by regulating p38 MAPK pathway.Moreover,liquid chromatography-tandem mass spectrometry analysis identified 2208 proteins encapsulated in iMSC-lEVs,including 134 new-found proteins beyond current Vesiclepedia database.By bioinformatics and Western blot analyses,we showed that DUSP2 and DUSP3,the negative regulator of p38 phosphorylation,were enriched in iMSC-lEVs and could be transported to macrophages.Further,the immunomodulatory effect of iMSC-lEVs on macrophages was validated in explant tendon tissue from tendinopathy patients.Taken together,our results demonstrate that iMSC-lEVs could reduce inflammation in tendinopathy by regulating macrophage heterogeneity,which is mediated via the p38 MAPK pathway by delivery of DUSP2 and DUSP3,and might be a promising candidate for tendinopathy therapy.
