Purpose:Familial exudative vitreoretinopathy(FEVR)is a genetically heterogeneous retinal vascular disorder,with nearly half of the cases attributed to mutations in genes involved in the Norrin/β-catenin signaling pat...Purpose:Familial exudative vitreoretinopathy(FEVR)is a genetically heterogeneous retinal vascular disorder,with nearly half of the cases attributed to mutations in genes involved in the Norrin/β-catenin signaling pathway.This study aimed to identify and functionally characterize novel FZD4 variants in patients with FEVR.Methods:Genetic sequencing of FZD4 was performed in a cohort of FEVR families,leading to the identification of five novel variants:c.434G>A,c.610T>G,c.844T>C,c.277C>T,and c.1155delC.Bioinformatic predictions,comprehensive clinical evaluations,and dual-luciferase reporter assays were conducted to assess the functional impact and pathogenicity of these variants.Results:All five FZD4 variants were found to significantly reduceβ-catenin signaling activity compared to wild-type FZD4.Among them,two variants previously classified as variants of uncertain significance(VUS)demon-strated functional impairment and clinical segregation consistent with pathogenicity,supporting their reclassi-fication as disease-causing mutations.Conclusions:These findings expand the known mutational spectrum of FZD4 in FEVR and highlight the critical role of functional validation in the interpretation of novel and uncertain variants.Incorporating experimental assays can improve diagnostic accuracy and inform clinical genetic counseling.展开更多
Objective:In the realm of Class I pathogens,Burkholderia pseudomallei(BP)stands out for its propensity to induce severe pathogenicity.Investigating the intricate interactions between BP and host cells is imperative fo...Objective:In the realm of Class I pathogens,Burkholderia pseudomallei(BP)stands out for its propensity to induce severe pathogenicity.Investigating the intricate interactions between BP and host cells is imperative for comprehending the dynamics of BP infection and discerning biomarkers indicative of the host cell response process.Methods:mRNA extraction from BP-infected mouse macrophages constituted the initial step of our study.Employing gene expression arrays,the extracted RNA underwent conversion into digital signals.The percentile shift method facilitated data processing,with the identification of genes manifesting significant differences accomplished through the application of the t-test.Subsequently,a comprehensive analysis involving Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway was conducted on the differentially expressed genes(DEGs).Leveraging the ESTIMATE algorithm,gene signatures were utilized to compute risk scores for gene expression data.Support vector machine analysis and gene enrichment scores were instrumental in establishing correlations between biomarkers and macrophages,followed by an evaluation of the predictive power of the identified biomarkers.Results:The functional and pathway associations of the DEGs predominantly centered around G protein-coupled receptors.A noteworthy positive correlation emerged between the blue module,consisting of 416 genes,and the StromaScore.FZD4,identified through support vector machine analysis among intersecting genes,indicated a robust interaction with macrophages,suggesting its potential as a robust biomarker.FZD4 exhibited commendable predictive efficacy,with BP infection inducing its expression in both macrophages and mouse lung tissue.Western blotting in macrophages confirmed a significant upregulation of FZD4 expression from 0.5 to 24 h post-infection.In mouse lung tissue,FZD4 manifested higher expression in the cytoplasm of pulmonary epithelial cells in BP-infected lungs than in the control group.Conclusion:Thesefindings underscore the upregulation of FZD4 expression by BP in both macrophages and lung tissue,pointing to its prospective role as a biomarker in the pathogenesis of BP infection.展开更多
基金supported by the National Natural Science Foundation of China(82271092).
文摘Purpose:Familial exudative vitreoretinopathy(FEVR)is a genetically heterogeneous retinal vascular disorder,with nearly half of the cases attributed to mutations in genes involved in the Norrin/β-catenin signaling pathway.This study aimed to identify and functionally characterize novel FZD4 variants in patients with FEVR.Methods:Genetic sequencing of FZD4 was performed in a cohort of FEVR families,leading to the identification of five novel variants:c.434G>A,c.610T>G,c.844T>C,c.277C>T,and c.1155delC.Bioinformatic predictions,comprehensive clinical evaluations,and dual-luciferase reporter assays were conducted to assess the functional impact and pathogenicity of these variants.Results:All five FZD4 variants were found to significantly reduceβ-catenin signaling activity compared to wild-type FZD4.Among them,two variants previously classified as variants of uncertain significance(VUS)demon-strated functional impairment and clinical segregation consistent with pathogenicity,supporting their reclassi-fication as disease-causing mutations.Conclusions:These findings expand the known mutational spectrum of FZD4 in FEVR and highlight the critical role of functional validation in the interpretation of novel and uncertain variants.Incorporating experimental assays can improve diagnostic accuracy and inform clinical genetic counseling.
基金The study was supported by Yuying Program Incubation Project of General Hospital of Center Theater(ZZYFH202104)Wuhan Young and Middle-Aged Medical Backbone Talent Project 2020(2020-55)Logistics Research Program Project 2019(CLB19J029).
文摘Objective:In the realm of Class I pathogens,Burkholderia pseudomallei(BP)stands out for its propensity to induce severe pathogenicity.Investigating the intricate interactions between BP and host cells is imperative for comprehending the dynamics of BP infection and discerning biomarkers indicative of the host cell response process.Methods:mRNA extraction from BP-infected mouse macrophages constituted the initial step of our study.Employing gene expression arrays,the extracted RNA underwent conversion into digital signals.The percentile shift method facilitated data processing,with the identification of genes manifesting significant differences accomplished through the application of the t-test.Subsequently,a comprehensive analysis involving Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway was conducted on the differentially expressed genes(DEGs).Leveraging the ESTIMATE algorithm,gene signatures were utilized to compute risk scores for gene expression data.Support vector machine analysis and gene enrichment scores were instrumental in establishing correlations between biomarkers and macrophages,followed by an evaluation of the predictive power of the identified biomarkers.Results:The functional and pathway associations of the DEGs predominantly centered around G protein-coupled receptors.A noteworthy positive correlation emerged between the blue module,consisting of 416 genes,and the StromaScore.FZD4,identified through support vector machine analysis among intersecting genes,indicated a robust interaction with macrophages,suggesting its potential as a robust biomarker.FZD4 exhibited commendable predictive efficacy,with BP infection inducing its expression in both macrophages and mouse lung tissue.Western blotting in macrophages confirmed a significant upregulation of FZD4 expression from 0.5 to 24 h post-infection.In mouse lung tissue,FZD4 manifested higher expression in the cytoplasm of pulmonary epithelial cells in BP-infected lungs than in the control group.Conclusion:Thesefindings underscore the upregulation of FZD4 expression by BP in both macrophages and lung tissue,pointing to its prospective role as a biomarker in the pathogenesis of BP infection.
