技术资料

ABSTRACTRAD50?interacting protein1 (RINT1) deficiency has been implicated in recurrent acute liver failure (RALF) triggered by fever or infections. RINT1,together with neuroblastoma amplified sequence and Zeste White 10 (forming the NRZ complex),localizes at the interface between the endoplasmic reticulum and Golgi apparatus,where it plays a key role in vesicular trafficking. However,the mechanisms by which RINT1 deficiency leads to RALF remain unclear. This study aimed to describe a woman with RALF harboring a homozygous missense mutation in RINT1. Induced pluripotent stem cells (iPSCs) were generated from the patient's mononuclear cells and differentiated into hepatocyte?like cells (HLCs). Upon exposure to high temperature (40°C),RINT1?deficient HLCs exhibited cellular damage characteristic of RALF. Furthermore,these cells also demonstrated heightened sensitivity to cytokines and viral mimetics while showing comparatively lower responsiveness to bacterial infection?related stimuli. Transcriptome sequencing revealed dysregulated gene expression associated with the extracellular matrix (ECM). Additionally,glycosaminoglycan disaccharide analysis revealed abnormal levels of chondroitin sulfate,heparan sulfate,and hyaluronan in RINT1?deficient HLCs. In conclusion,HLCs derived from RINT1?deficient iPSCs serve as a valuable model for investigating RINT1?related liver pathogenesis. The results suggest that cytokine responses,particularly those triggered by viral infections,play a central role in the development of RALF. Furthermore,ECM alterations provided novel insights into the potential role of RINT1 defects in RALF. RAD50?interacting protein1 (RINT1) deficiency causes recurrent acute liver failure (RALF) during fever or infections. To investigate its underlying mechanism,induced pluripotent stem cells were generated from a patient with RINT1 deficiency and differentiated into hepatocyte?like cells (HLCs). RINT1?deficient HLCs exhibited damage resembling RALF when exposed to high temperatures and were more susceptible to cytokines and viral mimetics than to bacterial infection?related factors. Furthermore,RNA?seq and disaccharide analyses revealed dysregulation of extracellular matrix?related genes and abnormalities in extracellular matrix levels. View Publication

Creatine transporter deficiency (CTD) is an X-linked disease caused by mutations in the Slc6a8 gene. The impaired creatine uptake in the brain leads to developmental delays with intellectual disability. We hypothesized that deficient creatine uptake in CTD cerebral cells impact methylation balance leading to alterations of genes and proteins expression by epigenetic mechanism. In this study,we determined the status of nucleic acid methylation in both Slc6a8 knockout mouse model and brain organoids derived from CTD patients’ cells. We also investigated the effect of dodecyl creatine ester (DCE),a promising prodrug that increases brain creatine content in the mouse model of CTD. The level of nucleic acid methylation was significantly reduced compared to healthy controls in both in vivo and in vitro CTD models. This hypo-methylation tended to be regulated by DCE treatment in vivo. These results suggest that increased brain creatine after DCE treatment restores normal levels of DNA methylation,unveiling the potential of using DNA methylation as a marker to monitor the drug efficacy. View Publication

A patient with the PSEN1 E280A mutation and homozygous for APOE3 Christchurch (APOE3Ch) displayed extreme resistance to Alzheimer’s disease (AD) cognitive decline and tauopathy,despite having a high amyloid burden. To further investigate the differences in biological processes attributed to APOE3Ch,we generated induced pluripotent stem (iPS) cell-derived cerebral organoids from this resistant case and a non-protected control,using CRISPR/Cas9 gene editing to modulate APOE3Ch expression. In the APOE3Ch cerebral organoids,we observed a protective pattern from early tau phosphorylation. ScRNA sequencing revealed regulation of Cadherin and Wnt signaling pathways by APOE3Ch,with immunostaining indicating elevated ?-catenin protein levels. Further in vitro reporter assays unexpectedly demonstrated that ApoE3Ch functions as a Wnt3a signaling enhancer. This work uncovered a neomorphic molecular mechanism of protection of ApoE3 Christchurch,which may serve as the foundation for the future development of protected case-inspired therapeutics targeting AD and tauopathies. View Publication

Human cortical neural progenitor cell transplantation holds significant potential in cortical stroke treatment by replacing lost cortical neurons and repairing damaged brain circuits. However,commonly utilized human cortical neural progenitors are limited in yield a substantial proportion of diverse cortical neurons and require an extended period to achieve functional maturation and synaptic integration,thereby potentially diminishing the optimal therapeutic benefits of cell transplantation for cortical stroke. Here,we generated forkhead box G1 (FOXG1)-positive forebrain progenitors from human inducible pluripotent stem cells,which can differentiate into diverse and balanced cortical neurons including upper- and deep-layer excitatory and inhibitory neurons,achieving early functional maturation simultaneously in vitro. Furthermore,these FOXG1 forebrain progenitor cells demonstrate robust cortical neuronal differentiation,rapid functional maturation and efficient synaptic integration after transplantation into the sensory cortex of stroke-injured adult rats. Notably,we have successfully utilized the non-invasive 18F-SynVesT-1 PET imaging technique to assess alterations in synapse count before and after transplantation therapy of FOXG1 progenitors in vivo. Moreover,the transplanted FOXG1 progenitors improve sensory and motor function recovery following stroke. These findings provide systematic and compelling evidence for the suitability of these FOXG1 progenitors for neuronal replacement in ischemic cortical stroke. Human NPCs show promise for stroke treatment,but challenges remain in neuron diversity and maturation time. Here,the authors describe the generation of FOXG1 progenitors from iPSCs that quickly mature into functional cortical neurons,enhancing stroke recovery in rats. View Publication

The generation of retinal models from human induced pluripotent stem cells holds significant potential for advancing our understanding of retinal development,neurodegeneration,and the in vitro modeling of neurodegenerative disorders. The retina,as an accessible part of the central nervous system,offers a unique window into these processes,making it invaluable for both study and early diagnosis. This study investigates the impact of the Frontotemporal Dementia-linked IVS 10?+?16 MAPT mutation on retinal development and function using 2D and 3D retinal models derived from human induced pluripotent stem cells. Our findings reveal that the MAPT mutation leads to delayed retinal cell differentiation and maturation,with tau-mutant disease models exhibiting sustained higher expression of retinal progenitor cell markers and a reduced presence of post-mitotic neurons. Both 2D and 3D tau-mutant retinal models demonstrated an imbalance in tau isoforms,favoring 4R tau,along with increased tau phosphorylation,altered neurite morphology,and impaired cytoskeletal maturation. These changes are associated with impaired synaptic development,reduced neuronal connectivity,and enhanced cellular stress responses,including the increased formation of stress granules,markers of apoptosis and autophagy,and the presence of intracellular toxic tau aggregates. This study highlights the value of retinal models derived from human induced pluripotent stem cells in exploring the mechanisms underlying retinal pathology associated with tau mutations. These models offer essential insights into the development of therapeutic strategies for neurodegenerative diseases characterized by tau aggregation.Supplementary InformationThe online version contains supplementary material available at 10.1186/s40478-024-01920-x. View Publication

Increasing shreds of evidence suggest that neurogenic-to-gliogenic shift may be critical to the abnormal neurodevelopment observed in individuals with Down syndrome (DS). REST,the Repressor Element-1 Silencing Transcription factor,regulates the differentiation and development of neural cells. Downregulation of REST may lead to defects in post-differentiation neuronal morphology in the brain of the DS fetal. This study aims to elucidate the role of REST in DS-derived NPCs using bioinformatics analyses and laboratory validations. We identified and validated vital REST-targeted DEGs: CD44,TGFB1,FN1,ITGB1,and COL1A1. Interestingly,these genes are involved in neurogenesis and gliogenesis in DS-derived NPCs. Furthermore,we identified nuclear REST loss and the neuroblast marker,DCX,was downregulated in DS human trisomic induced pluripotent stem cells (hiPSCs)-derived NPCs,whereas the glioblast marker,NFIA,was upregulated. Our findings indicate that the loss of REST is critical in the neurogenic-to-gliogenic shift observed in DS-derived NPCs. REST and its target genes may collectively regulate the NPC phenotype.Supplementary InformationThe online version contains supplementary material available at 10.1038/s41598-025-87314-y. View Publication

Peroxisomes are eukaryotic organelles that are essential for multiple metabolic pathways,including fatty acid oxidation,degradation of amino acids,and biosynthesis of ether lipids. Consequently,peroxisome dysfunction leads to pediatric-onset neurodegenerative conditions,including Peroxisome Biogenesis Disorders (PBD). Due to the dynamic,tissue-specific,and context-dependent nature of their biogenesis and function,live cell imaging of peroxisomes is essential for studying peroxisome regulation,as well as for the diagnosis of PBD-linked abnormalities. However,the peroxisomal imaging toolkit is lacking in many respects,with no reporters for substrate import,nor cell-permeable probes that could stain dysfunctional peroxisomes. Here we report that the BODIPY-C12 fluorescent fatty acid probe stains functional and dysfunctional peroxisomes in live mammalian cells. We then go on to improve BODIPY-C12,generating peroxisome-specific reagents,PeroxiSPY650 and PeroxiSPY555. These probes combine high peroxisome specificity,bright fluorescence in the red and far-red spectrum,and fast non-cytotoxic staining,making them ideal tools for live cell,whole organism,or tissue imaging of peroxisomes. Finally,we demonstrate that PeroxiSPY enables diagnosis of peroxisome abnormalities in the PBD CRISPR/Cas9 cell models and patient-derived cell lines. The array of tools to image peroxisome regulation is still limited. Here,the authors develop improved fatty acid-based probes with high peroxisome specificity and bright fluorescence in the red/far-red spectrum,which makes them ideal to study peroxisomes in live cells and whole organisms. View Publication

Somatic cell reprogramming into human induced pluripotent stem cells entails significant intracellular changes,including modifications in mitochondrial metabolism and a decrease in mitochondrial DNA copy number. However,the mechanisms underlying this decrease in mitochondrial DNA copy number during reprogramming remain unclear. Here we aimed to elucidate these underlying mechanisms. Through a meta-analysis of several RNA sequencing datasets,we identified genes responsible for the decrease in mitochondrial DNA. We investigated the functions of these identified genes and assessed their regulatory mechanisms. In particular,the expression of the thymidine kinase 2 gene (TK2),located in the mitochondria and required for mitochondrial DNA synthesis,is decreased in human pluripotent stem cells as compared with its expression in somatic cells. TK2 was significantly downregulated during reprogramming and markedly upregulated during differentiation. Collectively,this decrease in TK2 levels induces a decrease in mitochondrial DNA copy number and contributes to shaping the metabolic characteristics of human pluripotent stem cells. However,contrary to our expectations,treatment with a TK2 inhibitor impaired somatic cell reprogramming. These results suggest that decreased TK2 expression may result from metabolic conversion during somatic cell reprogramming. Mitochondrial DNA loss linked to stem cell reprogrammingInduced pluripotent stem (iPS) cells are special cells created by reprogramming regular body cells. Researchers explored how these cells change their energy production methods during reprogramming. The study focused on a protein called thymidine kinase 2 (TK2),which is important for maintaining mitochondrial DNA (mtDNA). Mitochondria are the cell’s powerhouses,and their DNA is crucial for energy production. Researchers used human cell lines to study how TK2 affects mtDNA during reprogramming. They found that,as cells become iPS cells,TK2 levels drop,leading to reduced mtDNA and a shift in energy production from oxidative phosphorylation to glycolysis. Results suggest that reducing TK2 and mtDNA is key for cells to gain pluripotency. This shift helps support the rapid growth and development of iPS cells. Understanding this process could improve stem cell therapies and regenerative medicine in the future.This summary was initially drafted using artificial intelligence,then revised and fact-checked by the author. View Publication