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Acute myeloid leukaemia (AML) is a disease of the haematopoietic stem cells(HSCs) that is characterised by the uncontrolled proliferation and impaired differentiation of normal haematopoietic stem/progenitor cells. Several pathways that control the proliferation and differentiation of HSCs are impaired in AML. Activation of the Wnt/beta-catenin signalling pathway has been shown in AML and beta-catenin,which is thought to be the key element of this pathway,has been frequently highlighted. The present study was designed to determine beta-catenin expression levels and beta-catenin-related genes in AML. In this study,beta-catenin gene expression levels were determined in 19 AML patients and 3 controls by qRT-PCR. Transcriptome analysis was performed on AML grouped according to beta-catenin expression levels. Differentially expressed genes(DEGs) were investigated in detail using the Database for Annotation Visualisation and Integrated Discovery(DAVID),Gene Ontology(GO),Kyoto Encyclopedia of Genes and Genomes(KEGG),STRING online tools. The transcriptome profiles of our AML samples showed different molecular signature profiles according to their beta-catenin levels(high-low). A total of 20 genes have been identified as hub genes. Among these,TTK,HJURP,KIF14,BTF3,RPL17 and RSL1D1 were found to be associated with beta-catenin and poor survival in AML. Furthermore,for the first time in our study,the ELOV6 gene,which is the most highly up-regulated gene in human AML samples,was correlated with a poor prognosis via high beta-catenin levels. It is suggested that the identification of beta-catenin-related gene profiles in AML may help to select new therapeutic targets for the treatment of AML. View Publication

Mesenchymal stem cells (MSCs) have long been postulated as an important source cell in regenerative medicine. During subculture expansion,mesenchymal stem cell (MSC) senescence diminishes their multi-differentiation capabilities,leading to a loss of therapeutic potential. Up to date,the extrachromosomal circular DNAs (eccDNAs) have been demonstrated to be involved in senescence but the roles of eccDNAs during MSC. Here we explored eccDNA profiles in human bone marrow MSCs (BM-MSCs). EccDNA and mRNA was purified and sequenced,followed by quantification and functional annotation. Moreover,we mapped our datasets with the downloading enhancer and transcription factor-regulated genes to explore the potential role of eccDNAs. Sequentially,gene annotation analysis revealed that the majority of eccDNA were mapped in the intron regions with limited BM-MSC enhancer overlaps. We discovered that these eccDNA motifs in senescent BMSCs acted as motifs for binding transcription factors (TFs) of senescence-related genes. These findings are highly significant for identifying biomarkers of senescence and therapeutic targets in mesenchymal stem cells (MSCs) for future clinical applications. The potential of eccDNA as a stable therapeutic target for senescence-related disorders warrants further investigation,particularly exploring chemically synthesized eccDNAs as transcription factor regulatory elements to reverse cellular senescence. View Publication

As autologous induced pluripotent stem cell (iPSC) therapy requires a custom-made small-lot cell production line,and the cell production method differs significantly from the existing processes for producing allogeneic iPSC stocks for clinical use. Specifically,mass culture to produce stock is no longer necessary; instead,a series of operations from iPSC production to induction of differentiation of therapeutic cells must be performed continuously. A three-dimensional (3D) culture method using small,closed-cell manufacturing devices is suitable for autologous iPSC therapy. The use of such devices avoids the need to handle many patient-derived specimens in a single clean room; handling of cell cultures in an open system in a cell processing facility increases the risk of infection. In this study,atelocollagen beads were evaluated as a 3D biomaterial to assist 3D culture in the establishment,expansion culture,and induction of differentiation of iPSCs. It was found that iPSCs can be handled in a closed-cell device with the same ease as use of a two-dimensional (2D) culture when laminin-511 is added to the medium. In conclusion,atelocollagen beads enable 3D culture of iPSCs,and the quality of the obtained cells is at the same level as those derived from 2D culture. View Publication

The roles of the transcriptional factor SIX2 have been identified in several tumors. However,its roles in gastric cancer (GC) progression have not yet been revealed. Our objective is to explore the impact and underlying mechanisms of SIX2 on the stemness of GC cells. Lentivirus infection was employed to establish stable expression SIX2 or PFN2 in GC cells. Gain- and loss-of-function experiments were conducted to detect changes of stemness markers,flow cytometry profiles,tumor spheroid formation,and tumor-initiating ability. ChIP,RNA-sequencing,tissue microarray,and bioinformatics analysis were performed to reveal the correlation between SIX2 and PFN2. The mechanisms underlying the SIX2/PFN2 loop-mediated effects were elucidated through tissue microarray analysis,RNA stability assay,IP-MS,Co-Immunoprecipitation,and inhibition of the JNK signaling pathway. The stemness of GC cells was enhanced by SIX2. Mechanistically,SIX2 directly bound to PFN2’s promoter and promoted PFN2 activity. PFN2,in turn,promoted the mRNA stability of SIX2 by recruiting RNA binding protein YBX-1,subsequently activating the downstream MAPK/JNK pathway. This study unveils the roles of SIX2 in governing GC cell stemness,defining a novel SIX2/PFN2 regulatory loop responsible for this regulation. This suggests the potential of targeting the SIX2/PFN2 loop for GC treatment (Graphical Abstracts). The online version contains supplementary material available at 10.1186/s12967-024-05618-5. View Publication

Hematopoietic stem progenitor cells (HSPCs) undergo phenotypical and functional changes during their emergence and development. Although the molecular programs governing the development of human hematopoietic stem cells (HSCs) have been investigated broadly,the relationships between dynamic metabolic alterations and their functions remain poorly characterized. In this study,we comprehensively described the proteomics of HSPCs in the human fetal liver (FL),umbilical cord blood (UCB),and adult bone marrow (aBM). The metabolic state of human HSPCs was assessed via a Seahorse assay,RT‒PCR,and flow cytometry-based metabolic-related analysis. To investigate whether perturbing glutathione metabolism affects reactive oxygen species (ROS) production,the metabolic state,and the expansion of human HSPCs,HSPCs were treated with buthionine sulfoximine (BSO),an inhibitor of glutathione synthetase,and N-acetyl-L-cysteine (NAC). We investigated the metabolomic landscape of human HSPCs from the fetal,perinatal,and adult developmental stages by in-depth quantitative proteomics and predicted a metabolic switch from the oxidative state to the glycolytic state during human HSPC development. Seahorse assays,mitochondrial activity,ROS level,glucose uptake,and protein synthesis rate analysis supported our findings. In addition,immune-related pathways and antigen presentation were upregulated in UCB or aBM HSPCs,indicating their functional maturation upon development. Glutathione-related metabolic perturbations resulted in distinct responses in human HSPCs and progenitors. Furthermore,the molecular and immunophenotypic differences between human HSPCs at different developmental stages were revealed at the protein level for the first time. The metabolic landscape of human HSPCs at three developmental stages (FL,UCB,and aBM),combined with proteomics and functional validations,substantially extends our understanding of HSC metabolic regulation. These findings provide valuable resources for understanding human HSC function and development during fetal and adult life. The online version contains supplementary material available at 10.1186/s13287-024-03930-x. View Publication

Fatty acid synthase (FASN)-catalyzed endogenous lipogenesis is a hallmark of cancer metabolism. However,whether FASN is an intrinsic mechanism of tumor cell defense against T cell immunity remains unexplored. To test this hypothesis,here we combined bioinformatic analysis of the FASN-related immune cell landscape,real-time assessment of cell-based immunotherapy efficacy in CRISPR/Cas9-based FASN gene knockout ( FASN KO ) cell models,and mathematical and mechanistic evaluation of FASN-driven immunoresistance. FASN expression negatively correlates with infiltrating immune cells associated with cancer suppression,cytolytic activity signatures,and HLA-I expression. Cancer cells engineered to carry a loss-of-function mutation in FASN exhibit an enhanced cytolytic response and an accelerated extinction kinetics upon interaction with cytokine-activated T cells. Depletion of FASN results in reduced carrying capacity,accompanied by the suppression of mitochondrial OXPHOS and strong downregulation of electron transport chain complexes. Targeted FASN depletion primes cancer cells for mitochondrial apoptosis as it synergizes with BCL-2/BCL-X L -targeting BH3 mimetics to render cancer cells more susceptible to T-cell-mediated killing. FASN depletion prevents adaptive induction of PD-L1 in response to interferon-gamma and reduces constitutive overexpression of PD-L1 by abolishing PD-L1 post-translational palmitoylation. FASN is a novel tumor cell-intrinsic metabolic checkpoint that restricts T cell immunity and may be exploited to improve the efficacy of T cell-based immunotherapy. Subject terms: Cancer metabolism,Oncogenesis View Publication

Aging is characterized by the accumulation of proteins that display amyloid-like behavior. However,the molecular mechanisms by which these proteins arise remain unclear. Here,we demonstrate that amyloid-like proteins are produced in a variety of human cell types,including stem cells,brain organoids and fully differentiated neurons by mistakes that occur in messenger RNA molecules. Some of these mistakes generate mutant proteins already known to cause disease,while others generate proteins that have not been observed before. Moreover,we show that these mistakes increase when cells are exposed to DNA damage,a major hallmark of human aging. When taken together,these experiments suggest a mechanistic link between the normal aging process and age-related diseases. Subject terms: Protein aggregation,Mechanisms of disease,Transcription View Publication

In Multiple Sclerosis (MS),inflammatory demyelinated lesions in the brain and spinal cord lead to neurodegeneration and progressive disability. Remyelination can restore fast saltatory conduction and neuroprotection but is inefficient in MS especially with increasing age,and is not yet treatable with therapies. Intrinsic and extrinsic inhibition of oligodendrocyte progenitor cell (OPC) function contributes to remyelination failure,and we hypothesised that the transplantation of ‘improved’ OPCs,genetically edited to overcome these obstacles,could improve remyelination. Here,we edit human(h) embryonic stem cell-derived OPCs to be unresponsive to a chemorepellent released from chronic MS lesions,and transplant them into rodent models of chronic lesions. Edited hOPCs display enhanced migration and remyelination compared to controls,regardless of the host age and length of time post-transplant. We show that genetic manipulation and transplantation of hOPCs overcomes the negative environment inhibiting remyelination,with translational implications for therapeutic strategies for people with progressive MS. Subject terms: Multiple sclerosis,Multiple sclerosis,Regeneration View Publication

Venetoclax plus azacitidine treatment is clinically beneficial for elderly and unfit acute myeloid leukemia (AML) patients. However,the treatment is rarely curative,and relapse due to resistant disease eventually emerges. Since no current clinically feasible treatments are known to be effective at the state of acquired venetoclax resistance,this is becoming a major challenge in AML treatment. Studying venetoclax-resistant AML cell lines,we observed that venetoclax induced sublethal apoptotic signaling and DNA damage even though cell survival and growth were unaffected. This effect could be due to venetoclax inducing a sublethal degree of mitochondrial outer membrane permeabilization. Based on these results,we hypothesized that the sublethal apoptotic signaling induced by venetoclax could constitute a vulnerability in venetoclax-resistant AML cells. This was supported by screens with a broad collection of drugs,where we observed a synergistic effect between venetoclax and PARP inhibition in venetoclax-resistant cells. Additionally,the venetoclax-PARP inhibitor combination prevented the acquisition of venetoclax resistance in treatment naïve AML cell lines. Furthermore,the addition of azacitidine to the venetoclax-PARP inhibitor combination enhanced venetoclax induced DNA damage and exhibited exceptional sensitivity and long-term responses in the venetoclax-resistant AML cell lines and samples from AML patients that had clinically relapsed under venetoclax-azacitidine therapy. In conclusion,we mechanistically identify a new vulnerability in acquired venetoclax-resistant AML cells and identify PARP inhibition as a potential therapeutic approach to overcome acquired venetoclax resistance in AML. Subject terms: Acute myeloid leukaemia,Acute myeloid leukaemia View Publication

Our study highlights the role of epigenetic machinery in transformation of normal pluripotent stem cells to variant pluripotent state. We demonstrate the importance of non-histone protein methylation,which underlie the EMT and abnormal differentiation behaviour of variant hESCs. View Publication

PRDX2 is significantly expressed in various cancers and is associated with the proliferation of tumor cells. Nonetheless,the precise mechanism of PRDX2 in tumor immunity remains incompletely understood. This study aims to investigate the impact of PRDX2,which is highly expressed in lung adenocarcinoma,on T cells in the tumor immune microenvironment,and its immune action target to promote the immune escape of lung cancer cells,to provide a theoretical basis for lung adenocarcinoma treatment with PRDX2 as the target. Mouse animal models to verify the effect of Conoidin A treatment on tumor growth and T cell infiltration. Flow cytometry and Western blot verified tumor cell apoptosis in the in vitro co-culture system as well as granzyme B and perforin expression in T cells. RNA-Seq was used to obtain the downstream immune molecule. si-RNA knockdown of Galectin-9 was co-cultured with T cells in vitro. Immunofluorescence and Western blot verified that PRDX2 regulates Galectin-9 expression through HDAC3. PRDX2 expression was negatively correlated with CD8 + T cell expression in LUAD patients. Inhibition of PRDX2 significantly enhanced T-cell killing of LUAD cells and reduced tumor load in both in vitro and in vivo models. Mechanistically,Conoidin A or shRNA_PRDX2 decreased Galectin-9 expression by down-regulating the phosphorylation of HDAC3,consequently enhancing the infiltration and function of CD8 + T cells. This study reveals the role of the PRDX2/HDAC3/Galectin-9 axis in LUAD immune escape and indicates Galectin-9 as a promising target for immunotherapy. The online version contains supplementary material available at 10.1186/s12967-024-05888-z. View Publication

GABA B receptor (GABA B R) activation is known to alleviate pain by reducing neuronal excitability,primarily through inhibition of high voltage‐activated (HVA) calcium (Ca V 2.2) channels and potentiating G protein–coupled inwardly rectifying potassium (GIRK) channels. Although the analgesic properties of small molecules and peptides have been primarily tested on isolated murine dorsal root ganglion (DRG) neurons,emerging strategies to develop,study,and characterise human pluripotent stem cell (hPSC)‐derived sensory neurons present a promising alternative. In this study,hPSCs were efficiently differentiated into peripheral DRG‐induced sensory neurons (iSNs) using a combined chemical and transcription factor‐driven approach via a neural crest cell intermediate. Molecular characterisation and transcriptomic analysis confirmed the expression of key DRG markers such as BRN3A,ISLET1,and PRPH,in addition to GABA B R and ion channels including Ca V 2.2 and GIRK1 in iSNs. Functional characterisation of GABA B R was conducted using whole‐cell patch clamp electrophysiology,assessing neuronal excitability under current‐clamp conditions in the absence and presence of GABA B R agonists baclofen and α‐conotoxin Vc1.1. Both baclofen (100 μM) and Vc1.1 (1 μM) significantly reduced membrane excitability by hyperpolarising the resting membrane potential and increasing the rheobase for action potential firing. In voltage‐clamp mode,baclofen and Vc1.1 inhibited HVA Ca 2+ channel currents,which were attenuated by the selective GABA B R antagonist CGP 55845. However,modulation of GIRK channels by GABA B Rs was not observed in the presence of baclofen or Vc1.1,suggesting that functional GIRK1/2 channels were not coupled to GABA B Rs in hPSC‐derived iSNs. This study is the first to report GABA B R modulation of membrane excitability in iSNs by baclofen and Vc1.1,highlighting their potential as a future model for studying analgesic compounds. View Publication