技术资料

Acute myeloid leukemia (AML) is a heterogenous disease characterized by the clonal expansion of myeloid progenitor cells. Despite recent advancements in the treatment of AML,relapse still remains a significant challenge,necessitating the development of innovative therapies to eliminate minimal residual disease. One promising approach to address these unmet clinical needs is natural killer (NK) cell immunotherapy. To implement such treatments effectively,it is vital to comprehend how AML cells escape the NK-cell surveillance. Signal transducer and activator of transcription 3 (STAT3),a component of the Janus kinase (JAK)-STAT signaling pathway,is well-known for its role in driving immune evasion in various cancer types. Nevertheless,the specific function of STAT3 in AML cell escape from NK cells has not been deeply investigated. In this study,we unravel a novel role of STAT3 in sensitizing AML cells to NK-cell surveillance. We demonstrate that STAT3-deficient AML cell lines are inefficiently eliminated by NK cells. Mechanistically,AML cells lacking STAT3 fail to form an immune synapse as efficiently as their wild-type counterparts due to significantly reduced surface expression of intercellular adhesion molecule 1 (ICAM-1). The impaired killing of STAT3-deficient cells can be rescued by ICAM-1 overexpression proving its central role in the observed phenotype. Importantly,analysis of our AML patient cohort revealed a positive correlation between ICAM1 and STAT3 expression suggesting a predominant role of STAT3 in ICAM-1 regulation in this disease. In line,high ICAM1 expression correlates with better survival of AML patients underscoring the translational relevance of our findings. Taken together,our data unveil a novel role of STAT3 in preventing AML cells from escaping NK-cell surveillance and highlight the STAT3/ICAM-1 axis as a potential biomarker for NK-cell therapies in AML. View Publication

Mycobacterium tuberculosis ( Mtb ),the causative agent of tuberculosis (TB),is the most common coinfection among people living with HIV-1. This coinfection is associated with accelerated HIV-1 disease progression and reduced survival. However,the impact of the HIV-1/TB coinfection on HIV-1 replication and latency in CD4 + T cells remains poorly studied. Using the acellular fraction of tuberculous pleural effusion (TB-PE),we investigated whether viral replication and HIV-1 latency in CD4 + T cells are affected by a TB-associated microenvironment. Our results revealed that TB-PE impaired T cell receptor-dependent cell activation and decreased HIV-1 replication in CD4 + T cells. Moreover,this immunosuppressive TB microenvironment promoted viral latency and inhibited HIV-1 reactivation. This study indicates that the TB-induced immune response may contribute to the persistence of the viral reservoir by silencing HIV-1 expression,allowing the virus to persist undetected by the immune system,and increasing the size of the latent HIV-1 reservoir. Subject areas: Immunology,Virology View Publication

Delivering macromolecules across biological barriers such as the blood–brain barrier limits their application in vivo. Previous work has demonstrated that Toxoplasma gondii,a parasite that naturally travels from the human gut to the central nervous system (CNS),can deliver proteins to host cells. Here we engineered T. gondii ’s endogenous secretion systems,the rhoptries and dense granules,to deliver multiple large (>100 kDa) therapeutic proteins into neurons via translational fusions to toxofilin and GRA16. We demonstrate delivery in cultured cells,brain organoids and in vivo,and probe protein activity using imaging,pull-down assays,scRNA-seq and fluorescent reporters. We demonstrate robust delivery after intraperitoneal administration in mice and characterize 3D distribution throughout the brain. As proof of concept,we demonstrate GRA16-mediated brain delivery of the MeCP2 protein,a putative therapeutic target for Rett syndrome. By characterizing the potential and current limitations of the system,we aim to guide future improvements that will be required for broader application. Subject terms: Parasitology,Biologics,Synthetic biology View Publication

Most studies used animal serum-containing medium for bioengineered-root regeneration,but ethical and safety issues raised by animal serum are a potentially significant risk for clinical use. Thus,this study aimed to find a safer method for bioengineered-root regeneration. The biological properties of human dental pulp stem cells (hDPSCs) cultured in animal component-free (ACF) medium or serum-containing medium (5%,10% serum-containing medium,SCM) were compared in vitro . hDPSCs were cultured in a three-dimensional (3D) environment with human-treated dentin matrix (hTDM). The capacity for odontogenesis was compared using quantitative real-time PCR (qPCR) and Western blot. Subsequently,the hDPSCs/hTDM complexes were transplanted into nude mice subcutaneously. Histological staining was then used to verify the regeneration effect in vivo . ACF medium promoted the migration of hDPSCs,but slightly inhibited the proliferation of hDPSCs in the first three days of culture compared to SCM. However,it had no significant effect on cell aging and apoptosis. After 7 days of 3D culture in ACF medium with hTDM,qPCR showed that DMP1,DSPP,OCN,RUNX2,and β-tubulin III were highly expressed in hDPSCs. In addition,3D cultured hDPSCs/hTDM complexes in ACF medium regenerated dentin,pulp,and periodontal ligament-like tissues similar to SCM groups in vivo . ACF medium was proved to be an alternative medium for bioengineered-root regeneration. The strategy of using ACF medium to regenerate bioengineered-root can improve clinical safety for tooth tissue engineering. View Publication

Besides neutralizing antibodies,which are considered an important measure for vaccine immunogenicity,Fc-mediated antibody functions can contribute to antibody-mediated protection. They are strongly influenced by structural antibody properties such as subclass and Fc glycan composition. We here applied a systems serology approach to dissect humoral immune responses induced by MVA-MERS-S,an MVA-vectored vaccine against the Middle East respiratory syndrome coronavirus (MERS-CoV). Building on preceding studies reporting the safety and immunogenicity of MVA-MERS-S,our study highlights the potential of a late boost,administered one year after prime,to enhance both neutralizing and Fc-mediated antibody functionality compared to the primary vaccination series. Distinct characteristics were observed for antibodies specific to the MERS-CoV spike protein S1 and S2 subunits,regarding subclass and glycan compositions as well as Fc functionality. These findings highlight the benefit of a late homologous booster vaccination with MVA-MERS-S and may be of interest for the design of future coronavirus vaccines. Subject areas: Cell biology,Immune response,Immunology,Virology View Publication

Streptococcus pneumoniae is a global priority respiratory pathogen that kills over a million people annually. The pore-forming cytotoxin,pneumolysin (PLY) is a major virulence factor. Here,we found that recombinant PLY as well as wild-type pneumococcal strains,but not the isogenic PLY mutant,upregulated the shedding of extracellular vesicles (EVs) harboring membrane-bound toxin from human THP-1 monocytes. PLY-EVs induced cytotoxicity and hemolysis dose-dependently upon internalization by recipient monocyte-derived dendritic cells. Proteomics analysis revealed that PLY-EVs are selectively enriched in key inflammatory host proteins such as IFI16,NLRC4,PTX3,and MMP9. EVs shed from PLY-challenged or infected cells induced dendritic cell maturation and primed them to infection. In vivo,zebrafish administered with PLY-EVs showed pericardial edema and mortality. Adoptive transfer of bronchoalveolar-lavage-derived EVs from infected mice to healthy recipients induced lung damage and inflammation in a PLY-dependent manner. Our findings identify that host EVs released during infection mediate pneumococcal pathogenesis. Subject areas: Microbiology,Bacteriology,Cell biology View Publication

Intrahepatic cholangiocarcinoma (iCCA) is a lethal primary liver tumor characterized by clinical aggressiveness,poor prognosis,and scarce therapeutic possibilities. Therefore,new treatments are urgently needed to render this disease curable. Since cumulating evidence supports the oncogenic properties of the Heat Shock Factor 1 (HSF1) transcription factor in various cancer types,we investigated its pathogenetic and therapeutic relevance in iCCA. Levels of HSF1 were evaluated in a vast collection of iCCA specimens. The effects of HSF1 inactivation on iCCA development in vivo were investigated using three established oncogene-driven iCCA mouse models. In addition,the impact of HSF1 suppression on tumor cells and tumor stroma was assessed in iCCA cell lines,human iCCA cancer-associated fibroblasts (hCAFs),and patient-derived organoids. Human preinvasive,invasive,and metastatic iCCAs displayed widespread HSF1 upregulation,which was associated with a dismal prognosis of the patients. In addition,hydrodynamic injection of a dominant-negative form of HSF1 (HSF1dn),which suppresses HSF1 activity,significantly delayed cholangiocarcinogenesis in AKT/NICD,AKT/YAP,and AKT/TAZ mice. In iCCA cell lines,iCCA hCAFs,and patient-derived organoids,administration of the HSF1 inhibitor KRIBB-11 significantly reduced proliferation and induced apoptosis. Cell death was profoundly augmented by concomitant administration of the Bcl-xL/Bcl2/Bcl-w inhibitor ABT-263. Furthermore,KRIBB-11 reduced mitochondrial bioenergetics and glycolysis of iCCA cells. The present data underscore the critical pathogenetic,prognostic,and therapeutic role of HSF1 in cholangiocarcinogenesis. The online version contains supplementary material available at 10.1186/s13046-024-03177-7. View Publication

Despite their role as innate sentinels,macrophages can serve as cellular reservoirs of chikungunya virus (CHIKV),a highly-pathogenic arthropod-borne alphavirus that has caused large outbreaks among human populations. Here,with the use of viral chimeras and evolutionary selection analysis,we define CHIKV glycoproteins E1 and E2 as critical for virion production in THP-1 derived human macrophages. Through proteomic analysis and functional validation,we further identify signal peptidase complex subunit 3 (SPCS3) and eukaryotic translation initiation factor 3 subunit K (eIF3k) as E1-binding host proteins with anti-CHIKV activities. We find that E1 residue V220,which has undergone positive selection,is indispensable for CHIKV production in macrophages,as its mutation attenuates E1 interaction with the host restriction factors SPCS3 and eIF3k. Finally,we show that the antiviral activity of eIF3k is translation-independent,and that CHIKV infection promotes eIF3k translocation from the nucleus to the cytoplasm,where it associates with SPCS3. These functions of CHIKV glycoproteins late in the viral life cycle provide a new example of an intracellular evolutionary arms race with host restriction factors,as well as potential targets for therapeutic intervention. View Publication

Monocytes,the circulating macrophage precursors,contribute to diseases like atherosclerosis and asthma. Long non-coding RNAs (lncRNAs) have been shown to modulate the phenotype and inflammatory capacity of monocytes. We previously discovered the lncRNA SMANTIS,which contributes to cellular phenotype expression by controlling BRG1 in mesenchymal cells. Here,we report that SMANTIS is particularly highly expressed in monocytes and lost during differentiation into macrophages. Moreover,different types of myeloid leukemia presented specific SMANTIS expression patterns. Interaction studies revealed that SMANTIS binds RUNX1,a transcription factor frequently mutated in AML,primarily through its Alu-element on the RUNT domain. RNA-seq after CRISPR/Cas9-mediated deletion of SMANTIS or RUNX1 revealed an association with cell adhesion and both limited the monocyte adhesion to endothelial cells. Mechanistically,SMANTIS KO reduced RUNX1 genomic binding and altered the interaction of RUNX1 with EP300 and CBFB. Collectively,SMANTIS interacts with RUNX1 and attenuates monocyte adhesion,which might limit monocyte vascular egress. Subject terms: Long non-coding RNAs,Transcription View Publication

Intervertebral disc disease (IDD) is a debilitating spine condition that can be caused by intervertebral disc (IVD) damage which progresses towards IVD degeneration and dysfunction. Recently,human pluripotent stem cells (hPSCs) were recognized as a valuable resource for cell-based regenerative medicine in skeletal diseases. Therefore,adult somatic cells reprogrammed into human induced pluripotent stem cells (hiPSCs) represent an attractive cell source for the derivation of notochordal-like cells (NCs) as a first step towards the development of a regenerative therapy for IDD. Utilizing a differentiation method involving treatment with a four-factor cocktail targeting the BMP,FGF,retinoic acid,and Wnt signaling pathways,we differentiate CRISPR/Cas9-generated mCherry-reporter knock-in hiPSCs into notochordal-like cells. Comprehensive analysis of transcriptomic changes throughout the differentiation process identified regulation of histone methylation as a pivotal driver facilitating the differentiation of hiPSCs into notochordal-like cells. We further provide evidence that specific inhibition of histone demethylases KDM2A and KDM7A/B enhanced the lineage commitment of hiPSCs towards notochordal-like cells. Our results suggest that inhibition of KDMs could be leveraged to alter the epigenetic landscape of hiPSCs to control notochord-specific gene expression. Thus,our study highlights the importance of epigenetic regulators in stem cell-based regenerative approaches for the treatment of disc degeneration. View Publication

Although bone marrow-derived cells with high aldehyde dehydrogenase activity (ALDH br ) have shown therapeutic potential against various diseases in animal studies,clinical trials have failed to show concurrent findings. We aimed to clarify the optimal conditions for the efficacy of ALDH br cells by using a murine bleomycin-induced pulmonary fibrosis model. We intravenously transferred male or female donor C57BL/6 mice-derived ALDH br cells into recipient C57BL/6 mice under various conditions,and used mCherry-expressing mice as a donor to trace the transferred ALDH br cells. Pulmonary fibrosis improved significantly when (1) female-derived,not male-derived,and (2) lineage (Lin)-negative,not lineage-positive,ALDH br cells were transferred during the (3) fibrotic,not inflammatory,phase. Consistent with the RNA-sequencing results,female-derived Lin − /ALDH br cells were more resistant to oxidative stress than male-derived cells in vitro,and transferred female-derived Lin − /ALDH br cells were more viable than male-derived cells in the fibrotic lung. The mechanism underlying the antifibrotic effects of Lin − /ALDH br cells was strongly associated with reduction of oxidative stress. Our results indicated that Lin − /ALDH br cell therapy could ameliorate pulmonary fibrosis by reducing oxidative stress and suggested that their efficacy was mediated by sex-related differences. Thus,sex-awareness strategies may be important for clinical application of bone marrow ALDH br cells as a therapeutic tool. The online version contains supplementary material available at 10.1186/s13287-024-03933-8. View Publication

Hematopoietic stem cells (HSCs) react to various stress conditions. However,it is unclear whether and how HSCs respond to severe anemia. Here,we demonstrate that upon induction of acute anemia,HSCs rapidly proliferate and enhance their erythroid differentiation potential. In severe anemia,lipoprotein profiles largely change and the concentration of ApoE increases. In HSCs,transcription levels of lipid metabolism-related genes,such as very low-density lipoprotein receptor ( Vldlr ),are upregulated. Stimulation of HSCs with ApoE enhances their erythroid potential,whereas HSCs in Apoe knockout mice do not respond to anemia induction. Vldlr high HSCs show higher erythroid potential,which is enhanced after acute anemia induction. Vldlr high HSCs are epigenetically distinct because of their low chromatin accessibility,and more chromatin regions are closed upon acute anemia induction. Chromatin regions closed upon acute anemia induction are mainly binding sites of Erg. Inhibition of Erg enhanced the erythroid differentiation potential of HSCs. Our findings indicate that lipoprotein metabolism plays an important role in HSC regulation under severe anemic conditions. Subject terms: Haematopoietic stem cells,Fat metabolism,Chromatin,Anaemia View Publication