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BackgroundDiabetic peripheral neuropathy (DPN) is one of the most prevalent and debilitating complications of diabetes,marked by chronic neuroinflammation,immune dysregulation,and progressive neuronal degeneration. Current treatments offer limited efficacy,largely focusing on symptomatic relief rather than addressing the underlying disease mechanisms. There is a critical need for disease-modifying therapies that target the molecular basis of DPN.ResultsIn this study,we developed a novel targeted nanotherapeutic system—ZH-1c-EVs@SIN—composed of neural stem cell-derived extracellular vesicles (NSC-EVs) modified with the ZH-1c aptamer and loaded with the anti-inflammatory compound sinomenine (SIN). This system was specifically designed to target microglia and inhibit the WNT5a/TRPV1 signaling pathway. Transcriptomic profiling of microglia revealed key gene networks implicated in DPN pathology and responsive to SIN treatment. Functional assays demonstrated that ZH-1c-EVs@SIN facilitated a shift in microglial phenotype from pro-inflammatory M1 to anti-inflammatory M2,significantly reduced inflammatory cytokine expression,and restored levels of neuronal regulatory proteins. Nanoparticle tracking analysis and transmission electron microscopy confirmed optimal vesicle size and morphology,while fluorescence imaging showed efficient uptake by microglia. In vivo studies in a murine model of DPN revealed marked improvements in pain-related behavior and histopathological signs of nerve damage.ConclusionZH-1c-EVs@SIN represents a promising therapeutic strategy for DPN,offering targeted immunomodulation and enhanced neural repair via regulation of the WNT5a/TRPV1 signaling axis. This nano-delivery platform introduces a novel and precise approach to intervening in diabetic neuropathy and may be applicable to other neuroinflammatory conditions.Graphical abstractMechanism of ZH-1c-EVs@SIN Mediating the WNT5a/TRPV1 Pathway to Improve Immune-Inflammatory Homeostasis in the Treatment of DPN in Mice. Supplementary InformationThe online version contains supplementary material available at 10.1186/s12951-025-03678-3. View Publication

Embryonic stem cells (ESC) are pluripotent,with the potential to differentiate into multiple cell types,making them a valuable tool for regenerative medicine and disease therapy. However,common culture methods face challenges,including strict operating procedures and high costs. Currently,Leukemia inhibitory factor (LIF),an indispensable bioactive protein for ESC culture,is typically applied to maintain self-renewal and pluripotency,but its instability and high cost limit its effectiveness in stable culture conditions. Hence,we have developed an innovative strategy using a soluble nanomaterial,metal-organic polyhedra (MOPs),to effectively maintain the self-renewal and pluripotency of ESC. The selected amino-modified vanadium-based MOP not only exhibits excellent biocompatibility and high stability but also possesses similar or even superior biological functions compared to commercial LIF. Due to the precise structure of MOPs,the active site responsible for maintaining ESC pluripotency has been identified and regulated at the molecular level. The new ESC culture method significantly reduces costs,simplifies preparation,and enhances the practicality of biopharmaceutical preparation and storage. This represents the first case of using MOPs to maintain self-renewal of ECS,opening an avenue for introducing advanced materials into the development of innovative ESC culture methods. Subject terms: Biomaterials - cells,Chemical biology View Publication

Craniosynostosis is a multigenic congenital condition in which one or more calvarial sutures have prematurely fused during the development of the fetus. Pathogenic variants in FGFR2 are associated with the development of syndromic craniosynostosis,such as Crouzon,Apert and Pfeifer syndromes. Investigation of FGFR2 -linked craniosynostosis is hindered by the lack of appropriate in vitro models. Patient-derived human induced pluripotent stem cell (hiPSC) in vitro disease models provide the opportunity to investigate the disease,identify molecular targets for pharmaceutical treatments,and enable the generation of autologous pluripotent stem cell catalogues. Here,we report three patient-derived hiPSC lines carrying the C342Y,S252W or E565G FGFR2 pathogenic variant. The patient hiPSC lines express characteristic pluripotency markers and display distinct phosphorylation profiles under unstimulated conditions. FGFR2 C342Y showed autophosphorylation in the absence of bFGF ligand,although downstream docking proteins PLCγ and FRS2α were not phosphorylated. FGFR2 S252W and FGFR2 E565G hiPSCs showed increased phosphorylation of docking proteins PLCγ and FRS2α,whereas FGFR2 was not phosphorylated. These patient hiPSC lines provide molecular and cellular options to investigate FGFR2 -linked craniosynostosis in the patient-specific genomic context and develop therapeutic modalities. View Publication

Transposable elements (TEs) are genomic elements present in multiple copies in mammalian genomes. TEs were thought to have little functional relevance but recent studies report roles in biological processes,including embryonic development. To investigate the expression dynamics of TEs during human early development,we generated long-read sequence data from human pluripotent stem cells (hPSCs) in vitro differentiated to endoderm,mesoderm,and ectoderm lineages to construct lineage-specific transcriptome assemblies and accurately place TE sequences. Our analysis reveals that specific TE superfamilies exhibit distinct expression patterns. Notably,we observed TE switching,where the same family of TE is expressed in multiple cell types,but originates from different transcripts. Interestingly,TE-containing transcripts exhibit distinct levels of transcript stability and subcellular localization. Moreover,TE-containing transcripts increasingly associate with chromatin in germ layer cells compared to hPSCs. This study suggests that TEs contribute to human embryonic development through dynamic chromatin interactions. Transposable elements are genetic parasites that have colonised genomes and they express as parts of coding and noncoding RNAs. Here,the authors explore how they are expressed in transcripts in normal human development,and how they alter transcript dynamics. View Publication

T-cell exhaustion contributes to immunotherapy failure in chronic lymphocytic leukemia (CLL). Here,we analyze T cells from CLL patients’ blood,bone marrow,and lymph nodes,as well as from a CLL mouse model,using single-cell RNA sequencing,mass cytometry,and tissue imaging. T cells in CLL lymph nodes show the most distinct profiles,with accumulation of regulatory T cells and CD8+ T cells in various exhaustion states,including precursor (TPEX) and terminally exhausted (TEX) cells. Integration of T-cell receptor sequencing data and use of the predicTCR classifier suggest an enrichment of CLL-reactive T cells in lymph nodes. Interactome studies reveal potential immunotherapy targets,notably galectin-9,a TIM3 ligand. Inhibiting galectin-9 in mice reduces disease progression and TIM3+ T cells. Galectin-9 expression also correlates with worse survival in CLL and other cancers,suggesting its role in immune evasion and potential as a therapeutic target. Multi-omics can be used to characterise tumour and immune cell populations. Here the authors use multi-omics to characterise CLL blood and tissue samples and use prediction models for CLL TCR specificity and implicate interactions between galectin-9 and TIM3 as involved in CLL immune escape and propose galectin-9 as a possible immunotherapy target. View Publication

AbstractObjectivesTumour necrosis factor inhibitors (TNFi) are widely used and effective as treatment for immune-mediated inflammatory diseases (IMIDs). However,TNFi therapy causes a faster waning of antibody responses following vaccination. The underlying cause by which TNFi affect humoral immunity remains to be elucidated. The formation of long-lasting,high-affinity antibodies after vaccination results from germinal centre (GC)-derived,T cell-dependent B-cell responses. Therefore,this study investigated how TNFi affect the formation and maintenance of antigen-specific B- and CD4+ T-cell responses following SARS-CoV-2 mRNA vaccination.MethodsSARS-CoV-2 spike-specific B-cell responses were characterised using spectral flow cytometry. Spike-specific CD4+ T cells were measured using an activation-induced marker assay. 15 patients with inflammatory bowel disease (IBD) treated with TNFi were compared with 9 IBD patients without systemic immunosuppression and 10 healthy controls.ResultsSpike-specific CD4+T-cell frequency and phenotype,including T follicular helper cells,were not affected by TNFi. Total spike-specific B-cell frequencies were reduced in TNFi-treated patients. Deep phenotyping revealed lower IgG+memory B-cell frequencies in TNFi-treated patients 3–6 months after vaccination. These data were confirmed in TNFi-treated rheumatoid arthritis patients. Interestingly,already at day 7 after the second vaccination,TNFi therapy reduced the induction of class-switched CD11c- CD71+activated B cells,which are believed to be GC-derived. Conversely,CD11c+B cells,associated with extrafollicular B-cell responses,were not affected by TNFi therapy.ConclusionsThese data suggest that TNFi therapy affects the differentiation of GC-derived B cells,which may explain its effect on humoral immune responses. View Publication

Induction of ferroptosis,an iron-dependent form of regulated cell death,holds promise as a strategy to overcome tumor resistance to conventional therapies and enhance immunotherapy responses. However,while the susceptibility of tumor cells to ferroptosis is extensively studied,limited data exists on the vulnerability of immune cells to disturbed iron balance and lipid peroxidation. Here,we found that T-cell stimulation rewires iron and redox homeostasis and by increasing levels of reactive oxygen species and labile iron promotes lipid peroxidation and T-cells’ ferroptosis. Upon stimulation,we detected changes in the balance of ferroptosis-suppressive proteins,including decrease of GPX4. Subsequently,we identified GPX4 as a master regulator orchestrating T/CAR-T-cells’ sensitivity to ferroptosis and observed that GPX4 inhibitors impair CAR-T cells’ antitumor functions. Our study demonstrated differential GPX4 expression and diverse susceptibility to ferroptosis between CD4⁺ and CD8⁺ T cells. Among analyzed subsets of naïve,central memory (CM),effector memory (EM),and terminally differentiated effector memory (TEMRA),CD8⁺ EM and CD8⁺ TEMRA cells exhibited the highest sensitivity to ferroptosis. We also showed that ferroptosis limited the anti-tumor efficacy of CAR-T cells,while ferroptosis inhibition improved their therapeutic effect,both in vitro and in vivo. Our findings are not only important to understand vulnerabilities of CAR-T cells but may also hold particular significance for their therapeutic development. In this context,future anticancer therapies should be carefully designed to selectively induce the ferroptosis of tumor cells without impeding cytotoxic cells’ antitumor efficacy. Additionally,we postulate that promoting less differentiated phenotype of CAR-T cells should be exploited therapeutically to create CAR-T products characterized by decreased sensitivity to ferroptosis within tumor microenvironment.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00262-025-04133-w. View Publication

AbstractBackgroundAdoptive transfer of chimeric antigen receptor (CAR)-expressing natural killer (NK) cells has demonstrated success against hematological malignancies. Efficacy against solid tumors has been limited by poor NK cell survival and function in the suppressive tumor microenvironment (TME). To enhance efficacy against solid tumors,stimulatory cytokines have been incorporated into CAR-NK cell therapeutic approaches. However,current cytokine strategies have limitations,including systemic toxicities,exogenous dependencies,and unwanted TME bystander effects. Here,we aimed to overcome these limitations by modifying CAR-NK cells to express a constitutively active interleukin (IL)-7 receptor,termed C7R,capable of providing intrinsic CAR-NK cell activation that does not rely on or produce exogenous signals nor activate bystander cells.MethodsWe examined persistence,antitumor function,and transcriptional profiles of CAR-NK cells coexpressing C7R in a novel tumor immune microenvironment (TiME) co-culture system and against hematologic and solid tumor xenografts in vivo.ResultsPeripheral blood NK cells expressing a CAR directed against the solid tumor antigen GD2 and modified with C7R demonstrated enhanced tumor killing and persistence in vitro compared with CAR-NK cells without cytokine support and similar functions to CAR-NK cells supplemented with recombinant IL-15. C7R.CAR-NK cells exhibited enhanced survival and proliferation within neuroblastoma TiME xenografts in vivo but produced poor long-term tumor control compared with CAR-NK cells supplemented with IL-15. Similar results were seen using C7R-expressing CD19.CAR-NK cells against CD19+leukemia xenografts. Gene expression analysis revealed that chronic signaling via C7R induced a transcriptional signature consistent with intratumor stressed NK cells with blunted effector function. We identified gene candidates associated with chronic cytokine-stressed NK cells that could be targeted to reduce CAR-NK cell stress within the solid TME.ConclusionC7R promoted CAR-NK cell survival in hostile TMEs independent of exogenous signals but resulted in poor antitumor function in vivo. Our data reveals the detrimental role of continuous IL-7 signaling in CAR-NK cells and provides insights into proper application of cytokine signals when attempting to enhance CAR-NK cell antitumor activity. View Publication

ABSTRACTCD8+ T cells are central to targeting and eliminating cancer cells. Their function is critically supported by type 1 conventional dendritic cells (cDC1s),which both prime antigen‐specific CD8+ T cells in tumour‐draining lymph nodes (tdLNs) and sustain primed CD8+ T cells within tumours. Despite their importance,the spatiotemporal organisation of cDC1s within tumours and their diverse functional roles remain poorly understood. Here,we use scRNAseq and unbiased spatial analysis to construct a detailed map of cDC1 states and distribution within immunogenic mouse tumours during CD8+ T‐cell‐mediated rejection. We reveal two distinct cDC1 activation states characterised by differential expression of genes linked to anti‐tumour immunity,including Cxcl9 and Il12b. Strikingly,Il12b‐expressing cDC1s are CCR7+ and enriched at tumour borders,where they closely associate with stem‐like TCF1+ CD8+ T cells. In contrast,CCR7– Cxcl9‐expressing cDC1s are preferentially found within the tumour parenchyma alongside effector CD8+ T cells. Analysis of a published dataset of human tumours similarly reveals a spatial association between CCR7+ cDC1 and stem‐like TCF1+ CD8+ T cells. These findings uncover a highly spatially coordinated interaction between cDC1s and CD8+ T cells within tumours,shedding light on the intricate cellular dynamics that underpin effective anti‐tumour immunity. Using scRNAseq and spatial analysis,we analyse cDC1 states and spatial distribution in tumours during immune‐mediated rejection. We identify two cDC1 activation states,each occupying different regions and associated with distinct CD8+ T cell populations. This reveals the spatial organisation of cDC1 states that may be key to anti‐tumour immunity. View Publication

IntroductionT cells are major components of the immune system. Their activation requires interaction between the T cell receptor and co-stimulatory molecules,crucial during infection,inflammation,and allogeneic rejection. Monomeric CRP (mCRP) is a known modulator of inflammation and particularly the innate immune response,however its interaction with T cells as part of the adaptive immune response remains unclear.MethodsPeripheral blood mononuclear cells (PBMC) and T cells were isolated. Flow cytometric analysis was conducted to evaluate Fcγ receptor CD16 expression on T cells,the binding of CRP to T cells,and its impact on proliferation and apoptosis. T cell activation was assessed after 1,2,3,5 and 7 days by assessing CD69 and CD25 expression,and under various conditions including coculture with monocytes and several inhibitory factors.ResultsT cells express CD16 that binds mCRP in a concentration-dependent manner,and particularly on activated T cells. While mCRP reduces apoptosis and accelerates proliferation in T cells,it does not independently activate them. However,activation of monocytes by mCRP leads to T cell activation,indicating a direct cell to cell interaction during CRP-induced activation. This effect could be alleviated by inhibition of the CD80/CD28 pathway.ConclusionCRP does not activate T Cells directly but via PI3-kinase-dependent activation of monocytes and subsequent CD80/CD28 cell to cell contact. The findings suggest the effects of CRP on T cells depend on their environment and the presence of other proinflammatory agents. View Publication