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p18 INK4 C (CDKN2C, encoded by p18 INK4c or Cdkn2c ) is an early G1-phase cyclin-dependent kinase inhibitor protein. Previous studies demonstrated enhanced self-renewal capacity of hematopoietic stem cells (HSCs) in p18 −/− mice compared to wild-type (WT) mice. Given the critical role of bone marrow niche cells-particularly mesenchymal stromal cells (MSCs)-in hematopoiesis, this study investigated the functional alterations of p18 −/− MSCs and their impact on hematopoietic support. Bone marrow derived MSCs were isolated from p18 −/− and WT mice. Their proliferation and differentiation capacities were assessed, followed by evaluation of hematopoietic support using cobblestone area-forming cell assay and long-term culture-initiating cell assay. RNA sequencing was performed to analyze the transcriptional profile of p18 −/− MSCs, with a focus on differentially expressed genes (DEGs). Key pathways associated with hematopoietic support were identified using Ingenuity Pathway Analysis. A candidate protein was quantified by ELISA, and its functional role in hematopoietic support was validated via a modified coculture system. p18 −/− MSCs displayed an increased proliferation rate, preferential differentiation toward osteogenesis over adipogenesis, and enhanced hematopoietic support. RNA sequencing analysis identified 137 DEGs, with secreted phosphoprotein 1 ( Spp1 , encoding osteopontin, Opn) being significantly upregulated in p18 −/− MSCs. Elevated Opn levels were confirmed in both bone marrow and MSC-conditioned media from p18 −/− mice. Functional validation further demonstrated that Opn enhanced the hematopoietic supportive capacity of MSCs in vitro. p18 deficiency promotes osteogenic differentiation and enhances the hematopoietic supportive function of MSCs, likely mediated by Opn upregulation. These findings suggest a potential therapeutic strategy for improving bone regeneration and HSC expansion. The online version contains supplementary material available at 10.1186/s13287-025-04402-6.

Nanoplastics (NPs) are emerging environmental pollutants that pose growing concerns due to their potential health risks. However, the effects of inhaled NP exposure during pregnancy on fetal brain development remain poorly understood. In this study, we investigated the impact of maternal exposure to polypropylene nanoplastics (PP-NPs) on fetal brain development and neurobehavioral outcomes in a mouse model and further explored its mechanism in human cerebral organoids. Maternal exposure to PP-NPs significantly impaired neuronal differentiation and proliferation in the fetal cortex. Neurobehavioral assessments revealed significant deficits in offspring following maternal exposure, including impaired spatial memory, reduced motor coordination, and heightened anxiety-like behavior. Furthermore, human brain organoids exposed to PP-NPs exhibited reduced growth and neuronal differentiation, with significant downregulation of key neuronal markers such as TUJ1, MAP2, and PAX6. Transcriptomic analysis identified alterations in gene expression, particularly in neuroactive ligand-receptor interaction pathway. Molecular docking and fluorescence co-localization analysis further suggested CYSLTR1 and PTH1R as key molecular targets of PP-NPs. These findings provide novel insights into the toxicological effects of NPs on the developing brain and emphasize the need for preventive measures to protect fetal neurodevelopment during pregnancy. The online version contains supplementary material available at 10.1186/s12951-025-03561-1.

Simvastatin, a 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitor, was used in cardiovascular diseases and could decrease low-density lipoprotein cholesterol, and may have a repurposed role in cancer therapy. However, the effects of simvastatin on endometrial cancer remain controversial. We aimed to elucidate the role and mechanisms of simvastatin in regulating previously identified endometrial cancer-associated mesenchymal stem cells (EmCaMSCs)-mediated immunosuppressive effects and anti-tumor progression. Coculture of EmCaMSCs and peripheral blood mononuclear cells (PBMC) was used to assay the population of CD8 + T cells, natural killer (NK) cells, and cytotoxicity of NK cells. The mechanisms were elucidated by applying recombinant proteins and inhibitors of candidate proteins, transforming growth factor-beta 2 (TGF-β2). Finally, the humanized mouse model was generated to study the effects of simvastatin-mediated immunotherapy in treating endometrial cancer. The protein expressions of TGF-β2, CD56, CD8, and PD-L1 in xenograft tumors were analyzed by Western blot or immunohistochemistry assay. In this study, simvastatin inhibited the proliferation of endometrial cancer cells (HEC-1 A and RL95-2) and EmCaMSCs, and the half-maximal inhibitory concentration (IC50) values of EmCaMSCs were much higher. Simvastatin rescued the proliferation and the population of CD8 + T cells and natural killer (NK) cells from PBMC coculturing with EmCaMSC. Simvastatin treatment reduced the expression of TGF-β2 in EmCaMSCs at both the gene and protein levels. TGF-β2 activated the downstream SMAD2/3 signaling, and their inhibition by simvastatin could enhance the cytotoxicity of NK cells against endometrial cancer cells in vitro. Additionally, a combination of simvastatin and NK cell therapy inhibited xenograft growth, potentially by reducing TGF-β2 expression. In conclusion, simvastatin could rescue the population of CD8 + T cells and NK cells from PBMC cocultured with EmCaMSCs. Furthermore, simvastatin could enhance the cytotoxicity of NK cells in vitro and inhibit tumor growth in vivo in a humanized mouse model. These results suggested that simvastatin may be considered as a repurposed and combination drug for treating endometrial cancer. The online version contains supplementary material available at 10.1038/s41598-025-08686-9.

The impact of exogenous stressors, such as cancer chemotherapies, on the genomic integrity and clonal dynamics of normal hematopoiesis is not well defined. We conducted whole-genome sequencing on 1,276 single-cell-derived hematopoietic stem and progenitor cell (HSPC) colonies from ten patients with multiple myeloma treated with chemotherapies and six normal donors. Melphalan treatment significantly increased the mutational burden, producing a distinctive mutation signature, whereas other chemotherapeutic agents had minimal effects. Consequently, the clonal diversity and architecture of post-treatment HSPCs resemble those observed in normal elderly individuals, particularly through the progression of oligoclonal hematopoiesis, thereby suggesting that chemotherapy accelerates clonal aging. Integrated phylogenetic analysis of matched therapy-related myeloid neoplasm samples traced their clonal origin to a single-HSPC clone among multiple competing clones, supporting a model of oligoclonal to monoclonal transformation. These findings underscore the need for further systematic research on the long-term hematological consequences of cancer chemotherapy. Subject terms: Genetics research, Acute myeloid leukaemia

Kidney organoids derived from human induced pluripotent stem cells lack a proper vasculature, hampering their applicability. Transplantation prevents the loss of organoid endothelial cells (ECs) observed in vitro, and promotes vascularization. In this study, we transplanted kidney organoids in chicken embryos and deployed single-cell RNA sequencing of ~12,000 organoid ECs to delineate their molecular landscape and identify key changes associated with transplantation. Transplantation significantly altered EC phenotypic composition. Consistent with angiogenesis, proliferating EC populations expanded 8 days after transplantation. Importantly, ECs underwent a major vein-to-arterial phenotypic shift. One of the transplantation-specific arterial EC populations, characterized by laminar shear stress response and Notch signalling, showed a similar transcriptome as human fetal kidney arterial/afferent arteriolar ECs. Consistently, transplantation-induced transcriptional changes involved proangiogenic and arteriogenic SOX7 transcription factor upregulation and regulon enrichment. These findings point to blood flow and candidate transcription factors such as SOX7 as possible targets to enhance kidney organoid vascularization. Subject terms: Nephrons, Transcriptomics, Angiogenesis, Angiogenesis, Stem cells, Stem-cell differentiation

Prenatal nicotine exposure impairs fetal cortical grey matter volume, but the precise cellular mechanisms remain poorly understood. This study elucidates the role of nicotinic acetylcholine receptors (nAChRs) in progenitor cells and radial glia (RG) during human cortical development. We identify two nAChR subunits—CHRNA7 and the human-specific CHRFAM7A—expressed in SOX2+ progenitors and neurons, with CHRFAM7A particularly enriched along RG endfeet. nAChR activation in organotypic slices and dissociated cultures increases RG proliferation while decreasing neuronal differentiation, whereas nAChR knockdown reduces RG and increases neurons. Single-cell RNA sequencing reveals that nicotine exposure downregulates key genes in excitatory neurons (ENs), with CHRNA7 or CHRFAM7A selectively modulating these changes, suggesting an evolutionary divergence in regulatory pathways. Furthermore, we identify YAP1 as a critical downstream effector of nAChR signaling, and inhibiting YAP1 reverses nicotine-induced phenotypic alterations in oRG cells, highlighting its role in nicotine-induced neurodevelopmental pathophysiology. Subject terms: Neuronal development, Developmental neurogenesis, Neural stem cells

The hematopoietic stem cell and multipotent progenitor (HSC/MPP) pool dynamically responds to stress to adapt blood output to specific physiological demands. In β-thalassemia (Bthal), severe anemia and ineffective erythropoiesis generate expansion of erythroid precursors and a chronic stress status in the bone marrow (BM) microenvironment. However, the response to the BM altered status at the level of the HSC/MPP compartment in terms of lineage commitment has not been investigated. Bulk and single-cell RNA-sequencing reveal that Bthal HSCs/MPPs are expanded and activated with enhanced priming along the whole Ery differentiation trajectory. Consistently, HSC/MPP showed an altered TGFβ expression and autophagy transcriptional signatures along with a declined dormancy state. We discovered that the altered TGFβ signaling fosters the Ery potential of HSCs by reducing their autophagic levels, and in vivo stimulation of autophagy is sufficient to rescue the imbalance of the HSC compartment. Our findings identify the interplay between TGFβ and HSC autophagy as a key driver in the context of non-malignant hematopoiesis. Subject terms: Haematopoietic stem cells, Haematological diseases, Autophagy

VEXAS syndrome is a clonal hematopoietic disorder characterized by hyperinflammation, bone marrow failure, and high mortality. The molecular hallmark of VEXAS is somatic mutations at methionine 41 (M41) in the E1 ubiquitin enzyme, UBA1. These mutations induce a protein isoform switch, but the mechanisms underlying disease pathogenesis remain unclear. Here, we developed a human cell model of VEXAS syndrome by engineering the male monocytic THP1 cell line to express the common UBA1 M41V mutation. We found that mutant UBA1 M41V cells exhibit aberrant UBA1 isoform expression, increased vacuolization, and upregulation of the unfolded protein response, recapitulating key features of VEXAS. Moreover, proteomic analyses revealed dysregulated ubiquitination and proteotoxic stress in UBA1 M41V cells, with alterations in inflammatory and stress-response pathways. Functional studies demonstrated that UBA1 M41V cells were highly sensitive to genetic or pharmacological inhibition of E1 ubiquitin enzymes. Treatment with the E1 enzyme inhibitor TAK-243 preferentially suppressed colony formation of UBA1 M41V cells as compared to WT cells. Moreover, UBA1 M41V cells exhibited greater sensitivity to TAK-243 in competition assays and showed increased apoptosis. Interestingly, TAK-243 preferentially inhibited UBA6 activity over UBA1, suggesting that UBA6 may compensate for UBA1 dysfunction in UBA1 M41V cells. Targeting UBA6 using shRNA or the UBA6-specific inhibitor phytic acid further revealed an acquired dependency on UBA6 in UBA1 M41V cells. Phytic acid selectively impaired growth and colony formation in UBA1 M41V cells while sparing WT cells, highlighting a potential therapeutic vulnerability. Together, these findings establish a novel human model of VEXAS syndrome, identify key roles for UBA1 and UBA6 in disease pathogenesis, and demonstrate that UBA6 inhibition represents a promising therapeutic strategy for selectively targeting UBA1 mutant clones. Subject terms: Haematological cancer, Cell signalling

Aplastic anemia (AA) is a life-threatening bone marrow (BM) failure syndrome characterized by pancytopenia. Recent studies revealed that dysfunctional endothelial progenitor cells (EPCs), critical components of the BM microenvironment, are involved in hematopoietic-dysfunction-related diseases, including AA. However, the mechanism underlying EPC damage in AA remains unknown. Here we find that transforming growth factor-β (TGF-β) signaling is hyperactive in dysfunctional AA EPCs with impaired hematopoietic support and immune regulatory ability, and TGF-β inhibition promotes hematopoiesis and immune rebalance by repairing dysfunctional EPCs. Through impaired EPC and AA murine models, we validated that TGF-β inhibition restores EPC dysfunction to improve hematopoiesis and immune status in vitro and in vivo. RNA sequencing and real-time quantitative polymerase chain reaction provided further validation. These results indicate that dysfunctional BM EPCs with hyperactive TGF-β signaling are involved in AA. TGF-β inhibition promotes multilineage hematopoiesis recovery and immune balance by repairing dysfunctional EPCs, providing a potential therapeutic strategy for AA. Subject terms: Experimental models of disease, Translational research

Cancer stem cells (CSCs) are responsible for colorectal cancer (CRC) chemoresistance, recurrence, and metastasis. Therefore, identifying molecular stemness targets that are involved in tumor growth is crucial for effective treatment. Here, we performed an extensive in vitro and in vivo molecular and functional characterization, revealing the pivotal role of the lysine methyltransferase SET and MYND Domain Containing 3 (SMYD3) in colorectal cancer stem cell (CRC-SC) biology. Specifically, we showed that SMYD3 interacts with and methylates c-MYC at K158 and K163, thereby modulating its transcriptional activity, which is implicated in stemness and colorectal malignancy. Our in vitro data suggest that SMYD3 pharmacological inhibition or its stable genetic ablation affects the clonogenic and self-renewal potential of patient-derived CRC-SCs and organoids by altering their molecular signature. Moreover, we found that SMYD3 stable knock-out or pharmacological inhibition drastically reduces CRC tumorigenicity in vivo and CRC-SC metastatic potential. Overall, our findings identify SMYD3 as a promising therapeutic target acting directly on c-MYC, with potential implications for countering CRC-SC proliferation and metastatic dissemination. Subject terms: Gastrointestinal cancer, Cancer stem cells

Several years after the COVID‐19 pandemic, the impact of SARS‐CoV‐2 on immunity and the potential protective role of Bacillus Calmette–Guérin (BCG) vaccination through trained immunity remain a subject of investigation. This study aimed to determine the long‐term impact of SARS‐CoV‐2 on immune cells and the association between BCG vaccination, latent infections and COVID‐19 severity and sepsis progression. We conducted a prospective analysis of patients who recovered from mild/severe/critical COVID‐19 ( n = 97, 3–17 months after COVID‐19) and sepsis patients ( n = 64). First, we assessed the impact of COVID‐19 and its severity on immune cell frequencies and expression of functional markers. Further, we analysed plasma titres of anti‐ Toxoplasma gondii /cytomegalovirus/BCG antibodies and their association with COVID‐19 severity and sepsis outcome. To examine monocyte responses to secondary challenge, monocytes isolated from COVID‐19 convalescent patients, BCG vaccinated and unvaccinated volunteers were stimulated with SARS‐CoV‐2 and LPS. Post‐COVID‐19 patients showed immune dysregulation regardless of disease severity characterised by altered expression of activation and functional markers in myeloid (CD39, CD64, CD85d, CD11b) and lymphoid cells (CD39, CD57, TIGIT). Strikingly, post‐critical COVID‐19 patients showed elevated expression of CD57 in CD8 + T cells compared to other severity groups. A trend toward improved outcomes in BCG‐seropositive COVID‐19/sepsis patients was observed, although this may be confounded by age differences between groups. In contrast, the monocyte response to stimulation appeared unaffected by COVID‐19 severity. These findings highlight the long‐term alterations of immune cells in post‐COVID‐19 patients, emphasising the substantial impact of COVID‐19 on immune function.

Ageing-dependent low-grade inflammation is a hallmark of central nervous system (CNS) diseases. Vascular and immune abnormalities are implicated in the progression of gliomas and occur in the early stages of Alzheimer's disease (AD); however, the mechanisms by which these alterations manifest in the brain parenchyma remain unclear. Using RNAseq, scRNAseq, bioinformatics tools and a cohort of patients with glioma and Alzheimer's disease for validation of results, we have established an analysis of blood–brain barrier (BBB) dysfunction and neuron loss. A mouse model for glioblastoma pathology was also used that reversed BBB disruption and neuron loss, with the incorporation of the IDH mutation. Finally, we established a characterization of the relevant immune populations with an IHC analysis and transcriptional profile. In this study, molecular analyses of the brain ecosystem revealed that blood–brain barrier dysfunction and neuronal synapse integrity exhibit significant threshold-dependent changes that correlate directly and inversely, respectively, with brain ageing (significant changes at 57 years) and the progression of AD and gliomas (survival of 1525 vs 4084 days for patients with High vs Low BBB dysfunction). Using human samples and mouse models, we identified immunoageing processes characterized by an imbalance between pro-inflammatory and anti-inflammatory signals. This dysregulation promotes the extravasation of monocyte-derived macrophages (85% increase of cells), particularly those with a suppressive phenotype, alongside an increase in inflammatory cytokine levels. Notably, our data show that vascular normalization in a glioma model can reverse neuronal loss and attenuate the aggressiveness of the tumours. Finally, tumour development can be prevented by reactivating the ageing immune system. We propose that the ageing brain represents a common, BBB dysfunction-associated process driving chronic inflammation. This inflammation is regulated by TREM2+/TIM3+ suppressive myeloid cells, which play a central role in disease progression. Our findings suggest that targeting these pathways could offer therapeutic strategies to mitigate CNS pathologies linked to ageing, characterized by toxic neuroinflammation and myeloid dysfunction. This study was funded by ISCIII and co-funded by the European Union.