A preeminent goal of virology is to discover cellular genes that mediate virus entry. Genome-wide loss-of-function screens can illuminate single genes necessary for virus entry, but are stymied by genetic redundancy. Here we report a genome-wide CRISPR activation screening strategy to discover single genes that are sufficient for viral entry into normally-uninfectable cells. Sequential rounds of viral infection vastly enhanced screening sensitivity. This sequential screening strategy was generalizable to two unrelated viruses-Ebola and rabies viruses-and could broadly accelerate the discovery of viral entry factors.

RNAi is a powerful cellular defense mechanism against genomic invaders that rely on dsRNA intermediates, such as viruses and mobile elements. Due to its specificity and ease of use, RNAi is also widely used as an experimental and therapeutic strategy to reduce levels of specific RNAs. For many systems however, delivery of the silencing agents into each individual cell is a major challenge. A mechanistic understanding of the processes involved in intercellular spread in systems with effective systemic RNAi thus is key to improve applications, and to increase our understanding of this defense mechanism. Remarkably, outside of C. elegans and plants, our molecular understanding of systemic RNAi remains very limited. We here investigated the highly-effective systemic RNAi of the planarian S. mediterranea , which can be triggered by introduction of dsRNA via food or injection and rapidly spreads through the entire body. Notwithstanding its efficiency, we find no evidence of an RNA amplification mechanism or of transgenerational effects as are found in C. elegans , and rather find that planarian RNAi effects are limited in time. We identify the biogenesis factors involved in the planarian RNAi mechanism, and find that these are independent of the miRNA pathway, enabling the separation of the effects from these small RNA pathways. Surprisingly, we find that planarian systemic RNAi relies on active stem cells. Further, we identify Argonaute-siRNA complexes as the mobile agent that effectuates systemic spread of RNAi throughout the tissue. These findings provide new insights into the mechanisms by which small RNAs spread between cells, and by which organisms can extend protection to all their cells upon encounter of a novel invading element. Additionally, our findings may have important implications for the design of effective applications in other systems.

Fasting enhances small intestinal regeneration after radiation but the contribution of the gut microbiome to this process remains uncharacterized. We identify Akkermansia muciniphila ( AKK ) as a key mediator of this response. AKK was enriched in fasted mice and its antibiotic depletion abrogated radioprotection whereas reintroduction restored both organismal survival and intestinal integrity. Fasting elevated propionic acid, consistent with AKK 's metabolic output. AKK -conditioned medium and propionate induced histone H3 acetylation in intestinal stem cell cultures while in vivo fasting induced AKK -dependent H3K27ac and H3K9ac, remodeling promoter-enhancer landscapes in crypt epithelial cells. Epigenetic profiling revealed a rewired core regulatory program enriched for pioneer transcription factors (Foxa, Gata, Klf), architectural organizers (Ctcf, Boris), and lineage-defining and metabolic regulators (Cdx2, Hnf4). This program supports expansion of a population of persister stem cells characterized by open chromatin accessibility at key stem and regenerative-associated loci including Clu , Olfm4 , Lgr5, Ascl2, Lrig1, Sox9, Rnf43, and Axin2. These findings define a fasting-induced microbiome-metabolite-chromatin axis that epigenetically primes highly plastic persister stem cells for rapid regeneration of the intestinal epithelium following radiation-induced injury.

The aim of this study was to study the role of transmission electron microscopy (TEM) in assessment of the phenotype of astrocytes obtained with the directed differentiation technique from induced pluripotent stem cells (iPSCs) from a healthy donor and from a patient with a hereditary form of Parkinson's disease (PD).

The aim of the study was to develop a novel approach to treatment of non-healing wounds by using a portable Biogan bioprinter and an ink based on fibrin-gelatin hydrogel and spheroids derived from mesenchymal stromal cells (MSCs) from adipose tissue in a model of ischemic pig wound.

Decellularized extracellular matrices (dECMs) have gained increasing attention in regenerative dentistry due to their ability to replicate aspects of the native cellular microenvironment while reducing immunogenicity. Dental-derived stem cells exhibit regenerative and immunomodulatory properties, making them promising candidates for tissue repair when combined with biologically derived scaffolds such as dECMs.

Cochlear supporting cells are glia-like cells critical for development, homeostasis, and regeneration. In the neonatal mouse cochlea, inner phalangeal cells (IPhCs), a supporting cell subtype surrounding inner hair cells, are uniquely regenerated by greater epithelial ridge (GER) cells. Here, we used fate-mapping and single-cell transcriptomic analysis to show that GER cells both mitotically and non-mitotically regenerate IPhCs. Mechanisms of regeneration are spatially distinct, with non-mitotic regeneration found throughout the cochlear spiral and mitotic regeneration restricted to the apical and middle turns. By profiling the regenerating cochlea at four time points, we revealed a damage-induced loss of medial-to-lateral specialization of GER cells. Moreover, in silico analysis predicted distinct lineages and associated pathways where mitotic and non-mitotic IPhC progenitors emerge in the lateral GER. Finally, inhibiting proliferation eliminated mitotic IPhC regeneration while sparing non-mitotic regeneration in vitro. Together, our study reveals dual mechanisms and molecular targets governing supporting cell regeneration.

Osteosarcoma is the most prevalent primary bone cancer affecting children and adolescents, characterized by its high invasiveness and the challenges in treatment posed by multidrug resistance. Cancer stem cells significantly contribute to poor clinical outcomes; however, current targeted therapies remain limited in their effectiveness. In this study, we developed a novel dual-responsive nanomedicine designed to target the microenvironment of osteosarcoma cancer stem cells (OCSCs) and osteosarcoma itself, which selectively releases all-trans retinoic acid and paclitaxel, termed mPCDAP. Our findings indicate that mPCDAP rapidly releases all-trans retinoic acid to suppress stemness under conditions of elevated glutathione, subsequently increasing intracellular reactive oxygen species levels in tumor cells. This mechanism further facilitates the release of paclitaxel under conditions of highly reactive oxygen species, thereby inducing apoptosis in osteosarcoma cells. Additionally, mPCDAP was effectively internalized by both osteosarcoma cells and OCSCs, with in vitro and in vivo results demonstrating significant synergistic antitumor effects, promoting apoptosis and markedly reducing tumor stemness. This study presents a novel approach and promising prospects for targeted therapy of osteosarcoma and OCSCs.

In this study, we aimed to construct an injectable hydrogel composed of gelatin methacryloyl/hyaluronic acid (GelMA-HA) and combine it with lyophilized concentrated growth factors (CGF) and dental pulp stem cells (DPSC) to promote the process of periodontal regeneration. We confirmed that bioactive lyophilized CGF can be anchored to hydrogels by Schiff base imine bonding, thereby achieving effective sustained release of various growth factors in the inflammatory microenvironment of periodontal bone destruction. Meanwhile, this hydrogel can provide a 3D cell growth microenvironment to improve the survival rate of dental pulp stem cells. GelMA-HA-CGF-DPSCs exhibit remarkable ability to promote cell proliferation and differentiation as well as excellent biocompatibility. In vitro experiments demonstrated that this system can increase alkaline phosphatase activity, enhance osteogenic mineralization capacity, and significantly upregulate the expression of osteogenesis-related genes and proteins. In the rat alveolar bone defect model, the experimental group significantly facilitated regeneration of periodontal bone. These findings suggest that this injectable composite material has the potential to overcome the limitations of CGF applications and stem cell injections. Specifically, through the chemotactic effects of growth factors and intercellular interactions, endogenous stem cells can also attract endogenous stem cells. Serving as a scaffold structure, it creates an osteogenic microenvironment and provides a space for bone regeneration. Moreover, its injectability and fluidity can achieve minimally invasive periodontal treatment in a noninvasive manner. The findings of this study are expected to provide valuable insights and references for the clinical translational repair treatment of periodontal bone defects caused by periodontitis.

Radical prostatectomy (RP) is a highly effective treatment for localized prostate cancer; unfortunately, post-prostatectomy urinary incontinence (UI) remains a prevalent and distressing complication, significantly diminishing patients' quality of life. Current therapeutic options often provide incomplete continence restoration and may lead to substantial morbidity. This review examines the rapidly advancing field of regenerative medicine, specifically focusing on stem cell and exosome-based therapies as innovative approaches to address post-RP UI. We go deeper into the unique pathophysiology of male post-prostatectomy UI, distinguishing it from other forms of UI, and present the compelling biological rationale for these regenerative interventions. Highlighting advancements from 2014 to 2025, we explore recent preclinical and clinical progress in this domain. Furthermore, we critically assess the persistent challenges crucial for widespread clinical application, including optimizing cell dose and source, ensuring long-term efficacy and safety, and interpreting complex regulatory environments. By bridging the understanding of sex-related differences between females and males in UI and tackling the specific challenges of male post-RP incontinence, this review emphasizes that while promising, the journey from laboratory bench to bedside for these innovative therapies demands rigorous scientific inquiry and collaborative efforts.

Glioblastoma (GBM) is the highly lethal intracranial tumor characterized by low survival rates and high recurrence, partly attributable to the challenges posed by the blood-brain barrier (BBB). To enhance therapeutic efficacy, the Exo-U2-Dox complex was engineered by functionalizing mesenchymal stem cell (MSC)-derived exosomes with the GBM-targeting aptamer U2 and integrating them with doxorubicin (DOX). This complex is designed to augment the sensitivity of GBM to chemo-radiotherapy. Here, it is found that Exo-U2 effectively accumulates in GBM-bearing mice, thereby inhibiting tumor progression. When administered in conjunction with DOX and radiation, Exo-U2-Dox increases DNA damage in GBM cells, and diminishes invasiveness. Mechanistically, Exo-U2 targets and inhibits the autophosphorylation of Epidermal growth factor receptor variant Ⅲ (EGFRvⅢ) in GBM cells, thereby activating the NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) inflammasome-mediated pyroptosis pathway, which leads to increased expression of Gasdermin D (GSDMD) and Cysteine-aspartic acid protease-1 (caspase-1), ultimately suppressing GBM cell proliferation, migration, and invasion. Furthermore, the combination of Exo-U2 with X-ray treatment inhibits the expression of p53-binding protein 1 (53BP1), reduces phosphorylation of the Ataxia-Telangiectasia Mutated/Checkpoint kinase 2 (ATM/Chk2) pathway, resulting in the accumulation of DNA damage. Collectively, these findings underscore the potential of aptamer-functionalized exosomes in conjunction with DOX as a promising strategy for GBM treatment. This approach not only broadens the therapeutic applications of DOX but also provides a novel direction for targeted GBM therapies.

The study of atrial arrhythmias has been hampered by the lack of anatomically relevant human cardiac tissue models and by the inability to perform targeted, functional perturbations in such models.

The clinical success of bone regeneration relies on developing a functional, vascularized bone. Insufficient vascularization of tissue constructs remains a challenge in stem cell-based approaches, leading to poor graft integration and necrosis of newly formed bone. To overcome current challenges, using pre-committed stem cells for endothelial lineages is a novel approach to developing a vascularized tissue construct. Human gingiva-derived mesenchymal stem cells (GMSCs) are a unique cell population that is readily accessible, has a high proliferation rate, and exhibits multipotent differentiation. This study aimed to investigate the in vitro differentiation potential of GMSCs into the endothelial lineage. GMSCs were induced with 0, 10, 50, or 100 ng/mL recombinant vascular endothelial growth factor (VEGF) for 1 week. The relative mRNA expressions of vascular cell adhesion molecule 1, protocadherin 12, VEGF receptor 1 (fms-like tyrosine kinase 1), VEGF receptor 2 (kinase insert domain receptor), and platelet endothelial cell adhesion molecule-1 were measured by quantitative reverse transcriptase polymerase chain reaction. The expression of all endothelial marker mRNAs was significantly upregulated in a dose-dependent manner in GMSCs induced by VEGF, and maximum expression was observed at 50 ng/mL VEGF induction. A Matrigel assay demonstrated the tube-forming ability of pre-differentiated GMSCs. Our findings demonstrated that GMSCs have the potential to differentiate into endothelial progenitor-like cells. Thus, our study identifies potential options for using GMSCs as an autologous or allogeneic stem cell source for craniofacial bone regeneration.

Treatment-resistant schizophrenia (TRS) affects 20%-30% of individuals diagnosed with schizophrenia and is effectively managed with clozapine. However, the molecular and cellular mechanisms that underlie its efficacy remain largely unclear. We previously generated induced pluripotent stem cell (iPSC) lines from a unique pair of monozygotic twins with TRS. One twin responded to clozapine (CLZ-res), while the other did not (CLZ-non-res). Building on our previous study of these twins, we included healthy controls and focused on the early developmental stages of neuronal differentiation. To investigate the phenotypic differences in the developmental stages of neural cells between patient-derived cells, we differentiated each iPSC line-from both patients and healthy individuals-into neural stem cells (NSCs) and subsequently induced the differentiation of NSCs into neurons. Our results demonstrated that NSCs derived from patients' iPSC lines exhibited impaired neuronal differentiation, with a more pronounced reduction in differentiation observed in CLZ-non-res cells than in CLZ-res cells. RNA sequencing analysis revealed significant differences in the expression of genes involved in neuronal development between CLZ-res and CLZ-non-res cells. These findings suggest that the differences in neuronal development may contribute to the variability in clozapine responsiveness. Although this study is limited to a single twin pair, this unique human model provides valuable insights into the molecular and cellular mechanisms underlying differential clozapine responses, offering a promising framework for the development of effective treatments for patients with TRS.

Juvenile systemic sclerosis (jSSc) is a rare, chronic, autoimmune disease in children/adolescents and is associated with significant morbidity, skin thickening/hardening (scleroderma), organ toxicity and sub-optimal therapeutic options. In this report, autologous stem cell transplantation is associated with clinical improvement in a 17-year-old with refractory jSSc.

Bone regeneration under diabetic conditions remains a formidable challenge, predominantly due to persistent oxidative stress, chronic inflammation, and impaired osteogenic potential of stem cells. Nanozymes have been widely employed to modulate oxidative stress and inflammation, and are considered promising candidates for diabetic bone regeneration. However, they have a limited osteogenic capacity, which substantially hinders their direct application in bone repair. Here, a series of osteogenic ion-doped Prussian blue (PB) nanozymes was designed to endow classical antioxidant PB with osteoinductive capacity, among which copper-doped PB (CuPB) exhibits optimal performance. The screened CuPB nanozyme effectively scavenges various types of excessive reactive oxygen species, suppresses inflammation, and simultaneously promotes osteogenic differentiation via activation of the phosphoinositide 3-kinase/protein kinase B (PI3K-Akt) pathway. In type II diabetic rats, both the CuPB nanoparticulate formulation and the CuPB-functionalized three-dimensional scaffold pronouncedly enhance bone regeneration in models of periodontitis-associated alveolar bone loss and cranial defects, respectively. This study establishes an integrated "osteogenic nanozyme" platform that synergistically couples antioxidant and osteoinductive functions.

Fibrinogen is a blood-derived protein involved in coagulation and can make its way into the central nervous system (CNS) following breakdown of the blood-brain barrier. This molecule has been implicated in multiple sclerosis (MS), a disease marked by inflammation and demyelination in the CNS. However, the effect of this molecule has not been studied on human myelinating cells. This study examines how fibrinogen influences human oligodendrocyte (OL) lineage cells at various stages of development. Using induced pluripotent stem cell-derived (iPSC) OL precursors and human primary OLs, we examined the effects of fibrinogen on cell differentiation, viability, and myelination-related function. Here we show the differential effect of fibrinogen, based on OL-lineage stage. While fibrinogen induced aberrant differentiation of early lineage OLs, by inhibiting their maturation and inducing an astrocytic phenotype, on mature OLs fibrinogen was found to promote myelination capacity, as shown by ensheathment assays as well as on the RNA level. These effects were associated with the activation of bone morphogenetic protein (BMP) signaling, both in early and mature OLs. We further found BMP signaling enrichment in OLs to be correlated with the inflammatory activity of an MS lesion and confirmed fibrinogen deposition on OLs in situ. Unlike previous rodent studies, these findings indicate that fibrinogen has a lineage-dependent effect, where it may be inhibitory earlier in the lineage while promoting OL function in later stages. Understanding this dual role will provide insight into remyelination failure in MS and highlights the importance of timing and target in future therapeutic strategies.

Flowering marks a pivotal transition in a plant's life cycle, signalling the shift from vegetative growth to reproductive development. Over the years, extensive research has uncovered key genes and regulatory networks governing this process. Central to this regulation is the Florigen Activation Complex (FAC), along with its interacting partners and upstream and downstream components, which have been well-characterized across numerous plant species. More recently, attention has turned to a lesser-known gene, FLOWERING PROMOTING FACTOR 1 (FPF1). Initially identified in Arabidopsis thaliana, FPF1 is a plant-specific gene lacking known functional domains, yet it plays a conserved and critical role in floral induction across diverse species. Despite its discovery in 1997, the molecular mechanism of FPF1 remained elusive until recent studies began to unravel the function of FPF and its homologs. One such study revealed that FPF1-Like Protein 1 (FLP1) in Arabidopsis is expressed in phloem companion cells sites of FLOWERING LOCUS T (FT) production. Like AtFT, AtFLP1 acts as a mobile florigenic signal, though it operates independently of the canonical AtFT pathway. AtFLP1 promotes flowering by activating the floral homeotic gene SEP3, suggesting an alternative regulatory route also influenced by photoperiod. Interestingly, studies in Brachypodium distachyon have highlighted a contrasting role for FLP-like genes, where they negatively regulate flowering by interfering with the FAC, underscoring species-specific diversity in its function. While initial studies have been majorly focused on their role in flowering, in recent years FPF1 family genes have also been implicated in other developmental processes, including stem and root elongation and shade avoidance responses. In this review, we explore these emerging insights into FPF1-like proteins, examining their multifaceted roles in flowering regulation and broader developmental functions, with a special emphasis on the most recent and impactful studies.

Enzalutamide resistance (EnzR) is a major challenge in the current treatment of castration-resistant prostate cancer, as tumors frequently progress to drug resistance after an initially effective treatment. Therefore, there is an urgent need to characterize the genes alterations that accompany EnzR in prostate cancer and to identify new therapeutic targets. In this study, we analyzed a total of 1273 publicly available transcriptomics datasets from patients who underwent prostate cancer surgery. We investigated transcriptomic changes after enzalutamide (ENZ) treatment, identified key genes involved in the process of EnzR, and developed EnzR scores to predict tumor progression. We further investigated the role of IGFBP3 in the regulation of EnzR in prostate cancer. The effect of IGFBP3 expression level on the malignant degree of EnzR cells was explored in vitro. In addition, we explored the downstream mechanism of IGFBP3 involvement in EnzR. We found that epithelial-mesenchymal transition (EMT), cancer stem cell-like properties, and neuroendocrine transformation occurred in tumor cells after ENZ treatment. Subsequently, we developed and validated EnzR scores to predict prostate cancer tumor progression. Furthermore, we experimentally confirmed that IGFBP3 promotes the proliferation of drug-resistant cells and enhances ENZ resistance via EMT signaling. Overall, we established a new EnzR scoring model through multidimensional analysis of EnzR patterns. This model can accurately predict the clinical prognosis of prostate cancer patients after surgery. Moreover, IGFBP3 can be used as a potential therapeutic target for ENZ resistance in prostate cancer.

Glioblastoma (GBM) is the most prevalent primary malignant tumor of the adult central nervous system, characterized by pronounced vascularity that facilitates tumor proliferation, invasion, and progression. Although anti-angiogenic therapy has emerged as a potential treatment strategy for GBM, currently available anti-angiogenic agents, such as bevacizumab targeting VEGF, have demonstrated limited efficacy in improving patient survival. This underscores the urgent need for novel therapeutic targets and strategies. In this study, we identified that the expression of maternally expressed gene 3 (MEG3), a tumor-suppressive long non-coding RNA (lncRNA), is negatively correlated with patient prognosis. Spatial transcriptomics sequencing and RT-qPCR analyses revealed that MEG3 is highly expressed in glioma stem cells (GSCs). In vitro tube formation assays further demonstrated that MEG3 in GSCs promotes angiogenesis in human brain microvascular endothelial cells (HBMECs). Transcriptome sequencing identified VGF nerve growth factor inducible (VGF), a secreted pro-angiogenic protein, as a downstream target, and showed that MEG3 regulates the pro-angiogenic activity of GSCs by modulating VGF expression. RNA pull-down assays revealed that MEG3 binds to far upstream element-binding protein 1 (FUBP3), which also regulates VGF expression. In vivo, knockdown of MEG3 significantly extended survival in orthotopic xenograft models of GSCs. Immunohistochemical analysis of mouse tumor tissues showed a corresponding reduction in VGF levels and microvessel density following MEG3 knockdown. In conclusion, this study demonstrates that MEG3 in GSCs promotes GBM angiogenesis through a FUBP3-dependent induction of VGF expression, highlighting MEG3 as a potential therapeutic target for anti-angiogenic intervention in GBM.