Perturbation of macropinocytosis triggers methuosis, a non-apoptotic cell death characterized by cytoplasmic vacuolization. However, the regulatory mechanisms of methuosis remain poorly defined. Lipid metabolism dysregulation is implicated in various cell death pathways, while its role in methuosis has remained elusive. Herein, LXX-8250, an isopropanolamine derivative of β-elemene, induced a vacuolization-associated cell death in breast and liver cancer cell lines. This process was accompanied by massive macropinocytosis, thereby confirming the occurrence of methuosis. Mechanistically, hypoxia-inducible lipid droplet-associated protein (HILPDA), a key regulator that promotes intracellular triacylglycerol (TAG) accumulation, was identified as the direct target of LXX-8250. By suppressing HILPDA, LXX-8250 inhibited diacylglycerol O-acyltransferase 1 (DGAT1) and activated adipose triglyceride lipase (ATGL). Consequently, lipid droplets and cellular TAG levels were reduced, while the subsequent increased diacylglycerol (DAG) stimulated macropinosome formation, leading to methuosis in these cells. In this study, we discover a novel methuosis agonist LXX-8250, and elucidate the critical role of HILPDA repression-dysregulated lipid metabolism in methuosis. Our study highlighted the potential of targeting this pathway as a therapeutic strategy to trigger cancer cell death.

Objective: To establish an in vitro fibrotic endometrial organoid model and investigate the early intervention effects of placenta-derived mesenchymal stem cell extracellular vesicles (Pd-MSC-EV) on fibrotic injury in endometrial organoids. Methods: Cell clusters were isolated from the endometrial tissues of healthy female mice and used to establish a three-dimensional endometrial organoid culture system. The structure and cellular composition of the organoids were characterized by histological staining and immunofluorescence staining. To construct an in vitro fibrosis model, the organoids were treated with different concentrations of transforming growth factor-β1 (TGF-β1), and model establishment was evaluated based on organoid morphological changes and the mRNA expression of fibrosis-related genes, including collagen type Ⅰ alpha 1 chain (COL1A1), α-smooth muscle actin (α-SMA), and fibronectin (FN). Pd-MSC-EVs were isolated and characterized by transmission electron microscopy, nanoparticle tracking analysis, and western blotting. The effects of Pd-MSC-EVs on TGF-β1-induced fibrotic changes in endometrial organoids were then assessed by morphological observation, real-time quantitative PCR (RT-qPCR) analysis of COL1A1, α-SMA, and FN mRNA expression, and immunofluorescence detection of COL1A1 and α-SMA protein expression. All experiments were repeated three times. Results: Mouse endometrial organoids with lumen-like structures were successfully established and expressed the epithelial markers E-cadherin and CK8 as well as the stromal marker Vimentin. Compared with the control group, TGF-β1 treatment disrupted organoid architecture and increased the mRNA expression levels of fibrosis-related genes, including COL1A1, α-SMA and FN, in a dose-dependent manner (all P<0.001). The isolated Pd-MSC-EV showed typical cup-shaped or disk-like vesicular morphology, with an average particle size of (163.8±50.0) nm, and expressed the exosomal markers CD63, CD81 and TSG101. Compared with the TGF-β1 group, co-treatment with Pd-MSC-EV alleviated organoid structural damage and reduced the mRNA expression levels of COL1A1, α-SMA and FN (all P<0.001). Immunofluorescence showed that COL1A1 protein expression was lower in the Pd-MSC-EV group than in the TGF-β1 group (P<0.005), while α-SMA protein expression showed a decreasing trend. Conclusion: Pd-MSC-EV attenuate TGF-β1-induced fibrotic changes in endometrial organoids in vitro and reduce organoid structural injury, providing experimental evidence for mechanistic studies and potential intervention strategies for endometrial fibrosis-related diseases such as intrauterine adhesions.

Ferroptosis has emerged as an important regulator of skeletal homeostasis, yet its role in osteoporosis (OP) remains incompletely understood. Accumulating evidence indicates that ferroptosis is not a uniform cell death program within bone, but rather a cell type-specific fate governed by distinct iron-handling capacities, redox buffering systems, and microenvironmental cues. This heterogeneity provides a new lens through which the complex pathogenesis of OP can be reinterpreted. In this review, we integrate recent advances to delineate ferroptosis-regulatory networks across major bone-resident cell populations, including osteoblasts, osteoclasts, osteocytes, and bone marrow mesenchymal stem cells (BMSCs). We highlight how ferroptosis suppresses osteogenic function in osteoblasts, amplifies differentiation and inflammatory signaling in osteoclasts, acts as an early vulnerability node in osteocytes, and reshapes lineage commitment in BMSCs. Importantly, ferroptosis in these cells is dynamically modulated by intercellular communication and niche-derived metabolic and mechanical signals. Building on this cell type-resolved framework, we propose that OP represents a disorder of multicellular ferroptotic dysregulation within the bone microenvironment rather than a simple imbalance of formation and resorption. Finally, we discuss translational implications, emphasizing ferroptosis-informed therapeutic strategies, including redox reprogramming, iron flux modulation, extracellular vesicle-based approaches, and microenvironment-responsive biomaterials. This integrative perspective provides a conceptual foundation for precision interventions targeting skeletal fragility across aging and disease contexts.

This study aimed to evaluate the cytocompatibility and transcriptional bioactivity profile of a calcium silicate-based endodontic sealer containing magnesium, in comparison with two widely used bioactive sealers, on immortalized human periodontal ligament stem cells (iPDLCs).

Structural remodeling of the dentate gyrus is a hallmark of temporal lobe epilepsy (TLE), yet the underlying molecular mechanisms remain incompletely understood. Nuclear distribution element-like 1 (Ndel1), a cytoskeleton-associated protein involved in neuronal migration and dendritic development, has not been characterized in dentate gyrus remodeling during epileptogenesis. Here, we investigated region- and cell-type-specific alterations in Ndel1 expression in a pilocarpine-induced mouse model of TLE and examined the effects of adeno-associated virus (AAV)-mediated Ndel1 expression on structural remodeling. Immunofluorescence was used to define Ndel1 localization across neural stem cells, granule lineage cells, mature neurons, and astrocytes, and dendritic architecture was assessed using Golgi staining and Sholl analysis. Total hippocampal Ndel1 expression increased after status epilepticus, whereas Ndel1-positive cells decreased selectively in the subgranular zone but increased among granule lineage cells in the hilus. Ndel1 was preferentially expressed in BLBP-positive neural stem cells and mature neurons, but not in neuroblasts. Activated astrocytic processes exhibited increased spatial association with Ndel1-positive cells during early remodeling. Ndel1 overexpression was associated with partial normalization of neuronal marker distribution, increased dendritic spine density, and reduced dendritic branching complexity. These findings suggest that Ndel1 is associated with region- and lineage-specific structural remodeling in the dentate gyrus during epileptogenesis.

Prostate cancer (PCa) is a leading cause of male cancer mortality worldwide. Its treatment failure is predominantly driven by the transition to castration-resistant phenotype (CRPC). Zinc finger E-box binding homeobox 1 (ZEB1) is a master inducer of epithelial-mesenchymal transition and has emerged as a central orchestrator of malignant phenotype. This review clarifies the complex interplay between ZEB1 and the tumor microenvironment, epigenetic modifications, and non-coding RNAs (lncRNAs and miRNAs), highlighting how these interactions collectively drive the progression of PCa. Moreover, ZEB1 is critical in driving the transformation of cancer cells to cancer stem cells and promoting neuroendocrine differentiation. This review highlights ZEB1 to resistance against standard-of-care treatments (such as taxane-based chemotherapies and next-generation androgen receptor pathway inhibitors) by metabolic reprogramming and modulation of androgen receptor signaling. Finally, it explores strategies to disrupt the regulatory circuitry of ZEB1 to overcome multidrug resistance, with the specific aim of evaluating whether ZEB1 can serve as a viable clinical biomarker and therapeutic target for PCa treatment.

Elastic cartilage is essential for preserving the structural integrity and functional flexibility of tissues such as the auricle and epiglottis. However, its limited capacity for self-repair poses significant challenges in the treatment of injuries resulting from trauma, congenital defects, or disease. This study investigates the in vitro and in vivo regenerative potential of a bioink designed for auricular cartilage repair. The primary polymeric components of the bioink were κ-carrageenan and silk fibroin (κ-CA/SF), while the cellular component consisted of adipose-derived stem cells (ADMSCs) isolated from rabbits, which were subsequently seeded onto poly(butylene adipate-co-terephthalate) (PBAT) microcarriers. These components were successfully printed utilizing an extrusion-based bioprinter. In vitro analyses demonstrated that ADMSCs were effectively encapsulated within the bioink, maintaining their viability for a duration of 21 days. Quantitative RT-qPCR analyses confirmed the expression of COL2A1, the major structural protein of cartilage and a critical indicator of the chondrocyte phenotype; its expression was observed to be approximately 35-fold higher in the κ-CA/SF + PBAT group compared to the κ-CA/SF group (***p < 0.001). Additionally, the presence of collagen II and aggrecan within the constructs was confirmed through immunostaining. For in vivo experiments, a circular defect in auricular cartilage, encompassing the perichondrium on both sides, was created in New Zealand rabbits to facilitate the implantation of tissue scaffolds. Cartilage regeneration was evaluated through histological staining at 4 and 8 weeks post-implantation. In the κ-CA/SF + PBAT group, distinct microcircular structures were identified within the connective tissue, and at later time points, the defect area was predominantly populated with mature cartilage cells, indicating advanced tissue development.

Controlled donation after circulatory determination of death (cDCD) has markedly expanded the lung donor pool; however, the biological consequences of donor warm ischemia on mitochondrial injury and systemic danger signalling remain uncertain. Cell-free mitochondrial DNA (cf-mtDNA) has been implicated in ischemia-reperfusion injury, yet its systemic behavior in ventilated human lungs is incompletely characterized.We conducted a prospective, multicenter observational study in matched adult lung transplant recipients from cDCD and donors after brain death (DBD) to characterize donor and recipient plasma cf-mtDNA kinetics and evaluate their relationship with warm ischemic exposure.Serial plasma samples were obtained from donors and from recipients.Donor plasma cf-mtDNA levels were comparable between groups.A divergence emerged at 72 hours after transplantation, reflected by a significant donor type-by-time interaction in adjusted longitudinal analysis. Neither total nor functional warm ischemia time was associated with cf-mtDNA concentrations. In multivariable analysis restricted to 72 hours, donor type was not independently associated with absolute systemic cf-mtDNA levels. Exploratory analysis of donor-derived cf-mtDNA demonstrated a decline in graft-derived mitochondrial DNA between reperfusion and 72 hours despite a concurrent increase in total cf-mtDNA. These findings suggest that post-transplant systemic cf-mtDNA dynamics reflect recipient-driven inflammatory processes rather than a direct surrogate of donor warm ischemic injury.

The autopod, the distal region of tetrapod limbs, represents a key evolutionary innovation requiring coordinated patterning, proliferation, and morphogenetic processes. While avian digit individualization has classically been described as driven almost exclusively by Programmed Cell Death (PCD) in the interdigital zone (IDZ), evidence from basal tetrapods indicates that differential growth between the digital zone (DZ) and IDZ originally mediated digit separation without apoptotic involvement. It remains unclear if this ancestral mechanism persists in amniotes or has been evolutionarily replaced by apoptosis. Here, we investigated growth dynamics in chicken (Gallus gallus) embryos by combining quantitative extension analyses, fluorescent DiI lineage tracing, and PH3-based proliferation assays. We demonstrate that differential growth is conserved in amniotes and arises through dynamic divergence in DZ and IDZ behaviour in chicken embryos. In chicken, although both regions expand comparably until HH32, IDZ extension declines markedly thereafter, while DZ outgrowth continues uninterrupted. DiI lineage tracing shows that the medial and proximal IDZ regions exhibit significantly greater extension at HH32 than at HH34, indicating a developmental decrease in growth that does not occur in the DZ, which maintains higher, stable extension rates across stages. Proliferation analyses align with these morphometric findings: the IDZ shows a sharp reduction in PH3-positive cells post-HH32, establishing a proliferative deficit relative to the DZ. Examination along the proximodistal axis revealed that the distal IDZ, influenced by the AER, shows the greatest proliferative activity, whereas medial and proximal regions contain fewer proliferative cells; this gradient is absent in the DZ, which displays relatively uniform proliferation. Together, these results indicate that avian digit separation is not governed solely by apoptosis but is initiated by an intrinsic decline in IDZ extension and cell-cycle activity. We propose that birds retain the ancestral differential growth mechanism, which works in concert with apoptosis to refine digit boundaries, bridging the evolutionary bridge between amphibian-like growth-driven individualization and amniote apoptotic refinement.

Gliomas are the most common primary tumors of the central nervous system. Among them, glioblastoma (GBM), a World Health Organization (WHO) grade 4 glioma, is the most aggressive form and remains one of the most lethal human cancers. Its poor prognosis is largely attributable to rapid progression, frequent recurrence, and profound therapeutic resistance, all of which are closely associated with glioma stem cells (GSCs). Although several signaling pathways sustaining GSC properties have been identified, upstream regulators that coordinate these pathways remain incompletely understood. TIMELESS, originally identified as a component of the circadian clock, has recently been implicated in the regulation of brain function and is aberrantly overexpressed in multiple cancer types; however, its role in glioma malignancy has not been defined. Here, we demonstrate that TIMELESS mRNA expression increases with glioma grade and is associated with poor prognosis across multiple glioma cohorts. Genetic depletion of Timeless suppressed tumor aggressiveness and prolonged survival in an orthotopic mouse glioma model by attenuating GSC properties. Mechanistically, Timeless enhanced stemness of glioma by upregulating Janus kinases (JAK) expression and promoting phosphorylation of signal transducer and activator of transcription 3 (STAT3). These effects were conserved in human GBM cells, supporting the relevance of TIMELESS-mediated signaling across species. Together, our findings uncover an unexpected, clock-independent function of TIMELESS in sustaining glioma stemness and malignancy and highlight the TIMELESS-JAK-STAT3 axis as a non-canonical mechanism that operates beyond the traditional circadian clockwork in treatment-resistant glioma.

Recent advances in immunotherapy have highlighted the need for clinically relevant immunocompetent models of metastatic neuroblastoma. However, such models of bone marrow (BM) metastasis remain limited. We previously developed a gene-modified mesenchymal stem cell (MSC)-based immunotherapeutic strategy that functions as a tumor-selective drug delivery system (DDS) and demonstrated its antitumor efficacy in a localized TH-MYCN neuroblastoma model. Therefore, this study aimed to establish a reproducible immunocompetent mouse model of neuroblastoma BM metastasis as a platform for evaluating novel therapeutic strategies.

Agonists of the lipid-sensing PPAR nuclear receptors, including PPARα, are being explored as therapies for metabolic disorders due to their roles in metabolic and anti-inflammatory processes. PPARα transcriptional programs have tissue-dependent features, yet there is a lack of genetic tools to study tissue-specific signaling activation without the addition of a systemic agonist. We aimed to investigate intestinal epithelial cell (IEC)-specific roles of PPARα signaling using a novel transgenic mouse that enables spatial and temporal control of Ppara overexpression. CAG-Ppara, -EGFP mice were bred to Villin-CreERT2 to establish the IEC-Ppara mouse, which was compared to littermate controls 2 weeks after tamoxifen exposure. IEC-Ppara mice had increased Ppara mRNA and PPARα protein in the intestinal epithelium. Transcriptional analysis of intestinal tissue from IEC-Ppara mice showed upregulation of PPARα target genes and functional enrichment for fatty acid catabolic processes. As expected, the enterocytes of IEC-Ppara mice were primed to absorb lipids following oral administration of an olive oil bolus. Unexpectedly, the enteroendocrine hormone Gip was among the most downregulated genes. GIP-positive cells were reduced in the intestines of IEC-Ppara mice and in mice treated with PPARα agonist WY-14643. Circulating GIP hormone, but not GLP-1 hormone, was reduced in IEC-Ppara mice. GLP-1-positive cells and hormone were unchanged. Consistent with reduced GIP function, IEC-Ppara mice consumed more food. These findings reveal PPARα as a regulator of GIP and support a new framework in which PPARα signaling influences systemic energy balance via a gut hormone axis. This study could have future impact on understanding responses to therapies targeting PPAR or incretin signaling.

Inducing white adipose tissue (WAT) browning is a promising strategy to combat obesity and metabolic disorders. However, current browning inducers often suffer from off-target effects and poor durability. Beige adipocytes, which can arise through de novo differentiation from adipose-derived stromal/stem cells (ASCs), offer a potential avenue for sustained thermogenesis. Here, we identified that forskolin (FSK) effectively induced the differentiation of C3H10T1/2 cells (a mesenchymal stem cell line) into beige adipocytes, evidenced by multilocular lipid droplets, enhanced mitochondrial biogenesis and function, and elevated uncoupling protein 1 expression. To overcome its poor bioavailability and lack of targeting specificity, we developed ASCs-targeting-peptide (ASP, sequence: GSWKYWFGEGGC) modified FSK nanoparticles (ASP@FSK NPs) by encapsulating FSK in the hybrid FDA-approved biodegradable polymers poly (lactic-co-glycolic acid) (PLGA) and polyethylene glycol (PEG)-ylated PLGA. ASP@FSK NPs selectively bind to glycanation site-deficient decorin (ΔDCN) receptors on ASCs, enabling prolonged retention and localized delivery within inguinal WAT (iWAT). In a high-fat diet-induced obesity model, local administration of ASP@FSK NPs significantly reduced body weight (∼30%), promoted de novo browning of iWAT, and alleviated liver steatosis in high-fat diet-induced obese mice, while expanding the therapeutic window of FSK and without apparent toxicity. Collectively, this study demonstrated that FSK could be developed as a potent inducer of de novo beige adipogenesis and targeted delivery of FSK to ASCs might provide a valuable strategy for the treatment of obesity. STATEMENT OF SIGNIFICANCE: Obesity and metabolic syndromes remain global health challenges, and strategies that induce white adipose tissue browning hold great therapeutic potential. However, existing browning agents are limited by systemic toxicity, off-target effects, and poor sustainability. This study addresses these critical gaps by demonstrating that forskolin (FSK) drives de novo beige adipocyte differentiation from mesenchymal stem cells. We further developed an adipose-derived stem cell-targeted nanoparticle system (ASP@FSK NPs) for localized and sustained delivery. In obese mice, ASP@FSK NPs efficiently promoted inguinal WAT browning, alleviated adiposity and hepatic steatosis, and expanded the therapeutic window of FSK without obvious toxicity. This work establishes a targeted and durable approach for WAT browning and obesity treatment by integrating a progenitor-directed biological strategy with precision nanomedicine.

Brain-metastatic breast cancer (BMBC) is a severe complication of advanced breast cancer, affecting 15-30% of metastatic patients, particularly those with HER2-positive or triple-negative subtypes, and is associated with dismal prognosis and median survival under 12 months. Therapeutic resistance, driven by the central nervous system's sanctuary role, poses a major barrier to effective treatment, often resulting in discordant intracranial versus extracranial responses. This comprehensive review highlights BMBC resistance mechanisms, drawing from preclinical models, clinical studies, and genomic analyses. Key drivers include genetic/epigenetic alterations, BBB-mediated drug exclusion via efflux transporters, and microenvironmental interactions with astrocytes and immune cells that promote survival signaling. Additional factors encompass cancer stem cell plasticity/dormancy enabling therapy evasion, metabolic reprogramming and extracellular matrix remodeling that shields tumor from drugs. We highlight how these interconnected pathways create a protective niche for metastatic cells. Promising strategies to overcome resistance include BBB-penetrant agents, antibody-drug conjugates, nanomedicine, and combination therapies targeting the tumor microenvironment and epigenetics. By integrating mechanistic insights with translational opportunities, this review emphasizes the potential for personalized, multi-targeted approaches to improve patient outcomes in BMBC.

Doxorubicin is a well-known cardiotoxic chemotherapeutic agent. Whether its combination with vincristine exacerbates cardiotoxicity and the underlying mechanisms remains unclear.

Neural organoids provide tractable models of human brain development, function, and disease, but adeno-associated viral vector (AAV)-based gene delivery is limited by inefficient transduction and insufficient comparative evaluation of the natural and engineered capsids. Here, we evaluated seven AAV capsids (AAV-2, -6, -9, -DJ, -2-retro, -PHP.eB, and -Cap-Mac) for transduction efficiency, cell type tropism, cytotoxicity, and functional integrity in human cortical and thalamic organoids. AAV-Cap-Mac demonstrated the highest transduction efficiency, with broad cellular tropism, but caused cytotoxicity and decreased neuronal activity. By contrast, AAV-DJ yielded efficient neuronal transduction, with preserved calcium dynamics and minimal cytotoxicity, which enabled rabies virus-based circuit tracing and all-optical physiology. Our findings reveal a capsid-dependent trade-off between transduction efficiency and functional integrity and identify AAV-DJ as an optimal serotype for robust gene delivery and physiological interrogation in organoid-based neuroscience. Combining AAV-DJ with emerging molecular and circuit-level technologies in organoids should facilitate mechanistic and translational studies of the human brain.

Natural Killer (NK) cells generally exhibit dysfunction in tumor microenvironment (TME), significantly limiting their efficacy in antitumor therapy. Tumor cells display enhanced glycolysis, leading to lactic acid (LA) secretion and accumulation in the TME. Using human-induced pluripotent stem cell (hiPSC)-derived NK cells (iNKs) as a model, we demonstrate that LA induces substantial iNK dysfunction, including reduced survival, impaired IFN-γ secretion, and diminished tumor-killing capacity due to severely compromised mitochondrial function. To overcome LA-induced dysfunction, we knocked in an expression cassette for lactate dehydrogenase B (LDHB) into hiPSCs (LDHB-hiPSCs), enabling conversion of cellular lactate to pyruvate. iNKs derived from LDHB-hiPSCs (LDHB-iNKs) largely resisted LA-induced dysfunction, exhibiting enhanced cytotoxicity and improved survival in high-LA tumor tissues. Importantly, LDHB-iNKs show superior suppression of solid tumor formation in vivo. Our study provides a novel strategy to enhance NK cell therapy against solid tumors by reshaping the metabolic pathways.

The pace of embryonic development differs between mammalian species, yet the molecular basis for this remains unknown. By comparing protein dynamics in mouse and human neural progenitors (NPs), we show that protein turnover is faster in mouse NPs, driven by higher rates of protein synthesis and degradation. Human NPs exhibit longer protein half-lives, reduced proteasomal activity, and lower proteasome abundance. These differences persist in post-mitotic neurons and are also observed in the embryonic spinal cord in vivo. Pharmacological inhibition of proteasomal activity slows differentiation in mouse NPs. Conversely, enhancing proteasomal activity accelerates neuronal output in human NPs. Moreover, accelerating the degradation of the key transcriptional repressor IRX3 in mouse NPs speeds the activation of its target gene. Together, these results provide evidence that species-specific regulation of proteasome-mediated proteolysis influences the timing of neural development and suggest that evolutionary tuning of proteasomal activity contributes to differences in embryonic developmental pace.

Chronic liver diseases of diverse etiologies, along with primary liver cancers, represent major global health burdens with limited curative options. CD8+ T-cell exhaustion is a central barrier to effective immune control in these pathological settings. However, precursor exhausted CD8+ T cells (Tpex), a stem-like subset with self-renewal and proliferative potential, retain the ability to generate effector-like progeny and sustain long-term immune surveillance. In this review, we first provide a definition of Tpex to distinguish them from other related CD8+ T-cell states, including terminally exhausted, effector-like exhausted, tissue-resident memory, and conventional memory T cells, in chronic liver diseases and liver cancer. We then summarize current knowledge on the fundamental biology, etiological dynamics, and therapeutic relevance of Tpex in both benign and malignant liver diseases. Specifically, we examine the transcriptional, metabolic, and microenvironmental networks that govern Tpex fate, with particular emphasis on how distinct etiologies, including viral hepatitis, metabolic dysfunction-associated steatotic liver disease, and autoimmune hepatitis, differentially shape Tpex abundance and functional state. In chronic viral hepatitis, Tpex dynamics are critically influenced by viral persistence, metabolic context, and co-infections, which in turn have direct implications for the responsiveness to immune checkpoint blockade. In hepatocellular carcinoma and intrahepatic cholangiocarcinoma, Tpex signatures are not only correlated with clinical outcomes but also modulated by tumor genetic landscapes and the composition of the immune microenvironment. Finally, we highlight emerging Tpex-targeted therapeutic strategies, including co-stimulatory agonists, metabolic reprogramming, vaccines, CAR-T cell engineering, and gut microbiota modulation, as promising avenues to overcome immunotherapy resistance.