Skeletal muscle stem cells (SkMSCs) are essential for muscle regeneration and represent a promising therapeutic target for muscle disorders. However, effective strategies to precisely regulate SkMSC fate by integrating biochemical and mechanical cues remain limited.

Temozolomide (TMZ) resistance in glioblastoma (GBM) remains a critical barrier to treatment success, driven by O6-methylguanine-DNA methyltransferase (MGMT) overexpression, glioma stem cell (GSC) persistence, and redox adaptation.

Information on the maintenance of tissue homeostasis is important for developing effective therapeutic methods. However, reports on the cellular composition and tissue stem cells of the larynx are scarce. Therefore, we analyzed mouse laryngeal mucosa using single-cell RNA- sequencing and spatial transcriptomics by photo-isolation chemistry, and we also generated laryngeal organoids as an in vitro model. Consequently, we found a SOX9-positive basal cell subpopulation and a Lgr5-positive cell subpopulation in the mouse vocal fold, and obtained three types of epithelial organoids from laryngeal epithelium. We also confirmed the differences in pseudostratified ciliated columnar epithelium characters between the supraglottis and subglottis of the mouse larynx. These findings provide valuable insights and tools for future research in laryngology and stem cell biology.

FLT3-ITD inhibitors are approved for acute myeloid leukemia (AML) treatment but relapse is common. In this study, the combined inhibition of FLT3-ITD signal and protein translation by QUIZartinib and Omacetaxine Mepesuccinate (QUIZOM) synergistically suppressed the most critical FLT3-ITD survival signals including mitochondrial respiration and proteostasis, which induced apoptosis and pro-inflammatory response. In a Phase 2 trial (NCT03135054) involving 40 chemo-refractory/unfit FLT3-ITD AML patients, QUIZOM achieved a composite complete remission (CRc) of 83%, a median leukemia-free survival (LFS) of 10 months (Range: 0.7-68.2 months) and a median overall survival (OS) of 12.9 months (Range: 1.8-69.2 months). 13/33 (39%) received allogeneic HSCT after a median of 143 days (Range: 53-367 days). Higher CRc rates were observed in patients with NPM1 mutations, DNMT3A mutations, and wild-type WT1. Single-cell RNA-sequencing of QUIZOM cohort revealed positive correlation between pro-inflammatory response in blasts, CD8 + T activation and clinical responsiveness. Further, we identified a leukemic stem cell (LSC) subpopulation with activated JNK/JUN/HSPA1B axis via PLD1-driven phosphatidylcholine metabolism, which promoted proteostasis and drove QUIZOM resistance. PLD1-inhibitor remodeled phospholipid metabolism, induced ferroptosis and restored QUIZOM response in LSC. Our findings provided the therapeutic and resistant mechanisms of QUIZOM and paved the way for targeted interventions in this AML subtype.

While genetic screens have facilitated the dissection of protein function in animal development, advances in systematic point mutagenesis open new opportunities for forward genetics in mammalian cells. Here, we develop a CRISPR/Cas9-mediated base editing screen that allows functional screening of extensive collections of single amino acid substitutions of endogenous proteins. We demonstrate the application on the X-chromosomal Hprt and the autosomal Msh2 gene in diploid male and haploid mouse embryonic stem cells, respectively. Finally, we use this methodology to generate a sequence-function map of the transcriptional co-repressor SPEN in X chromosome inactivation. We demonstrate that the substitution of the SPEN RRM4-residue W522 abrogates X-linked gene repression by Xist RNA and impairs the establishment of H3K27me3 deposition. Our results demonstrate that screening in haploid cells allows efficient identification of mutations that would be recessive in diploid cells, suggesting applications across a wide range of areas.

Hematopoietic stem cells (HSCs) survive many types of cellular stress but often lose their regenerative and lymphopoietic capacities as a result. Such functional decline also occurs with age, and dysfunctional HSCs with impaired mitochondria accumulate during aging. However, the molecular link between HSC stress response and age-related functional decline remains poorly understood. Here we show that multiple stress responses converge on the RIPK3-MLKL axis to induce age-related changes in HSCs. The necroptosis effector MLKL is readily activated by inflammation and replication stress and accumulates in HSC mitochondria. Consequently, activated MLKL does not cause cell death but impairs HSC self-renewal and lymphoid differentiation. Such MLKL-mediated functional decline also occurs in HSCs during organismal aging, with activated MLKL primarily mediating age-related mitochondrial damage and reduced glycolytic flux. Collectively, our results establish the RIPK3-MLKL axis as a key mediator of HSC aging and identify a necroptosis-independent role of MLKL in mitochondrial damage.

Protein tyrosine phosphatase receptor type Z1 (PTPRZ1) is one of the most abundantly expressed and enriched genes in astrocytes during development, yet its function in astrocytes is unknown. Using an astrocyte-neuron co-culture system, we found that knockdown of Ptprz1 in astrocytes significantly impaired astrocyte branching morphogenesis. To investigate the function of PTPRZ1 in astrocytes during brain development, we generated a Ptprz1 conditional knockout mouse and deleted Ptprz1 from astrocytes postnatally, after the bulk of astrogenesis is complete. At postnatal day 21, we found subtle changes in astrocyte morphology and a reduction in the density of co-localized pre and post synaptic excitatory synapse markers across multiple layers of the visual cortex in both male and female mice, suggesting important functions for astrocytic PTPRZ1 in both astrocyte morphogenesis and synaptogenesis. PTPRZ1 is expressed in several neural cell types, including radial glial stem cells and oligodendrocyte progenitor cells (OPCs), and regulates critical aspects of neurodevelopment, including neurite outgrowth, neuronal differentiation, myelination, and extracellular matrix (ECM) development. Moreover, altered PTPRZ1 expression is associated with schizophrenia and glioblastoma. Therefore, this mouse model is a valuable resource for investigating cell-type-specific PTPRZ1 function in numerous neurodevelopmental and neuropathological mechanisms.Significance Statement PTPRZ1 is an abundant, astrocyte-enriched protein linked to neurological dysfunction; however, its astrocyte-specific functions are unknown. We generated a Ptprz1 conditional knockout mouse and found that astrocyte-specific deletion of Ptprz1 reduces the density of co-localized excitatory synapse markers in the developing mouse cortex, with mild impact to astrocyte morphology. PTPRZ1 is an emerging therapeutic target for glioblastoma and neurodegeneration. This study provides a new tool to study PTPRZ1 function in neurodevelopment and neuropathology.

Osteogenesis depends on the self-renewal and differentiation of mesenchymal stem cells (MSCs). Emerging research underscores the regulatory functions of RNA methylation on bone homeostasis. Here, we show PCIF1, the N6,2'-O-dimethyladenosine (m6Am) methyltransferase, is essential for maintaining bone mass and promoting osteogenic differentiation of MSCs. Multiple complementary analyses-including GWAS, TWAS, and single-cell transcriptomics-collectively point to PCIF1 as a regulator of human bone mineral traits and early-stage mesenchymal differentiation. Global or MSC-specific Pcif1 deletion elicits osteoporotic pathology in mice, although myeloid cell-specific Pcif1 knockout does not induce femur bone alterations. Mechanistically, Pcif1 knockout decreases m6Am signals of Wnt-related genes (Wnt11, Fzd4, and Fgfr2) and accelerates mRNA degradation. This down-regulates active β-Catenin protein, and thus impairs osteogenic function of MSCs. Additionally, the WNT agonist attenuates the osteoporosis-like phenotype induced by Pcif1 deletion. These findings highlight the crucial role of PCIF1-mediated m6Am modification in regulating osteogenesis and suggest potential therapeutic implications for bone disorders.

Bone maintains a dynamic and stable state through the orchestration of osteoclasts and osteoblasts. Osteoblasts are derived mainly from mesenchymal stem cell (MSC) and are responsible for bone formation. The inhibition of osteoblast proliferation and differentiation is involved in many diseases, including osteoporosis, osteoarthritis, infected bone defects, and inflammatory aseptic loosening of implants. Given the currently limited treatment options, exploring new methods to promote bone formation is an important focus significant for orthopedists. Platelet-rich plasma (PRP), an autologous substance that is rich in various growth factors, is widely used in regenerative medicine. However, the effect of PRP on inflammatory bone destruction remains unclear. We investigated the effects of PRP on the viability of MSC, and the impact of different concentrations of PRP (1% and 3%, respectively) on cell death, proliferation, and differentiation. Furthermore, we tested the therapeutic effect of different concentrations of PRP (1% and 3%) on LPS-induced inflammatory bone destruction in vivo. PRP enhanced the cellular activity of MSC and promoted osteogenesis. A higher concentration of PRP (3%) primarily reduced the death and increased the proliferation of in LPS-treated MSC via the PI3K/AKT pathway, while a lower concentration of PRP (1%) promoted MSC differentiation into osteoblasts through the MAPK pathway. Consistent with in vitro experiments, we validated the protective effect of PRP against LPS-induced bone loss by increasing bone formation in vivo. These results suggest that different concentrations of PRP can ameliorate LPS-induced inflammatory bone loss through distinct mechanisms.

Nucleic acid-based therapeutics, which involve the manipulation of genetic materials to treat or prevent diseases, have gained considerable attention, leading to the approval of medicines such as COVID-19 vaccines, patisiran (Onpattro), and nusinersen (Spinraza). However, their clinical application is hindered by challenges such as nuclease degradation, poor biodistribution, limited cellular uptake, and inefficient endosomal escape. Extracellular vesicles (EVs), which are natural nanoscale drug delivery systems derived from various eukaryotic and prokaryotic cells, offer a safe, efficient, specifically targeted, and non-pathogenic method for nucleic acid delivery. In this review, we summarize the classical methods and the latest research advances in EV preparation and nucleic acid loading. Additionally, we review the primary administration routes for nucleic acid-loaded EVs, such as intravenous, local, oral, intranasal, and inhalation delivery. By addressing these aspects, this review aims to guide the optimal design and clinical application of nucleic acid-loaded EVs.

Necroptosis is an inflammatory programmed cell death pathway linked to diverse physiological and pathological disorders, yet its role in Enterotoxigenic Escherichia coli (ETEC)-induced intestinal inflammation and mucosal injury remains poorly understood. This study aimed to elucidate the contribution of intestinal epithelial cell necroptosis to the development of intestinal inflammation and injury induced by ETEC infection in piglets. Following ETEC challenge in piglets, key proteins involved in necroptosis, including phosphorylated receptor-interacting protein kinase 3 (p-RIP3) and high-mobility group box 1 (HMGB1), were upregulated in jejunal crypt epithelial cells, which are primarily composed of Paneth cells and stem cells, in a time-dependent manner. In addition, ETEC challenge triggered time-dependent pyroptosis in jejunal lamina propria lymphocytes, a population that includes macrophages, as demonstrated by elevated levels of NLRP3, Caspase-1, GSDMD-N, and Cleaved -IL-1β (p17) proteins in lamina propria lymphocytes. Necroptosis of jejunal crypt epithelial cells occurred prior to pyroptosis of lamina propria lymphocytes, indicating that epithelial cell necroptosis may contribute to the induction of pyroptosis in lamina propria lymphocytes. ETEC challenge induced progressive TNF-α and IL-1β upregulation in plasma, jejunal crypt epithelial cells, and lamina propria lymphocytes of piglets. These changes coincided with intestinal injury and barrier loss, which were indicated by increased plasma i-FABP and decreased jejunal ZO-1 and Occludin. Notably, Nec-1 pretreatment mitigates ETEC-induced intestinal inflammation and tissue damage in piglets by inhibition of crypt epithelial cells necroptosis and the ensuing pyroptosis of lymphocytes. These results indicate that targeting upstream epithelial-cell necroptosis is an important strategy to attenuate inflammation and preserve barrier integrity.

Gliomas are the most common primary malignant tumors of the adult central nervous system, characterized by rapid growth, high recurrence rates, and limited response to standard treatments, with median survival under 15 months. The SIX transcription factor family has been implicated in tumor development, but the role and regulatory mechanism of SIX5 in glioblastoma (GBM) remain unclear. This study systematically investigates the biological function of SIX5 and its regulatory network in GBM. Differential expression and weighted gene co-expression network analyses of GSE4290 and GSE50161 datasets, combined with machine learning algorithms including LASSO, identified SIX5 as a core candidate gene. Functional enrichment analyses and evaluation using TCGA and UALCAN databases revealed that SIX5 is highly expressed in GBM and associated with poor prognosis. Single-cell RNA sequencing and spatial transcriptomics showed enrichment of SIX5 in the tumor core and in astrocyte-like and stem cell-like subsets at the invasion front. In vitro, U87 and U251 cells with lentivirus-mediated SIX5 knockdown or overexpression were assessed for proliferation, migration, invasion, apoptosis, and colony formation. SIX5 knockdown significantly inhibited proliferation, migration, invasion, epithelial-mesenchymal transition, and tumorigenicity, while promoting apoptosis. Mechanistically, KDM5C positively regulates SIX5, which directly binds the UBE2C promoter to activate its transcription, enhancing AKT/mTOR signaling and promoting aerobic glycolysis via upregulation of GLUT1, HK2, PGK1, and LDHA. Rescue experiments showed that UBE2C overexpression partially restored malignant phenotypes under SIX5 downregulation. In vivo xenograft studies confirmed that the KDM5C-SIX5-UBE2C axis drives GBM growth. In conclusion, SIX5 functions as a critical oncogenic driver in GBM, regulated by KDM5C and promoting tumor progression through UBE2C-mediated activation of AKT/mTOR signaling and glycolytic reprogramming. The KDM5C-SIX5-UBE2C regulatory axis represents a potential prognostic biomarker and therapeutic target in glioblastoma.

ARG1 catalyzes the conversion of L-arginine to L-ornithine, urea, polyamines, and L-proline, thereby balancing nitrogen detoxification with tissue-specific roles in proliferation and immunity. This review delineates the context-dependent functions of ARG1 across diverse cell types-including tumor cells, immune cells, endothelial cells, keratinocytes, and stem cells. In tumors, ARG1 drives immunosuppression and metabolic reprogramming but can paradoxically suppress tumorigenesis. Immune modulation via ARG1-polyamine crosstalk regulates T cell differentiation, macrophage polarization, and microbiota interactions, influencing infection and autoimmunity. Endothelial ARG1 exacerbates obesity-related vascular dysfunction, while keratinocyte ARG1 impacts wound healing and psoriasis. Emerging therapies-such as ARG1 inhibitors, engineered extracellular vesicles, and microbiome interventions-show preclinical promise in cancer, cardiovascular, and neurodegenerative diseases. By mapping ARG1's spatiotemporal metabolic networks, this work highlights its dual roles and positions ARG1 as a central player for precision medicine in complex pathologies.

A diverse range of innovative biological therapies is being developed to treat various human diseases. The safety assessment of these biologics is a critical factor determining clinical success. Enhanced humanised mouse models have the potential to revolutionise immunotoxicological profiling by refining procedures for effective in vivo safety assessments.

Both primary and metastatic brain tumors rely on signals from the surrounding environment for their survival and progression. In particular, the most common and lethal brain cancer, glioblastoma (GBM), derived from glial cells (astrocytes or microglia), has been shown to integrate into synaptic networks and to receive paracrine signals from neighbouring tumor microenvironment (TME) cells. There is increasing evidence that metastatic disease in the brain exhibits similar behavior. The TME both maintains malignant cells and is maintained by them, a process that relies on cancer stem cells (CSCs). These stem cells and their signaling mechanisms, including in the case of GBM, "GSCs," provide possible novel targets for immunotherapy. In this review, we will discuss the integration of primary and malignant brain tumors into normal synaptic networks, the role of tumor stem cells and the TME in this integration, and the potential for immunotherapeutic targeting of these processes.

The management of critical-sized bone defects has attracted heightened interest due to its challenging nature. To date, numerous engineered tissues incorporating nano-hydroxyapatite (nHap) have been proposed; however, nHap continues to encounter limitations, particularly regarding its inadequate immunomodulatory effects on bone. Therefore, needlelike nHap (NnHap)-based scaffolds were fabricated using a polylactic acid derivative and carboxymethyl chitosan. We hypothesize that NnHap@CP can not only promote bone immunomodulatory effects and angiogenesis in human umbilical vein endothelial cells through M2 subtype polarization but also directly promote osteogenesis in rat bone-marrow-derived mesenchymal stem cells (rBMSCs). Furthermore, mass spectrometry was employed to determine that osteoprotegerin/RANK/RANKL may represent a potential signaling pathway through which NnHap@CP enhances the osteogenesis of rBMSCs. In our study, NnHap@CP demonstrated a satisfactory effect on M2 subtype polarization in macrophages and enhanced osteogenesis in rBMSCs, as observed in an in vitro study. We employed NnHap@CP for the in vivo examination of a rat model with cranial critical-sized bone defects. We discovered that NnHap@CP significantly enhances new bone regeneration and neovascularization, potentially serving as an innovative treatment strategy for critical bone defects.

This review synthesizes evidence from recent clinical and mechanistic studies published between 2015 and 2024 to provide updated insights into the prevention and management of adverse drug reactions (ADRs) associated with first-line anti-tuberculosis drugs (ATDs)-namely isoniazid (INH), rifampicin (RIF), pyrazinamide (PZA), and ethambutol (EMB)-which are essential for tuberculosis (TB) treatment but frequently cause significant ADRs that threaten therapeutic success. We examine four major toxicities: hepatotoxicity (primarily from INH and RIF, mediated by oxidative stress, mitochondrial dysfunction, and cytochrome P450 induction); peripheral neuropathy (driven by INH-induced pyridoxine depletion and EMB-related copper chelation leading to optic and axonal damage); central nervous system (CNS) toxicity (notably INH-induced seizures due to GABAergic disruption); and myelosuppression (mainly RIF- or PZA-related, involving oxidative injury to hematopoietic stem cells and impaired DNA synthesis). Key risk factors include advanced age, malnutrition, pre-existing organ dysfunction, and pharmacogenetic variations (eg, NAT2 acetylator status). Management strategies emphasize protocol-driven monitoring-including baseline and serial liver function tests (LFTs), complete blood counts (CBC), neurologic exams, and monthly visual assessments for EMB-and graded interventions based on severity thresholds (eg, temporary discontinuation if ALT >3× upper limit of normal (ULN) with symptoms or >5× ULN asymptomatic), alongside targeted therapies such as pyridoxine for neuropathy and N-acetylcysteine for hepatotoxicity. Proactive measures, including pretreatment risk stratification, patient education, and multidisciplinary coordination, are critical to optimizing adherence and outcomes. Effective management of first-line anti-TB drug toxicity requires mechanism-informed monitoring, individualized interventions, and proactive patient education to maintain treatment adherence and improve global TB outcomes.

To systematically evaluate the efficacy and safety of mesenchymal stem cell (MSC) therapy for patients with Multiple System Atrophy (MSA) by synthesising available clinical trial evidence and clarifying signals of disease modification.

The use of animal-derived reagents in biomedical research poses challenges for reproducibility due to batch-to-batch variability and inter-species differences, along with ethical concerns related to their origin. In pursuing a human-relevant in vitro model, an animal-free and defined cell culture process is preferred to improve relevance and reproducibility. We investigated the use of serum replacement (SR) consisting of human hepatocyte-derived proteins in cell culture and recombinant antibodies with a plant-derived blocking solution (animal-free blocker, AFB) in immunocytochemical staining of cells. Human serum (HS) instead of animal-derived serum was used in this study for comparison with SR. We showed that bone marrow stem/stromal cells (BMSCs) maintain their proliferation capacity and cell-specific morphology in SR-supplemented medium, whereas human umbilical vein endothelial cells (HUVECs) show compromised growth under similar conditions. In a more complex co-culture, BMSCs + HUVECs formed a stable vascular network in SR-supplemented medium. In immunocytochemical staining, we compared the performance of recombinant antibodies with animal-derived antibodies and an AFB solution with a bovine serum albumin (BSA)-based blocking solution. Adipose stem/stromal cells (ASCs) showed their typical spindle-shaped morphology when stained with recombinant antibodies against alpha-smooth muscle actin (αSMA) in both AFB and BSA-based blocking solutions. We detected partial non-specific binding of recombinant antibodies and animal-derived antibodies against β-tubulin III in ASC. In contrast, we did not observe non-specific binding on these neuronal antibodies in HUVECs in any tested condition. While protocol optimization depends on the cell type used, our findings indicate that animal-derived materials can reliably be replaced.