PEG-MGF

P2Y12 receptor in trigeminal ganglion contributes to CFA-induced mechanical allodynia in mice.

Zhishan Zou, Qianyi Shi, Lijia Mai +4 more

Chronic orofacial inflammatory pain remains a major clinical challenge, and peripheral sensitization within the trigeminal ganglion (TG) is thought to contribute to its development. This study aimed to investigate whether P2Y12 receptor signaling contributes to orofacial inflammatory pain, with particular focus on the TG as a potential peripheral site of action. A mouse model of orofacial pain was established through the subcutaneous injection of complete Freund's adjuvant (CFA) into the right whisker pad. The expression and distribution of P2Y12 receptor were characterized using single-cell RNA sequencing, immunohistochemistry, and immunofluorescence. The functional role of P2Y12 signaling was examined by systemic clopidogrel treatment in vivo and by ADP stimulation with or without the selective antagonist PSB-0739 in primary satellite glial cells (SGCs) and macrophages in vitro. P2Y12 expression in the TG was significantly upregulated 7 days after CFA injection and was predominantly expressed on SGCs and macrophages. Systemic pharmacological inhibition of P2Y12 alleviated mechanical allodynia and was associated with reduced c-Fos expression in the spinal trigeminal nucleus caudalis (SpVc). In addition, the P2Y12 inhibition was accompanied by decreased SGC activation, fewer CD86-positive macrophages, and lower interleukin-6 (IL-6) expression in the TG. In vitro assays further showed that P2Y12 activation promoted pro-inflammatory cytokine expression in both SGCs and macrophages, and this effect was attenuated by PSB-0739. Collectively, these findings suggest that P2Y12 receptor may contribute to the regulation of orofacial inflammatory pain, in part through neuroimmune alterations within the TG.

Tripartite warfare: decoding the cell-virus-virophage arms race.

Ting Chu, Yongjie Wang

Giant viruses constitute a remarkable group of large double-stranded DNA (dsDNA) viruses distinguished by their exceptional structural complexity and genomic features. Their genomes can reach 2.8 Mb, encoding hundreds of proteins, and virion diameters up to 1.5 μm. They infect diverse eukaryotic hosts and establish viral factories within host cells. Virophages are small dsDNA viruses (17-34 kb; 50-75 nm) that parasitize giant viruses. These satellite viruses hijack giant virus replication machinery while suppressing giant virus progeny, benefiting the host cell and creating a parasitic-symbiotic dynamic. This review examines the tripartite relationship between host cells, giant viruses, and virophages (CVv systems), focusing on: (1) virus-host interactions in amebae, marine flagellates, and unicellular algae; (2) molecular mechanisms of these interactions; and (3) ecological and evolutionary implications. We also identify current research challenges and propose future directions, particularly the molecular basis of viral interactions in CVv systems.

Genomic structural equation modeling reveals the shared genetic architecture of osteosarcopenia across five musculoskeletal phenotypes.

Weiqiang Lai, Kaiqin Gong, Ronghao Zhong +9 more

Osteosarcopenia-the concurrent presence of osteoporosis and sarcopenia-affects approximately 18.5% of older adults globally, yet its shared genetic basis remains poorly understood. We applied genomic structural equation modeling (Genomic SEM) to integrate genome-wide association study (GWAS) summary statistics across five phenotypes spanning the pathophysiological spectrum of osteosarcopenia: appendicular lean mass (ALM), bone mineral density (BMD), handgrip strength (HGS), walking pace, and fracture. Fine-mapping, transcriptome-wide association study (TWAS), pathway enrichment, cell-type enrichment, and spatial transcriptomic mapping were performed to functionally annotate the identified loci. A single-factor model (CFI = 0.976) captured the shared genetic liability, with HGS and ALM loading most strongly. A two-factor sensitivity analysis confirmed partial separability of muscle and bone dimensions, though the single common factor was retained for integrated downstream annotation. We identified 58,696 genome-wide significant single-nucleotide polymorphisms (SNPs) condensed into 1078 independent lead variants, including 29 novel loci. Fine-mapping prioritized 317 high-confidence causal variants, encompassing key genes including BMP6, ACAN, IHH, LRP5, and SOX5. TWAS and MAGMA converged on IGF1R, FOXO3, and IRS1 as dual susceptibility genes. Pathway analysis revealed significant enrichment in endochondral ossification and growth hormone/insulin-like growth factor-1 signaling. Cell-type enrichment localized genetic risk to mesenchymal stem cells and skeletal muscle satellite cells, while spatial mapping identified cartilage primordium as the most enriched developmental context. This study systematically elucidates the shared genetic architecture of osteosarcopenia, highlighting developmental, endocrine, and stem cell-related pathways as core mediators. These findings provide a theoretical foundation for precision geroscience and the development of dual-target therapeutic strategies.

Regulation of Satellite Cells and Myogenesis in Response to Eccentric Resistance Exercise in Hypoxic Conditions in Healthy Young Men.

Sophie van Doorslaer de Ten Ryen, Geoffrey Warnier, Nancy Antoine +5 more

Satellite cells participate in myogenesis and contribute to skeletal muscle regeneration and hypertrophy. Amongst other stimuli, satellite cells can be activated by exercise and hypoxia. However, the cumulative effect of exercise on hypoxia on myogenesis is not well understood, certainly in humans. Furthermore, whether satellite cell activation and myogenesis differ between environmental hypoxia and blood flow restriction is not known. The purpose of this study was to analyze satellite cell and myogenic markers in response to acute eccentric resistance exercise in normoxia, normobaric environmental hypoxia, and with blood flow restriction (local hypoxia). Thirty-eight healthy young men were allocated to one of the three experimental conditions: normoxia (n = 13), normobaric environmental hypoxia (n = 12), and blood flow restriction (n = 13). They all performed 5 series of 15 repetitions at 60°/s for the knee extension and 30°/s for the knee flexion on an isokinetic dynamometer. Vastus lateralis muscle biopsies and blood samples were taken before, 1, 24, and 72 h after exercise. Myogenic regulatory factor expression was upregulated after exercise similarly in the normoxic and hypoxic groups and attenuated in the blood flow restriction group. Despite differential regulation of myogenic regulatory factor expression and circulating creatine kinase levels after eccentric resistance exercise, none of the investigated hypoxia markers and immediate early genes, inflammatory markers, growth factors, except insulin-like growth factor-1, and mitogen-activated protein kinase members were differently regulated between the groups. Contrary to our hypothesis, satellite cell activation and myogenesis were not potentiated by the combination of eccentric resistance exercise and hypoxic conditions.

Artificial Crystalline-Amorphous Architecture Enables Continuous Ion Transport in Poly(Vinylidene Fluoride)-Based Solid-State Electrolytes.

Yue-Ming Chen, Min Zuo, Hanghua Wu +9 more

Solid-state polymer electrolytes (SPEs) hold promises for next-generation batteries but are hindered by low ionic conductivities. This issue stems from crystalline regions that block long-range ion transport in amorphous phases, while fully amorphous polymers are mechanically unstable. Here we design an "artificial crystalline-amorphous" architecture that converts the isolated amorphous conduction zones into continuous, long-range pathways. This is achieved by infiltrating a fully amorphous poly (vinylidene fluoride‑co‑chlorotrifluoroethylene) (PVT) into an oriented electrospun fibrous framework of polar semi‑crystalline PVT. The framework serves as an "artificial crystalline phase," offering robust mechanical support and facilitating lithium‑salt dissociation. Simultaneously, the amorphous phase utilizes this microscale network to enable long-range ion conduction. The resulting SPEs exhibit an extremely high ionic conductivity of 1.23 mS cm-1 at 25°C, outperforming most reported all-polymeric-SPEs (10-7 ∼ 10-5 S/cm). Li//Li symmetric cells demonstrate stable cycling over 1300 h at 0.2 mA cm-2, in clear contrast to 60 h for the controlled cell. Furthermore, assembled high-voltage Ni0.8Co0.1Mn0.1O2 (NCM811)//Li full cells also deliver a stable cycling performance at 25°C. This work opens a new route for improving ion transport efficiency by constructing an artificial crystalline-amorphous structure.

A phosphorylated variant of the mast/stem cell growth factor receptor KIT is upregulated in dorsal root ganglia of Friedreich ataxia.

Arnulf H Koeppen, Joseph E Mazurkiewicz, Paul J Feustel +4 more

Friedreich ataxia (FA) causes hypoplasia of nerve cells in dorsal root ganglia (DRG). Beyond hypoplasia, however, the lesion in DRG includes disorganization and proliferation of satellite cells, formation of residual nodules, and neuronophagia. Antibody microarray analyses of DRG lysates with 878 validated antibodies that target cell signaling proteins reveal prominent up-regulation of the mast/stem cell growth factor (SCF) receptor-tyrosine kinase KIT. In its bioactive form upon engagement with SCF, KIT undergoes dimerization and autophosphorylation at Tyr-936 amongst other sites, and is hereafter called KIT-pY936. By immunohistochemistry and double-label laser scanning confocal immunofluorescence, KIT-pY936 is located in satellite cell cytoplasm. Electrophoresis and Western blots of DRG lysates show that the strong immunoreactivity of KIT-pY936 is due to a new truncated KIT variant of 50 kDa. KIT is a proto-oncogenic protein with prominent roles in hematopoiesis including mast cell proliferation. In conclusion, proteomic analysis confirms the prominent participation of a new truncated KIT in satellite cells in the pathogenesis of FA in DRG.

Primary Neuroendocrine Carcinoma of the Breast: A Diagnostic Challenge and Case Report.

Hamid Zeinali Nezhad, Shima Yaghoobi, Nazanin Zeinali Nezhad +3 more

Primary neuroendocrine carcinoma of the breast (NECB) represents an exceedingly rare entity, comprising less than 0.5% of all breast carcinomas and approximately 1% of all neuroendocrine neoplasms. The 2019 WHO classification defines NECB as tumors expressing neuroendocrine markers in > 90% of tumor cells with morphological features resembling neuroendocrine tumors of other anatomical sites.

Reply to the comment on: Differential effects of human fibromyalgia sera on murine satellite glial cells.

Francisco Mercado, Angélica Almanza, Laura-Aline Martínez-Martínez +1 more

Shaping the microvascular network: insights into skeletal muscle angiogenesis.

Thomas Gustafsson, Emmanuel Nwadozi, Andrea Tryfonos +1 more

Angiogenesis, the formation of new blood vessels from pre-existing vasculature, is a complex and tightly regulated biological process that plays a fundamental role in both physiological and pathological tissue remodeling by facilitating the delivery of oxygen and nutrients. Over recent decades, extensive research has identified a wide array of factors that regulate the balance between endothelial cell quiescence and activation. This review discusses the cellular events and molecular mechanisms that regulate angiogenesis within skeletal muscle, considering dynamic interactions with the extracellular matrix and highlighting the critical involvement of multiple resident and infiltrating cell types-including myofibres, satellite cells, fibro-adipogenic progenitors, immune cells and pericytes. The current understanding of these regulatory networks is examined in both healthy muscle tissue as part of the phenotype changes that occur during exercise and in pathological conditions that affect skeletal muscle angiogenesis. Particular attention is given to introduce data of emerging high-resolution techniques, especially omics-based approaches such as single-cell RNA sequencing (scRNA-seq) of skeletal muscle tissue. These methodologies hold significant promise for elucidating cell-type-specific roles and intercellular interactions that drive angiogenic processes in both physiological and disease contexts. Despite substantial progress, the precise mechanisms governing angiogenesis in skeletal muscle remain only partially understood.

In vivo base editing via single myotrophic adeno-associated viruses in dystrophic mouse muscle and satellite cells.

Kuan-Hung Lin, Amy Lam, Samuel van Ooijen +7 more

Duchenne muscular dystrophy (DMD) is the most common, lethal X-linked neuromuscular disorder of childhood and is caused by mutations in the Dmd gene that disrupt dystrophin expression. Although adeno-associated virus-mediated gene therapies hold tremendous promise for DMD treatment, their clinical applications have been limited by dose-dependent vector and genome-level toxicities. Here, we developed and tested a single-vector adenine base editing strategy as a potentially safer genome editing approach to recode the pathogenic nonsense mutation into a benign missense mutation in mdx 4cv DMD mouse model. Delivered using a muscle-tropic adeno-associated virus (MyoAAV) at a clinically-feasible dose (4E13 VG/kg), this strategy enabled detectable molecular recoding of the mdx 4cv mutation in mice ranging in age from 3 days to 6 months. Yet, the overall efficiency and therapeutic impact of in vivo base editing with this system was highest in mice treated at the juvenile stage, with animals administered MyoAAV vectors at 3 weeks of age showing robust recovery of dystrophin expression and significant improvement in muscle contractile properties only one month later. Notably, introduction of adenine base editors either earlier in development, in neonatal mice, or later, in adulthood, yielded substantially lower editing efficiencies, particularly in muscle satellite cells whose editing is essential to ensure durable rescue of dystrophin expression in growing and regenerating muscle. Taken together, these results demonstrate the therapeutic potential of single-vector adenine base editing for DMD and underscore the importance of recipient age and disease stage in achieving optimal treatment outcomes for this and other genetic muscle disorders.

Nuclear Speckles Regulate Splicing During Muscle Stem Cell Activation and Aging.

Steve D Guzman, Pamela Duran, Yu Xiao +4 more

Skeletal muscle contains a population of adult stem cells called satellite cells or muscle stem cells (MuSCs) that are responsible for regeneration after injury. MuSCs utilize gene expression programs to maintain quiescence and differentiate after injury and a key regulator of gene expression is splicing, which uniquely changes when transcripts interact with nuclear speckles (NS). NS are membrane-less biomolecular condensates that phase separate proteins, RNAs and chromatin, but how these organelles regulate molecular processes in MuSCs remains unknown. Herein, we build a comprehensive and systems-level understanding of NS influence on alternative splicing, transcriptional regulation and stem cell function before and after injury and in aging. We establish that NS increased in size and number in MuSCs following injury and influence MuSC activation dynamics. We generated a catalog of isoform-resolved splicing events and linked how RNA interactions with NS amplify splicing completion during the injury response. In old age, MuSCs lose NS, yet shifted towards longer, more completely spliced isoforms enriched for RNA binding protein motifs and multivalency. Our studies unveil evidence that RNA interactions with NS shape stem cell state and regenerative responses but are attenuated in old age.

A complete human pancreatic cancer genome.

Justin Wagner, Ayse G Keskus, Keisuke K Oshima +76 more

Cancer genome sequencing is essential for understanding tumor evolution and advancing precision medicine. 1 However, reference gaps and germline variants obscure detection of small and large somatic variants and methylation in repetitive regions. 1-3 It is common for tumor cells to gain or lose chromosome arms due to somatic structural changes that occur inside highly repetitive satellite DNA sequences in the centromeres. 4 To identify the full spectrum of somatic variants, including complex rearrangements, we construct and curate near-complete, haplotype-resolved assemblies of the most recent common ancestor of an early-passage broadly-consented hypodiploid pancreatic cancer cell line and matched normal tissues. The tumor assembly completely recapitulates all 35 tumor chromosomes observed with karyotyping, with multiple translocation-induced hybrid chromosomes. The hybrid chromosomes contain putative functional dicentric and fused centromeres, nested foldback inversions causing 14 breakpoints with a haplotype switch in a single event, and centromeric satellite tandem duplications up to 136 kbp. Direct comparison of tumor and normal assembly haplotypes uncovers >7,000 variants altering >1 Mbp of sequence in repetitive regions that have been hidden by reference gaps and germline variants. 44 % of somatic small variants change representation because they alter germline variants on GRCh38, impacting mutational signatures and kataegis/omikli clusters. Most somatic LINE insertions originate from two hypomethylated non-reference germline LINE insertions, highlighting their impact on insertion mutation burden. These assemblies demonstrate that centromeric, acrocentric, and telomeric regions conventionally excluded from analysis harbor extensive somatic and epigenetic changes. Resolving complete tumor genomes enables a deeper understanding of cancer structural plasticity and the endpoints of breakage-fusion-bridge cycles. These assembled, curated paired normal-tumor benchmarks will serve as a critical foundation for developing future algorithms to characterize the most intractable regions of cancer genomes.

The cellular ecosystem of skeletal muscle regeneration: molecular mechanisms, pathological disorders, and potential therapeutic strategies.

Jiahuan Gong, Hongyi Xu, Xinlei Yao +5 more

Skeletal muscle regeneration is a highly coordinated physiological process. It relies on the intricate collaboration of a complex cellular ecosystem. This ecosystem includes muscle stem cells, immune cells, stromal cells, vascular cells, neural cells, and the extracellular matrix. Research has recently expanded beyond focusing solely on satellite cells. It now delves into the multi-level regulatory networks within this ecosystem. These networks encompass key signaling pathways, such as Wnt/β-catenin, TGF-β, Hippo/YAP, and AMPK. They also include epigenetic regulation, cellular metabolic reprogramming, and extracellular vesicle-mediated intercellular communication. However, under pathological conditions, this regenerative program is severely impaired. This leads to failed repair, fibrosis, and fatty infiltration, ultimately resulting in loss of muscle function. This review aims to systematically outline recent advances in the field of skeletal muscle regeneration. First, from the perspective of the "cellular ecosystem," we will elaborate on the dynamic behaviors and regulatory mechanisms of various cell types during regeneration. Second, we will dissect the core mechanisms underlying regenerative failures in various pathological states. Third, we will comprehensively evaluate the most promising current intervention strategies. Finally, considering the limitations of current research, we will provide future perspectives. This review aims to systematically integrate existing knowledge and provide a clear roadmap for future research, ultimately offering a robust theoretical foundation and innovative insights for the development of clinical treatments targeting skeletal muscle regenerative disorders.

FAS-controlled T cells drive lymphoproliferation through glycolysis without effector differentiation.

Maria Elena Maccari, Christoph König, Geoffroy Andrieux +38 more

Lymphoproliferation in autoimmune lymphoproliferative syndrome (ALPS) due to FAS deficiency is driven by highly proliferative FAS-controlled T cells (FCT) with a distinct molecular signature. Activating signals and metabolic fuels of their proliferation are poorly understood. Lymphoproliferation caused by proliferative T cells is also a hallmark of acute EBV infection. In these antiviral T cells, a metabolic switch to glycolysis underpins effector differentiation and IFNγ translation. Here, we used EBV-induced CD8 effector T cells as a benchmark to characterize FCT metabolism. Metabolic assays, RNA sequencing, and in silico computational analysis revealed that FCT are as highly glycolytic as EBV-induced effector T cells, but this metabolic program is uncoupled from T-BET expression and IFNγ production. In contrast to virus-activated T cells, FCT showed mitochondrial hyperpolarization and elevated reactive oxygen species production. These findings support a model of FCT lymphoproliferation, in which activating signals strongly enhance glycolysis but do not induce classical effector differentiation.

Designing the MyoFusion Media: A Serum-Free Medium Optimized for Bovine Satellite Cell Differentiation.

Aysenaz Tavsanli, Viktor Milkevych, Jette Feveile Young +1 more

Cultivated meat production requires efficient differentiation of muscle progenitor cells into myotubes without relying on animal-derived serum, which poses ethical and scalability challenges. This study aimed to develop a chemically defined, serum-free medium optimized for bovine satellite cell differentiation. Using a data-driven design of experiments approach, we systematically screened 13 growth factors and supplements previously associated with myogenesis. Fusion index served as the primary metric for differentiation efficiency, complemented by additional morphological traits. Initial screening identified platelet-derived growth factor BB (PDGF-BB), cytosine arabinoside, and linoleic acid as key contributors, with transforming growth factor beta (TGF-β) included for its biological relevance. Subsequent optimization employed full factorial and central composite designs combined with response surface modeling to refine concentration ranges and evaluate interactions. The resulting formulation, termed MyoFusion, consists of DMEM supplemented with 4.5 × 10⁻⁶ mg/mL TGF-β1, 4.66 × 10⁻⁶ mg/mL PDGF-BB, 0.73 × 10⁻⁵ mg/mL cytosine arabinoside, and 2.07 × 10⁻⁴ mg/mL linoleic acid. Experimental validation demonstrated that MyoFusion achieved a fusion index of 65.08 ± 4.97, representing an improvement of up to 52.7% compared to serum-based controls (2% FBS). Correlation analyses confirmed strong associations between fusion index and other differentiation traits under serum-free conditions, indicating robust myotube formation. These findings establish MyoFusion as a promising serum-free alternative for cultivated meat applications, supporting ethical and scalable production while maintaining differentiation efficiency comparable to or exceeding traditional serum-containing media.

Glutamine-driven reductive TCA cycle metabolism supports aged muscle stem cell function via de novo lipogenesis.

David E Lee, Lauren K McKay, Akshay Bareja +8 more

Sarcopenia and the age-related decline in muscular strength and regenerative capacity contribute directly to loss of autonomy, greater risk for hospitalization and healthcare utilization. One contributing cellular phenotype associated with skeletal muscle aging is a loss in the function and number of resident muscle stem cells (MuSCs) or satellite cells. MuSC activation leads to dramatic changes in cellular architecture and metabolic reprogramming, including both mitochondrial biogenesis and increased glycolysis. Despite these changes to increase energy production, high energy demands may not be fully met during periods of MuSC activation. Here we used in vitro and in vivo approaches in mice to demonstrate the function of glutaminase for age-related changes in MuSC function. By combining fluorescence-activated cell sorting (FACS) isolation with metabolomics and stable isotope tracing, we show an age-related decline in reductive (counterclockwise) flux of glutamine through the tricarboxylic acid (TCA) cycle, a pathway by which MuSCs build cellular fatty acid stores as necessary biomass for MuSC function.

Phage satellites induced by virulent phages are mobilized by natural competence leading to phage resistance in a new host.

Carlee Morency, Geneviève M Rousseau, Zacharie Morneau +1 more

A phage satellite (PS) typically resides within repeat regions (attL and attR sites) of a bacterial genome. Its genome ranges from 7 to 20-kb and includes genes encoding an integrase along with regulatory and DNA replication functions. However, it lacks genes associated with viral structural proteins. Streptococcus thermophilus (S.t.) is extensively used to produce yogurt and specialty cheeses. Intriguingly, the majority of S.t. strains harbor a PS while very few possess a complete prophage, suggesting that PSs may confer advantages to their hosts. In this study, we showed that PSs of S.t. can excise from the bacterial chromosome, at a very low rate, without any phage interaction. Furthermore, we found that they can also be induced by virulent phages. By leveraging CRISPR-Cas9, we selected S.t. cells devoid of any PS (delta-PS strain). Then, we mobilized a PS from one strain to a delta-PS strain, using only natural competence, bypassing the need for a helper phage. The resulting strain exhibited increased resistance to virulent phages. Through the isolation of phage mutants escaping the resistance phenotype, we pinpointed a specific phage protein responsible for the induction of a PS. Lastly, we demonstrated that a PS can be significantly induced by a virulent phage, which, in turn, greatly promote their transfer and specific integration into new cells through natural competence. Our study introduces a novel natural approach to develop phage-resistant strains.

Modulation of Bovine Muscle Satellite Cells Myogenesis by Paracrine Fibroblast-Derived Signaling in Three-Dimensional Spinner Flask Culture System.

Karolina Zygmunt, Katarzyna Piórkowska, Julia Adamiak +1 more

Studies examining the paracrine effect of fibroblasts on the myogenesis of bovine muscle satellite cells (MuSCs) have confirmed their stimulating effect on proliferation and early differentiation. However, traditional two-dimensional (2D) cell culture models fail to accurately represent the complexity of in vivo muscle tissue. This study aims to investigate the paracrine effect of fibroblasts on myogenesis in a three-dimensional (3D) cell culture model. Cells were cultured in monoculture and co-culture with fibroblasts in a spinner flask using gelatin microcarriers, Cultishper, which maximizes the growth surface area for adherent cells. Then, muscle cells from the co-culture were sorted by the FACS method based on negative expression of CD90, a fibroblast-associated marker. The progress of myogenesis was assessed based on qPCR analysis for selected muscle markers and at the protein level for skeletal myosin. Fluorescent staining and luminescent metabolic assays were performed to control culture conditions. The obtained results revealed up-regulation of genes involved in cell activation and proliferation (PAX7, MYF5, and MYOD) and differentiation (MYOG, EGR1, and MYH). Metabolism analyses did not show changes between mono- and co-culture conditions. To summarize, fibroblasts through paracrine signaling promote early differentiation of MuSCs and potentially proliferation, which provides valuable insights for the advancement of cultured meat production.

Isolation and Therapeutic Application of Extracellular Vesicles Derived from Self-Assembled Mesenchymal Stem Cell Aggregates.

Jun-Xi He, Si-Yu Lan, Li-Heng Ren +10 more

Extracellular vesicles (EVs) are vital mediators of intercellular communication and promising cell-free agents for regenerative medicine. Among EV subpopulations, large extracellular vesicles (LEVs) are particularly suited for tissue repair due to their enriched pro-regenerative cargo. However, standardized and reproducible methods to generate functional dental stem cell-derived LEVs for downstream characterization and therapeutic evaluation remain limited. Utilizing the exceptional osteogenic potential of stem cells from the apical papilla (SCAPs), we developed a 3D culture model under low-adhesion conditions to induce self-assembled SCAP aggregates. This model mimics physiological mesenchymal condensation, significantly enhancing cell-cell interactions and boosting LEV secretion. Key culture parameters and handling notes are provided to improve aggregate consistency and maximize LEV output while maintaining cell viability. Here, we present a standardized protocol for the efficient production of functional SCAP-derived LEVs, encompassing 3D aggregate generation, isolation via differential centrifugation, nanoparticle tracking analysis (NTA)-based particle size distribution and concentration analysis, and therapeutic delivery using a thermally crosslinked hydrogel for mandibular bone defect repair. This approach streamlines LEV production for downstream mechanistic studies and preclinical evaluation in craniofacial regeneration. This optimized workflow ensures the reproducible preparation of bioactive LEVs with consistent yields and enriched pro-osteogenic cargo, facilitating fundamental research and accelerating the clinical translation of EV-mediated craniofacial reconstruction. In representative applications, hydrogel-loaded LEVs provide a localized depot at defect sites and facilitate convenient handling during implantation. Collectively, this method provides a scalable platform to prepare SCAP-derived LEVs and highlights key considerations for standardized reporting and translational study design. This approach supports broader adoption across craniofacial EV laboratories.

Glial cells in neuropathic pain.

Temugin Berta, Arkady Khoutorsky, Jing Xu +1 more

Neuropathic pain is a chronic, debilitating condition that affects approximately 78% of the global population and remains difficult to treat due to limited therapeutic options. Over the past two decades, mounting preclinical evidence has highlighted the crucial role of glial cells in the development, maintenance, and resolution of neuropathic pain. Conditions such as peripheral nerve injury are associated with reactive responses of glial cellsnamely microglia and astrocytes in the central nervous system, and satellite glial cells in the peripheral ganglia. Under physiological conditions, glial cells contribute to homeostatic regulation of spinal nociceptive circuits, preserving normal pain sensitivity. However, neuropathic pain is increasingly recognized as a form of gliopathy, wherein dysregulated glial activity contributes to pain hypersensitivity through enhanced excitability of primary sensory neurons (peripheral sensitization) and neuronal plasticity in the central pain circuits (central sensitization). In this review, we provide a comprehensive overview of glial cell research in neuropathic pain, with a specific focus on microglia, astrocytes, and satellite glial cells, the most extensively studied glial types in this context. We also highlight the critical involvement of glial cells in neuroinflammation and neuro-glial interactions and evaluate emerging therapeutic strategies targeting glial mechanisms of neuropathic pain.

Sodium tanshinone IIA sulfonate modulates proliferation and differentiation of human skeletal muscle satellite cells via the PI3K/AKT pathway.

Zongyu Zhang, Zhijing Zhou, Peng Zhang +2 more

Human skeletal muscle satellite cells (HSkMSCs) are the primary stem cells responsible for skeletal muscle regeneration and repair. The balance between their proliferation and differentiation is essential for maintaining muscle homeostasis. Sodium tanshinone IIA sulfonate (STS) has been reported to possess anti-inflammatory, antioxidant, and antifibrotic effects. However, the regulatory role and underlying mechanisms of STS in governing the fate of HSkMSCs remain largely unexplored. This study aimed to investigate the effects of STS on the proliferation and differentiation of HSkMSCs and to elucidate the potential molecular mechanisms. HSkMSCs were treated with different concentrations of STS. Cell proliferation was assessed using EdU assay. The expression levels of the proliferation marker Pax7 and the myogenic differentiation-related genes MyoD and fast-twitch skeletal myosin heavy chain (MyHC -II) were determined by RT-qPCR and western blotting. To verify the involvement of PI3K/AKT signaling pathway, the inhibitor LY294002 was co-administered with STS to further evaluate the regulatory role of this signaling in mediating the biological effects of STS. EdU staining revealed that STS significantly reduced the proliferation of HSkMSCs in a dose-dependent manner. STS markedly downregulated the expression of Pax7, while upregulating the expression of MyoD and MyHC-II. Furthermore, STS treatment significantly enhanced the phosphorylation levels of PI3K and AKT. Notably, co-treatment with LY294002 effectively attenuated the regulatory effects of STS on Pax7, MyoD, and MyHC-II expression. In conclusion, STS inhibits the proliferation and upregulates myogenic differentiation markers expression in HSkMSCs by activating the PI3K/AKT signaling pathway.

Impacts of genetic selection on satellite cell function in poultry.

Joseph Yimiletey, Sandra G Velleman, Hui Yu

Over the past several decades, intensive genetic selection has markedly increased growth rate, breast muscle yield, and production efficiency in commercial broilers. These gains have been accompanied by substantial changes in muscle organization, physiology, metabolism, and cellular regulation. Satellite cells, the resident muscle stem cells, play a key role in posthatch muscle growth by supplying nuclei to adjacent growing myofibers, thereby supporting the muscle hypertrophy process. This review summarizes the role of satellite cells in poultry muscle development and discusses how genetic selection for rapid muscle growth has reshaped satellite cell proliferation, differentiation, molecular profiles, and functional heterogeneity, with important consequences for muscle growth and quality. Moreover, increasing evidence suggests that changes in satellite cell function associated with genetic selection may contribute to the development of breast muscle myopathies, particularly wooden breast in broiler chickens. Moving forward, continued refinement of poultry production systems will benefit from deeper insights into satellite cell biology to support efficient muscle growth while reducing the occurrence of muscle myopathies.

Single-cell transcriptional landscape of muscle-derived stem/progenitor cells reveals hallmarks of aging and rejuvenation.

Kavitha Mukund, Seth David Thompson, Chelsea L Rugel +5 more

Muscle-derived stem/progenitor cells (MDSPCs) are an adult stem cell population with demonstrated regenerative and rejuvenative potential distinct from other muscle progenitor cells. However, their molecular identity and developmental status remain poorly defined. Using single-cell transcriptomics and proteomics, we comprehensively profiled murine MDSPCs across age groups. We show that MDSPCs exist along a transcriptional continuum of maturation-ranging from metabolically active, proliferative early-stage cells to late-stage, lineage-committed myogenic populations. While lacking canonical pluripotency markers, early-stage MDSPCs express gene programs associated with embryonic progenitor identity, suggesting a non-canonical, multipotent-like state. These features distinguish them from both satellite cells and committed myoblasts. Aging reshapes this continuum by reducing stemness-associated signatures while enhancing differentiation programs and oxidative stress. Our identification of distinct MDSPC states provide critical insights into mechanisms that underly tissue regeneration and aging. These findings offer a blueprint for development of future regenerative therapies to combat age-related functional decline.

Curcumin attenuates uterine pain in mice through suppression of neuroinflammation in the DRG and spinal cord.

Ya-Ru Yang, Ji-Tao Tang, Bing-Qiang He +4 more

Uterine pain associated with labor, dysmenorrhea, or endometriosis is often driven by inflammation, which enhances nociceptive signaling and contributes to hyperalgesia. Although nonsteroidal anti-inflammatory drugs (NSAIDs) and glucocorticoids are commonly prescribed, their clinical application is limited by side effects, and a subset of patients exhibit inadequate or no response to NSAID therapy. Curcumin, a natural polyphenol extracted from Curcuma longa, exhibits well-documented anti-inflammatory and analgesic properties. In this study, we investigated the analgesic efficacy and underlying mechanisms of curcumin in a mouse model of uterine pain. Both intraperitoneal and intrathecal administration of curcumin significantly reduced writhing responses and improved locomotor performance in open-field tests. Molecular analyses revealed that estrogen and oxytocin treatment induced activation of glial cells in the dorsal root ganglia (DRG) and spinal cord. Intrathecal curcumin attenuated the activation of satellite glial cells and macrophages in the DRG, suppressed MAPK signaling (ERK, JNK, and p38), and decreased the expression of proinflammatory mediators, including IL-6, TNF-α, IL-1β, CCL2, and CXCL1. Similarly, in the spinal cord, curcumin reduced astrocyte and microglia activation, MAPK phosphorylation, and inflammatory cytokine and chemokine levels. Moreover, curcumin diminished p-ERK expression in DRG neurons and reduced c-Fos expression in the spinal dorsal horn. Collectively, these findings demonstrate that curcumin alleviates uterine pain by suppressing glial activation, MAPK signaling, and inflammatory mediators' production, ultimately reduces markers of neuronal activation and plasticity in the DRG and spinal cord. This provides mechanistic support for curcumin's potential in managing uterine pain.

One half of the sprinting coin: The oxygen transport cascade of a 91-year-old female world-record sprinter.

Marta Colosio, Andrea M Pilotto, Massimiliano Ansaldo +8 more

A 91-year-old female set the W90+ world record in the 200m sprint in 2024 surpassing her previous record by 1.13s. This study characterized her cardiorespiratory fitness, skeletal muscle oxidative capacity, fiber type distribution, capillarization, and satellite cells and compared these outcomes, where possible, to published reference data. Cardiorespiratory responses were assessed during a ramp cycling test to exhaustion, and muscle oxidative capacity (mV ̇O₂) was determined using near-infrared spectroscopy during repetitive arterial occlusions. Fiber type distribution, cross-sectional area, capillarization, satellite cell content and localization, and mitochondrial respiratory capacity were analyzed in a vastus lateralis biopsy. Peak oxygen uptake (V ̇O₂peak) was 23ml·kg⁻¹·min⁻¹ at 98W, with a maximal heart rate of 141beats·min⁻¹ and cardiac output of 13.6L·min⁻¹. The mV ̇O₂ recovery rate constant (k) was 1.83min⁻¹. Fiber composition was 57% fast MyHC II fibers (50%IIa, 5%IIa-IIx, 2%IIx), 38% slow MyHC I fibers and 5% hybrid I-IIa fibers. MyHC I fibers were larger, more vascularized and had satellite cells located closer to capillaries (4267±2181μm²; 1.67 individual capillary-to-fiber ratio (C/Fi), 5.58 capillaries×1000μm-1 capillary-to-fiber perimeter exchange index (CFPE), 1.8µm satellite cell-to-capillary distance) than MyHC II fibers (2752±1608μm²; 1.03 C/Fi, 4.00capillaries×1000μm-1 CFPE and 10.4µm, respectively). Mitochondrial O₂ flux was 58 and 68pmol·(s·mg)⁻¹ during coupled and uncoupled respiration, respectively. The athlete's cardiorespiratory and oxidative capacity resembled those of females' in their 50s or younger. Collectively, her large, well-vascularized slow fibers, high proportion of fast fibers, and preserved muscle oxidative capacity likely contributed to her world record performance, illustrating the remarkable plasticity of skeletal muscle even in very advanced age.

Identifying Quiescent Satellite Cells: A Scoping Review of Transcriptomic Markers and Limitations.

Anika L Syroid, Alexandra P Steele, Kevin A Murach +1 more

Skeletal muscle regeneration relies on the resident stem cell population, termed satellite cells. Mechanistically, understanding the quiescence and activation dynamics of muscle satellite cells are essential for regenerative therapies and emerging applications such as cellular agriculture. Quiescent satellite cells (QSCs) are typically identified by expression of PAX7 and functional characteristics including a lack of proliferation. However, with the rapidly growing body of transcriptomic data, there is a lack of consensus regarding what markers can be used to identify quiescent satellite cells across transcriptomic studies. The purpose of this review was to evaluate the transcripts currently used to identify QSCs using transcriptomics and to establish an evidence-based foundation that could be used for future analyses. After surveying published single-cell transcriptomic studies, we identified Pax7 and/or Myf5 as the most used markers of general satellite cell identity, while Spry1, Cd34, and Calcr, together with the absence of Myod1, Mki67, and Cdk1 were most commonly used to identify QSC clusters in murine studies. In contrast, there is currently insufficient literature to make a confident conclusion on quiescence markers in larger mammals, including humans, pigs, and cattle. We also highlight the conceptual and technical challenges associated with transcriptomic analysis of satellite cell subpopulations, including continuum-based cell states, isolation induced transcriptional changes, and inconsistent terminology. As a field, greater consistency in language, standardized analyses, and cross-species validation will be required to progress the study of satellite cell quiescence and its translational utility.

Chondrolectin regulates the sublaminar localization and regenerative function of muscle satellite cells in mice.

Lijie Gu, Kun Ho Kim, Xiyue Chen +5 more

Skeletal muscle satellite cells (SCs) reside between the myofiber sarcolemma and basal lamina, where extracellular matrix (ECM) interactions maintain stemness and regenerative function. Here, we identify chondrolectin (CHODL), a type I transmembrane protein with a C-type lectin domain, as a critical regulator of SC biology. Single-cell RNA-seq analysis reveals that Chodl is highly enriched in quiescent SCs but downregulated in proliferating myoblasts. The conditional deletion of Chodl in embryonic myoblasts (Chodl MKO ) or adult SCs (Chodl PKO ) leaves muscle development intact yet delays injury-induced regeneration in young and aged mice. Chodl-deficient SCs exhibit reduced self-renewal and diminished proliferation, leading to defective myofiber repair. In silico network perturbation further predicts disrupted ECM-ligand interactions and Notch signaling, consistent with SC mislocalization outside the basal lamina and precocious activation in Chodl PKO muscle. Together, these findings establish CHODL as a determinant of SC niche localization and function, linking ECM interactions to muscle stem cell maintenance and repair.

Curcumin Alleviates Bone Cancer Pain by Inhibiting Satellite Glial Cell Activity via Janus Kinase 1/Signal Transducer and Transcription 3 Pathway Activator.

Xinchao Jiang, Yi Song, Mei Fang +4 more

Curcumin is the main active component of Curcuma longa L and has anti-inflammatory, antitumor, and neuroprotective properties, making it a potential candidate for bone cancer pain management. Network pharmacology, molecular docking, and molecular dynamics simulations were used to screen and validate core therapeutic targets of curcumin in bone cancer pain. An in vivo mouse model of bone cancer pain was established via intrafemoral injection of Lewis lung cancer cells. A series of in vivo assays was conducted to evaluate pain sensitivity, bone microstructure, inflammatory cytokines, pain-related neuropeptides, and protein expression levels. In silico analysis verified that curcumin has a strong binding affinity for JAK1 and STAT3. In vivo results demonstrated that curcumin dose-dependently elevated the paw withdrawal threshold and latency, as well as ameliorated bone mineral density and bone destruction. Curcumin also suppressed pro-inflammatory cytokines, substance P, calcitonin gene-related peptides, and inhibited satellite glial cell activation by blocking the JAK1/STAT3 pathway. Notably, JAK1 agonist (RO8191) co-administration markedly reversed the analgesic effects of curcumin. Curcumin targets the JAK1/STAT3 signaling pathway, inhibits satellite glial cell activation, downregulates pro-inflammatory cytokine release, and pain-related neuropeptide expression. Therefore, curcumin exerted a significant analgesic effect against bone cancer pain. Elucidating the analgesic mechanism revealed the core therapeutic targets of curcumin in treating bone cancer pain. This study provides experimental evidence for further research on bone cancer pain.

Overexpression of IGF2 Alters the Transcriptional Profile of Goose Skeletal Muscle Satellite Cells.

Cui Wang, Yi Liu, Yunzhou Yang +2 more

Insulin-like growth factor 2 (IGF2) plays a pivotal role in regulating growth and development; however, its functional involvement in skeletal muscle satellite cells (SMSCs) remains incompletely understood. To elucidate the regulatory role of IGF2, goose SMSCs were engineered to overexpress IGF2 via lentiviral transduction, followed by comprehensive transcriptomic profiling. Comparative analysis revealed 2802 differentially expressed genes (DEGs) in IGF2-overexpressing cells relative to controls, comprising 1202 upregulated and 1600 downregulated genes. IGF2 overexpression markedly activated fibrogenic programs, as evidenced by the upregulation of AP-1 complex components (FOS, JUN), extracellular matrix-related genes (COL1A1, COL5A3), and Wnt signaling receptors (FZD1, FZD7). In contrast, genes involved in myogenic differentiation and contractile function were broadly suppressed, including key myogenic transcription factors (MEF2C, MEF2D), sarcomeric structural proteins (MYBPC1, ACTN2, MYOM3), and metabolic enzymes. Through the construction of protein-protein interaction networks coupled with functional enrichment analysis, we observed a concerted suppression of myogenic regulatory networks critical for myofiber formation. Quantitative real-time PCR validation further confirmed the reliability of the transcriptomic data. Collectively, these findings suggest that overexpression of IGF2 induces a phenotypic shift from myoblasts toward a fibroblast-like state, uncoupling proliferation from differentiation while enhancing fibrogenic identity. This study provides novel insights into IGF2-mediated regulatory mechanisms underlying skeletal muscle development and fibrotic processes.

Skeletal Fiber Type in Muscle Pain and Dysfunction.

Maria Lopes Cardia, Bruno Daniel Carneiro, Isaura Tavares +1 more

Different types of skeletal muscle fibers display marked heterogeneity in metabolic, mechanical, and regenerative properties. However, their role in chronic musculoskeletal pain remains insufficiently integrated into clinical models. Chronic pain is associated with altered neuromuscular control, prolonged low-level activation, and reduced recruitment of high-threshold motor units. These factors may promote fiber type-specific remodeling. This narrative review critically synthesizes current evidence on the relationship between musculoskeletal pain and muscle fiber types. The focus was on metabolic vulnerability, mechanical susceptibility, and regenerative capacity. A structured literature search was conducted in PubMed, Scopus, and Web of Science, focused on human studies and key translational models. Chronic musculoskeletal pain is characterized by acquired fiber type-specific adaptations rather than a fixed unfavorable profile. In chronic pain scenarios, Type I fibers present features of chronic overload, including hypertrophy with insufficient capillarization and increased satellite cell activity. Type II fibers exhibit relative disuse, atrophy, and reduced satellite cell content, resembling accelerated muscle aging. Symptom duration, neuromuscular control strategies, and task-specific loading patterns modulate these adaptations, with interindividual variation. Muscle dysfunction in chronic pain reflects maladaptive but potentially reversible neuromuscular and histological plasticity. These findings indicate that rehabilitation strategies should be individualized, involving context-specific exercise strategies to restore muscle structure, function, and regenerative potential in chronic musculoskeletal conditions.

Time-of-day, satellite cells, and velocity collectively influence ex vivo isovelocity force production in mouse extensor digitorum longus muscle.

Ryan E Kahn, Sudarshan Dayanidhi, Richard L Lieber

Skeletal muscles are exquisitely designed to produce force that facilitate movement. Circadian "molecular clocks" residing in muscle play a role in regulating force production with muscle stem cell (satellite cells, SC) molecular clocks modulating isometric and eccentric force according to time-of-day. However, many tasks of daily living and exercise (i.e., walking/running) involve force and power produced during muscle shortening. Thus, the purpose of this study was to determine whether isovelocity forces and power are also modulated by SCs according to time-of-day. Using previously published samples (a mouse model capable of SC ablation), we evaluated isovelocity forces across a range of velocities (1-11 Lf/s) at two different times of day ZT1, ZT9 in the presence and absence of SCs. The main finding of this investigation was that isovelocity force production is regulated by a third-order interaction effect between time-of-day × SCs × velocity (p < 0.001). Additionally, a significant effect of time-of-day was observed for isovelocity force and power when comparing ZT1 vs. ZT9 SC+ mice whereas this effect was absent in SC- animals. These results suggest SCs harbor a time-of-day and velocity dependent effect on isovelocity force production and power. Further work is required to elucidate the underlying mechanisms of this phenomenon.

Ageing-Associated Dysregulation of Myogenic Differentiation in Inclusion Body Myositis.

Geert M de Vries, Willem De Ridder, Jonathan Baets

Skeletal muscle is a postmitotic tissue dependent on a complex and tightly regulated regeneration process involving numerous intracellular and extracellular factors, including myogenic regulatory factors (MRFs), cytokines and myokines. Quiescent satellite cells are activated by physiological stimuli, injury or other traumatic insults for the repair of injuries or growth of the tissue. Activation of satellite cells induces proliferation and expression of MRFs, which in turn activate myogenic differentiation transcription programmes. Transitioning into and committing to terminal differentiation are regulated by myogenin and cell cycle exit markers, notably Rb1 and p21. Differentiation is then complete with the formation of new muscle fibres which incorporate into existing fibres. Upon ageing, the efficiency of differentiation is reduced as a consequence of a loss in the physiological balance between pathways regulating satellite cell quiescence and activation, notably the Notch and Wnt pathways, and increased senescence of the satellite cell pool. Extracellular factors involved in the dysregulation of differentiation upon ageing include low-grade chronic inflammation and remodelling of the extracellular matrix by fibro-adipogenic progenitor cells, thereby negatively affecting the differentiation capacity of satellite cells, resulting in either premature differentiation or senescence. These ageing-associated alterations in muscle homeostasis appear to be amplified in inclusion body myositis (IBM), an idiopathic inflammatory myopathy that almost exclusively manifests in individuals over 45 years of age, making it a prototypical age-related muscle disease. IBM is characterised by chronic inflammation, progressive muscle degeneration and premature ageing of both muscle tissue and the satellite cell niche. Studied with immunohistochemical techniques and multi-omics, muscle biopsy tissue demonstrated increased expression of MRFs as well as increased expression of senescence and genomic stress markers. IBM primary myoblasts demonstrated premature ageing and senescence and increased activity of the Wnt pathway, though differentiation into multinucleated myotubes did not show notable aberrations in signalling pathways or differentiation efficiency. In conclusion, ageing and chronic inflammation lead to dysregulation of key pathways that, in turn, alter the capacity of satellite cells to activate and proliferate, leading to prematurely aged satellite cells that still retain their capacity to differentiate into myofibres. Though in IBM there is an increased abundance of active differentiation markers, reflecting a regenerative response to the massive, sustained muscle atrophy, senescence of the satellite cell niche may impair effective regeneration of the lost muscle tissue.

Cadmium-induced stress memory regulates Bacillus subtilis transport in porous media: Evidence from multi-scale analysis.

Yijun Mo, Jialin Huang, Zihan Xiao +8 more

Accurately predicting microbial transport under environmental stress in subsurface and aquatic environments is essential for assessing microbial risks, from modeling pathogen spread to optimizing in-situ bioremediation. While the biological foundations of microbial heavy metal tolerance are established, the effects of toxic heavy metals like cadmium (Cd) on microbial-mineral interactions and subsequent transport mechanisms remain poorly understood. To address these challenges, we explored how Cd stress reconfigures the deposition behavior of Bacillus subtilis in porous media. We report, for the first time, that Cd pre-exposure induces a "stress memory" that attenuates the acute escape response from toxic surfaces, thereby enhancing deposition. We employed a multi-scale approach, integrating column experiments, quartz crystal microbalance with dissipation (QCM-D), whole-genome resequencing, and proteomics. This integrated analysis determined that the history-dependent shift from active avoidance to tolerant colonization may be driven by enhanced EPS secretion (Increased by 100.0-139.0%) and a reprogrammed signal transduction pathway (increased from 0 to 5). Consequently, B. subtilis subjected to higher prior Cd stress exhibited more attenuated migration responses, resulting in greater deposition and colonization while encountering Cd on the contact surface, with maximum deposited mass being 49.1-91.2% higher than that of the unstressed cells. These findings highlight the necessity of incorporating microbial active behavior and stress adaptation strategies into existing transport theory and provide insights for the prevention and control of pathogenic bacteria in polluted environments.

iPSC-derived skeletal muscle spheroids for Duchenne Muscular Dystrophy modeling.

Joyce Esposito, Felipe de Souza Leite, Igor Neves Barbosa +10 more

The progressive skeletal muscle degeneration observed in Duchenne Muscular Dystrophy (DMD) patients requires multiple cycles of satellite cells (SCs) activation to promote tissue regeneration. Dystrophic SCs present intrinsic defects, and the disrupting fibrotic niche hinders appropriate muscle recovery. Traditional 2D culture systems face challenges in modeling the DMD muscle niche and SCs behavior. Our aim was to validate a 3D culture of skeletal muscle spheroids (iSMS) for DMD modeling, as compared to the traditional 2D culture, while investigating the pathophysiological mechanisms of dystrophin deficiency in vitro.

Uncovering immune dysfunction in ACLF: cellular mechanisms, molecular pathways, and therapeutic frontiers.

Marti Ortega-Ribera, Robert Brenig, Christine Bernsmeier +1 more

Acute-on-chronic liver failure (ACLF) is a life-threatening condition characterized by acute hepatic decompensation, multi-organ failure, and high short-term mortality in patients with liver cirrhosis. A hallmark of ACLF is profound deterioration of the immune system, which contributes to organ-specific excessive inflammation and immune dysfunction, predisposing patients to infection and multi-organ failure. This review aims to elucidate the cellular and molecular mechanisms underlying systemic immune dysfunction in ACLF, highlighting key pathophysiological pathways and their clinical significance. We provide an overview of ACLF including its global prevalence and clinical significance, against the background of the underlying immune dysfunction in its pathogenesis. The discussion focuses on innate immune alterations, such as impaired neutrophil and monocyte phagocytosis, excessive neutrophil extracellular trap (NET) formation, and monocyte/macrophage dysfunction contributing to immuneparesis and exaggerated inflammation, respectively, which evolve in an organ-specific manner. Dysregulation of natural killer (NK) cell cytotoxicity and adaptive immune dysfunction, including changes in T cell subpopulations and B cell antibody production in ACLF, are discussed. We further dissect the emerging evidence of molecular pathways driving dysfunction of immune cells and their impaired ability to control infections in ACLF, emphasizing the roles of pathogen- and damage-associated molecular patterns (PAMPs/DAMPs), toll-like receptor (TLR) signaling, oxidative stress, mitochondrial dysfunction, epigenetic/metabolic reprogramming and immune checkpoint molecules. The review expands on immune cell communication within the immune system (innate and adaptive), with other non-parenchymal and parenchymal cells and at the inter-organ level, detailing interactions between immune cells of key organs and compartments affected during ACLF, including the liver, circulation, brain, gut and kidney. Finally, we summarize the latest preclinical and clinical findings exploring biomarkers of immune dysfunction and immunomodulatory therapeutic strategies aimed at restoring immune homeostasis in patients with ACLF.

Damage-induced i-loops generate eccDNA from repetitive elements.

Elia Zanella, Michele Giannattasio, Sara Bisi +2 more

Extrachromosomal circular DNA (eccDNA) drives genome instability and tumorigenesis, warranting thorough investigations of its biogenesis. Here, we report a mechanism of eccDNA formation in human cells, distinct from the one mediated by joining of broken DNA ends. We show that repeats such as telomeres and centromeric alpha satellite form internal loops (i-loops) as a consequence of single-stranded DNA (ssDNA) breaks or gaps rather than double-stranded breaks (DSBs). I-loops are precursors for the excision of eccDNA, visible by electron microscopy (EM) and detectable via rolling-circle amplification. Apoptosis triggers the formation of i-loops and eccDNA at telomeric and alpha satellite repeats. Nanopore sequencing revealed other repetitive elements, including rDNA and retrotransposons, as sources of eccDNA in apoptosis. Based on the prevalence of SSBs over DSBs and the abundance of repeats in the human genome, we propose that the i-loop mechanism contributes substantially to all forms of eccDNA, with implications for tumor biology and genome evolution.

γ-Aminobutyric acid enhances myogenesis and heat/cold stress resistance in bovine muscle satellite cells.

Abid Manzoor, Sajida Naseem, Zhiqi Fu +5 more

γ-Aminobutyric acid (GABA) is the principal inhibitory neurotransmitter in the central nervous system and is involved in the development of neural tissue as well as the regulation of its functions. Meanwhile, GABA has also been demonstrated to confer multiple physiological benefits, including alleviating stress and improving metabolic homeostasis. This study investigated GABA effects on proliferation, differentiation, and temperature stress protection of bovine skeletal muscle satellite cells (BSCs).

Mechanobiological landscape of muscle stem cells.

Kotaro Hirano, Yuji Hara

Pervasive and programmed nucleosome distortion on single chromatin fibres.

Marty G Yang, Hannah J Richter, Simai Wang +19 more

Despite decades of biochemical and structural studies of the nucleosome1, researchers lack genome-scale methods to determine variability in nucleosome structure along individual chromatin fibres. To address this, here we present Iteratively Defined Lengths of Inaccessibility (IDLI), a computational method that maps the single-molecule co-occupancy of structurally distinct nucleosomes, subnucleosomes and other protein-DNA interactions through long-read single-molecule footprinting2,3. IDLI classifies methylase-inaccessible footprints on individual chromatin fibres into (i) linker-histone-associated nucleosomes; (ii) nucleosomes with focal DNA accessibility along the nucleosome wrap; (iii) unwrapped nucleosomes; and (iv) subnucleosomal species such as hexasomes, tetrasomes and other short DNA protections. Applying IDLI to chromatin from mouse embryonic stem cells, we discover that more than 85% of nucleosomes exhibit intranucleosomally accessible DNA (nucleosome 'distortion'). We observe epigenomic-domain- and expression-level-specific patterns of distortion, including at promoters and mouse satellite repeat sequences. Transcription factor (TF) motif occurrence correlates significantly with distinct types of distortion, and degron experiments provide evidence of direct regulation by TFs. We apply IDLI to in vitro endoderm differentiation in human induced pluripotent stem cells and primary mouse hepatocytes. In both cases, we observe distortion at pioneer TF FOXA2 binding sites, demonstrating that distortion is developmentally encoded and present in vivo. Finally, genetic experiments in mice show that a nucleosome-binding domain of FOXA2 directly affects nucleosome structure in vivo, implicating these protein-nucleosome interactions as direct mediators of distortion. Our work suggests extreme but regulated nucleosome structural variability at the single-molecule level. Furthermore, our approach offers opportunities to model TF binding, nucleosome remodelling and cell-type-specific chromatin regulation across biological contexts.

CircMAPK1-encoded micropeptide circMAPK1-110aa inhibits proliferation and differentiation of goat skeletal muscle satellite cells by suppressing MAPK1/3 phosphorylation.

Qianqian Pan, Mengyu Lou, Mengkang Zhu +4 more

Skeletal muscle development directly determines growth efficiency and meat quality in livestock. Circular RNAs (circRNAs) and their encoded micropeptides are emerging regulators of this process, yet their roles in goats remain poorly understood. Through integrated ribosome profiling (Ribo-seq) and circRNA-seq of goat skeletal muscle satellite cells (SMSCs), we identified a MAPK1-derived protein-coding circRNA, circMAPK1 (novel-circ-0039015). Functional analyses revealed that circMAPK1 overexpression suppressed SMSCs proliferation and myogenic differentiation. Mechanistically, circMAPK1 encodes a 110-amino-acid micropeptide (circMAPK1-110aa) that binds Mitogen-Activated Protein Kinase Kinase 1 (MAP2K1) and reduces MAPK1/3 phosphorylation, thereby attenuating Mitogen-Activated Protein Kinase (MAPK) signaling. Importantly, pharmacological activation of the MAPK pathway with C16-PAF partially restored muscle cell growth and differentiation. These findings establish circMAPK1 as a negative regulator of goat skeletal muscle development and highlight its encoded micropeptides as a potential molecular target. By uncovering a new regulatory mechanism of muscle growth, this study provides valuable insights for breeding strategies aimed at enhancing meat yield and quality in goats.

Hemoglobin Nanofibrils as Electrospun Cell Scaffolds to Enhance Primary Satellite Cell Proliferation and Differentiation for Muscle Regeneration.

Qun Chen, Jin Kyo Oh, Vaughan Feisst +3 more

Stem cells are usually sensitive to extracellular matrix (ECM), and an effective synthetic ECM to mimic there in vivo growth surroundings is always desired. Poly(ɛ-caprolactone) (PCL) is a common synthetic material extensively employed for ECMs in medical regeneration applications. However, it lacks inherent bioactivity, which poses limitations to its utility. Our study provides a solution to address this drawback by incorporating hemoglobin nanofibrils (HbFs) into PCL. Leveraging the economical and easily formed HbFs, the resulting electrospun cell scaffolds exhibited improved cell adhesion of muscle satellite cells. Furthermore, these scaffolds facilitated enhanced cell proliferation, cell infiltration into the scaffold, and higher levels of expression of differentiation-related proteins. This study demonstrates the feasibility of HbFs-incorporated electrospun scaffolds as a promising ECM substitute for muscle stem cell-based regeneration therapies.

Establish a cultured meat model with adjustable muscle-to-fat ratios based on muscle satellite cells and fibroadipogenic progenitor cells in an optimized gelatin-mTG scaffold.

Liu Shuqin, Tao Shaoying, Bo Chunjie +10 more

Cell-cultured meat offers a sustainable alternative to conventional meat; however, existing monocellular models fall short in replicating its texture and sensory qualities. This study developed an economical gelatin/microbial transglutaminase scaffold to construct a multicellular beef analogue. Muscle satellite cells (MuSCs) and Fibroadipogenic progenitor cells (FAPs) were isolated by flow cytometry. Under 2D culture, MuSCs exhibited enhanced myogenesis (MYH3, MyoD, MyoG upregulated 1.5-2.4-fold), while FAPs showed adipogenic potential (ADIPOQ, FABP4 elevated). Scaffold variants were optimized: GOS (Gelatin-Oriented Scaffold) (muscle-supportive) and GAS (Gelatin-Sodium Alginate Scaffold) (fat-supportive), both supporting high proliferation. Transition to 3D culture further promoted differentiation, increasing myogenic/adipogenic gene expression (2-2.3-fold, p < 0.01) and extracellular matrix secretion (∼2-fold). After 14 days, constructs displayed uniform pink coloration. Post-frying, color parameters (a* [redness], L* [lightness]) were comparable to conventional beef (p > 0.05). This scalable approach creates structured, sensory-comparable cultured meat.

L-Carnosine Enhances the Proliferation and Myogenic Differentiation of Yanbian Cattle Skeletal Muscle Satellite Cells for Cultured Meat Production via Activating the Akt/mTOR/P70S6K Signaling Pathway.

Bin Sun, Huaina Jin, Xuanying Xin +4 more

The composition of muscle fiber types and the development of skeletal muscle are critical determinants of cultured meat quality. L-carnosine, a dipeptide abundant in ruminant muscle, is known to influence meat quality, yet its regulatory mechanisms in bovine skeletal muscle satellite cells (BSCs) for cultured meat production remain unclear. This study aimed to elucidate the effects of L-carnosine on the proliferation, differentiation, and muscle fiber type transformation of Yanbian cattle BSCs. We identified 10 mM as the optimal concentration for enhancing cell proliferation (p < 0.05), a key finding established by screening L-carnosine treatments from 0 to 40 mm. This enhancement was mediated by the upregulation of cell cycle genes (Pax7, Ki67, CDK1, CDK2, PCNA) and the suppression of inhibitors (p21, p53, p16). Furthermore, L-carnosine robustly promoted myotube formation and specifically upregulated fast-twitch muscle fiber markers (MyHC2a, MyHC2b, MyHC2x) while downregulating the slow-twitch marker MyHC1 (p < 0.05). Transcriptomic analysis identified 449 differentially expressed genes, which were significantly enriched in the PI3K-Akt signaling pathway. Western blotting confirmed that L-carnosine activates the Akt/mTOR/P70S6K signaling pathway to drive myogenesis. Additionally, L-carnosine demonstrated significant antioxidant capacity by reducing reactive oxygen species (ROS) and lipid peroxidation (MDA) while enhancing antioxidant enzyme activities (SOD and GSH-Px). In conclusion, this study provides the first evidence that L-carnosine promotes BSC proliferation and fast-twitch fiber differentiation via the Akt/mTOR/P70S6K pathway, suggesting its potential as a highly effective, natural additive for cultured meat production.

The Muscle Tissue Environment Limits Muscle Stem Cells in Aged Mice.

Alicia A Cutler, Tenaya K Vallery, Thomas O Vogler +11 more

Frailty arising from loss of muscle function and mass is a significant health concern impacting quality of life and dramatically increasing health care costs as our population ages. Ameliorating frailty derived from reduced muscle function is thus a critical research priority to improve health span. Cell intrinsic defects in muscle stem cells (MuSC), or satellite cells, occur as skeletal muscle ages, reducing the capacity of MuSCs to maintain and repair skeletal muscle and are accompanied by cell nonautonomous changes. Although rejuvenating stem cells in aged tissues or organs has potential to improve muscle aging phenotypes, we found that the extracellular environment in aged mice abrogates rejuvenated muscle stem cell potential. MuSCs from young mice were unable to grow on extracellular matrix derived from aged mice that contains elevated collagen protein levels, establishing a critical role for the environment in contributing to muscle phenotypes in aging. Combining an inducible FGF receptor 1 (FGFR1) to rescue MuSC intrinsic aging defects with a drug to reduce fibrosis partially rescued muscle mass loss in aged mice. We conclude that aging affects tissues, and particularly skeletal muscle tissue, via complex multifactorial processes requiring multifaceted interventions to improve aging phenotypes.

ARID1A deficiency unleashes centromeric RNA transcription to drive chromosomal instability and boosts PKMYT1 inhibitor efficacy via RNA sensing.

Chengguo Li, Xueqian Cheng, Weizhen Liu +15 more

Cancer gene-associated mutations and molecular hallmarks of chromosomal instability (CIN) are unexpectedly common in histologically normal cells and tissues. These emerging findings challenge the binary distinction between "normal" and "cancerous" cells and suggest that early tumorigenesis may commence against a background of widespread yet largely tolerated genomic instability. However, it remains largely unexplored how a cancer gene-associated mutation can initiate the development of CIN-like states in non-malignant cells and drive tumor evolution. ARID1A , a chromatin remodeling factor, was identified as the most frequently mutated gene in both gastric normal epithelium and tumors. This distinctive molecular convergence presents an opportunity to elucidate the mechanisms by which a cancer-associated gene facilitates the initiation of early CIN phenotypes and develop effective antitumor strategies. In the present study, using primary human gastric organoids, we employed optical genome mapping (OGM) and live-imaging technologies to demonstrate that ARID1A depletion induced a wide spectrum of structural variants (SVs), copy number variants (CNVs), and chromosomal segregation errors, characteristic features of CIN at a very early stage of gastric tumorigenesis. Mechanistically, ARID1A bound centromere repetitive satellite DNA (satDNA) sequences. Its SWI/SNF-associated chromatin remodeling activity was required for suppressing satDNA transcription and the production of α-SatRNA, through restricting RNAPII elongation. Consequently, ARID1A depletion led to overexpression of α-SatRNA, and a higher incidence of sister chromatid exchange (SCE), a sensitive indicator of CIN. Importantly, the elevated α-SatRNA expression in ARID1A -deficient cells further established a dual therapeutic vulnerability for G2/M checkpoint blockade, such as PKMYT1 inhibitor (PKMYTi), by concurrently aggregating CIN-induced cell death and activating self-dsRNA sensing-mediated innate immune response. Notably, PKMYTi markedly promoted α-SatRNA expression, aberrant release of these self-derived dsRNAs into the cytosol and a robust activation of the RIG/MDA5-MAVS-depenent type-I interferon response in ARID1A -depleted cells. As expected, PKMYTi potentiated the efficacy of immunotherapy in ARID1A -deficient gastric tumors. Together, our findings reveal that ARID1A deficiency unleashes centromeric α-SatRNA transcription, which sets the molecular stage for tumor evolution and targeted therapy by coordinately inducing CIN and self-dsRNA-induced innate immune responses.

Defective transcription of AAGAG satellite DNA causes sex-ratio meiotic drive in Drosophila.

Tomohiro Kumon, Mami Nakamizo-Dojo, Amelie A Raz +3 more

Male germ cells have complex transcriptomes, with a large fraction of the genome being transcribed. This includes protein-coding genes (often not translated), non-coding DNA, and repetitive DNA, such as transposons and satellite DNA, which are normally silenced as heterochromatin. The significance of such widespread transcription remains unknown. Here, we show that a heterochromatin protein, HP2, is required for the transcription of AAGAG satellite DNA in Drosophila spermatocytes. HP2 depletion leads to abnormal retention of heterochromatin histone marks (H3K9me3) and spermatid death during sperm DNA packaging, leading to a model that transcription of AAGAG satellite DNA facilitates the remodeling of its heterochromatic nature in preparation for sperm DNA packaging. Strikingly, the severity of the spermatid death correlates with the amount of AAGAG satellite DNA carried by the spermatids, leading to preferential death of Y-chromosome-containing spermatids over X-containing spermatids, and hence sex-ratio meiotic drive phenotype. We propose that widespread spermatocyte transcription may reflect the process of chromatin remodeling to allow sperm DNA packaging. We further propose that differential composition and amount of satellite DNA on chromosomes may underlie naturally occurring male meiotic drive.

Mechanisms and Efficacy of Massage Therapy for Post-Exercise Muscle Repair: A Narrative Review.

Peter M Tiidus

Although widely used, massage has not been reported to be effective in enhancing recovery from exercise-induced muscle damage in humans. Studies using massage-like interventions in animal models have, in contrast, consistently demonstrated a significant enhancement of muscle repair, reduction in muscle inflammation and enhanced return of muscle force following muscle damage. The physiology of muscle damage and repair and the putative physiological mechanisms of potential massage-induced muscle repair and post-damage recovery, including soreness sensation, edema, inflammation, protein synthesis and other related mechanisms, are reviewed in this context. Animal models have demonstrated that massage effectiveness in enhancing post-damage muscle repair is dictated by the timing, duration, force and technique of its application and may also be modified by age and sex effects. The potentially very narrow "window of effectiveness" of massage application for the enhancement of post-damage muscle repair in humans has yet to be defined. And the lack of demonstrated effectiveness for massage on post-damage muscle recovery may be due to the wide range and inconsistency of massage techniques, timing and methodologies applied in human studies. Until a specific massage application protocol is defined for massage efficacy in post-damage human muscle recovery, therapists will continue to work blind, using a variety of techniques which lack empirical validity and have an undemonstrated effectiveness for enhancing muscle repair.

Differential effects of human fibromyalgia sera on murine satellite glial cells: comment on the article by Mercado et al.

Suhail Aamar, Emil Aamar

Single cell analysis of muscle contracture in cerebral palsy reveals pro-fibrotic and anti-myogenic stem cell populations with altered cell-cell interactions.

Madison Stewart, Lin-Ya Hu, Taryn Loomis +8 more

Development of muscle contractures are common in cerebral palsy (CP) and characterized by high muscle stiffness that limits function and mobility. However, the state of stem cells within contracture, particularly muscle stem cells and fibro-adipogenic progenitors, are largely unknown. This study leverages single cell RNA sequencing technology to determine how specific cell types are altered in the contracture environment. Skeletal muscle biopsies were collected from children with CP or typically developing (TD) children undergoing surgery. The 10X Genomics platform was used on tissue from n=3 patients per condition. Significant changes in CP compared to TD were investigated within individual cell types for differentially expressed genes, gene ontologies, cell subpopulations and predicted interactions. CP muscle stem cells demonstrated significant upregulation of fibrotic genes and down regulation of myogenic genes compared to typically developing. Fibro-adipogenic progenitors in CP showed the emergence of a significant proportion of a highly profibrotic subpopulation, leading to the most dramatically up-regulated genes in CP also being extracellular matrix constituents. Interacting signals between fibro-adipogenic progenitors, muscle stem cells, and immune cells were identified that support contracture progression. Contracture reduces myogenic muscle stem cells and enhances fibrotic signals in muscle stem cells and fibro-adipogenic progenitors that perpetuate contracture. The study reveals specific genes and signaling pathways as therapeutic targets to reduce muscle contracture in children with CP.

Single Fiber Isolation Assay for the Assessment of Oxidative Myofiber Behavior.

Huascar Pedro Ortuste Quiroga, Yoshitaka Mita, Yasuko Manabe +2 more

Skeletal muscle is composed of multinucleated myofibers whose integrity and function are essential for movement and overall health. In neuromuscular disorders, muscle groups can be affected differentially across disease types and stages, making it important to isolate viable single fibers from representative muscles for mechanistic studies. The objective of this protocol is to provide a reliable method for isolating a high yield of intact single myofibers from the murine soleus (SOL) muscle, which is challenging to dissociate due to fiber length and fragility. By optimizing collagenase concentration and digestion time, the protocol minimizes fiber loss and preserves viability during dissociation. The optimized workflow markedly reduces residual tissue attachment to isolated fibers, thereby limiting carryover of non-fiber cells and minimizing cross-cell contamination. This technique is suitable for single-fiber analyses as well as satellite cell isolation and subsequent expansion under conditions with minimal contamination. Using this optimized workflow, we routinely obtain ~300-500 intact SOL-derived single myofibers that can be immunolabelled with high efficiency and also yield a high-quality satellite cell population with minimal fibroblasts contamination. Using comparative analyses with Extensor Digitorum Longus (EDL) muscle we underscore the value of a tailored, muscle-specific approach to evaluating therapies in neuromuscular disease research, enabling candidate pharmaceutical interventions to be tested within small, tunable experimental windows.

Satellite glial GLRX3 drives ageing-biased neuropathic pain via HMGB1.

Yang Yang, Bing Zhao, Xinyu Liu +6 more

Chronic neuropathic pain disproportionately affects older individuals, particularly in the context of persistent oxaliplatin-induced peripheral neuropathy (OIPN); however, the molecular mechanisms sustaining this ageing-biased chronicity remain elusive. In this study, we integrated age-stratified murine models and a multicentre longitudinal cohort of patients receiving oxaliplatin-based chemotherapy for colorectal cancer to investigate a glia-to-neuron redox circuit in the dorsal root ganglion. Using single-nucleus RNA sequencing and redox proteomics, we identified selective upregulation of the deglutathionylase glutaredoxin-3 (GLRX3) in satellite glial cells in aged mice during the chronic phase of OIPN. This upregulation leads to a pronounced loss of protein S-glutathionylation (PSSG) within the dorsal root ganglion, a pattern absent in young mice and during acute stages. Mechanistically, GLRX3, via its catalytic Cys148 residue, catalyses the deglutathionylation of high-mobility group box 1 (HMGB1) at the Cys106 site. This modification converts HMGB1 into a potent agonist for the toll-like receptor 4 (TLR4)-myeloid differentiation factor 2 (MD2) complex, triggering neuronal nuclear factor-κB signalling and the subsequent upregulation of transient receptor potential ankyrin 1 and vanilloid 2 channels in PACAP-positive (C1 subtype) peptidergic nociceptors, thereby sustaining long-term mechanical and cold hypersensitivity. Satellite glial cell-targeted knockdown of GLRX3 restored HMGB1 glutathionylation and reversed the pain phenotype specifically in aged mice. In the clinical cohort, advanced age was significantly associated with a higher incidence of chronic neuropathy. Longitudinal serum analysis revealed that systemic levels of PSSG and glutathionylated HMGB1 declined progressively and correlated inversely with pain duration, particularly among older individuals. Furthermore, oral γ-glutamylcysteine or pharmacologic TLR4 blockade (TAK-242) effectively alleviated refractory hypersensitivity in aged models. These findings define the satellite glial GLRX3-HMGB1-TLR4 redox axis as a critical driver of age-biased neuropathic pain. Circulating PSSG represents a novel age-stratified clinical biomarker, and targeting this redox-sensitive pathway offers a promising therapeutic strategy for geriatric and chemotherapy-related neuropathies.

HuR-mediated regulation of mTOR mRNA stability promotes the commitment of satellite cells towards myogenesis.

Anne-Marie K Tremblay, Brenda Janice Sánchez, Bianca Colalillo +6 more

The RNA-binding protein HuR has been shown to promote the differentiation of cultured muscle cells into muscle fibers. HuR mediates this process by differentially regulating, at different stages of this process, mRNA targets encoding pro-myogenic factors. Despite these advancements, the role of HuR, in vivo, at various stages of the myogenic process and its impact on muscle formation and function remain elusive. Towards this end, we used the Myf5-Cre loxP system to knock out HuR at a stage where muscle precursor cells (satellite cells, SCs) commit to myogenesis. Using these mice, we found that the muscle-specific depletion of HuR impairs, physiologically, its formation during embryogenesis and in response to injury. These mice exhibited smaller skeletal muscles and reduced exercise endurance. We demonstrate, using these mice, that this effect is due, in part, to the HuR-mediated regulation of mTOR (mechanistic target of rapamycin) mRNA expression. Using primary and cultured muscle cells, we show that HuR associates with this message, regulating its stability. In doing so, HuR facilitates the commitment of satellite cells toward myogenesis, thus preventing their transdifferentiation toward adipogenesis. These findings thus identify HuR as a master regulator of SCs' commitment to myogenesis and uncover a potential target for manipulating muscle myogenic capacity in both normal and pathological conditions.

Cy3-Conjugated Biocompatible Polymer Nanoparticles for Long-Term Mitochondrial Imaging.

Souma Kawashima, Mitsuo Inui, Izumi Takaba +2 more

The cyanine dye Cy3 has recently gained attention as a membrane potential-responsive, small-molecule mitochondrial imaging probe. However, the efficiency of Cy3 for mitochondrial imaging is relatively low, as diffusion of Cy3 leads to staining of the entire cell over time. This problem could be due to the passive diffusion-based, concentration-dependent uptake into cells and molecular diffusion inherent to small-molecule compounds. Therefore, we hypothesized that polymer chemistry approach could be applied to overcome these limitations. In this study, we designed modified dye in which biocompatible polymers were covalently attached to Cy3. This enabled control of intracellular dynamics that could not be achieved using Cy3 alone. Toward this objective, we synthesized a Cy3-modified amphiphilic dextran with phenylalanine ethyl ether side chains. The amphiphilic polymers self-assembled into nanoparticles. We then characterized the synthesized polymer in terms of intracellular uptake and the mitochondria translocation capacity of the nanoparticles using human skeletal muscle satellite cells. Our approach changed the cellular internalization process, resulting in increased cellular uptake efficiency, enhanced mitochondrial imaging capability, and improved mitochondrial retention over the long term. These nanoparticles, which can stain mitochondria regardless of cell type, have the potential to become a new type of mitochondrial imaging reagent.

Renalase knockdown inhibits proliferation of mouse satellite cells.

Yuri Kato, Katsuyuki Tokinoya, Kai Aoki +1 more

Editorial: Lifetime achievements in avian physiology.

Colin G Scanes, Sandra G Velleman

STAT3 isoform dynamics reveal robust splice ratio maintenance across cytokine-activated human immune cells.

Nils Ott, Bodo Grimbacher, Virginia Andreani

Alternative splicing of STAT3 produces two principal isoforms, STAT3α and STAT3β, that differ in transactivation capacity and DNA-binding behavior. Whereas STAT3α is thought to be a transcriptional activator, STAT3β - due to the lack of the transactivation domain - is thought to be a transcriptional repressor. Therefore, the relative abundance of STAT3α and STAT3β molecules within a leukocyte will be critical for immune homeostasis and for gene-therapeutic strategies targeting STAT3. This study investigates whether short-term exposure to IFN-α, IL-6/sIL-6Rα, or IL-10 alters STAT3α/STAT3β stoichiometry in purified human CD4+, CD8+, CD14+ and CD19+ cells. We evaluated in healthy donors PBMCs the STAT3α and STAT3β mRNA and protein levels after stimulation with IFN-α, IL-6/sIL-6Rα or IL-10. For mRNA analysis, twelve candidate reference genes were assessed for stability across subsets and stimuli, with UBE2D2 identified as the most stable reference gene. We demonstrate that at mRNA level, cytokine treatment induced STAT3α and STAT3β mRNA in a subset-dependent manner. STAT3β mRNA largely paralleled STAT3α, and crucially, the STAT3α/STAT3β mRNA ratio remained constrained within a narrow range. At protein level, STAT3α predominated markedly, yielding a pronounced transcript-protein decoupling. Together, these data indicate that ex vivo short-term cytokine signaling with IFN-α, IL-6/sIL-6Rα, and IL-10 co-induces STAT3 isoforms without broadly reprogramming their splice balance in primary human leukocytes and underscore the importance of preserving physiological isoform ratios in gene-therapeutic strategies targeting STAT3. These findings suggest that short-term inflammatory signals alone are insufficient to shift STAT3 isoform production, supporting the idea that additional or more prolonged regulatory inputs are required to modulate STAT3 splicing in vivo and that physiologically, the STAT3α/STAT3β ratio may be modulated at protein level.

Pharmacological Inhibition of N-terminal methyltransferase 1 promotes myogenic differentiation and muscle regeneration in a mouse model of Duchenne muscular dystrophy.

Haoyuan Zhang, Ming He, Muhammad Asif +3 more

Duchenne muscular dystrophy (DMD), caused by mutations of the DMD gene, is a lethal degenerative disease with no cure. Stimulating myogenesis of muscle stem cells (MuSCs) represents a promising strategy to ameliorate muscle pathology in DMD patients. Although previous work has revealed a role of N-terminal methyltransferase 1 (NTMT1) in myogenesis, its potential as a therapeutic target to ameliorate muscular dystrophy remains unexplored.

Isolation, Culture, and Differentiation of Bovine Muscle Resident Stem Cells.

Perri Gish, Madison W Stewart, Maykal Tsonov +4 more

Bovine muscle satellite cells (MuSC) and fibro-adipogenic progenitor cells (FAP) are muscle resident stem cells that are responsible for postnatal muscle growth, intramuscular fat deposition, and extracellular matrix generation. These cells are of increasing interest for the cultivated meat community due to their ability to generate all the major components of meat; additionally, these cells are of interest to conventional animal science research to elucidate mechanisms to improve meat quality. To use these cells for these goals, efficient and accurate cell isolation, culture, and differentiation are essential to evaluate their cell fate decisions and behaviors. In this protocol, we detail a simultaneous isolation of both MuSCs and FAPs with multiple intermediate stopping points, allowing for flexibility for day-of time constraints. We also detail improved growth conditions to maximize cell expansion and procedures to assess cell differentiation. This protocol provides a flexible isolation procedure that is compatible with sampling in modern slaughterhouses or from biopsies. Additionally, the differentiation procedures provide improved differentiation but still allow in vitro treatment and assessment. Key features • This protocol offers a flexible in-lab procedure to isolate bovine FAPs and MuSCs from tissue collected post-slaughter with multiple pause points. • The protocol demonstrates successful conditions to grow, expand, and differentiate bovine FAPs with an optimized adipogenic differentiation medium. • Strategies for planning your primary cell isolation, choosing the sampling location, and characterizing differentiation of bovine FAPs and MuSCs are included.

The role of mast cells in simvastatin-induced myopathy and the possible protective role of omega-3 fatty acids in adult male albino rats: light and electron microscopic study.

Dalia Ahmed Baha Eldein, Shereen Hamed, Shireen Mazroa +1 more

Simvastatin is antihyperlipidemic drugs that inhibit hydroxy methylglutaryl CoA reductase to decrease cholesterol synthesis; however, it causes skeletal myopathy. Omega-3 are polyunsaturated fatty acids with anti-inflammatory and anti-oxidant effects. Mast cells have an important role in the response to tissue injury by secreting different mediators and cytokines. The current study was performed to assess the role of mast cells in simvastatin-induced myopathy and the possible protective role of omega-3 fatty acids. Forty adult male albino rats were divided into four main groups received the drugs orally for of 21 days. Group 1: 1 ml distilled water, group 2: 300 mg/kg/day omega-3, group 3: 88 mg/kg/day simvastatin, and group 4: 88 mg/kg/day simvastatin plus 300 mg/kg/day omega-3. Light and electron microscopic studies were done along with morphometric and statistical studies. Group 1 and 2 showed regular histological structure. By light microscope, group 3 revealed muscle fiber disorganization, splitting, loss of striations, and cytoplasmic fragmentation. Some nuclei were centrally displaced. Toluidine blue sections showed a large number of mast cells. By transmission electron microscope, there was loss of the regular banding pattern with irregular mitochondria, nuclei, and sarcolemma. The concurrent administration of simvastatin with omega-3 showed less myopathic changes, decrease in mast cell numbers and more activation of the satellite stem cells to accomplish the process of muscle regeneration. In conclusion, the administration of omega-3 can decrease the myopathic changes of simvastatin. Further studies are recommended to explore the mast cells' role in the progression and repair of statin-induced myopathy.

Effects of Glial Cell Line-Derived Neurotrophic Factor and Ciliary Neurotrophic Factor on Extraocular and Limb Muscle Precursor Cells.

Austin J Winker, Laura L Johnson, Linda K McLoon

SummaryMyogenic precursor cells within skeletal muscles are responsible for the maintenance of skeletal muscle over a lifetime. Neurotrophic and growth factors play critical roles in this maintenance and in responses of myogenic precursor cells. Both glial cell line-derived neurotrophic factor (GDNF) and ciliary neurotrophic factor (CNTF) play roles in the maintenance and/or development of strabismus, yet few studies have examined their roles in the control of myogenic precursor cell proliferation and differentiation. Two populations of myogenic precursor cells were isolated from extraocular and leg muscle by fluorescence-activated cell sorting: EECD34 cells, largely PITX2-positive, and PAX7-positive cells. Cultures were treated with GDNF or CNTF and processed immunohistochemically to determine proliferation and differentiation rates. Neither GDNF nor CNTF affected cell proliferation rates for either muscle. Both treatments impacted cell differentiation by increasing multinucleated cell number, with TA-derived precursor cells producing cells containing large numbers of nuclei and EOM-derived precursor cells producing shorter multinucleated fibers with fewer nuclei. These differences may explain the presence of extremely short myofibers within normal adult EOM compared with limb muscle. As GDNF and CNTF are downregulated in strabismic muscles, data suggest that myofiber length homeostasis may be disrupted in strabismic EOM and suggest possible approaches for strabismus treatment.

Decellularized extracellular matrix enhances hydrogel printability for bioprinting functional muscle constructs in a volumetric muscle loss model.

Shabnam Sabetkish, Yuxuan Luo, Yen-Zhen Lu +3 more

Volumetric muscle loss (VML) remains a major clinical challenge due to the limited regenerative capacity of skeletal muscle. Effective repair requires the use of biomaterials that support cell viability, promote myogenic differentiation and enable vascularisation to allow the formation of structurally aligned, functional muscle. Here, we have integrated a biomimetic bioink based on gelatin methacryloyl (GelMA) and methacryloyl-modified decellularized extracellular matrix (dECM-MA) with a micropost tension-assisted bioprinting strategy to engineer skeletal muscle constructs for VML repair. We synthesized and characterized GelMA and dECM-MA bioinks, demonstrating well-preserved ECM components, reproducible gelation, mechanical properties and rheological properties suitable for extrusion bioprinting. In vitro, printed GelMA + dECM-MA scaffolds supported high cell viability, alignment and robust myogenic differentiation of C2C12 myoblasts, human satellite cells (hSkMSCs) and human umbilical vein endothelial cells (HUVECs). Co-culture of satellite cells with HUVECs enhanced endothelial network formation and improved myotube maturation. Printing around PDMS microposts provided passive mechanical tension, producing aligned fibres and greater contraction velocity and micropost displacement in co-culture constructs under electrical stimulation. To assess regenerative potential, 3-day-matured constructs were implanted in a mouse VML model. Constructs containing both hSkMSCs and HUVECs showed the greatest tissue regeneration, higher myofiber density, improved organization, and enhanced functional recovery compared with acellular or monoculture constructs. No immune response towards the presence of the human cells or porcine ECM was observed, suggesting a protective role for dECM-MA. Together, this integrated bioink-biomechanical platform resulted in the generation of vascularised, aligned, and functional muscle tissue with strong translational potential for VML therapy.

Defective transcription of AAGAG satellite DNA causes sex-ratio meiotic drive in Drosophila.

Tomohiro Kumon, Mami Nakamizo-Dojo, Amelie A Raz +3 more

Male germ cells have complex transcriptomes, with a large fraction of the genome being transcribed. This includes protein-coding genes (often not translated), non-coding DNA, and repetitive DNA, such as transposons and satellite DNA, which are normally silenced as heterochromatin. The significance of such widespread transcription remains unknown. Here, we show that a heterochromatin protein, HP2, is required for the transcription of AAGAG satellite DNA in Drosophila spermatocytes. HP2 depletion leads to abnormal retention of heterochromatin histone marks (H3K9me3) and spermatid death during sperm DNA packaging, leading to a model that transcription of AAGAG satellite DNA facilitates the remodeling of its heterochromatic nature in preparation for sperm DNA packaging. Strikingly, the severity of the spermatid death correlates with the amount of AAGAG satellite DNA carried by the spermatids, leading to preferential death of Y chromosome-containing spermatids over X-containing spermatids, and hence sex-ratio meiotic drive phenotype. We propose that widespread spermatocyte transcription may reflect the process of chromatin remodeling to allow sperm DNA packaging. We further propose that differential composition and amount of satellite DNA on chromosomes may underlie naturally occurring male meiotic drive.

Haplotype-resolved centromeric chromatin organization from a complete diploid human genome.

Yuan Xu, Hailey Loucks, Julian Menendez +23 more

Centromeres ensure proper chromosome segregation during cell division, yet the organization and regulation of centromeric chromatin within satellite DNA arrays remain incompletely understood. Here, we leverage the complete diploid human genome benchmark (T2T-HG002) to provide a detailed study of centromeric sequence and chromatin architecture on individual haplotypes. Using adaptive-sampling-enriched, ultra-long-read DiMeLo-seq, we achieve single-molecule chromatin profiling across all centromeres, revealing that along single chromatin fibers, CENP-A, the histone variant specifying centromere identity, forms multiple discrete subdomains within hypomethylated centromere dip regions (CDRs) that are flanked by H3K9me3-enriched heterochromatin. Despite underlying sequence variation, CDRs localize to sequence-homogeneous domains and maintain relatively balanced CENP-A dosage and aggregate length across all chromosomes and between haplotypes. Further, we show that bidirectional changes to centromeric and pericentromeric DNA methylation are accompanied by changes to centromeric chromatin architecture. In passaged cells with centromeric hypomethylation, subdomain boundaries are eroded, and adjacent CENP-A domains tend to merge and expand. Conversely, in pluripotent stem cells with centromeric hypermethylation, CDRs are fundamentally reorganized, such that discrete hypomethylated domains are frequently consolidated into broader contiguous tracts. These methylation-associated CDR restructuring events suggest that DNA methylation acts as a principal regulator of human centromere organization, with implications for understanding centromere plasticity, epigenetic inheritance, and chromosomal instability in development and disease.

Preoperative systemic inflammation and muscle fatty infiltration are prognostic factors for quadriceps atrophy following anterior cruciate ligament reconstruction.

Hanyi Wang, Yuqi Li, Hao Zheng +5 more

Muscle atrophy following anterior cruciate ligament reconstruction (ACLR) significantly impedes functional recovery, yet its underlying prognostic factors and potential cellular mechanisms remain poorly understood.

EXPRESS: CXCL12/CXCR4 Axis in Neuropathic Pain: Insights from Preclinical Models and Translational Implications.

Ming Li, Xiao-Xiao Lu, Lu-Yao Cai +2 more

Neuropathic pain affects approximately 7%-10% of the global population, significantly impairing patients' quality of life and placing a substantial burden on public health systems. Current pharmacological treatments have limited efficacy and are often accompanied by notable side effects, highlighting the urgent need for novel therapeutic targets. Increasing evidence supports the important role of chemokines and their receptors in neuro-immune interactions underlying pain sensitization. Among these pathways, the CXCL12/CXCR4 axis has emerged as an important regulator of both the initiation and maintenance of neuropathic pain. Beyond its canonical function in immune cell trafficking, the CXCL12/CXCR4 axis modulates neuronal excitability, glial activation, synaptic plasticity, and nociceptive sensitization. Notably, this axis is frequently upregulated in both peripheral and central neurons, as well as in multiple glial populations, including astrocytes, microglia, and satellite glial cells, across diverse neuropathic pain models. Importantly, CXCR4 is one of the few chemokine receptors with a clinically approved antagonist, highlighting its unique translational potential. This review systematically summarizes the expression patterns, biological functions, and pain-related mechanisms of the CXCL12/CXCR4 axis in preclinical models of neuropathic pain and discusses current limitations and potential future therapeutic strategies targeting this pathway.

Dual roles of syndecan-4 in regulating chicken fibrosis in vitro.

Lucie Pejšková, Nina Therese Solberg, Marianne Lunde +3 more

Wooden Breast (WB) is a myopathy affecting the skeletal breast muscle (Pectoralis major) in broiler chickens and is characterized by muscle fiber damage and varying degrees of fibrosis, ECM remodeling and inflammation. Several key factors such as pro-inflammatory cytokines like TGF-β1 and IL-1β, drive fibrosis in WB myopathy. We have previously shown that the expression of syndecan-4 (SDC4), a transmembrane proteoglycan, was increased in WB poultry skeletal muscle tissue. Furthermore, the ectodomain shedding of SDC4 by matrix metalloproteinases (MMPs) differed in the skeletal muscle satellite cells from isolated affected chickens compared with normal. While SDC4 has been previously implicated as a key driver for regulating myofibroblast activity in mechanically induced fibrosis in cardiac tissue, its specific role and shedding activity in chicken fibroblasts in relation to WB myopathy remain poorly understood.

DNA O-MAP uncovers the molecular neighborhoods associated with specific genomic loci.

Yuzhen Liu, Christopher D McGann, Conor P Herlihy +14 more

The accuracy of crucial nuclear processes such as transcription, replication, and repair depends on the local composition of chromatin and the regulatory proteins that reside there. Understanding these DNA-protein interactions at the level of specific genomic loci has remained challenging due to technical limitations. Here, we introduce a method termed 'DNA O-MAP', which uses programmable peroxidase-conjugated oligonucleotide probes to biotinylate nearby proteins. We show that DNA O-MAP can be coupled with label-free or sample multiplexed quantitative proteomics, targeted chemical perturbations, and next-generation sequencing to quantify DNA-proximal proteins and DNA-DNA interactions at specific genomic loci in human and murine cells. Furthermore, we establish that DNA O-MAP is applicable to both repetitive and unique genomic loci of varying sizes, from kilobase HOX gene clusters to megabase alpha-satellite repeats, and that DNA O-MAP can measure proximal molecular effectors in a homolog-specific manner.

Mitochondrial transfer as a therapeutic target for peripheral neuropathy.

Junlin Wei, Fang Wang

Satellite glial cells transfer mitochondria to sensory neurons via myosin 10-dependent tunneling nanotubes. Ji et al. show that this transfer is impaired in diabetic neuropathy, causing energy failure. Restoring it via cell or mitochondrial transplantation alleviates pain and promotes nerve regeneration, revealing a new therapeutic strategy for peripheral neuropathy.

Plant-derived exosome-like vesicles enhance exercise-induced muscle recovery and sleep quality.

Emrah Aykora, Damla Aykora

Exercise-induced muscular stress triggers a complex cascade of adaptive responses, including micro-injury, inflammation, activation of satellite cells, mitochondrial remodeling, and myofibrillar repair. The efficiency of recovery processes is crucial for athletic performance, especially among elite athletes, where rapid restoration of muscle function, reduction of inflammation, and improved sleep quality influence training results. Beyond traditional recovery methods, EVs and, more recently, plant-derived exosome-like nanovesicles (PELNs) have emerged as promising bioactive mediators of intercellular communication and tissue regeneration. PELNs contain various biomolecules such as lipids, proteins, small RNAs, and plant-specific metabolites that may affect oxidative stress, inflammatory signaling, and cellular repair pathways. While most research has focused on mammalian or cell-line sources, growing evidence indicates that PELNs may improve muscle regeneration and recovery through cellular modulation and enhanced sleep-related recovery. Notably, PELNs represent a multi-target strategy that may simultaneously modulate neuroendocrine pathways involved in sleep regulation and metabolic-inflammatory mechanisms governing skeletal muscle repair. By influencing circadian rhythm signaling, mitochondrial dynamics, and redox homeostasis, PELNs may bridge the sleep-muscle recovery axis, an emerging concept in exercise physiology. This dual regulatory capacity distinguishes PELNs from conventional recovery interventions and highlights their innovative and translational potential in sports science. This review aims to compile current evidence linking PELNs to exercise-induced muscle recovery, highlighting potential mechanisms, including the regulation of inflammatory and redox balance, microRNA-driven signaling, and neurometabolic adaptation. By combining insights from exercise physiology and molecular regenerative biology, we propose that PELNs offer a natural approach to enhancing recovery and performance in athletes.

Centriolar satellites regulate CEP350 mRNA localization and centrosome amplification.

Abraham Martinez, Chad G Pearson

Messenger RNAs (mRNAs) accumulate at centrosomes in mitosis and interphase, yet the mechanisms governing their localization and the functional significance of centrosomal localization remain poorly understood. Here, we investigate the regulation and function of the centrosome-localized mRNA, CEP350 . We find that CEP350 mRNA localizes to centrosomes during S phase via the centriolar satellite protein CEP131 and the RNA binding protein (RBP) Unkempt (UNK), in a microtubule (MT)-dependent manner. CEP131 and UNK stabilize CEP350 mRNA to maintain CEP350 mRNA steady-state levels. Furthermore, CEP131 and UNK promote normal CEP350 protein levels at centrosomes. CEP350 is required for PLK4-induced centriole overduplication but is less important for canonical centriole duplication. Moreover, CEP131, UNK, and CEP350 are important for centrosome amplification in triple-negative breast cancer cells. Together, these findings reveal a centriolar satellite-RBP pathway regulating CEP350 mRNA localization to centrosomes.

TRPV2 is essential for calcium signalling in the early stages of myogenesis.

Yanzhu Chen, Kimiaki Katanosaka, Yuki Katanosaka

Skeletal muscle responds to stressors such as exercise and muscle injury by adaptive remodelling. The resilience of skeletal muscle involves not only mature muscle fibres but also the adjacent muscle satellite cells (MuSCs). We previously found that transient receptor potential vanilloid type 2 (TRPV2) is expressed in MuSCs and is essential for MuSC proliferation and activation in MuSC-specific conditional knockout mice. These mice show no mechanical-load-induced muscle hypertrophy and delayed injury-induced muscle regeneration. The effect of TRPV2 on Ca2+ signalling during early myogenesis is unknown; however, here, we demonstrate that tranilast, an inhibitor of TRPV2, suppressed IP3R-derived Ca2+ oscillations in early myogenesis. The addition of adenovirus (Ad)-TRPV2 or Ad-Cre recombinase to floxed-TRPV2 cells modulated TRPV2 expression, and demonstrated the TRPV2 dependence of IP3R and MEF2c expression, nuclear translocation of MEF2c, and Ca2+ oscillations. These findings indicate that TRPV2 regulates intracellular Ca2+ signalling during early myogenesis and highlight its potential as a target for the prevention and treatment of muscle disorders.

Self-assembled mesoporous bioglass polyphenol nanozymes for repairing musculoskeletal trauma.

Shuao Zhao, Yesheng Jin, Zhihao Jia +7 more

Volumetric Musculoskeletal Trauma (VMST), which is characterized by volumetric muscle loss accompanied by bone injury, poses a significant challenge to regenerative medicine. While current therapies primarily focus on the individual regeneration of muscle or bone, there is no systematic and integrated treatment strategy. In this study, we developed a CuMBG-PA, a copper-doped nanozyme based on mesoporous bioactive glass (MBG) and procyanidin (PA), for integrated muscle and bone repair of VMST. CuMBG-PA self-assembles into a stable polyphenol network via Cu-PA coordination bonds, enhancing PA stability and reactive oxygen species-scavenging capacity. In vitro and in vivo experiments demonstrated that CuMBG-PA promoted osteogenesis and myogenesis while exhibiting high biocompatibility and antibacterial activity. Single-cell RNA-sequencing results revealed that CuMBG-PA synergistically induces stem cell differentiation and promotes tissue repair through multiple myoskeletal shared metabolic pathways. Mechanistically, CuMBG-PA exerts its beneficial effects by increasing the number of Proteoglycan 4 (Prg4) + fibro/adipogenic progenitor cells (FAPs), which highly express fibronectin and insulin-like growth factor. In addition, PRG4 regulates immune cells, removes overactivated muscle satellite cells, reduces muscle fibrosis, and promotes functional recovery during regeneration. In summary, this work demonstrates that the novel self-assembled CuMBG-PA nanozyme represents a potential biomaterial-based strategy for integrated muscle and bone repair in VMST.

snRNA sequencing-based skeletal muscle analysis of Jiangquan black pigs with different average daily growth rates.

Hongzhen Cao, Jing Wang, Yunzhou Wang +7 more

The Jiangquan black pigs, a new breed of swine obtained by introducing traits from Duroc pigs into Yimeng black pigs, exhibits fast growth rates and high meat quality. To understand how daily weight gain influences muscle development in this breed, we analyzed longissimus dorsi muscle cell subpopulations from Jiangquan black pigs using snRNA and bulk RNA sequencing. Thirteen distinct cell types (e.g., muscle stem cells, satellite cells, fibroblasts) were identified, and marker genes (PAX7, MYOD, MYOG) were found to exhibit stage-specific expression during differentiation. Pseudotime analysis revealed the differentiation trajectories of these cell populations, while cell cycle analysis uncovered the higher mitotic activity in satellite cells of the fast-growth versus slow-growth groups. Furthermore, cell communication analysis highlighted the interactions between muscle cells and other cell types. Finally, intergroup analysis revealed that 2,466 and 2,597 genes were differentially expressed in muscle stem cells and muscle satellite cells, respectively. These genes were enriched in disease-related pathways. This study provides a single-cell resolution atlas of porcine muscle development, offering insights into the genetic regulation of growth and potential targets for breeding optimization.

Telocytes in skeletal muscle: Emerging players in homeostasis and repair/regeneration.

Irene Rosa, Eloisa Romano, Mirko Manetti

Telocytes (TCs) have recently emerged as novel components of the skeletal muscle interstitium. They are distinguished from other stromal cells by their immunophenotypic profiles and, especially, unique ultrastructural traits. Specifically, TCs feature a small cell body and very long, thin telopodes with a moniliform appearance conferred by the alternation of slender segments (podomers) and small dilated portions (podoms). Experimental evidence suggests that, as part of the skeletal muscle stem cell niche, TCs may be involved in orchestrating satellite cell activation and myogenic differentiation through both direct physical interactions and paracrine signaling. Yet, further in-depth research is needed to uncover specific immunophenotypic signatures for skeletal muscle TCs within the niche, as well as to identify the signaling pathways by which they influence neighboring satellite cells and, possibly, other cellular components of the niche. In the present review, particular emphasis is placed on the putative strategic role of TCs in maintaining skeletal muscle tissue homeostasis, their involvement in muscle pathological alterations, and, most importantly, their possible role in the coordination of the regenerative response following injury. In perspective, the promising therapeutic potential of TC-based strategies to enhance skeletal muscle tissue repair/regeneration and restrain post-injury fibrosis is also discussed.

The Crucial Role of Exercise in Promoting Adolescent Muscle Development.

Xilong Hu, Haiwang Shi, Rui Duan

Skeletal muscle, as one of the largest organ system in the human body, exerts a determining influence on adolescents' mastery of motor skills and their lifelong health. Puberty represents a critical window for muscle development, during which the quality of myogenesis not only shapes athletic potential but also profoundly influences long-term health outcomes in adulthood. Under pathological conditions, such as obesity, an aberrant metabolic environment can compromise muscle function in youth, impede the progression of motor abilities, and increase susceptibility to metabolic disorders. It is well established that scientifically prescribed exercise interventions effectively unlock adolescents' muscle-building potential, thereby laying a solid foundation for enduring physical performance and overall well-being.This chapter offers a systematic overview of the key biological principles that regulate skeletal muscle development during puberty and provides an in-depth analysis of the mechanisms and signaling pathways by which structured exercise drives muscle growth and functional adaptation. In addition, it examines the challenges posed by muscle structural and functional impairments under pathological states and evaluates how targeted exercise regimens can restore and enhance muscle health. By mastering the conceptual framework presented here, coaches and parents will be better equipped to identify early warning signs of impaired muscle development, support individualized training plans that optimize motor skill acquisition while minimizing injury risk, and implement proactive interventions to prevent metabolic dysregulation and age-related muscle decline.

Global, regional and national burden of ischemic heart disease attributable to suboptimal diet, 1990-2023: a Global Burden of Disease study.

Sooji Lee, Hayeon Lee, Yejun Son +1036 more

Ischemic heart disease (IHD) remains a leading cause of death worldwide, with dietary risks being its most significant modifiable factor. Here, using the Global Burden of Diseases, Injuries and Risk Factors Study 2023, we estimated the mortality and disability-adjusted life years from diet-related IHD across 204 countries. In 2023, a suboptimal diet was responsible for 4.06 million (95% uncertainty interval (UI) 0.74-6.22) IHD deaths and 96.84 million (18.82-142.52) IHD disability-adjusted life years. The global age-standardized death rate of IHD attributable to suboptimal diet decreased by 43.92% (95% UI 34.44-53.23) per 100,000 population from 1990 to 2023. Among dietary factors, low intake of nuts and seeds (9.87, 95% UI 2.84-17.12 deaths per 100,000 population), low whole grains (9.22, 4.73-13.67), low fruits (7.25, 1.54-13.34) and high sodium (7.15, 0.92-17.97) were primary contributors to IHD deaths. The burden was particularly pronounced in low- and middle-sociodemographic index countries. By disentangling dietary risk factors, we identified the portion of IHD burden directly modifiable through food interventions.

CIMT combined with BoNT-A regenerates skeletal muscle and improves upper limb function through activating IGF-1/FGFR2 axis in hemiplegic cerebral palsy.

You Wang, Qihong Wu, Xiuying Zhao +16 more

Hemiplegic cerebral palsy (HCP) is a prevalent cause of pediatric motor disability. Constraint-induced movement therapy (CIMT), when combined with botulinum neurotoxin type A (BoNT-A), improves upper limb function and social participation in individuals with HCP. However, the mechanisms underlying the combined interventions remain unclear.

Single-Cell RNA-Sequencing Reveals Cachectic Satellite Cell Population in Muscle of Male Mice With Cancer Cachexia.

Alex Brown, Nicolás Collao, Aisha Saleh +3 more

Cancer cachexia leads to decreases in body mass, lean mass and fat mass, decreased therapeutic potential and ~20% of cancer-related deaths. While several studies have demonstrated changes to components of the muscle microenvironment with cancer cachexia, none have comprehensively assessed changes to cellular dynamics across the duration of cachexia development.

Review of the Pathology of Muscle in Amyotrophic Lateral Sclerosis.

Matthew Katz, Thomas Robertson, Shyuan T Ngo +4 more

In amyotrophic lateral sclerosis (ALS), a central event is the withdrawal of the motor nerve terminal from its target muscle. Whether this defect is driven by faults in the motor neuron or faults that originate within the muscle remains an area of investigation. In this review, we focus on the pathological abnormalities that are found in skeletal muscle, focusing, when possible, on human ALS, with support from ALS animal models. We begin with an overview of skeletal muscle, including a review of muscle fiber type, motor units and the neuromuscular synapse. Next, we provide a description of the clinical and biomarker changes that occur in the muscles of patients with ALS. We provide an extensive account of the histopathological changes that are evident in ALS muscle, such as fiber type grouping, muscle inflammation, protein misfolding, mitochondrial dysfunction, and alterations in neuromuscular junctions and muscle satellite cells. Our review then concludes with an update of metabolic and molecular-genetic changes that are found in ALS muscle. The evidence shows that muscle can be an additional target for therapy in ALS, in combination with therapies targeting neurons and glia within the central nervous system (CNS).

Satellite Glial Cells in Peripheral Nerve Injury and Regeneration.

Linjia Hu, Haimin Lu, Yufan Shen +4 more

Satellite glial cells (SGCs) are morphologically unique peripheral glial cells that surround neuronal somas in sensory, sympathetic, and parasympathetic ganglia. Satellite glial cells communicate with neurons that they ensheathe and form a distinct structural and functional unit. Following peripheral nerve injury, satellite glial cells undergo remarkable morphological changes, including gliosis, and help regulate the microenvironment surrounding neuronal somas. The expression of many satellite glial cell markers such as glial fibrillary acidic protein (GFAP) and connexin-43, pro-inflammatory cytokines, and growth factors in satellite glial cells is altered in these cells. Injury responses of satellite glial cells, particularly the activation of peroxisome proliferator-activated receptor α (PPARα), contribute to enhanced axonal regeneration. Targeting satellite glial cells may therefore offer novel therapeutic strategies for the treatment of peripheral nerve injury.

Co-cultured spheroids of piscine cells as building blocks for cultured fish meat.

Yingfei Su, Shengliang Zhang, Yingqi Jiang +3 more

Cultured meat has emerged as a promising approach to enhancing global food sustainability. Current production methods, however, rely mainly on single-cell-type culture, which fails to replicate the complex composition of conventional meat. In this study, a scaffold-free co-culture system was established to generate structured microtissue comprising highly differentiated muscle and fat cells. Co-cultured spheroids with the ratios of piscine adipose-derived stem cells (PADSCs) to piscine satellite cells (PSCs) being 5:5, 7:3 and 9:1 were investigated, and the result showed that all ratios remained robust and highly viable throughout the differentiation process and successfully induced the formation of bionic tissue containing both myotubes and adipocytes. Based on transcriptome data, the cluster analysis indicated that co-cultured spheroids (the ratio of PADSCs to PSCs was 7:3) were similar to the PADSC spheroids, and the expression levels of cell cycle-related and extracellular matrix-related genes were biased towards those of PADSC spheroids. These results indicated that these co-cultured spheroids held promise as building blocks for further tissue assembly and offered a promising approach for scaling up the production of structured cultured fish meat.

Leuprolide Acetate Promotes Sensory Recovery and Modulates Dorsal Root Ganglion Responses After Sciatic Nerve Transection in Rats.

Irma Hernández-Jasso, Denisse Calderón-Vallejo, José Ávila-Mendoza +5 more

Background/Objectives: Sciatic nerve injuries are among the most common classes of peripheral nerve harm and have a strong impact on quality of life, as well as a significant negative economic impact for patients, society, and governments, since they represent a frequent cause of work-related disabilities and sick leave applications. Following nerve injury, neurons, Schwann, and satellite cells undergo marked changes in phenotype, metabolic activity, neuronal survival, nervous transmission, and an exacerbated activation of the inflammatory response. Leuprolide acetate (LA), a clinically available agonist of gonadotropin-releasing hormone (GnRH), has shown clear neurotrophic properties and is considered a novel potential candidate for treating neural injuries, including sciatic nerve pathologies. This study aimed to analyze the effect of LA treatment on sensory function and dorsal root ganglia (DRG) changes in a rat sciatic nerve full-transection (SNT) model. Methods: Variations in cold and heat sensitivity were assessed using the thermal plate test, while DRG tissue sections were examined for modifications in reactive gliosis by immunofluorescence analysis, and axonal transport using a retrograde tracer. Also, changes in the expression of pro-regenerative genes Stat3, Socs3, Fos, Jun, Atf4, and Limk1 were quantified by qPCR. Results: Our results showed that LA treatment exerted a distinct neurotrophic effect, since it promoted the specific recovery of cold sensitivity, improved axonal transport, regulated the inflammatory response, and modulated the exacerbated expression of pro-regenerative genes in the SNT model. Conclusions: These findings indicate that LA therapy may have the potential to improve sensory recovery in patients with sciatic nerve injuries.

Transcriptome Analysis of miRNAs Involved in the Myogenic Differentiation of Goat Skeletal Muscle Satellite Cells.

Runxiao Luo, Tao Zhong, Linjie Wang +4 more

Skeletal muscle myogenesis is a crucial factor influencing meat production in livestock. MicroRNAs (miRNAs) play a significant role in skeletal muscle myogenesis. The objective of this study was to identify key miRNAs involved in the process of goat skeletal muscle satellite cell (MuSC) differentiation into myotubes. We performed miRNA expression profiling analysis during the proliferation phase (cultured in growth medium, GM) and the differentiation phase (cultured in differentiation medium for 1 day and 5 days, classified as DM1 and DM5, respectively) of goat skeletal muscle satellite cells (MuSCs). A total of 1846 miRNAs were identified in MuSC samples, of which 677 differentially expressed miRNAs (DEmiRNAs) were screened through pairwise comparisons across three groups (GM vs. DM1, GM vs. DM5, and DM1 vs. DM5), and the results were further confirmed by a quantitative real-time PCR assay. Time-series expression profiling facilitated the categorization of the DEmiRNAs into eight distinct clusters, one of which demonstrated a significantly downregulated expression pattern (p < 0.05). Functional enrichment analysis revealed that the target genes of DEmiRNAs are involved in several pathways that are critical for myogenesis, including Hippo, TGF-β, MAPK and cell adhesion molecules. Interaction network analysis identified 19 miRNAs and 56 mRNAs associated with muscle cell development. Notably, novel-m0047-5p emerged as a key regulator, exhibiting strong negative correlations (r = -0.88 to -0.89, q < 0.01) with muscle-related target genes FOSB, CPT1B, and MYOZ2. These findings elucidate miRNA-mediated regulatory networks in goat myogenesis and provide candidate molecular targets for genetic improvement of meat production traits.

Inflammaging-induced TRAF3 degradation impairs AMP biosynthesis to drive sarcopenia.

Jinbo Li, Yaning Xing, Jinxiao Fan +10 more

Inflammaging is a recognized driver of age-related pathologies, yet its specific mechanistic link to sarcopenia remains poorly understood. Here, we identified a significant reduction of TNF receptor-associated factor 3 (TRAF3) in myoblasts exposed to aged serum and in skeletal muscles from both aging mice and humans. Genetic deletion of TRAF3 in myocytes or satellite cells induced early-onset sarcopenia and impaired regeneration, independent of non-canonical NF-κB signaling. Mechanistically, TRAF3 maintains energy homeostasis by stabilizing the key metabolic enzyme, adenylosuccinate lyase (ADSL), and its loss impairs AMP biosynthesis and ATP production. Muscle-specific TRAF3 restoration or AMP supplementation rescued sarcopenic phenotypes in TRAF3-deficient mice. Notably, neutrophil-derived transforming growth factor β1 (TGFβ1) caused IAP-mediated ubiquitination and degradation of TRAF3 in aged mice--a process reversible by the IAP inhibitor SM-164. Inducible neutrophil-specific TGFβ1 deletion prevented age-related sarcopenia. Our study establishes that TRAF3 is a key protective factor in muscle aging, and its loss mechanistically links inflammaging to bioenergetic deficits, suggesting new strategies to prevent age-related muscle wasting.

Cellular Senescence in Skeletal Muscle: Myogenic and Non-Myogenic Cell Populations, Mechanisms, and Therapeutic Opportunities.

Konstantinos Papanikolaou, Angad Yadav, Robert T Mankowski +5 more

Skeletal muscle plays a central role in systemic metabolism, physical function, and overall health. Aging and disease diminish the ability of myogenic and non-myogenic skeletal muscle cells to coordinate adaptation and repair, but the mechanisms underlying this decline are not fully understood. Growing evidence implicates cellular senescence, a stress response marked by irreversible cell-cycle arrest and pro-inflammatory signaling, as a key contributor to muscle pathology. In this review, we synthesize current insights into the molecular mechanisms that govern cellular senescence in skeletal muscle, its effects on myogenic and non-myogenic cell populations, and recent technologies that have clarified key aspects of senescence biology. We further explore emerging therapeutic strategies aimed at targeting senescent cells and discuss key knowledge gaps that must be addressed to advance our understanding of senescent myogenic and non-myogenic cells in skeletal muscle.

The expression characteristics of miR-206-3p in musculoskeletal tissue and its clinical significance.

Khan Akhtar Ali, Xuefeng Yuan, Tianxiang Cui +3 more

Osteosarcopenia, a comorbidity of osteoporosis and sarcopenia in the elderly, involves bone-muscle crosstalk, but its core molecular mechanism remains unclear. miR-206 is traditionally considered muscle-specific; this study explores miR-206-3p's expression in musculoskeletal tissue and correlation with clinical parameters.

Involvement of peripheral and central sensitization in prolonged mechanical allodynia of the tongue in a rat.

Saki Kishimoto, Sho Katsura, Yoshie Okamoto +3 more

This study aimed to characterize peripheral and central sensitization in mechanical allodynia of the tongue induced by sleep-related disorders, neuropathic pain, and inflammatory pain. Male rats were exposed to chronic intermittent hypoxia (CIH) for 16 days using an obstructive sleep apnea model. Lingual nerve injury (LNI) was induced to establish a neuropathic tongue pain model, while complete Freund's adjuvant was injected into the tongue to establish a tongue inflammation (TI) model. The expression levels of calcitonin gene-related peptide (CGRP), hypoxia-inducible factor (HIF)-1α, piezo-type mechanosensitive ion channel component 2 (Piezo2), transient receptor potential cation channel subfamily V member 4 (TRPV4), and glial fibrillary acidic protein (GFAP) in the trigeminal ganglion (TG) and cFos in the trigeminal spinal subnucleus were determined using immunohistochemistry on day 16. All CIH, LNI, and TI rat models exhibited prolonged mechanical allodynia of the tongue. CIH and TI increased the number of CGRP-immunoreactive (IR) neurons and HIF-1α-IR cells. However, only CIH increased the number of Piezo2-IR neurons and GFAP-positive satellite glial cells. The number of TRPV4-IR neurons was elevated in the LNI and TI groups but not in the CIH group. Only CIH induced persistent cFos expression in the trigeminal spinal subnucleus caudalis, indicating long-lasting central sensitization. These findings indicate that CIH-induced tongue pain arises through distinct peripheral and central sensitization processes, highlighting the diverse mechanisms underlying chronic mechanical allodynia of the tongue.

Metabolic and cellular physiological differences between embryonic breast and leg muscle satellite cells in chickens.

Jongryun Kim, Jeongeun Lee, Dongjin Yu +6 more

This study aimed to compare the anatomical characteristics of embryonic chicken muscle satellite cells (CMSCs) derived from breast and leg muscles of 18-day-old embryos. We analyzed the cellular behaviors related to proliferation, metabolism, and differentiation capacity. Breast-derived CMSCs exhibited significantly higher proliferation rates and accelerated cell cycle progression, as evidenced by a higher S phase distribution and lower G2/M phase distribution compared to leg-derived CMSCs. In addition, immunofluorescence staining for myogenic regulatory factors revealed that breast-derived CMSCs exhibited higher expression levels of paired box protein 7 (PAX7), consistent with elevated PAX7 and myogenic differentiation 1 (MYOD) mRNA expression compared with leg-derived CMSCs. In contrast, leg-derived CMSCs showed significantly higher expression of myogenin (MYOG). Moreover, leg-derived CMSCs exhibited significantly higher mitochondrial respiratory activity indices, including oxygen consumption rate and basal respiration. In differentiation capacity analysis, the leg-derived CMSCs formed structurally more developed myotubes, and the expression of muscle-specific genes (MYOD, MYOG, MYH1E, MYH7, TNNI1, TNNI2) was also significantly higher. These findings suggest that breast-derived CMSCs exhibit superior proliferative capacity, while leg-derived CMSCs possess enhanced myogenic differentiation potential, as supported by their increased oxidative phosphorylation activity and elevated expression of differentiation-related markers. In conclusion, the anatomical origin of CMSCs significantly influences their proliferative and differentiation capacities as well as their metabolic properties, providing a valuable basis for selecting optimal cell sources for cultured meat production and meat quality improvement.

Transcriptional competence defines the heterochromatin nucleating potential of isolated MSR units.

Yi-Hsuan Lo, Nicholas Shukeir, Galina Erikson +6 more

In mouse cells, constitutive heterochromatin is associated with underlying arrays of A/T-rich DNA repeat elements, called the major satellite repeats (MaSat or MSR). We examine >18,000 MSR copies in mouse ES cells and identify that heterochromatin forms only at transcriptionally competent MSR units. To directly dissect the function of MSR DNA, we insert isolated MSR units into an inert genomic region that is repeat- and gene-free. Insertion of three or more intact MSR units induces heterochromatic histone marks, recruitment of HP1 and incorporation of histone H1. Only transcriptionally competent MSR units, but not permutated MSR variants or LINE1 5'UTR elements, nucleate de novo heterochromatin. MSR-derived transcription is bi-directional and MSR-originating transcripts are attenuated by the RNAPII-associated Integrator complex. Instructively, multi-copy intact MSR units impart an unwound DNA template that facilitates RNAPII engagement. Together, this study uncovers a DNA/RNA-based logic and transcription-coupled mechanism for the nucleation of heterochromatin.

Interaction of Sepsis, Disuse, and Aging on Skeletal Muscle Function and Remodeling in Male and Female Mice.

Diana C Muller, Franccesco P Boeno, Gisienne Reis +7 more

Sepsis is associated with skeletal muscle weakness and atrophy, particularly in older and immobilized patients; however, how sepsis interacts with disuse, reloading, aging, and biological sex remains poorly defined.

Enhancing cultured meat production with ginseng leaf-stem extract: a novel supplementation approach to promote porcine muscle stem cell growth.

Su Min Park, Do Hyun Kim, Hyuk Cheol Kwon +4 more

Ginsenosides, including ginsenoside Re (G-Re), exert muscle-protective effects, and ginseng leaves and stems are rich in G-Re. Cultured meat has garnered attention as a sustainable alternative to conventional meat production methods. The proliferation and differentiation of muscle satellite cells (MuSCs) are essential for meat production. Herein, we examined the effects of ginseng leaf-stem extract (G-LSE) on the growth, differentiation, and myotube formation of MuSCs. The G-Re content of G-LSE was quantified using LC-MS/MS. MuSCs were treated with G-LSE (0, 0.25, 0.5, 1, 2, and 4 mg/mL for 48 h). Cell proliferation and cell cycle analyses were performed. Myogenic factors, the AKT/mammalian target of rapamycin (mTOR) signaling pathway, and FoxO3a were detected using real-time PCR, Western blotting, and immunofluorescence analyses. At concentrations of 0.25, 0.5, and 1 mg/mL, G-LSE enhanced cell proliferation over 7 days in a concentration-dependent manner. G-LSE also promoted cell cycle progression. Treatment with G-LSE increased mRNA levels of myogenic differentiation 1 (MyoD), myogenin (MyoG), and myosin heavy chain after 7 days in a concentration-dependent manner. G-LSE also elevated cellular protein levels of MyoD and MyoG. A 7-day treatment with 1 mg/mL G-LSE maintained high MyoG levels and morphological transition to myotubes. Myogenic proliferation and differentiation were mediated via AKT/mTOR activation and inhibition of FoxO3a nuclear translocation. G-LSE promotes porcine MuSC proliferation and differentiation by activating AKT/mTOR signaling and inhibiting FoxO3a nuclear translocation. G-LSE could be a novel plant-derived supplement for promoting MuSC growth and differentiation, particularly in cultured meat production.