Background Perineural invasion (PNI) is a hallmark of malignancy in solid tumors, including oral squamous cell carcinoma (OSCC), and is closely associated with poor prognosis. Emerging evidence suggests a critical association between PNI and the tumor immune microenvironment (TME). Methods In this study, we used spatial transcriptomics, in vitro and in vivo experiments, and clinical specimen validation to systematically study the interplay between neural infiltration and immune modulation in OSCC. Results Our results suggest that intratumoral nerves may enhance the recruitment of regulatory T cells (Tregs) through C-C motif chemokine ligand 5 (CCL5)-mediated chemotaxis and further promote the acquisition of an immunosuppressive phenotype in Tregs by activating the RAMP1 signaling pathway. Through these coordinated mechanisms, neural components in the TME contribute to immune suppression and facilitate tumor progression. Therapeutically, both combination of CCL5 blockade with anti-CTLA-4 antibody and the combination of RAMP1 blockade with anti-PD-1 antibody exhibited significantly enhanced anti-tumor efficacy. Conclusions This study highlights a previously underappreciated neural-Treg axis in the TME and provides new insights into potential combinatorial strategies for cancer immunotherapy.

Periodontitis is a multifactorial disease characterized by bacterial infection, inflammation, oxidative stress, and dysregulated bone remodeling, for which current treatments remain limited. Effective treatment requires not only antimicrobial action but also coordinated modulation of immune and regenerative pathways within the periodontal microenvironment. Here, this study reports a bioinspired "octopus-like" nanomicelle platform with mucoadhesive tentacles and immune-targeting surfaces, designed to enhance drug retention, selective delivery, and multifunctional therapeutic performance. Phenylboronic acid modification enables high-affinity binding to sialic acids in mucosal glycoproteins, prolonging local retention, while fucoidan decoration directs nanomicelles to scavenger receptors on macrophages and neutrophils, the key effector cells in periodontitis. As cannabidiol was used as a potential therapeutic payload, the nanomicelles exhibited potent antibacterial activity against Staphylococcus aureus and Porphyromonas gingivalis, immune microenvironment remodeling and enhanced osteogenic differentiation in vivo. This dual-targeting strategy suppresses macrophage inflammation and oxidative stress via modulating Toll-like receptor, janus kinase-signal transducer and activator of transcription (JAK-STAT), and advanced glycation end products-Receptor for AGEs (AGE-RAGE) pathways, while also reducing neutrophil extracellular traps formation via regulation of apoptosis, mitochondrial depolarization, and calcium signaling. In a mouse periodontitis model, the platform achieved superior therapeutic outcomes compared to free drug. Together, this multifunctional adhesive nanoplatform integrates mucoadhesion, immune modulation, antimicrobial activity, and bone regeneration, offering a versatile and generalizable strategy for periodontitis therapy and broader microenvironmental reprogramming in chronic inflammatory diseases.

Chronic osteomyelitis presents substantial prognostic challenges due to its high recurrence risk and complex postoperative recovery trajectories. Traditional assessment often relies on manual scoring systems, which limit scalability, efficiency, and consistency in clinical practice. Furthermore, the heterogeneous nature of clinical data poses challenges for current multimodal learning approaches that require aligned inputs and large annotated datasets. In this work, we propose RAG4Outcome, a retrieval-augmented generation (RAG) framework for prognostic prediction in chronic osteomyelitis. Our method integrates multimodal clinical data, including PET-CT imaging reports, structured surgical and diagnostic records, and unstructured follow-up notes, into a unified prediction pipeline. By combining a domain-specific retrieval corpus with expert-guided prompting, the framework enables more interpretable, evidence-grounded, and clinically reliable prognosis. Preliminary results on real-world cases demonstrate promising effectiveness and clinical alignment, highlighting the potential of RAG4Outcome for AI-assisted infection management and postoperative decision support.

Angiogenesis after ischemic brain injury contributes to the restoration of blood supply in the ischemic zone. Strategies to improve angiogenesis may facilitate the function recovery after stroke. Recent researches have demonstrated that dysfunction of long non-coding RNAs are associated with angiogenesis. We have previously reported that long non-coding RNAs (lncRNAs) are aberrantly expressed in ischemic stroke. However, little is known about long non-coding RNAs and theirs role in angiogenesis after stroke. In this study, we identified a rat lncRNAs, Meg3, and found that Meg3 was significantly decreased after ischemic stroke. Overexpression of Meg3 suppressed functional recovery and decreased capillary density after ischemic stroke. Downregulation of Meg3 ameliorated brain lesion and increased angiogenesis after ischemic stroke. Silencing of Meg3 resulted in a proangiogenic effect evidenced by increased endothelial cell migration, proliferation, sprouting, and tube formation. Mechanistically, we showed that Meg3 negatively regulated notch pathway both in vivo and in vitro. Inhibition of notch signaling in endothelial cells reversed the proangiogenic effect induced by Meg3 downregulation. This study revealed the function of Meg3 in ischemic stroke and elucidated its mechanism in angiogenesis after ischemic stroke.

As we know, Cervical cancer is one of the leading causes of mortality among women worldwide. Magnesium ions (Mg2+) are intricately involved in virtually all biological processes, demonstrating antitumor properties. However, the pathways through which Mg2+ mediates its antitumor activity in cervical carcinoma require further clarification. This research investigated the effects of varying Mg2+ levels on the growth and programmed cell death of cervical cancer cells (SiHa and HeLa). Mg2+ effectively suppressed cellular proliferation, migration, and invasive capacity, and induced the proportion of the G0/G1 phase. Meanwhile, Apoptosis was also promoted in cervical cancer cell lines following Mg2+ exposure, proceeding through mitochondrial dysfunction characterized by an elevated Bax/Bcl-2 ratio, reduced membrane potential, cytochrome c leakage, and caspase-3 activation. Significant downregulation of phosphorylated PI3K, AKT, and FoxO1 following Mg2+ treatment was observed, while PI3K activator administration attenuated Mg2+-induced apoptosis. Mg2+-mediated suppression of HeLa xenograft tumor growth in nude mice was demonstrated, with immunohistochemical confirmation of elevated TdT-mediated dUTP Nick-End Labeling(TUNEL) and cleaved caspase-3 expression. These results suggest that Mg2+, which was highly concentrated in the region, exerts its anti-cervical cancer function through downregulation of the PI3K/AKT/FoxO1 pathway, which offers a novel method and mechanism to therapeutic strategies for managing cervical cancer.

Adoptive natural killer (NK) cell therapy for solid tumors faces critical challenges, including tumor antigen heterogeneity, impaired tumor infiltration, and suboptimal activation imposed by the immunosuppressive microenvironment. Here we developed an engineered nanoplatform featuring transmembrane DNA nanochannel-engineered artificial receptors (NCAR) to direct NK cells against solid tumors through two synergistic mechanisms: 1) Tumor Microenvironment (TME) Reprogramming: leveraging cholesterol-mediated insertion, NCAR incorporates into tumor membranes to disrupt phospholipid bilayers, inducing immunogenic cell death with the release of damage-associated molecular patterns (DAMPs; e.g., HMGB1, CRT), which remodels immunosuppression TME and recruits/activates NK cells. 2) Precision Targeting: NCAR forms programmable synthetic immune synapses with DNA nanoartificial ligands (NAL) engineered on NK cells via base-pairing. This antigen-independent assembly network establishes a universal membrane interface, enabling sustained tumor-targeted NK cell activation. The dual-component system enables sustained intratumoral accumulation of NK cells (>96 h), with a 15.1-fold increase in activated NKP46(+)GZB(+) NK cells compared to controls. By bridging DNA nanotechnology with cell immunotherapy, our nanoplatform provides a universal strategy for navigating tumor-immune interactions, addressing key limitations of adoptive NK cell immunotherapy in solid tumors.

DNA origami holds great potential for advancing therapeutics, but the lack of methods for the precise assessment of structural integrity in vivo prevents its translation. Here we introduce proximity ligation assay for structural tracking and integrity quantification (PLASTIQ) for resolving origami structural integrity with only 1 µl of blood sample and with a detection limit of 0.01 fM. Through PLASTIQ, we could observe and quantify the dynamics of DNA origami degradation during blood circulation and evaluate the effectiveness of PEGylation for slowing this process in a murine model. Additionally, by using a double-layered barrel-like origami structure, we found distinct degradation kinetics of DNA helices depending on their specific location, revealing the slower degradation of internal helices compared with the outer ones. Our results suggest that PLASTIQ offers a quantitative approach for assessing DNA origami integrity in vivo by longitudinal sampling, providing dynamic pharmaceutical-level insights for accelerating the development of DNA-nanostructure-based therapeutic molecules and drugs. A label-free, DNA-based proximity ligation assay that uses ligatable staple pairs enables the longitudinal quantification of DNA origami structural stability dynamics in vivo, with single-helix resolution for both wireframe and lattice designs.

BACKGROUND:CPT1 (carnitine palmitoyltransferase 1) is a rate-limiting enzyme for long-chain fatty acid oxidation. In adult hearts, CPT1b predominates, while CPT1a is coexpressed at lower levels. Pathological stress on the heart induces CPT1a expression, coinciding with a reduction in fatty acid oxidation, yet the role of CPT1a in pathological remodeling is unknown. METHODS:CPT1 isoform expression was assayed in the myocardium of patients with heart failure with nonischemic cardiomyopathy and a preclinical mouse model of heart failure. Mice were subjected to afterload stress via transverse aortic constriction (TAC) or sham surgery (sham) with cardiac-specific CPT1a knockdown or cardiac-specific, adeno-associated virus serotype 9 (AAV9)-mediated CPT1a overexpression (AAV9.cTnT [cardiac troponin T].Cpt1a) versus empty virus or PBS infusions as controls. MicroRNA 370, known to suppress hepatic CPT1a, was assayed and overexpressed to determine if microRNA 370 regulates cardiac CPT1a expression. RESULTS:CPT1a protein was elevated and microRNA 370 reduced in the myocardium of male and female patients with nonischemic cardiomyopathy, as well as in failing mouse hearts. AAV9-mediated microRNA 370 overexpression in mouse hearts suppressed CPT1a expression and attenuated the response of CPT1a to TAC. Preventing CPT1a upregulation in response to TAC in cardiac-specific CPT1a knockout mice exacerbated adverse remodeling, severe dysfunction, and increased mortality. In contrast, CPT1a overexpression (2.8-fold) attenuated impaired ejection fraction (by 54%) versus control TAC hearts (P<0.05). Delivery of AAV9.cTnT.Cpt1a 4 weeks after TAC surgery led to significant rescue of ejection fraction and mitigated the exacerbated dysfunction of cardiac-specific CPT1a knockout mice TAC hearts. RNA-seq revealed a novel function of CPT1a in suppressing hypertrophic, profibrotic, and cell death gene programs in both sham and TAC hearts, irrespective of changes in fatty acid oxidation, with reduced histone acetylation. CONCLUSIONS: The effects of CPT1a in the heart extend beyond fatty acid oxidation including noncanonical regulation of gene programs. CPT1a upregulation occurs in nonischemic cardiomyopathy and is a critical cardioprotective adaptation to pathological stress.

Gene expression is governed by dynamic switches between repressive and activating transcriptional states. Among the molecules mediating these transitions, chromatin readers and transcription factors play pivotal roles. However, how they assemble with regulatory machineries to enable crosstalk between gene repression and activation remains unknown. Here, we use an integrative structural dynamics approach - combining cryo-EM, crosslinking mass spectrometry, fragment-resolved protein interactome mapping and crystallography - to show how the dual-role chromatin reader Cti6 and transcription factors Ash1 and Ume6 engage the Sin3 deacetylase complex, a major regulatory hub in eukaryotes. We find that Cti6 competes with Ash1 to drive its dynamic recruitment to a shared peripheral module, while Ume6 engages the Sin3 scaffold through a defined, minimal interface. Using high-throughput mutational scanning, we reveal deleterious and gain-of-function mutations in Sin3, identifying evolutionarily conserved residues essential for anchoring transcription factors. Together, these results provide structural and functional insights into how dual-role regulators engage the central Sin3 complex, revealing subtle assembly principles that may facilitate crosstalk between gene repression and activation. They also establish an integrative multidisciplinary framework to dissect the dynamics of macromolecular assemblies across biological systems.

Abstract Background: Previous MCED studies have shown differences in detection performance across different aggressiveness cancer subtypes. However, the methylation profiles and ctDNA shedding patterns across different aggressiveness subtypes have not been analyzed. We systematically compared cell-free DNA (cfDNA) tumor fraction and methylation profiles across aggressiveness subtypes in three cancers to identify epigenetic biomarkers that differentiate tumor aggressiveness. Methods: Blood samples from a case-control study (NCT06217900) including 757 cancer cases were analyzed using a targeted methylation-based MCED test. We compared cfDNA tumor fraction and methylation profiles between aggressiveness subtypes, stage-matched small cell lung cancer (SCLC) (n=137) versus (vs) non-small cell lung cancer (NSCLC) (n=137), stage I invasive lung adenocarcinoma (IAC) (n=81) vs minimally invasive adenocarcinoma (MIA) (n=81), triple-negative breast cancer (TNBC) (n=151) vs non-TNBC (n=151), and intermediate/high-grade prostate cancer (Gleason Grade Group[GG] 2-5)(n=14) vs low-grade prostate cancer(GG1)(n=6). Tumor fraction was estimated using zero-inflated Negative Binomial model based on the distribution of methylation signals, and methylation profiles were assessed by calculating methylated/unmethylated (M/U) ratios between different aggressiveness subtypes. Marker comparisons were conducted using the Mann-Whitney U test with multiple testing corrections. Results: The sensitivity is higher in more aggressive subtypes (lung cancer 93.73% vs SCLC 99.27%; breast cancer 70.76% vs TNBC 81.46%; prostate cancer 40% vs intermediate and high-grad prostate cancer 42.86%). In lung cancer, SCLC exhibited a significantly higher cfDNA tumor fraction than NSCLC (Wilcoxon p = 2.24×10-24), with 41,136 markers (79.18% hypomethylated) showing elevated M/U ratios. In stage I lung adenocarcinoma, IAC demonstrated a higher cfDNA tumor fraction (Wilcoxon p = 2.55×10-6) and systematic methylation differences compared to MIA. In breast cancer, TNBC had a significantly higher cfDNA tumor fraction than non-TNBC (Wilcoxon p = 2.55×10-6), with 25,717 markers (80.32% hypomethylated) displaying increased M/U ratios. Among prostate cancers of the same stage, GG2-5 cases exhibited higher cfDNA tumor fraction (Wilcoxon p = 0.038) and systematic methylation alterations compared to GG1. Conclusion: Aggressive tumor subtypes consistently display elevated cfDNA tumor fraction and characteristic methylation profiles marked by increased M/U ratios. These findings suggest cfDNA methylation patterns are promising epigenetic biomarkers for distinguishing tumor aggressiveness, potentially improving cancer screening precision and reducing overdiagnosis. Citation Format: Kezhong Chen, Jian Huang, Dahong Zhang, Shu Wang, Hongxu Liu, Wenzhao Zhong, Xiangnan Li, Qiang Zhang, Zhigao Li, Jiaqi Liu, Ziqing Tian, Fei Zhou, Gongsheng Jin, Xudong Xiang, Zhigang Li, Hui Xie, Ya Wei, Guochun Zhang, Guolin Ye, Ming Cai, Junfeng Wang, Yan Zhang, Chao Cheng, Hefei Li, Desong Yang, Jianhong Lian, Sheng Huang, Tao Xu, Zengjun Wang, Xi Guo, Zhuowei Liu, Minfeng Chen, Yang Wang, Yue An, Yanzhan Yang, Min Li, Jing Liu, Baoliang Zhu, Yonghui Li, Xiaohui Wu, Fan Yang, Jun Wang. Epigenetic profiling identifies markers of aggressive cancer subtype using a targeted methylation-based multi-cancer early detection (MCED) blood test [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1100.

Breast-conserving therapy is currently the preferred treatment option for early-stage breast cancer; however, it can still cause breast deformity and asymmetry, compromising aesthetics. Cell-free tissue engineering, a strategy with high translational potential, offers a promising solution to this issue. To fabricate cell-free scaffolds for adipose tissue regeneration, this study developed a novel composite bioactive hydrogel based on methacrylated hyaluronic acid (HAMA), methacrylated silk fibroin (SFMA), and platelet-rich plasma (PRP). The integration of HAMA and SFMA imparted the composite hydrogel with excellent mechanical strength and printability, while the incorporation of PRP further enhanced the bioactivity. The HAMA/SFMA/PRP (HSP) composite bioactive hydrogels exhibit favorable biocompatibility, cell adhesion, and the ability to promote proliferation, migration, adipogenic differentiation, and angiogenesis. In vivo evaluation using rabbit adipose tissue defect models showed that scaffolds printed with the HSP hydrogel containing 50% PRP safely and effectively promoted adipose tissue regeneration. Overall, the HSP hydrogel scaffold demonstrates excellent capability for adipose tissue regeneration and holds significant potential in breast defect repair.

BACKGROUND:Glucocorticoid (GC)-induced osteonecrosis of the femoral head (ONFH) involves bone marrow-derived mesenchymal stem cell (BMSC) apoptosis and dysregulated osteo-adipogenic differentiation. While aberrant H19 promoter methylation and expression have been linked to various bone metabolic disorders such as osteoporosis and osteosarcoma, their specific role in the pathogenesis of GC-induced ONFH remains largely unexplored. METHODS:We analyzed H19 promoter methylation, DNMTs, and H19 expression in human ONFH BMSCs. Roles of Dnmt1 and H19 in osteogenic/adipogenic differentiation were assessed using staining (Alizarin Red/Oil Red O) and pathway analysis. Effects of Dnmt1 knockdown or H19 overexpression were tested via BMSC implantation in a GC-induced ONFH rat model. RESULTS:H19 promoter hypomethylation caused H19 overexpression in undifferentiated GC-ONFH BMSCs; expression decreased upon differentiation. H19 and Dnmt1 expression were negatively correlated. Dnmt1 predominated among DNMTs in epigenetically regulating H19 and reciprocally modulated differentiation (inhibiting osteogenesis, promoting adipogenesis). Conversely, H19 promoted osteogenesis and inhibited adipogenesis by suppressing GSK-3β, activating Wnt/β-catenin signaling. In the rat model, implanted BMSCs with Dnmt1 knockdown or H19 overexpression reduced empty lacunae, corrected the osteo-adipogenic imbalance, and delayed progression. CONCLUSION:The Dnmt1/H19/GSK-3β axis reciprocally regulates BMSC osteogenic and adipogenic differentiation in GC-induced ONFH, representing a novel epigenetic mechanism underlying GC-induced ONFH and a promising MSC-based therapeutic strategy for early-stage disease.

BACKGROUND:Approximately 10% of the patients with infantile hemangioma (IH) may exhibit resistance to propranolol (PRN) therapy, and thus alternative strategies are required. Our previous studies reported that oxymatrine (OMT) could inhibit the growth of hemangiomas, however the underlying pharmacological actions have not been fully addressed. METHODS:In this study, a murine IH model was constructed by implantation of EOMA cells into nude mice. OMT was administrated (50 mg/kg; i.p) for 21 days. Metabolic changes were examined by proteomics and metabolomics, followed by in vitro experimental validation using EOMA cells. RESULTS:OMT significantly suppressed the growth of hemangioma in vivo without significant adverse effects. A total of 869 differentially expressed proteins and 38 metabolites were identified. In addition to canonical apoptosis regulation, OMT also caused significant metabolic disturbances, particularly in purine and pyrimidine metabolism. Furthermore, ferroptosis may be involved in the therapeutic effect of OMT. In the validation experiments in vitro, we found that OMT dose-dependently reduced the viability of EOMA cells, concomitant with increased production of lipid reactive oxygen species (ROS) and Fe2 + accumulation. CONCLUSIONS:In conclusion, these findings suggested that treatment with OMT could suppress the growth of hemangiomas through metabolic disturbances and inducing ferroptosis, which may provide new insights to the management of IH.

Interpreting missense variants in highly conserved, paralog-rich gene families remains a major barrier to understanding protein function and advancing precision medicine. Here, we combine comprehensive mutagenesis, high-content live-cell imaging, and automated quantitative analysis to generate a complete functional atlas of all 2,683 single-nucleotide coding variants in human α-tubulin TUBA1A. Systematic profiling of microtubule assembly revealed distinct classes of mutations that disrupt folding, chaperone engagement, and protofilament geometry, defining structural constraints that govern tubulin function. This approach complements conservation-based predictors and enables functional reinterpretation of disease-associated variants. Molecular dynamics simulations further revealed how local perturbations in GTP (guanosine triphosphate) binding, dimer contacts, and lateral interfaces propagate to alter filament architecture. Integrating these datasets produced a predictive framework that generalizes across tubulin isotypes and species, enabling accurate inference of variant effects. This work establishes a scalable, high-resolution strategy for functional annotation of conserved proteins and provides mechanistic insight into cytoskeletal assembly and flexibility.

Genotype-phenotype relationships are mediated through intricate networks of physical and functional interactions among macromolecules. Knowledge of the interactome is vital to understand and model genetics and cellular biology. Recent advances in accurately predicting tertiary protein structures using artificial intelligence (AI) approaches such as AlphaFold1 have revived the vision that the protein-protein interactome might be fully predictable through computational modeling of quaternary structures. Here we present a comprehensive experimental framework to systematically assess the impact of AI-driven interactome predictions for yeast2 and human3. We find that the quality of high-confidence predictions is on par with established experimental approaches. However, in proteome-wide screening, the tested AI approaches underperform in the discovery of strictly novel protein-protein interactions (PPIs) compared to experimental reference interactome maps. In particular, the yeast interactome map described here identifies >40-fold more novel PPIs than its AI counterpart. Strikingly, AlphaFold provides structural models for a substantial number of experimentally identified PPIs missed by the virtual screens. Our results suggest that, at this stage, the main contribution of AI predictions is to provide quaternary structure models for experimentally identified PPIs.

Hirschsprung disease (HSCR) is a life-threatening congenital disorder characterized by defective migration and proliferation of enteric neural crest cells (ENCCs), a process in which the receptor tyrosine kinase RET plays a central regulatory role. However, the regulatory mechanisms governing RET expression remain incompletely understood. In the present study, we identify the circular RNA circRET (hsa_circ_0093651) as a critical translational regulator of its parental gene RET and explore its role in HSCR pathogenesis. We demonstrate that circRET is significantly downregulated in the colonic tissues from patients with HSCR and directly binds RET mRNA, modulating RET protein synthesis at the translational level without altering mRNA abundance. Notably, circRET interacts with the coding region of RET mRNA, potentially facilitating its translation. circRET knockdown suppresses RET/AKT signaling and impairs ENCC migration, proliferation, and distal colon colonization. Our study uncovers a novel circRET-mediated regulatory mechanism of its parental gene RET, with potential implications for therapeutic targeting in HSCR.NEW & NOTEWORTHY Hirschsprung disease (HSCR) results from impaired enteric neural crest cell development, in which RET plays a central role. Here, we identify circRET as a translational regulator of RET. circRET is downregulated in HSCR tissues and modulates RET protein by directly binding RET mRNA without affecting mRNA levels. Loss of circRET suppresses RET/AKT signaling and ENCC migration, highlighting a novel regulatory mechanism with therapeutic relevance.