👤 Guangfeng Zhao

MicroRNAs (miRNAs) are associated with amyloid-β (Aβ) dysmetabolism, a pivotal factor in the pathogenesis of Alzheimer's disease (AD). This study unveiled a novel miRNA, microRNA-32533 (miR-32533), featuring a distinctive base sequence identified through RNA sequencing of the APPswe/PSEN1dE9 (APP/PS1) mouse brain. Its role and underlying mechanisms were subsequently explored. Bioinformatics and confirmatory experiments revealed that miR-32533 had a novel 23-base sequence with minimal coding potential, functioning within the Drosha ribonuclease III (Drosha)/Dicer 1, ribonuclease III (Dicer)-dependent canonical pathway and identifiable via northern blot. miR-32533 was abundantly brain-distributed and downregulated in diverse AD-related models, including APP/PS1 and five familial AD (5×FAD) mouse brains and AD patient plasma. Overexpression or inhibition of miR-32533 led to improvements or exacerbations in cognitive dysfunction, respectively, by modulating Aβ production, apoptosis, oxidation, and neuroinflammation through targeting cAMP-responsive element binding protein 5 (CREB5), which interacted with α disintegrin and metalloproteinase 10 (ADAM10), beta-site amyloid precursor protein cleaving enzyme 1 (BACE1), and presenilin 1 (PS1) promoters, thereby enhancing Aβ production through BACE1 and PS1 upregulation while suppressing non-amyloidogenic amyloid precursor protein (APP) processing via ADAM10 downregulation. Furthermore, modulation of the miR-32533/CREB5 axis ameliorated or worsened cognitive impairment by inhibiting or amplifying Aβ overproduction through the BACE1-involved amyloidogenic and ADAM10-involved non-amyloidogenic pathways. Overall, the findings suggest miR-32533 as a regulator of Aβ metabolism, oxidative stress, and neuroinflammation, establishing the miR-32533/CREB5 signaling pathways as potential therapeutic targets for combating Aβ accumulation and cognitive deficits in AD. Show less

Interferon-induced transmembrane protein 3 (IFITM3) modulates γ-secretase in Alzheimer's Disease (AD). Although IFITM3 knockout reduces amyloid β protein (Aβ) production, its cell-specific effect on AD remains unclear. Single nucleus RNA sequencing (snRNA-seq) was used to assess IFITM3 expression. Adeno-associated virus-BI30 (AAV-BI30) was injected to reduce IFITM3 expression in the cerebrovascular endothelial cells (CVECs). The effects on AD phenotypes in cells and AD mice were examined through behavioral tests, two-photon imaging, flow cytometry, Western blot, immunohistochemistry, and quantitative polymerase chain reaction assay (qPCR). IFITM3 expression was increased in the CVECs of patients with AD. Overexpression of IFITM3 in primary endothelial cells enhanced Aβ generation through regulating beta-site APP cleaving enzyme 1 (BACE1) and γ-secretase. Aβ further increased IFITM3 expression, creating a vicious cycle. Knockdown of IFITM3 in CVECs decreased Aβ accumulation within cerebrovascular walls, reduced Alzheimer's-related pathology, and improved cognitive performance in AD transgenic mice. Knockdown of IFITM3 in CVECs alleviates AD pathology and cognitive impairment. Targeting cerebrovascular endothelial IFITM3 holds promise for AD treatment. Interferon-induced transmembrane protein 3 (IFITM3) expression was increased in the cerebrovascular endothelial cells (CVECs) of patients with Alzheimer's Disease (AD). Cerebrovascular endothelial IFITM3 regulates amyloid β protein (Aβ) generation through regulating beta-site APP cleaving enzyme 1 (BACE1) and γ-secretase. Knockdown of IFITM3 in CVECs reduces Aβ deposits and improves cognitive impairments in AD transgenic mice. Cerebrovascular endothelial IFITM3 could be a potential target for the treatment of AD. Show less

The β-site amyloid precursor protein-cleaving enzyme 1 (BACE1) gene polymorphism (rs638405) has been widely reported to be associated with Alzheimer's disease (AD) risk. However, studies on the relationship between BACE1 gene polymorphism (rs638405), brain volume, and cognition in AD patients remain scarce. To investigate the effect of genetic polymorphism in BACE1 on gray matter volume (GMV) and cognition in AD, this study recruited 111 cognitively unimpaired (CU) controls and 144 AD patients. The effect of BACE1 rs638405 polymorphism on cognition was explored in CU and AD groups. Then the interaction effect of the diagnosis and BACE1 rs638405 polymorphism on GMV was performed, following the post-hoc analysis of regions of interest (ROIs) in interaction analysis. Mediation analysis was used to elucidate the relationship among genotypes, ROIs and cognition. BACE1 rs638405 G carriers (BACE1 G+) showed significantly lower scores in global cognition and memory function than noncarriers (BACE1 G-) in AD group. Genotypes (G+/G-) and diagnosis (CU/AD) have interaction on GMV of medial temporal lobe (MTL) including the left parahippocampus and right hippocampus. Post-hoc analysis revealed that BACE1 G+ exhibited significantly lower GMV in ROIs compared to BACE1 G- in AD. Finally, mediation analysis further demonstrated that the GMV of ROIs mediated the effect of BACE1 rs638405 polymorphism on cognition in AD. Our results emphasize the BACE1 rs638405 gene polymorphisms may affect the GMV of MTL and cognition in AD, deepening the understanding of AD pathogenesis. Show less

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by hyperphosphorylation of tau, neuroinflammation, and amyloid-beta (Aβ) plaques. Lead (Pb) exposure has been linked to an increased risk of AD and neuroinflammation. The purpose of this study is to determine if black soybean peptide (BSP1) may reduce neuroinflammation caused by Pb and associated AD-like pathology. Pb exposure was given to mouse hippocampus HT22 cells in the presence or absence of BSP1, positive control resveratrol (Rsv), or the SIRT1 inhibitor EX-527. Our findings suggest that BSP1 downregulates the expression of beta-secretase (BACE1) and amyloid precursor protein (APP), inhibits tau phosphorylation, and reduces Aβ1-42 deposition. In addition, BSP1 effectively alleviated Pb-induced neuroinflammation by reducing the phosphorylation of NF-κB and the expression of pro-inflammatory cytokines (IL-1β, TNF-α, NLRP3, and IL-18). BSP1 provides neuroprotective effect via phosphorylating LKB1 and AMPK, inhibiting mTOR signaling, and activating the AMPK/SIRT1 pathway. These results suggest that BSP1 may be therapeutically beneficial for preventing or treating AD by reducing Pb-induced neuroinflammation. Show less

Osteosarcoma, an aggressive bone malignancy predominantly affecting children and adolescents, is characterized by a poor prognosis and high mortality rates. The development of reliable prognostic tools is critical for advancing personalized treatment strategies. However, identifying robust gene signatures to predict osteosarcoma outcomes remains a significant challenge. In this study, we analyzed gene expression data from 138 osteosarcoma samples across two multicenter cohorts and identified 14 consensus prognosis-associated genes via univariate Cox regression analysis. Using 66 combinations of 10 machine learning (ML) algorithms, we developed a machine learning-derived prognostic signature (MLDPS) optimized by the average C-index across TARGET, GSE21257, and merged cohorts. The MLDPS effectively stratified osteosarcoma patients into high- and low-risk score groups, achieving strong predictive performance for 1-, 3-, and 5-year overall survival (AUC range: 0.852 - 0.963). The MLDPS, comprising seven genes (CTNNBIP1, CORT, DLX2, TERT, BBS4, SLC7A1, NKX2-3), exhibited superior predictive accuracy compared to 10 established gene signatures. The findings of the MLDPS carry significant clinical implications for osteosarcoma treatment. Patients with a high-risk score demonstrated worse prognosis, increased metastasis risk, reduced immune infiltrations, and greater sensitivity to immunotherapy. Conversely, low-risk patients exhibited prolonged survival and distinct drug sensitivities. These findings underscore the potential of MLDPS to guide risk stratification, inform personalized therapeutic strategies, and improve clinical management in osteosarcoma. Show less

By integrating single-cell and bulk RNA-sequencing data for esophageal cancer (ESCA), we developed and validated a seven-macrophage-gene prognostic signature (FCN1, SCARB2, ATF5, PHLDA2, GLIPR1, CHORDC1, and BCKDK). This signature effectively stratified patients into high- and low-risk groups with significantly different overall survival, achieving area under the curve (AUC) values greater than 0.7 for 1-, 2-, and 3-year survival prediction. A high-risk status correlated with an immunosuppressive tumor microenvironment, characterized by lower infiltration of B cells and CD8 + T cells, and was associated with reduced sensitivity to multiple chemotherapeutic agents, including Cisplatin and 5-Fluorouracil. Conversely, a low-risk status was linked to greater immune cell infiltration and higher predicted chemosensitivity. At the single-cell level, pseudotime analysis revealed that macrophage maturation significantly correlated with a decreasing risk score, suggesting that mature macrophages may contribute to a favorable prognosis. Furthermore, cell communication analysis identified high-risk macrophages as dominant drivers of a pro-tumorigenic microenvironment via signaling pathways, such as SPP1 and complement. In conclusion, this seven-gene signature is a robust prognostic biomarker that offers a new strategy for personalized risk assessment and treatment selection in ESCA. The online version contains supplementary material available at 10.1007/s13205-025-04452-w. Show less

Rubia cordifolia L. (RCL) is a widely used medicinal with a long history. It exhibits anti-inflammatory and antioxidant properties and prevents apoptosis. While there is growing evidence that exhausted exercise (EE) might cause cardiac damage, RCL has been shown to provide cardioprotective effects. The effects and mechanisms of RCL on exercise-induced myocardial injury remain unclear. In this study, we tested the RCL extract using a rat model of exhausted swimming. We evaluated the therapeutic effect of RCL on exercise-induced myocardial damage using PCR, ELISA, hematoxylin-eosin (H&E) staining, DHE staining, and other methods. UPLC-Q-TOF-MS was employed to identify the components of the RCL extract and its blood-entry components, and network pharmacology was constructed. LC-MS was utilized to investigate left ventricular metabolomics. These two approaches were combined to predict the possible metabolic pathways regulated by RCL. Finally, the targets of the metabolic pathway were verified using molecular docking and western blot analysis. The findings suggest that rubioncolin B, 4-hydroxy-2-carbexyanthraquinone, and 9-Oxo-9H-xanthene-4-carboxylic acid may be the primary active compounds of RCL. RCL promotes the degradation pathway of branched-chain amino acids (BCAA), including valine, leucine, and isoleucine, regulates the proteins BCAT2 and BCKDK, reduces pathological injuries, inflammation, oxidative stress, and collagen deposition, and mitigates the effects of exhaustion-induced myocardial injuries by influencing the key target AKR1C1 and the metabolite L-Valine. This study provides a foundation for the development of RCL as a sports supplement to alleviate EE-induced myocardial injury. Show less

Neuroblastoma (NB) is the most common solid tumor in children, characterized by high recurrence rates, drug resistance, and significant mortality. In this study, we analyzed the proteomic profiles of NB tissue samples alongside other pathological categories, including ganglioneuroma (GN) and ganglioneuroblastoma (GNB). Using weighted gene co-expression network analysis (WGCNA), the core prognostic gene models associated with histopathology of NB were identified. Furthermore, by mapping our core prognostic gene models onto drug-perturbed transcriptome profiles from the L1000FWD and CMap databases, repurposing drug candidates were screened and validated for NB. Our proteomic analysis reveals that pathways associated with the cell cycle and DNA replication are significantly upregulated in NB, while oxidative phosphorylation, pyruvate metabolism, and the TCA cycle are notably downregulated compared to GNB and GN. By applying WGCNA, we identified a core prognostic gene model strongly associated with the unfavorable subtype and high MKI of NB and primarily related to chromatin binding and mRNA metabolic process. Protein-protein interaction network analysis identified 15 hub genes in this core prognostic module: SMARCA4, SMARCA5, SMARCC2, SMARCC1, PBRM1, BRD3, ARID1A, BRD2, ARID1B, KDM1A, TP53BP1, ALYREF, CBX1, SF3B1, and ADNP, which mainly related to chromatin remodeling. Notably, SMARCA4 and ALYREF are also high-risk genes of mortality and validated as potential prognostic biomarkers for NB. Through repurposing drugs screening, mocetinostat and clofarabine were validated as effective treatments in two NB cell lines. Mocetinostat and clofarabine offer valuable insights for the development of novel targeted therapies in neuroblastoma. Show less

Obesity and autoimmune disorders represent a significant comorbidity burden, yet their shared genetic architecture is not fully understood. Elucidating the pleiotropic genetic basis underlying both conditions is crucial for unraveling the mechanisms driving their co-occurrence and advancing therapeutic strategies. We conducted a large-scale cross-trait analysis integrating genome-wide association study (GWAS) summary data for obesity and 17 autoimmune diseases. Genetic correlations were assessed using LD score regression and high-definition likelihood. Cross-trait pleiotropic analysis was performed using Stratified Pleiotropic Locus Mapping (PLACO) to identify shared loci, followed by Bayesian colocalization to confirm shared causal variants. Gene-level and tissue-specific heritability analyses were conducted, and drug targets were prioritized via summary-based Mendelian randomization (SMR). Finally, immune co-localization and bidirectional Mendelian randomization were employed to elucidate immunological mechanisms and causal relationships. Our analysis identified eight autoimmune diseases with significant genetic correlations to obesity. We discovered 10,324 pleiotropic SNPs, which mapped to 52 independent risk loci, with nine loci confirmed as shared causal variants by colocalization. Gene-level analysis revealed 133 unique pleiotropic genes, including CLN3, SH2B1, and MMEL1, enriched in pathways of hematopoietic cell differentiation and immune homeostasis. Tissue-specific heritability was most prominent in the spleen, whole blood, and EBV-transformed lymphocytes. Immuno-co-localization implicated six IgD+ CD38- %B cell-related traits as key pathological conduits. Bidirectional Mendelian randomization established a causal role of obesity in hypothyroidism, psoriasis, and multiple sclerosis, while revealing an inverse causal association of type 1 diabetes with obesity risk. This study demonstrates a robust shared genetic foundation between obesity and multiple autoimmune diseases, pinpointing specific pleiotropic loci, genes, and immune cell subsets. Our findings provide a mechanistic framework for their comorbidity and highlight potential targets for therapeutic intervention. The online version contains supplementary material available at 10.1186/s12967-025-07422-1. Show less

Ochratoxin A (OTA) is a core environmental toxin that induces kidney injury by interfering with glomerular filtration, antioxidant defense, and tubular transport functions. Alginate oligosaccharides (AOS), as active substances from marine, carry natural antioxidant, anti-inflammatory and other biological activities. The purpose of this study is to explore the molecular network of AOS against nephrotoxicity caused by OTA. A total of 36 5-week male mice were randomly divided into three groups: the CON group, the OTA group (250 μg/kg B.W. OTA) and the AOS + OTA group (400 mg/kg B.W. AOS +250 μg/kg B.W. OTA). The treatment was continued for 21 d. OTA induced renal injury in mice, manifested by glomerular capsule blurring, lymphocytic infiltration, and mitochondrial damage in tubular epithelial cells. Treatment with AOS significantly alleviated these pathological changes. Multi‑omics analysis revealed that AOS activated the PPAR signaling pathway, upregulating key genes (Aldehyde Dehydrogenase 1 Family Member A3 ( This study reveal that AOS antagonizes OTA-induced nephrotoxicity in mice through PPAR signaling axis, thus providing new insight into the renal protection mechanism of marine active substances. Show less

Lung adenocarcinoma (LUAD) is a major cause of cancer-related morbidity and mortality globally, with challenges in prognosis and treatment due to its complex pathogenesis and heterogeneous tumor microenvironment (TME). Neutrophil extracellular traps (NETs) and oxidative stress play critical roles in tumor progression: NETs promote tumor cell adhesion, migration, and immune suppression, while oxidative stress induces DNA damage and activates pro-tumor signaling pathways. Moreover, oxidative stress is an important inducer of NETs, and their crosstalk shapes the LUAD immune microenvironment. However, systematic exploration of LUAD immunotherapeutic response prediction based on NETs and oxidative stress-related genes remains lacking. The gene set related to oxidative stress was obtained from MSigDB. The gene set related to NETs was sourced from relevant literature. Transcriptomic and clinical data were integrated from The Cancer Genome Atlas (TCGA)-LUAD (training set) and GSE31210 (validation set). Weighted Gene Co-Expression Network Analysis (WGCNA) was employed to screen gene modules and characteristic scores related to NETs and oxidative stress signatures. Differentially expressed genes (DEGs) were screened, and prognostic model was established using univariate and LASSO Cox regression. Immune infiltration was analyzed using ESTIMATE algorithm, MCP-counter and ssGSEA methods. And we developed a nomogram incorporating clinicopathological features and RiskScore model, and performed drug sensitivity analysis. Finally, the biological role of CPS1 in lung cancer cells was investigated through CCK-8, wound-healing, and Transwell experiments. 22 co-expression modules were screened, among which the brown module showed significant correlations with NETs and oxidative stress signature scores. This module was intersected with DEGs, yielding 624 overlapping genes implicated in immune-relevant pathways (like leukocyte differentiation, neutrophil activation involved in immune response). A prognostic model was established utilizing 8 key genes (ADGRE3, ARHGEF3, CD79A, CLEC7A, CPS1, EPHB2, LARGE2, and OAS3). In the TCGA database, the model demonstrated robust prognostic discrimination (area under the curve (AUC) > 0.6), with high-risk patients exhibiting shorter overall survival (OS) (p < 0.05). Its stability was validated in GSE31210 (AUC > 0.6). The RiskScore showed negative correlations with immune infiltration (like T cells, CD8 T cells, and natural killer cells) as well as immune/stromal scores. A nomogram model combining RiskScore with N staging was developed and validated, demonstrating strong predictive accuracy through calibration and decision curve analyses. High-risk patients were more sensitive to drugs like BI-2536, BMS-509744, and Pyrimethamine. Finally, in vitro tests showed that CPS1 knockdown markedly decreased the viability, migration, and invasion of lung cancer cells. The constructed prognostic model by NETs and oxidative stress-relevant genes effectively predicts LUAD prognosis, correlates with immune microenvironment characteristics, and guides drug sensitivity, providing novel insights for LUAD prognostic assessment and personalized therapy. Show less

Lung adenocarcinoma (LUAD) is the most prevalent subtype of lung cancer. Previous studies have highlighted the critical roles of complement and coagulation cascades in tumor development, maintenance, and therapeutic response. However, the overall impact of complement and coagulation cascade-related (CCCR) genes on LUAD progression and their role in the tumor microenvironment (TME) remain insufficiently explored. Therefore, we screened CCCR genes with important roles in LUAD using RNA sequencing data from The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO). Subsequently, a prognostic model, based on 8 hub genes (IGFBP1, TUBB, PLEK2, CNTNAP2, CPS1, EREG, CENPE, HBEGF) identified using the Lasso-Cox algorithm, was developed to stratify LUAD patients into high- and low-risk groups. This model demonstrated strong predictive capability and calibration, with an AUC of 0.816 in the external validation cohort. Multiomics clustering revealed that 2 cancer subtypes (CSs) are associated with prognosis, with CS2 demonstrating the most favorable prognostic outcome and validating the validity of the prognostic model. Additionally, we analyzed the immune infiltration, tumor mutation burden (TMB) and immunophenoscore (IPS) of the riskscore in the models. Through this analysis, we have identified for the first time CCCR genes are highly associated with clinical characteristics, immune cell infiltration patterns, and immune therapeutic responses of LUAD. This prognostic model constructed based on CCCR genes represents a valid tool for the prognosis of LUAD patients. Our findings provide valuable insights into the prognostic and immunological relevance of CCCR genes in LUAD, offering a robust foundation for personalized treatment strategies and future research. Show less

This study aims to explore how CPS1 influences the progression of lung adenocarcinoma by affecting the ammonia-induced ROS/AMPK/P53/LKB1 signaling pathway. Bioinformatics analysis was conducted to identify differential gene expression in lung adenocarcinoma patients. A549 cells were infected with control (NC) or CPS1 knockdown (CPS1-KD) lentivirus. Cells were treated with or without AMPK agonists, AMPK inhibitors, P53 agonists, or P53 inhibitors, followed by Western blot analysis of CPS1, NOX2, NOX4, p-AMPK, p-P53, and LKB1 protein levels. The content of MDA and SOD was measured, and the expression of AMPK, caspase-3 and P53 in tumor cells was detected through immunofluorescence. Apoptosis-related protein expression and tumor cell apoptosis were assessed using Western blot and flow cytometry. Tumor cell proliferation was evaluated using CCK-8 assays and colony formation experiments. Tumor size was measured in xenograft models using nude mice. Bioinformatics analysis indicated that LKB1 positively regulates AMPK activity. CPS1 knockdown results in increased ammonia levels, with upregulated expression of NOX2, NOX4, p-AMPK, p-P53, and LKB1 in tumor cells. Elevated P53 levels, along with significant increases in Bax, Caspase-8,and Caspase-12 expression, were observed, promoting apoptosis and inhibiting tumor cell proliferation. AMPK and P53 act to inhibit lung adenocarcinoma progression. CPS1 promotes the progression of lung adenocarcinoma by suppressing ammonia-induced activation of the ROS/AMPK/P53/LKB1 signaling pathway. Show less

Metabolic pathways are known to significantly impact the development and advancement of lung cancer. This study sought to establish a signature related to butyrate metabolism that is specifically linked to lung adenocarcinoma (LUAD). For the purpose of identifying butyrate metabolism-related differentially expressed genes (BMR-DEGs) in the TCGA-LUAD dataset, we introduced transcriptome data. This was followed by the implementation of the univariate Cox and LASSO analyses in order to construct a LUAD gene signature. We performed a comprehensive analysis of gene function enrichment between the two populations at risk, thoroughly examined their immune microenvironment characteristics, and assessed the effectiveness of immunotherapy. Finally, the function of CDKN3 in LUAD was verified by in vitro experiments. Through a comprehensive analysis of the TCGA-LUAD dataset, 51 significant BMR-DEGs were confirmed. Subsequently, five characteristic genes, CPS1, ABCC2, CDKN3, SLC2A1, and IGFBP1 were identified to create prognostic features for butyrate metabolism related outcomes in LUAD. Cox regression analysis determined that the pathological T stage, tumor stage, and RiskScore could serve as independent prognostic indicators. Analysis of the abundance of 22 immune infiltrating cells revealed that 15 immune cell types exhibited substantial differences and were strongly associated with risk ratings and prognosis. An important correlation exists between risk ratings and immunological checkpoints, which can be utilized to forecast the efficacy of treatment. In the high-risk group, there was an upregulation of the expression of PD-L2, PD-L1, and PD-1. Additionally, the risk score showed a positive correlation with TIDE and Exclusion score, while showing a negative correlation with Dysfunction score. Furthermore, the IC We identify and validate a novel BMR-related prognostic signature comprising 5 DEGs for LUAD patients. Our data might provide a new molecular target for LUAD treatment. Show less

Glioblastoma is the most aggressive form of primary brain tumor, characterized with poor prognosis and resistance to conventional therapies. Increasing evidence points to oxidative stress and redox dysregulation as important contributors to glioblastoma progression. Previously, chloride intracellular channel protein 4 (CLIC4), a redox-sensitive protein, has been implicated in cancer biology. However, its roles in glioblastoma remain poorly understood. Here, we found that CLIC4 expression is upregulated in glioblastoma tissues and cell lines, and is positively correlated with tumor malignancy and poor survival outcomes in patients with glioblastoma. Gene silencing of CLIC4 significantly reduces glioblastoma cell viability, migration, and proliferation in vitro and suppress tumor growth in vivo. Mechanistically, CLIC4 appears to maintain redox homeostasis by regulating mitochondrial functions, including membrane potential, mass, ROS production, and the activity of complexes III and IV. Moreover, a G-quadruplex (G4) structure located in CLIC4 promoter region is related to CLIC4 upregulation by oxidative stress in glioblastoma. This G4 structure can be readily oxidized to a parallel conformation, thereby enhancing its binding with DHX36 protein to promote gene transcription. Collectively, these findings position CLIC4 as a pivotal modulator of oxidative stress in glioblastoma and a potential target for developing therapeutic approaches for the treatment of glioblastoma. Show less

RNA G-quadruplexes (rG4s) are non-canonical secondary nucleic acid structures found in the transcriptome. They play crucial roles in gene regulation by interacting with G4-binding proteins (G4BPs) in cells. rG4-G4BP complexes have been associated with human diseases, making them important targets for drug development. Generating innovative tools to disrupt rG4-G4BP interactions will provide a unique opportunity to explore new biological mechanisms and potentially treat related diseases. Here, we have rationally designed and developed a series of rG4-based proteolytic targeting chimeras (rG4-PROTACs) aimed at degrading G4BPs, such as DHX36, a specific G4BP that regulates gene expression by binding to and unraveling rG4 structures in messenger RNAs (mRNAs). Our comprehensive data and systematic analysis reveals that rG4-PROTACs predominantly and selectively degrade DHX36 through a proteosome-dependent mechanism, which promotes the formation of the rG4 structure in mRNA, leading to the translation inhibition of rG4-containing transcripts. Notably, rG4-PROTACs inhibit rG4-mediated APP protein expression, and impact the proliferative capacity of skeletal muscle stem cells by negatively regulating Gnai2 protein expression. In summary, rG4-PROTACs provide a new avenue to understand rG4-G4BP interactions and the biological implications of dysregulated G4BPs, promoting the development of PROTACs technology based on the non-canonical structure of nucleic acids. Show less

Porcine circovirus type 3 (PCV3) is an emerging pathogen that causes porcine dermatitis, and reproductive failure. PCV3 Cap interacts with DExD/H-box helicase 36 (DHX36), a protein that functions primarily through regulating interferon (IFN)-β production. However, how the interaction between DHX36 and PCV3 Cap regulates viral replication remains unknown. Herein, we observed impaired PCV3 proliferation after DHX36 overexpression as indicated by decreased Rep protein expression and virus production. In contrast, PCV3 replication increased upon small interfering RNA-mediated DHX36 depletion. Furthermore, DHX36 positively regulated IFN-β production and interferon-stimulated genes (ISGs) expression. Mechanistically, PCV3 Cap interacted with DHX36, and the PCV3 Cap-NLS and DHX36-NTD were essential for the interaction. Furthermore, DHX36 may get degraded because its binding cellular partners became ubiquitinated and then reduced, and PCV3 Cap-(35-100aa) also promoted the degradation of DHX36 through the K48-linked ubiquitination. Taken together, these results show that DHX36 antagonizes PCV3 replication by interacting with PCV3 Cap and activating IFN-β response, which provides important insight on the prevention and controlling of PCV3 infection. IMPORTANCE: Porcine circovirus type 3 (PCV3) is a newly discovered pathogen associated with multiple clinicopathological signs. Clarifying the mechanisms that host factors modulate PCV3 replication helps understanding of the viral pathogenesis. The PCV3 capsid (Cap) protein has been shown to interact with DExD/H-box helicase 36 (DHX36) (Zhou et al., 2022b), a crucial protein that regulates virus replication. Herein, we further demonstrated that DHX36 protein is degraded in PCV3-infected cells and antagonizes the replication of PCV3 and that DHX36 increases interferon-β and interferon-stimulated gene levels by binding to PCV3 Cap. In addition, PCV3 infection could decrease DHX36 expression levels to antagonize its antiviral activity. These results reveal a molecular mechanism by which DHX36 antagonizes PCV3 replication by binding to PCV3 Cap protein and activating IFN signals, thereby providing important targets for preventing and controlling PCV3 infection. Show less

The Mpox virus (MPXV) has emerged as a formidable orthopoxvirus, posing an immense challenge to global public health. An understanding of the regulatory mechanisms of MPXV infection, replication and immune evasion will benefit the development of novel antiviral strategies. Despite the involvement of G-quadruplexes (G4s) in modulating the infection and replication processes of multiple viruses, their roles in the MPXV life cycle remain largely unknown. Here, we found a highly conservative and stable G4 in MPXV that acts as a positive regulatory element for viral immunodominant protein expression. Furthermore, by screening 42 optically pure chiral metal complexes, we identified the Λ enantiomer of a pair of chiral helical compounds that can selectively target mRNA G4 and enhance expression of the 39-kDa core protein encoded by the MPXV Show less

This study identified fibroblast-specific genes to develop a RiskScore model to improve prognostic accuracy and guide personalized treatment in glioblastoma (GBM). We analyzed fibroblast-specific signatures in the GSE273274 cohort using "Seurat" R package for scRNA-seq data processing. Fibroblast-related gene modules were identified via WGCNA, and functional enrichment was assessed with "clusterProfiler" package. A RiskScore model was established using univariate, Lasso Cox regression analysis, and "survival" package, validated by "timeROC" for receiver operator characteristic (ROC) curve. Finally, immune infiltration and drug sensitivity was evaluated applying "ESTIMATE," "TIMER," "MCPcounter," and "pRRophetic" packages. Experimental validation included qPCR for gene expression detection, and CCK-8, wound healing, and Transwell assays for functional measurement. The scRNA-seq analysis identified nine cell types of cells, with fibroblasts elevated in the GBM group. Fibroblast signatures were linked to tumorigenesis, cytoskeleton remodeling, and regulation of neuronal development process that affected GBM invasion. A 6-gene RiskScore divided GBM patients into high- and low-risk groups in training and validation sets, with high-risk patients exhibiting poorer survival, elevated StromalScore, and negative correlations with the infiltration of neutrophils and B_cells. Moreover, high-risk patients demonstrated heightened sensitivity to Cisplatin, MG-132, AZ628, Dasatinib, CGP-60474, A-770041, TGX221, and Bortezomib. Finally, qPCR showed that the VWA1 was upregulated in GBM cells, while knock-down of VWA1 inhibited the cell proliferation, migration, and invasion activity. We constructed a RiskScore model for predicting the survival outcomes based on fibroblasts-related genes. These findings highlighted the role of fibroblasts in GBM development and offered six potential therapeutic targets (VWA1, DUSP6, LOXL1, IGFBP4, CYGB, and ZIC3) for GBM treatment. Additionally, immune infiltration analysis and drug sensitivity prediction further supported the model's utility in guiding personalized treatment of GBM. Show less

Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that certain of the β‑actin control data shown in the western blots in Fig. 3E on p. 6 were strikingly similar to data appearing in different form in other articles written by different authors at different research institutes that had already been published elsewhere prior to the submission of this paper to Show less

Glioma, characterized by its cellular and molecular heterogeneity, presents formidable challenges in treatment strategy and prognostic assessment. The tumor microenvironment (TME) profoundly influences tumor behavior and treatment response, with tumor-associated neutrophils (TANs) playing a complex but understudied role. This study aimed to investigate the heterogeneity and role of TANs in glioma and to develop a prognostic model. Analysis of scRNA-seq data identified cellular subpopulations and differentially expressed neutrophil-related genes (DE-NRGs). Bulk RNA-seq was obtained from four independent datasets. Molecular subtypes of glioma samples were determined by consensus clustering. WGCNA was conducted to elucidate the association between gene modules and subtypes. We developed a risk score model. Expression of selected genes was confirmed using immunohistochemistry (IHC). In vitro experiments were also performed for functional verification, including CCK8, EdU, Transwell, and TUNEL assays. A total of 108 DE-NRGs for TANs were identified based on scRNA-seq data. Two molecular subtypes were characterized, showing significant differences in prognosis and clinical features. Immune-related analyses demonstrated varied immunological characteristics between subtypes. The risk score model was constructed with 7 genes, including AEBP1, CAVIN1, DCTD, DEPP1, DUSP6, FKBP9, and UGCG. It showed significant prognostic value and was validated across three external datasets. The mutation landscape highlighted higher IDH mutation prevalence in low-risk groups. Drug sensitivity analysis indicated TMZ resistance in high-risk groups. In vitro experiments showed that UGCG could promote glioma cell proliferation, migration, and invasion, while decreasing apoptosis. This study explored the heterogeneity of TANs and developed a prognostic model, providing insights for prognostic prediction and guiding personalized treatment strategies in glioma. Declaration of Generative AI in Scientific Writing: The authors declare nonuse of generative AI and AI-assisted technologies in the writing process. Show less

Cervical cancer remains one of the leading causes of cancer-related deaths among women globally, and there is still a need to research molecular targets that can be used for prognosis assessment and personalized molecular therapies. Here, we investigate the role of potential molecular target ribosomal L22-like 1 (RPL22L1) on cervical cancer, identify its potential mechanisms, and explore its related applications in prognosis and molecular therapies. Multiple cervical cancer cohorts online, tissue microarrays and clinical tissue specimens were analyzed for the association between RPL22L1 expression and patient outcomes. Functional and molecular biology studies of cell and mice models were used to clarify the effects and potential mechanisms of RPL22L1 on cervical cancer. RPL22L1 is highly expressed in both cervical adenocarcinoma and squamous cell carcinoma, and its expression is significantly associated with histology grade, clinical stage, recurrence, vascular space involvement, tumor sizes and poor prognosis. In vitro and in vivo experiment revealed that RPL22L1 overexpression significantly promoted cervical cancer cell proliferation, migration, invasion, tumorigenicity and Sorafenib resistance, which were attenuated by RPL22L1 knockdown. Mechanistically, RPL22L1 competitively binds to ERK phosphatase DUSP6, leading to excessive activation of ERK. The combined application of ERK inhibitors can effectively inhibit RPL22L1 overexpressing cervical cancer cells both in vivo and in vitro. RPL22L1 promotes malignant biological behavior of cervical cancer cells by competitively binding with DUSP6, thereby activating the ERK pathway. The combined use of Sorafenib and an ERK inhibitor is a potentially effective molecular targeted therapy for RPL22L1-high cervical cancer. Show less

Here, we identified a type of hypothetical T7SS effector in This alternative strategy facilitates effectors' delivery, even for fragmented substrates, highlighting its importance in ensuring the functionality of T7SS. Show less

Heat stress (HS) severely significantly reduces milk yield and causes substantial economic losses of dairy cows. TMT-based proteomes and an untargeted metabolomics approach were used to conduct the proteomics and metabolomics in heat-stressed (HS, Show less

The malignant transformation of odontogenic keratocysts (OKC) into cancerous odontogenic keratocysts (COKC) is exceedingly rare, and its mechanisms remain poorly understood. Studies exploring the cellular heterogeneity, molecular pathways, and clinical features of COKC are limited. In this study, we performed single-cell RNA sequencing (scRNA-seq) on three COKC samples and integrated the data with a public OKC dataset, identifying 22,509 single cells. Two COKC-specific epithelial subpopulations, Basal-C0-EXT1 and Basal-C3-HIST1H3B, were identified. These subpopulations exhibited enhanced stemness and invasive potential, respectively, suggesting their roles as key drivers of OKC carcinogenesis. Fibroblasts underwent phenotypic transitions, particularly from inflammation-associated fibroblasts (IFBs) to myofibroblasts (MFBs). Similarly, macrophage phenotypic transformation may also play a role in OKC carcinogenesis. Clinical observations of severe lesion-area pain in COKC patients suggest potential neuroinvasiveness, Supported by single-cell transcriptomic data, imaging findings, and histopathological evidence. A review of clinical data revealed that none of the COKC patients exhibited cervical lymph node metastasis. Single-cell transcriptomics suggests that this phenomenon may be associated with an active immune microenvironment in COKC, reduced epithelial-mesenchymal transition (EMT) activity, lower VEGFC expression, and upregulated MAST4 expression as a potential regulator of lymphatic metastasis. In conclusion, COKC exhibits distinct molecular, cellular, and clinical characteristics compared to OKC, featuring potent neuroinvasiveness and low lymph node metastatic potential. These findings provide important insights into the mechanisms underlying COKC development and may guide novel diagnostic and therapeutic strategies. Show less

Men, despite having a lower likelihood of longevity compared to women, generally exhibit better health status when they achieve longevity. The role of DNA methylation in this paradox remains unclear. We performed whole-genome bisulfite sequencing on long-lived men (LLMs), long-lived women (LLWs), younger men (YMs) and younger women (YWs) to explore specific methylation characteristics in LLMs. Despite an accelerated methylation aging rate in LLMs compared to LLWs, we identify thousands of differentially methylated genomic units (DMUs) in LLMs independent of age and sex. These DMUs, validated by an elastic net classifier, can serve as methylation markers for discriminating longevity potential in men. Many are located near health-related genes. Genes like PIWIL1 and EXT1, with promoters featuring DMUs, exemplify the potential role of LLM-specific methylation patterns in suppressing age-related diseases by regulating gene transcription. Our findings provide evidence of a distinct methylation feature contributing to healthy aging and longevity of LLMs. Show less

Gallstone disease (GD) is a common gastrointestinal disorder with a significant genetic component. Despite known risk factors, the genetic basis of GD remains incompletely understood. We aimed to identify novel genetic loci associated with GD, explore their clinical implications and investigate their therapeutic potential. We conducted a genome-wide association study from the UK Biobank followed by a meta-analysis, integrating summary statistics from the FinnGen R11, with further replication from Biobank Japan. Using systematic bioinformatic approaches, we performed gene prioritisation, colocalisation analysis, transcriptome-wide association study, Mendelian randomisations, cross-trait genetic correlations, phenome-wide association study, clinical investigations and gene-environment interactions by leveraging data from the FinnGen, Genotype-Tissue Expression project and Liver Cell Atlas single-cell transcriptomics data set. Our study highlighted novel susceptibility loci near candidate genes (ie, This study provides new insights into the genetic basis of GD and highlights the role of hepatocytes in GD pathogenesis. These findings have implications for the personalised prevention strategies and new therapeutic interventions in individuals predisposed to GD. Show less

Fatty acid desaturase 1 (FADS1) is significantly and specifically upregulated following diabetic corneal injury. However, its role in diabetic keratopathy remains unclear. This study aimed to investigate the impact of FADS1 on wound healing and functional recovery of the diabetic corneal epithelium and explore its potential mechanisms. Using high-glucose-induced corneal epithelial cells and a streptozotocin-induced type 1 diabetic mouse model, FADS1 expression was suppressed via FADS1 small interfering RNA (siRNA). Cell migration was assessed using scratch and transwell assays. Wound healing and functional recovery of the corneal epithelium were evaluated using sodium fluorescein staining, anterior segment optical coherence tomography, hematoxylin and eosin staining, and immunofluorescence staining. FADS1 knockdown promoted wound healing and functional recovery of the diabetic corneal epithelium both in vivo and in vitro. Suppression of FADS1 enhanced high-glucose-induced corneal epithelial cell migration, which was dependent on elevated levels of the upstream metabolite γ-linolenic acid. This effect was mediated through the activation of the mitogen-activated protein kinase signaling pathway and the accumulation of autophagosomes. After diabetic corneal epithelial injury, FADS1 expression is specifically upregulated. Knockdown of FADS1 promotes wound healing and functional recovery, suggesting a novel therapeutic strategy for diabetic keratopathy. Show less