Nasopharyngeal carcinoma (NPC) remains a major clinical challenge, particularly at advanced stages. RNA-binding proteins (RBPs) are important post-transcriptional regulators of tumorigenesis, but their roles in NPC remain poorly understood. This study investigated whether RNA-binding motif protein 4 (RBM4) stabilizes homeobox B13 (HOXB13) mRNA and thereby activates oncogenic signaling to promote NPC progression. Bioinformatics analysis identified differentially expressed genes in NPC. ENCORI was used to predict potential RNA-binding proteins (RBPs) targeting the candidate gene, and the predicted interaction was validated by RNA immunoprecipitation. Functional effects of HOXB13 and RBM4 in NPC cells were examined using colony formation and Transwell assays, along with modulating RBM4, HOXB13, nuclear factor kappa B (NF-κB) p65, and β-catenin expression. In vivo, effects of the RBM4/HOXB13 axis on NPC tumor growth and metastasis were evaluated using murine xenograft and lung metastasis models. HOXB13 was among the most strongly upregulated genes in NPC, and RBM4 was predicted to be a candidate RBP targeting HOXB13 mRNA. RBM4 was also upregulated in NPC models and increased HOXB13 mRNA stability. RBM4 silencing suppressed NPC cell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT), while increasing apoptosis and reducing HOXB13 expression and NF-κB and Wnt/β-catenin signaling. Overexpression of HOXB13, NF-κB p65, or β-catenin partially reversed these effects. In vivo, RBM4 silencing inhibited xenograft growth and experimental lung metastasis, and these effects were partially reversed by HOXB13 overexpression. RBM4 acts as a pro-tumorigenic RBP in NPC by stabilizing HOXB13 mRNA and activating the NF-κB and Wnt/β-catenin pathways. The RBM4/HOXB13 axis therefore warrants further investigation as a potential molecular target in NPC.
This study aimed to investigate whether vestitol ameliorates osteoarthritis (OA) and the mechanisms involved. We employed destabilization of the medial meniscus (DMM)-constructed OA mouse model and IL-1β-induced chondrocyte model to evaluate vestitol’s effects on OA. In vivo, pathological alterations in cartilage tissues were assessed via safranin O and H E staining. In vitro, chondrocyte viability and apoptosis were detected utilizing CCK-8 and flow cytometry. Vestitol’s mechanisms in OA were explored using network pharmacology analysis. Factors related to extracellular matrix (ECM) degeneration, inflammation, oxidative stress, ferroptosis, and pathway were checked using western blot, RT-qPCR, immunohistochemistry, and corresponding commercial reagent kits. In OA mice model, vestitol decreased cartilage destruction and OARSI score. Vestitol enhanced the levels of COL2A1, Aggrecan, SOD, FTH1, but reduced the levels of MMP-13, ADAMTS5, IL-6, IL-1β, COX-2, TNF-α, MDA, Fe2+, ROS, ACSL4 and 4-HNE. In IL-1β-induced chondrocyte model, vestitol enhanced cell viability and inhibited cell apoptosis. The changes in indicators related to ECM degradation, inflammation, oxidative stress, and ferroptosis were consistent with in vivo experiments. Moreover, vestitol increased p-GSK3B (Ser9), Nrf2, and GPX4 levels both in vivo and in vitro. The inhibition of vestitol on chondrocyte apoptosis, ECM degradation, inflammation, and ferroptosis-associated features was reversed by GSK3B activation. Vestitol mitigates OA by suppressing chondrocyte apoptosis, ECM degradation, inflammation, and ferroptosis-associated features, with pharmacological evidence implicating the PI3K/AKT/GSK3B axis and downstream modulation of the Nrf2/GPX4 pathway.
Acute myeloid leukemia (AML) is an aggressive hematologic malignancy with poor prognosis. Although chemotherapy and targeted therapies have advanced significantly, relapse and chemoresistance continue to be the major clinical challenges, which emphasizes the need for novel treatments. Brusatol, a natural quassinoid compound, has been reported to exert anti-tumor effects in various solid tumors; however, its anti-leukemic activity and potential mechanism in AML remain poorly characterized. THP-1 and KG-1a cells were treated with brusatol alone or combined with venetoclax. Proliferation was evaluated by CCK-8, cell counting and morphological observation. Apoptosis was detected by flow cytometry, Western blot, morphological changes and TUNEL staining. RNA-seq with GO, KEGG, and GSEA was used to identify key pathways. Cell cycle was analyzed by flow cytometry. DNA synthesis was detected by EdU assay. mRNA expression was measured by qPCR. Protein levels were determined by Western blot. Brusatol significantly suppressed the proliferation of AML cells. Moreover, brusatol notably increased the apoptotic rate, upregulated the expression of cleaved‑PARP and Bax, downregulated MCL1, and induced typical apoptotic morphological changes. Transcriptomic analysis demonstrated that brusatol primarily modulated pathways related to the cell cycle and steroid biosynthesis. Brusatol downregulated the expression of MCM2, PCNA, and CDK2, thereby inducing cell cycle arrest and inhibiting DNA replication. Brusatol also significantly reduced the expression of key genes involved in steroid biosynthesis, including DHCR7, SQLE, EBP, and FDFT1. Furthermore, co-treatment with brusatol and venetoclax strengthens cell cycle arrest, proliferation inhibition and apoptosis induction in AML cells compared with single drug treatment. Brusatol inhibits AML cell proliferation, induces apoptosis, triggers cell‑cycle dysregulation, and downregulates sterol‑biosynthesis‑related gene expression. Brusatol also enhances the anti-leukemic activity of venetoclax. This study preliminarily elucidates the underlying mechanisms and provides experimental evidence supporting the potential therapeutic value of brusatol for acute myeloid leukemia.
Sodium ion (Na⁺), the predominant cation in the extracellular space, is essential for maintaining fundamental cellular activities via preservation of the transmembrane Na⁺ gradient. While intracellular Ca²⁺ overload has long been recognized as the core driver of cell injury and death, accumulating evidence indicates that Na⁺ overload, defined as abnormal intracellular Na⁺ accumulation, may act as an independent trigger of irreversible cellular damage, rather than merely a preceding event or concomitant phenomenon of Ca²⁺ overload. In 2025, the term Necrosis by Na⁺ Overload (NECSO) was first coined to describe a novel form of regulated necrosis directly triggered by sustained and excessive Na⁺ influx, which is primarily mediated by the transient receptor potential melastatin 4 (TRPM4) channel. Distinct from apoptosis and other established forms of regulated cell death, NECSO is characterized by early cellular and organellar swelling, mitochondrial energy metabolism collapse, ionic homeostasis disruption, and eventual loss of plasma membrane integrity, without the hallmark features of caspase activation and chromatin condensation. The core execution pathway of NECSO involves TRPM4-mediated Na⁺ influx, subsequent mitochondrial Na⁺ accumulation, impaired tricarboxylic acid cycle and oxidative phosphorylation, and catastrophic cellular energy depletion, which further aggravates Na⁺ overload via inhibition of Na⁺/K⁺-ATPase and forms a fatal vicious cycle. In pathophysiological contexts, NECSO serves as a central mediator of myocardial injury in ischemia-reperfusion, and is also implicated in the progression of heart failure and neuronal death in stroke. In cancer biology, NECSO and its core mediator TRPM4 exhibit highly tissue-specific dual roles, acting as either oncogenic drivers or tumor suppressors in different cancer types. Meanwhile, NECSO-related genes have shown promising potential as diagnostic and prognostic biomarkers, as well as predictors of therapeutic response to chemotherapy and immunotherapy across multiple malignancies. Targeted strategies against NECSO, including direct modulation of TRPM4 activity, intervention of Na⁺ homeostasis, and combination therapeutic regimens, have exhibited favorable efficacy in preclinical models of cardiovascular diseases and cancer. This review systematically summarizes the definition, core molecular mechanisms, and pathophysiological functions of NECSO, discusses the application value of NECSO-related biomarkers, and outlines current challenges and future directions in this emerging field, to provide a novel perspective for the development of therapeutic strategies against major human diseases.
Bleomycin (BLM) is a widely used chemotherapeutic agent associated with cutaneous toxicity characterised by inflammation, oxidative stress, and progressive fibrosis. However, effective strategies to mitigate these adverse tissue responses remain limited. This study aimed to evaluate potential protective effects of alpha-lipoic acid (ALA) against the BLM-induced skin model, focusing on fibrotic, inflammatory, and proliferative alterations. Thirty adult Wistar albino rats were randomly assigned to four groups: Control, ALA, BLM, and BLM + ALA. BLM (30 mg/m²) was administered intramuscularly on Days 1, 8, and 15, whereas ALA (50 mg/kg/day, subcutaneously) was administered once daily from Days 1–14. Skin tissues were collected immediately after the final scheduled treatment on Day 15 for histopathological and immunohistochemical analyses. Histopathological evaluation was performed using H E, Masson’s trichrome, and Toluidine blue staining. Immunohistochemical staining was conducted to evaluate the expression levels of collagen types I and III, interleukin-6 (IL-6), inducible nitric oxide synthase (iNOS), caspase-1, and the proliferation marker Ki-67, followed by quantitative image analysis. BLM exposure resulted in marked epidermal thickening, inflammatory cell infiltration, increased collagen deposition, and elevated mast cell density compared with controls. These histopathological alterations were significantly attenuated in the BLM + ALA group. Consistently, BLM significantly increased the expression of collagen I/III, IL-6, iNOS, caspase-1, and Ki-67, whereas ALA treatment significantly reduced these elevations. Overall, ALA attenuated BLM-induced dermal alterations. ALA treatment was associated with reduced collagen deposition and decreased IL-6, iNOS, caspase-1, and Ki-67 immunoreactivity in the experimental skin model. Further mechanistic studies are required to establish the causal relationships underlying these observations. α-Lipoic Acid (ALA) reduces bleomycin-induced skin in a rat model. ALA decreases collagen deposition and dermal fibrotic remodelling. ALA treatment was associated with reduced IL-6, iNOS and caspase-1 immunoreactivity. ALA suppresses epidermal hyperproliferation and mast cell accumulation. Findings support the protective histopathological and immunohistochemical effects of ALA treatment in experimental dermal fibrosis.
Thymoquinone, the principal bioactive constituent of Nigella sativa, has been extensively investigated for its anti-inflammatory, antioxidant, and metabolic effects. However, these activities are often examined separately, limiting understanding of their broader biological integration. This review examines thymoquinone through three interconnected immunometabolic axes: NF-κB–mediated inflammation, AMPK/mTOR-driven metabolic regulation, and ROS/Nrf2-dependent redox balance. Preclinical evidence indicates that thymoquinone modulates each of these pathways and may influence the crosstalk linking oxidative stress, metabolic reprogramming, and inflammation. Based on these findings, we propose a conceptual framework positioning thymoquinone as a multi-target immunometabolic modulator. This framework is hypothesis-generating and requires further validation through integrated experimental and clinical studies.
CD38 is a type II transmembrane glycoprotein and serves as the primary ectoenzyme responsible for regulating cellular nicotinamide adenine dinucleotide (NAD+) levels in mammalian tissues. Initially identified as a lymphocyte activation marker, recent research has revealed its significant role in immunosuppression within oncological processes, NAD+ depletion associated with aging, and exosomal signaling. This review provides a comprehensive analysis of the enzymatic functions of CD38, its pathophysiological impacts on both hematological and solid tumors, and the mechanisms of resistance that emerge against anti-CD38 therapies. Furthermore, the potential of CD38 as an exosomal biomarker in liquid biopsy procedures and its function as a “metabolic resetter” for promoting healthy aging are explored.
Glycation, a spontaneous reaction between sugars and biomolecules, is a central driver of oxidative stress and protein dysfunction in metabolic and neurodegenerative disorders. α-Synuclein (α-Syn), a key protein implicated in Parkinson’s disease, is particularly susceptible to modification by reactive carbonyl species such as methylglyoxal (Me) due to its lysine-rich structure. These modifications result in the formation of advanced glycation end products (AGEs), which promote protein aggregation, impair cellular clearance, and induce oxidative and inflammatory damage via the AGE–RAGE axis. This cascade, characterised by increased reactive oxygen and nitrogen species, mitochondrial dysfunction, and metabolic imbalance, accelerates neuronal degeneration and disease progression.This study investigates trans-ferulic acid (TF) as a multifunctional therapeutic candidate targeting this interconnected network. TF effectively inhibited glycation in both ribose and Me-induced α-Syn systems, demonstrating strong antiglycation potential. Additionally, TF exhibited significant scavenging activity against hydrogen peroxide and nitric oxide, underscoring its capacity to reduce oxidative and nitrosative stress. TF also inhibited α-glucosidase, indicating a further mechanism for controlling glucose availability and limiting the formation of reactive intermediates. Collectively, these findings identify TF as a promising multi-target agent capable of disrupting the glycation-oxidative stress cycle at multiple levels. By concurrently reducing reactive species, suppressing carbonyl formation, and limiting protein aggregation, TF represents a compelling strategy to mitigate glycation-driven toxicity and its pathological consequences.
While the persistent inflammation in chronic rhinosinusitis (CRS) is closely linked to M1 macrophage polarization, the mechanisms that control this polarization have not been fully elucidated. A positive correlation was observed between UCHL1 (a significantly upregulated deubiquitinating enzyme in CRS tissues) and M1 macrophage markers in our bioinformatic analysis. Suppression of UCHL1, via genetic knockdown or pharmacologic inhibition (6RK73), attenuated M1 macrophage polarization and curtailed the production of IL‑1β, TNF‑α and IL‑6 in both cellular and animal models. Mechanistically, UCHL1 interacts with HIF‑1α and stabilizes it by deubiquitinating it, thereby blocking its proteasomal degradation. This stabilization is functionally relevant, as HIF‑1α activation reversed the suppression of M1 polarization induced by UCHL1 knockdown. Furthermore, conditioned medium from UCHL1‑deficient macrophages mitigated epithelial cell damage-an effect that was abolished by HIF‑1α agonism. In a murine CRS model, pharmacological inhibition of UCHL1 significantly ameliorated histopathological injury, reduced behavioral symptoms such as sneezing and nasal scratching, and lowered IgE levels, and pro‑inflammatory cytokines, while downregulating the UCHL1/HIF‑1α axis and M1 marker expression. Collectively, these findings indicate that UCHL1 drives M1 macrophage polarization and exacerbates CRS inflammation by stabilizing HIF‑1α through deubiquitination, highlighting UCHL1 as a potential therapeutic target in CRS.
Neuroblastoma remains a challenging malignancy in which redox imbalance and mitochondrial dysfunction represent potential cellular vulnerabilities. This study investigated the redox-associated cytotoxic effects and mitochondrial responses of Pervari honey (PH), a region-specific natural product from Türkiye, in SH-SY5Y neuroblastoma cells. MTT-based viability was assessed in SH-SY5Y neuroblastoma cells and L929 fibroblasts, used as a non-cancerous control cell model, after 48 h of PH exposure. Redox status was evaluated by measuring intracellular reactive oxygen species (ROS), total antioxidant capacity (TAC), total oxidant status (TOS), oxidative stress index (OSI), glutathione (GSH), malondialdehyde (MDA), and 8-hydroxy-2’-deoxyguanosine (8-OHdG). Mitochondrial membrane potential (ΔΨm) was analyzed using the JC-10 assay. Expression levels of apoptosis- and antioxidant-related genes were determined by RT-qPCR. Comparative viability analysis showed that L929 fibroblasts were less affected by PH exposure than SH-SY5Y cells, and the IC50 value of PH was not reached in L929 cells within the tested concentration range. Treatment increased ROS production and shifted the oxidant/antioxidant balance toward oxidative stress, as indicated by increased TOS and OSI and decreased TAC and GSH levels. Elevated MDA and 8-OHdG levels confirmed oxidative damage to lipids and DNA. Additionally, PH reduced mitochondrial membrane potential and altered the expression of apoptosis- and antioxidant-related genes, including cytochrome c (Cyt-c), p53, SOD1, and CAT. PH exerted cytotoxic effects in SH-SY5Y cells through redox imbalance and mitochondrial dysfunction. These findings suggest that PH may influence redox-sensitive pathways in neuroblastoma cells. However, further studies including compositional analysis and appropriate controls are required to clarify the underlying mechanisms.
Nanobody technology is a promising approach in cancer research and treatment. Monoclonal antibodies have long been central to targeted therapies; however, their large size, complex production, and limited tissue penetration restrict their clinical performance. Nanobodies, derived from camelid heavy-chain antibodies, possess unique properties, including small size ( 15 kDa), high stability, and the ability to access otherwise inaccessible epitopes. At the molecular and cellular levels, these characteristics arise from a defined structure-function relationship that governs antigen binding, receptor modulation, and downstream signaling. Beyond affinity, nanobody function is influenced by binding kinetics and intracellular trafficking. Their ease of production in bacterial systems further enhances cost-effectiveness compared to conventional antibodies. Nanobody-based strategies have evolved from diagnostic and imaging tools to multifunctional therapeutic platforms, including nanobody-drug conjugates, bispecific and multispecific engagers, immune checkpoint modulators, and engineered cell-based therapies such as CAR-T systems. Preclinical studies have demonstrated improved tumor targeting, enhanced immune activation, and reduced off-target effects. Emerging clinical evidence, including approved nanobody therapeutics and ongoing trials, supports their safety, feasibility, and translational potential in cancer patients. Together, these findings highlight nanobodies as versatile platforms capable of overcoming key limitations of conventional therapies and facilitating clinical translation in precision oncology.
Cerebral ischemia/reperfusion injury (CIRI) is a major cause of neurological dysfunction after ischemic stroke, and effective therapies remain limited. Gomisin A, a bioactive lignan isolated from Schisandra chinensis, exhibits antioxidant and anti-apoptotic activities, but its role in CIRI remains unclear. Here, the protective effects of Gomisin A were investigated using middle cerebral artery occlusion (MCAO) rats and oxygen-glucose deprivation/reoxygenation (OGD/R) cell models. Gomisin A significantly reduced cerebral infarct volume and improved neurological outcomes in MCAO rats. In OGD/R models, it enhanced endothelial cell migration and tube formation, accompanied by increased VEGF expression and activation of the VEGF/PI3K/AKT/NOS3 signaling pathway. In addition, Gomisin A alleviated oxidative stress and suppressed apoptosis in both cellular and animal models. Pharmacological inhibition of VEGF with axitinib partially reversed these protective effects, supporting the involvement of VEGF-dependent signaling. Collectively, these findings suggest that Gomisin A exerts neuroprotective effects against CIRI by enhancing angiogenic activity while attenuating oxidative stress and apoptosis, partly through activation of the VEGF/PI3K/AKT/NOS3 pathway. Gomisin A may therefore represent a promising candidate for the treatment of ischemic stroke.
Although Rheumatoid arthritis (RA) management has improved substantially with modern pharmacological therapies, many patients still fail to achieve sustained treatment targets. This study investigated the therapeutic effects and potential mechanisms of asparagus polysaccharide (APS) in tumor necrosis factor-α (TNF-α)-induced MH7A cell dysfunction and collagen-induced arthritis (CIA) in mice. In vitro, MH7A cells were treated with different concentrations of APS, and cell viability, apoptosis, cell cycle distribution, and related protein and gene expression were assessed. In vivo, CIA model was established in mice, followed by APS administration. Paw volume, joint histopathology, serum cytokine levels, and the splenic Th17 cell proportion were evaluated. APS (≥ 3 mg/mL, 48 h) decreased MH7A cell viability. APS (3, 4, 5 mg/mL, 24 h) promoted apoptosis and inhibited proliferation in TNF-α-treated MH7A cells. APS (5 mg/mL, 24 h) downregulated Bcl-2 and cell cycle-related genes and suppressed NF-κB pathway activation. In the CIA mouse model, APS reduced paw volume, alleviated joint inflammation and tissue damage, decreased serum TNF-α, interleukin-6 (IL-6), IL-17A levels, increased IL-10 levels, and inhibited splenic Th17 cell expansion in a dose-dependent manner, without a significant effect on Treg cell proportion. APS exerts anti-arthritic effects in vitro and in vivo by regulating synoviocyte proliferation and apoptosis, inflammatory cytokine balance, and Th17 cell differentiation. The in vitro mechanism was associated with inhibition of NF-κB signaling. These findings provide preliminary experimental evidence supporting further investigation of APS as a candidate therapeutic agent for RA.
It is known that prolonged cell passaging may affect the outcomes of cell-based experiments. This study aimed to comprehensively investigate the consequences of long-term culture of U87MG glioblastoma cells, focusing on cell metabolism, morphology, tumorigenicity and drug response. U87MG (L) cells subjected to long-term culturing (20 passages) displayed altered morphology and significantly enhanced autofluorescence of lipofuscin, a well-established marker of oxidative stress and cellular senescence, compared with control U87MG cells maintained for only 5 passages. Fluorescence-lifetime imaging microscopy (FLIM) of NADH revealed the metabolic shift likely associated with glycolysis in long-term cultured U87MG (L) cells. Using the genetically encoded sensor HyPer7, the increase in basal intracellular levels of hydrogen peroxide in U87MG (L) cells was detected. In vivo studies employing orthotopic intracranial xenotransplantation in immunodeficient NSG mice revealed that, unlike short-term cultured U87MG cells, which formed visible tumor nodules with clearly delineated margins, long-term cultured U87MG (L) cells infiltrated diffusely into brain tissues, invaded bone tissue, and exhibited perineural growth. Such an aggressive behavior resulted in worse survival outcomes of xenograft-bearing mice. Notably, U87MG (L) cells became less susceptible to temozolomide, but acquired sensitivity to death receptor 5 (DR5)-selective variant of cytokine TRAIL in vitro and in vivo due to increased DR5 expression on the cell surface and downregulation of cFLIP expression. Our findings with the U87MG cell line indicate that culture duration can alter cellular responses, thereby impacting experimental outcomes. It should be carefully considered when establishing tumor models and evaluating efficacy of potential drug candidates.
Triple-negative breast cancer (TNBC) is an aggressive malignancy defined by the absence of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). These molecular characteristics render targeted therapies less effective and contribute to poor clinical outcomes. Towards this, we studied the electroporation-mediated delivery of resveratrol, a polyphenolic compound with pleiotropic anticancer activities, on MDA-MB-231, human TNBC cell line. Both conventional two dimensional (2D) monolayer cell culture and three-dimensional (3D) hyaluronic acid HA-EAbuK-IKVAV scaffold-based cell cultures were used. Two electroporation (EP) protocols (EP1: 8 pulses, 800 V/cm, 100 µs at 1 Hz; EP2: 8 pulses, 1000 V/cm, 100 µs at 1 Hz) were evaluated for their effects on cytotoxicity and reactive oxygen species (ROS) generation. In 3D, the combined treatment similarly reduced viability to 20
Osteoarthritis (OA) is a chronic degenerative joint disorder characterized by articular cartilage deterioration, synovial inflammation, and progressive loss of joint function. Current treatments predominantly address symptoms without modifying structural disease progression, highlighting the need for disease-modifying strategies. Mesenchymal stem cells (MSCs), particularly umbilical cord-derived MSCs (UC-MSCs), exert therapeutic effects primarily through paracrine mechanisms rather than direct engraftment. The UC-MSCs secretome, comprising soluble bioactive factors and extracellular vesicles, modulates pro-inflammatory signaling, inhibits chondrocyte apoptosis, and promotes cartilage matrix biosynthesis. In vitro studies demonstrate restoration of anabolic chondrocyte phenotype, suppression of catabolic enzymes (MMP-13, ADAMTS-5), and attenuation of NF-κB and MAPK pathway activation. Preclinical models show preservation of cartilage architecture, reduced synovial inflammation, and improved joint function following intra-articular administration. Early clinical investigations report reductions in pain scores and functional improvements with favorable short-term safety profiles, though evidence remains largely derived from MSCs-based rather than secretome-specific interventions. Current findings support the biological plausibility and translational potential of UC-MSCs secretome therapy; however, clinical evidence remains limited and heterogeneous. Adequately powered randomized controlled trials with standardized protocols and extended follow-up are required to establish long-term efficacy and safety.
Diminished ovarian reserve decreases the chance of fertility, but the mechanisms regulating ovarian function remain unclear. This study aimed to explore the mechanism through which FOXO1 regulates mitochondrial function in ovarian granulosa cells. KGN cells were transfected with FOXO1, METTL3, and/or SMAD4 vectors, and then their proliferation and apoptosis were detected using CCK-8 and EdU assays and flow cytometry, respectively. Molecular binding was detected by immunoprecipitation. The effect of FOXO1 on METTL3 promoter activity was analyzed using luciferase reporters. SMAD4 m6A was quantified using MeRIP-qPCR. ROS and MMP were measured using immunofluorescence, ATP and mtDNA levels were determined using their respective kits, and the expression of mitophagy-related proteins, including DRP1, PINK1, and parkin, was quantified using Western blotting. FOXO1 overexpression or SMAD4 knockdown reduced the proliferation of KGN cells and enhanced their apoptosis. FOXO1 inhibited METTL3 transcription via binding. METTL3 promoted SMAD4 m6A modification. FOXO1 repressed METTL3 and SMAD4 expression, while METTL3 upregulated SMAD4 expression. In FOXO1-overexpressing cells, METTL3 or SMAD4 overexpression enhanced proliferation, suppressed apoptosis, reduced ROS production, elevated ATP, mtDNA, and MMP levels, and downregulated DRP1, PINK1, and parkin expression. In conclusion, FOXO1 targeted METTL3 to downregulate SMAD4 and modulated the expression of mitophagy-related proteins, thereby promoting mitophagy and mitochondrial damage in granulosa KGN cells.
Cell-surface glycosylation plays an important role in lectin-mediated molecular recognition and is frequently altered in malignant cells. In this study, a fixation-modified cell-ELISA was developed to enable stable immobilization of non-adherent cells and to compare the binding behavior of Phaseolus vulgaris leucoagglutinin (PHA-L) toward SR lymphoma cells and normal human peripheral blood mononuclear cells (PBMCs). Saturation binding and time-course assays were performed at 5, 15, 25, and 37 °C, and the resulting whole-cell ELISA data were analyzed using Langmuir, Van’t Hoff, Arrhenius, and Eyring models to obtain comparative apparent kinetic and thermodynamic descriptors. Saturation binding analysis showed lower apparent equilibrium dissociation constants (Kdapp) and higher maximum binding responses (ODmax) for SR lymphoma cells than for normal PBMCs under identical experimental conditions, indicating a stronger apparent PHA-L binding response. Time-course analysis demonstrated faster apparent binding kinetics in SR lymphoma cells, characterized by higher apparent observed and association rate constants (kobsapp and konapp), lower apparent dissociation rates (koffapp), and shorter observed association half-times. Model-based thermodynamic analysis suggested that the apparent binding process was endothermic and predominantly entropy-favored in both cell types. Arrhenius and Eyring analyses further provided comparative apparent energetic descriptors, with SR lymphoma cells exhibiting lower apparent Arrhenius-derived energetic parameters and lower apparent activation enthalpy than normal PBMCs under the experimental conditions. Because these parameters were derived from mathematical analysis of whole-cell ELISA measurements rather than direct real-time biophysical techniques, they should be interpreted as apparent comparative descriptors rather than intrinsic molecular kinetic or thermodynamic constants. Overall, the fixation-modified cell-ELISA provided a feasible platform for exploratory comparative evaluation of PHA-L binding to suspension cells and revealed measurable differences in the apparent binding behavior of SR lymphoma cells and normal PBMCs. These findings support the potential utility of this approach for comparative lectin–cell interaction studies and future investigations of cell-surface glycosylation.
Lung cancer remains a major cause of cancer-related mortality, and reliable molecular targets with therapeutic relevance are still needed. This study aimed to identify key genes associated with lung cancer progression and explore the potential regulatory effect of resveratrol. The GSE43458 dataset, including 80 lung cancer samples and 30 control samples, was used for differential expression analysis and weighted gene co-expression network analysis (WGCNA). Intersecting genes were subjected to protein–protein interaction network construction, Cytoscape-based hub gene screening, functional enrichment analysis, and immune infiltration analysis. Single-cell RNA sequencing data from GSE131907 were further analyzed to characterize cellular heterogeneity and hub gene distribution. Resveratrol was predicted as a candidate compound, followed by molecular docking with hub proteins. Finally, CCK-8, Western blotting, and qRT-PCR assays were performed in BEAS-2B and A549 cells. A total of 1,655 differentially expressed genes were identified, and WGCNA identified the 252-gene turquoise module as the module most strongly associated with the lung cancer phenotype (r = − 0.86, P = 6.0 × 10⁻³⁵). Intersection analysis yielded 250 candidate genes, representing 15.1
