Supplementary Figure S3. Immunophenotypic analysis of myeloid, B and T cells from bone marrow and splenic preps of Control or K2 KO 4T1 injected mice, two- and five-weeks following cell infusion.

Supplementary Figure S4. Pathway analyses of the scRNA-seq data using Gene Ontology biological functions analyses.

OBJECTIVE:Tongue squamous cell carcinoma (TSCC) is characterized by high invasiveness and early lymph node metastasis, leading to a poor prognosis.Current surgical and chemoradiotherapy regimens often cause functional impairment, drug resistance, and other drawbacks. Thus, developing highly effective, lowtoxicity treatments is a key research priority. Traditional Chinese medicine (TCM) offers unique advantages via multitarget synergistic effects, enabling innovative therapeutic approaches. METHOD:In this study, we developed Astragaloside IV-Brucea javanica oil nanoemulsion (AS/BJO-NEs) and characterized its stability, polydispersity, and pharmacokinetic profile. The effects of AS/BJO-NEs on TSCC cells were evaluated through a series of functional assays, including CCK-8 (viability), colony formation (proliferation), and scratch wound healing (migration). Both in vitro and in vivo experiments were performed to investigate whether its mechanism involves regulation of the MTFR2-HIF-1α-EZH2/FoxM1 signaling axis. RESULTS:The optimized AS/BJONEs exhibited uniform spherical morphology at the nanoscale and demonstrated favorable stability. In both in vitro and in vivo models, AS/BJONEs significantly suppressed the malignant phenotype of TSCC cells. Mechanistic investigation further revealed that the anti-tumor effects of AS/BJONEs are mediated, at least in part, through targeting the MTFR2-HIF-1α-EZH2/FoxM1 signaling axis, as confirmed by establishing stable MTFR2/HIF-1α knockdown and overexpression cell models. CONCLUSION:This study demonstrates that AS/BJONEs inhibit TSCC progression by targeting the MTFR2-HIF-1α-EZH2/FoxM1 signaling axis. Our findings provide experimental evidence supporting the development of multi-target therapeutic strategies derived from traditional Chinese medicine for the treatment of TSCC.

Objective This study aimed to investigate whether Astragaloside-Brucea javanica oil nanoemulsion (AS/BJO-NEs) inhibits the malignant progression of oral squamous cell carcinoma (OSCC) and to further explore its potential regulatory mechanisms. Methods Immunohistochemistry (IHC) was used to evaluate the expression of related pathway proteins in human OSCC and adjacent normal tissues. Stable OSCC cell lines with knockdown or overexpression of CDK1/HOXC10 were established. The effects of AS/BJO-NEs and the underlying mechanisms were assessed in vitro through colony formation, wound healing, and Transwell invasion assays, as well as RT-qPCR, western blot, chromatin immunoprecipitation (ChIP), and dual-luciferase reporter assays. An OSCC subcutaneous xenograft model in nude mice was constructed for in vivo validation using RT-qPCR, western blot, hematoxylin and eosin (H&E) staining, and IHC. Results Analysis of clinical samples revealed upregulated expression of CDK1, P-EZH2, HOXC10, MTFR2, and N-cadherin, alongside downregulated expression of H3K27me3 and E-cadherin in OSCC tissues. In vitro experiments confirmed that AS/BJO-NEs downregulated CDK1 in a concentration-dependent manner, subsequently reducing the expression of P-EZH2, HOXC10, and MTFR2, increasing H3K27me3 levels, and inhibiting cell proliferation, migration, and invasion. H3K27me3 was enriched in the HOXC10 promoter region, and HOXC10 directly bound to and activated MTFR2 transcription. In vivo experiments demonstrated that AS/BJO-NEs effectively inhibited tumor growth, regulated molecules within this pathway and epithelial-mesenchymal transition (EMT) markers, whereas CDK1 overexpression counteracted these effects Conclusion This study demonstrates that AS/BJO-NEs exert anti-OSCC effects by inhibiting CDK1, downregulating HOXC10, thereby reducing MTFR2 expression, and suppressing cell proliferation, migration, invasion, and the EMT process.

Supplementary Figure S1. Immunophenotypic analysis of myeloid, B and T cells from bone marrow and splenic preps of naïve, or 4T1 injected mice, two- and five-weeks following cell infusion.

The incidence of tongue squamous cell carcinoma (TSCC) has been increasing annually. Type 2 diabetes mellitus (T2DM) is one of the risk factors for TSCC, with hyperglycemia being a hallmark of T2DM that promotes the development of oral cancer and results in poorer prognoses for TSCC patients who also have T2DM. Oleic acid (OA), a component of Brucea javanica oil emulsion, exhibits both anti-cancer and anti-diabetic properties. However, the precise regulatory mechanisms of OA on tongue squamous cell carcinoma (TSCC) under high glucose conditions have yet to be fully elucidated. Our findings demonstrated that high glucose significantly enhanced the migratory, invasive, and proliferative capabilities of TSCC cells. Notably, OA treatment effectively reversed these high glucose-induced enhancements in TSCC cellular activities. Furthermore, our research revealed that OA exerted its anti-tumor effects by stabilizing its binding with S100A9, which modulated the glycolytic pathway. Ultimately, we confirmed that the anti-tumor mechanism of OA in TSCC patients with T2DM was associated with the inactivation of the S100A9-HK2/PKM2-SOD2 pathway. In summary, OA inhibited the proliferation, migration, and invasion of TSCC cells under high glucose conditions through the regulation of the S100A9-HK2/PKM2-SOD2 pathway.

The 14 members of Eph receptor tyrosine kinases (RTK) bind to membrane-tethered ligand called ephrins and mediate cell contact signaling where the receptors and ligands engage in trans on adjacent cells. Previous studies reveal that some Eph and ephrin pairs are coexpressed on the same cells, including EphA3-ephrin-A3 and EphA4/ephrin-A5, can also interact with each other in cis. However, significant discrepancies persist as to the molecular basis and functional significance of the cis interactions, owning to the difficulties to directly interrogate the interactions. Here, we utilize time-resolved live cell fluorescence spectroscopy to demonstrate direct in cis interactions between EphA2 and ephrin-A1. Structure-guided mutagenesis mapped interactions to two salt bridges between the ligand- and receptor-binding domains of EphA2 and ephrin-A1. Interestingly, the same interface is shared with in trans interaction. Consequently, EphA2-ephrin-A1 interaction in cis competes with their interaction in trans, which leads to attenuation of EphA2 canonical signaling and inhibition of cell rounding when ligand is presented in trans. EphA2 and ephrin-A1 are widely coexpressed in many epithelial tissues, and dysregulation of their expression is known to contribute to tumor initiation and progression. The detailed molecular characterization of the mutually exclusive cis and trans interactions uncovers a new mechanism underpinning their unique roles in oncogenesis.

Triple-negative breast cancer (TNBC) presents significant clinical challenges because of its limited treatment options and aggressive behavior, often associated with poor prognosis. This study focuses on kindlin-2, an adapter protein, and its role in TNBC progression, particularly in hematopoiesis-mediated immune evasion. TNBC tumors expressing high levels of kindlin-2 induce a notable reshaping of hematopoiesis, promoting the expansion of myeloid cells in the bone marrow and spleen. This shift correlated with increased levels of neutrophils and monocytes in tumor-bearing mice over time. Conversely, genetic knockout (KO) of kindlin-2 mitigated this myeloid bias and fostered T-cell infiltration within the tumor microenvironment, indicating the pivotal role of kindlin-2 in immune modulation. Further investigations revealed that kindlin-2 deficiency led to reduced expression of PD-L1, a critical immune checkpoint inhibitor, in TNBC tumors. This molecular change sensitized kindlin-2-deficient tumors to host antitumor immune responses, resulting in enhanced tumor suppression in immunocompetent mouse models. Single-cell RNA sequencing, bulk RNA sequencing, and IHC data supported these findings by highlighting enriched immune-related pathways and increased infiltration of immune cells in kindlin-2-deficient tumors. Therapeutically, targeting PD-L1 in kindlin-2-expressing TNBC tumors effectively inhibited tumor growth, akin to the effects observed with genetic kindlin-2 KO or PD-L1 KO. Our data underscore kindlin-2 as a promising therapeutic target in combination with immune checkpoint blockade to bolster antitumor immunity and counteract resistance mechanisms typical of TNBC and other immune-evasive solid tumors.Implications: Kindlin-2 regulates tumor immune evasion through the systemic modulation of hematopoiesis and PD-L1 expression, which warrants therapeutic targeting of kindlin-2 in patients with TNBC.

The aim of this study was to investigate the potential mechanisms of Rhaponticin (Rha) in the treatment of periodontitis. Network pharmacology and molecular docking techniques were used to identify potential targets of Rha for the treatment of periodontitis and its ability to bind to the targets. Next, in vitro as well as in vivo experiments were conducted to validate Rha's potential role in treating periodontitis. We found in network pharmacology and molecular docking that the HIF-hypoxia signaling pathway is involved in the potential mechanism of Rha treatment of periodontitis and that it can bind stably to HIF1A. In vitro experiments, based on the hypoxia-induced inhibition of proliferation, migration, and osteogenic differentiation of hPDLSCs, we found that Rha inhibited the expression of HIIF1A, promoted the expression of PCNA, CXCR4, and OCN, and enhanced their proliferation, migration, and osteogenic differentiation. In in vitro experiments, Rha promoted alveolar bone repair and inhibited gingival inflammation in periodontitis rats. Rha is a potential drug for the treatment of periodontitis. Therefore, this study provides new insights into the potential mechanisms of Rha in periodontitis treatment.

OBJECTIVE:This study aims to investigate the core target of Astragaloside-Brucea javanica oil nanoemulsion (AS/BJO-NEs) against oral squamous cell carcinoma (OSCC) through network pharmacology, bioinformatics, and in vivo/in vitro experiments, elucidating its effects on epithelial-mesenchymal transition (EMT) mediated via MTFR2 and the underlying molecular mechanisms. METHODS:By integrating network pharmacology and weighted gene co-expression network analysis, critical gene modules linked to tumor phenotypes, EMT-related adverse prognosis, and elevated MTFR2 expression were identified, pinpointing the core target gene. Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, Gene Set Enrichment Analysis, molecular docking and molecular dynamics simulation were utilized for analysis and verification. OSCC cells (SCC9 and CAL27) were cultured, and stable cell lines with CDK1 knockdown or overexpression were established. The effects of AS/BJO-NEs on cell proliferation, migration, invasion, as well as the expression levels of related genes and proteins were evaluated. In vivo, OSCC xenograft models were established in nude mice. Following treatment with AS/BJO-NEs, tumor growth inhibition and changes in the expression of relevant genes were assessed. RESULTS:CDK1 is the core target gene of AS/BJO-NEs against OSCC, and the two can bind stably. CDK1 and MTFR2 showed highly expressed in OSCC and strongly correlated with poor prognosis and EMT. In vitro experiments revealed that AS/BJO-NEs could suppress the proliferation, migration and invasion of OSCC, and down-regulate CDK1, MTFR2 and N-cadherin, whereas up-regulate E-cadherin expression. Knockdown and overexpression of CDK1 further confirmed its effects on OSCC cell phenotype, its regulatory relationship with MTFR2, and the intervention effects of AS/BJO-NEs. In vivo experiments confirmed that AS/BJO-NEs significantly inhibited tumor growth and reduced the expression of CDK1 and MTFR2 in tumor tissues. CONCLUSION:This study demonstrated that AS/BJO-NEs inhibited the proliferation, migration, invasion and EMT process of OSCC by down-regulating CDK1 and subsequently reducing MTFR2 expression, exerting anti-tumor effects and providing new possibilities and a theoretical foundation for OSCC treatment.

Background: Oral squamous cell carcinoma (OSCC) is a common malignant tumor of the head and neck, and glycolysis plays a key role in its development. In the early stages of the study, we prepared a nanoemulsion containing Astragaloside IV (AS-IV) and Brucea javanica oil (BJO). This Astragaloside–Brucea Javanica Oil nanoemulsion (AS/BJO-NE) demonstrated a stronger inhibitory effect on the proliferation, invasion, and migration of OSCC cells than either AS-IV or BJO alone. Preliminary experiments also showed that AS/BJO-NEs inhibited glycolysis in OSCC cells. The aim of this study was to investigate how AS/BJO-NEs act against OSCC by targeting glycolysis-related genes and pathways. Methods: Prepare AS/BJO-NEs and determine their particle size, PDI, and potential. Network pharmacology and bioinformatics analysis were employed to identify the core genes and pathways of AS/BJO-NEs involved in regulating glycolysis in OSCC. In vitro and vivo experiments were performed to investigate the effects of AS/BJO-NEs on OSCC tumor development and core gene expression levels. Results: Aurora kinase A (AURKA) is a critical target through which AS/BJO-NEs regulate glycolytic metabolism in OSCC. Combined in vitro and in vivo experiments revealed that AS/BJO-NEs suppress glycolysis-related enzymes HK2 and PKM2 through the AURKA/PI3K/AKT/HIF-1α signaling axis, consequently inhibiting OSCC proliferation, invasion, metastasis, and subcutaneous tumorigenesis. Conclusions: Bioinformatics analysis combined with in vitro and vivo experiments demonstrated that AS/BJO-NEs downregulate OSCC glycolysis via the AURKA/PI3K/AKT/HIF-1α pathway at the metabolic level, thereby inhibiting OSCC progression. Elucidation of this mechanism provides theoretical support and experimental evidence for the anti-OSCC effects of AS/BJO-NEs.

Rhaponitin (Rha) possesses anti-tumor activity and mediates the transcriptional activity of hypoxia-inducible factor (HIF)-1α that affects cisplatin (Cis) resistance. However, whether Rha can lessen Cis resistance in tongue squamous cell carcinoma (TSCC) by mediating HIF-1α activity is unclear. Cis-resistant SCC9 (SCC9-CisR) cells were treated with Cis, Rha, or Cis plus Rha to explore the effect of Rha on Cis resistance using a cell counting kit-8, flow cytometry, and tumor sphere formation assays. Stemness markers CD44 and SOX2 and HIF-1α mRNA levels were detected by quantitative PCR. The GSE115119 database and plugin iRegulon were employed to select target genes mediated by HIF-1α. Protein levels of HIF-1α, monocarboxylate transporter 4 (MCT4), and the Wnt/β-catenin pathway were measured by western blot. Subcutaneous xenograft models were constructed to explore the efficacy of Rha in combating Cis resistance. Rha repressed the growth and stemness of SCC9-CisR cells in vitro and in vivo. HIF-1α protein levels were markedly elevated in SCC9-CisR cells, yet Rha treatment attenuated the transcriptional activity of HIF-1α but not HIF-1α mRNA levels. Rha plus Cis repressed the viability and stemness of SCC9-CisR cells, but not HIF-1α-knockdown SCC9-CisR cells, compared with Cis alone. Rha-induced stemness inhibition and apoptosis in SCC9-CisR cells were overridden after HIF-1α overexpression. Rha inhibited the Wnt/β-catenin signaling by regulating the HIF-1α/MCT4 axis. In conclusion, Rha reduced cell stemness and enhanced Cis sensitivity in TSCC, which was achieved possibly via suppressing the Wnt/β-catenin signaling through mediation of the HIF-1α/MCT4 axis.

IntroductionOral squamous cell carcinoma (OSCC) is one of the most common malignant tumors in oral and maxillofacial region. The development of new chemotherapy agents and new drug combinations may improve patient survival and quality of life, but both surgery and radiotherapy have significant functional side effects and drug resistance, ultimately resulting in a 5-year survival rate of no more than 60% for OSCC patients. Studies have shown that Brucea javanica oil (BJO) extracts have anti-cancer effects against a variety of cancers, but little research has been reported on OSCC. MethodsCCK8, Colony formation, Scratch test and Transwell invasion assays were applied to determine the effects of BJO on the proliferation, migration, and invasion ability of OSCC cells in vitro. MTFR2 knockdown (shRNA) and overexpression (cDNA) OSCC cells were constructed to evaluate the effect of MTFR2 on the proliferation and invasion of OSCC cells. The nude mouse model of subcutaneous xenograft tumor was used to evaluate the effect of BJO on OSCC cells in vivo. PCR, western blot and immunohistochemistry were used to verify the expression of MTFR2, glycolysis markers and related pathway molecules after BJO treatment.ResultsIn vivo experiments using nude mice with xenografted OSCC cells and in vitro experiments with OSCC cell lines demonstrated that BJO treatment significantly inhibited the proliferation, migration, and invasiveness of OSCC cells. WB and PCR proved that BJO could effectively reduce the expression levels of MTFR2 and SOD2/H2O2 related signal transduction pathways. At the same time, the expression of oxidative phosphorylation markers increased, the expression of glycolytic markers decreased, and glycolysis-mediated decomposition of reactive oxygen species decreased, and H2O2 and oxygen levels decreased.In addition, when MTFR2 expression increased or decreased, SOD2/H2O2 expression also increased or decreased.DiscussionIn this study, we concluded through in vitro and in vivo experiments that BJO may affect the SOD2/H2O2 signaling pathway by down-regulating MTFR2-mediated aerobic glycolysis, thereby inhibiting cell proliferation, Migration, and Invasion. The elucidation of this mechanism helps us to understand the molecular mechanism ofinhibiting OSCC invasion and metastasis by BJO, which has important clinical value or improving the survival rate of OSCC patients.

Kindlin-2 is a cytoskeletal adapter protein that is present in many different cell types. By virtue of its interaction with multiple binding partners, Kindlin-2 intercalates into numerous signaling pathways and cytoskeletal nodes. A specific interaction of Kindlin-2 that is of paramount importance in many cellular responses is its direct binding to the cytoplasmic tails of integrins, an interaction that controls many of the adhesive, migratory and signaling responses mediated by members of the integrin family of cell-surface heterodimers. Kindlin-2 is highly expressed in many cancers and is particularly prominent in prostate cancer cells. CRISPR/cas9 was used as a primary approach to knockout expression of Kindlin-2 in both androgen-independent and dependent prostate cancer cell lines, and the effects of Kindlin-2 suppression on oncogenic properties of these prostate cancer cell lines was examined. Adhesion to extracellular matrix proteins was markedly blunted, consistent with the control of integrin function by Kindlin-2. Migration across matrices was also affected. Anchorage independent growth was markedly suppressed. These observations indicate that Kindlin-2 regulates hallmark features of prostate cancer cells. In androgen expressing cells, testosterone-stimulated adhesion was Kindlin-2-dependent. Furthermore, tumor growth of a prostate cancer cell line lacking Kindlin-2 and implanted into the prostate gland of immunocompromised mice was markedly blunted and was associated with suppression of angiogenesis in the developing tumor. These results establish a key role of Kindlin-2 in prostate cancer progression and suggest that Kindlin-2 represents an interesting therapeutic target for treatment of prostate cancer.

Purpose: This study aims to assess the therapeutic potential of combining Shen-Ling-Bai-Zhu-San (SLBZS) or prebiotics with intermittent fasting (IF) in type 2 diabetes mellitus (T2DM) mice and to investigate the synergistic effects and underlying mechanisms. Methods: Type 2 diabetic mouse models were induced using high-fat diet (HFD) and streptozotocin (STZ), followed by IF treatment. Mice were then grouped for combined therapy with different doses of SLBZS and prebiotics. Fasting blood glucose (FBG) levels, body weight variations, and oral glucose tolerance tests were assessed to elucidate metabolic alterations. The hepatic and renal parameters were evaluated to determine systemic changes in T2DM mice, while the insulin levels were quantified by ELISA to assess glucose homeostasis. Gut microbiota alterations were examined via 16S rRNA sequencing. Alterations of the genes in relevant signaling pathways were analyzed using RT-qPCR. Results: IF improved FBG, body weight, insulin levels, and other diabetes indicators. Combined IF with SLBZS or prebiotics yielded similar effects. Furthermore, it ameliorated dyslipidemia and mitigated hepatic and renal parameters in T2DM mice. Pancreatic tissue histopathology showed islet cell restoration post-intervention. IF therapy reduced the abnormally elevated GSK-3 beta gene expression and increased the abnormally reduced GLUT2 genes. Further analysis indicated that the combination of IF with prebiotics and high doses of SLBZS upregulated the expression of the INSR and IRS] genes. Gut microbiota analysis revealed restored diversity and structure, with notable changes in specific bacterial families. At the family level, the contents of Akkermansiaceae and Bifidobacteriaceae were restored. Phylogenetic investigation of communities by reconstruction of unobserved states (PICRUSt2) analysis suggested metabolic pathway alterations. Conclusion: IF improved type 2 diabetic symptoms, with combined SLBZS and prebiotics showing similar effects. IF with high concentration of SLBZS and prebiotics doses upregulated the INSR and IRS] genes and had superior effects on gut microbiota compared to IF alone.

Abstract Prostate cancer (PC) is the leading type of cancer in men accounting for more than 25% of all newly reported cancer cases and remains the second leading cause of cancer-related deaths in men in USA in 2023. The success of early treatments has resulted in a 98% 5-year survival rate for early-stage PC. However, resistance to androgen ablation can lead to metastatic castration-resistant prostate cancer (mCRPC), with a significant drop in the survival rate to ~30%. The metastatic cascade involves changes in adhesive, metabolic and signaling properties of PC cells, allowing them to survive, proliferate and thrive in their new microenvironment. Kindlin-2 (K2), a cytoskeletal adapter protein, is highly expressed in cancer, including PC cells and tumors, contributing to their tumorigenic properties. The present study focuses on three K2 functions relevant to PC metastasis: 1)Reduction in K2 levels, achieved through CRISPR/Cas9, profoundly affected the tumorigenic properties of PC cell lines LNCaP (androgen dependent), DU145, and PC3 (both androgen independent), influencing colony formation, invasion, and adhesion to integrin ligands. While reduction in K2 was observed, K1 and K3 levels in these cell lines was not altered by K2KO. 2)The E3 ubiquitin ligase, Parkin, known for its role in Parkinson’s disease, has emerged as a tumor suppressor inhibiting various cancer hallmarks. Our recently published studies showed that Parkin induces ubiquitination of specific lysines in K2, leading to the hypothesis that Parkin acts as a tumor suppressor by targeting K2 for degradation. Stable over-expression of Parkin inhibited adhesion to fibrinogen (Fg), while loss of function Parkin mutants failed to do so, supporting a role of Parkin in K2 turnover and its impact on mitochondrial dynamics. We are currently investigating the effect of K2-lysines mutants as well as loss of function Parkin mutants on the progression and metastasis of PC tumors in vivo.3)K2's established function is the regulation of integrin activation, affecting adhesion receptors' transition from low to high affinity/avidity states. The K2-integrin axis is pivotal for understanding oncogenic responses, and its role in the prostate microenvironment in PC progression and metastasis is under exploration using our newly established K2-floxed mouse. The K2-floxed mice are currently being crossed with mice harboring prostate-epithelial specific cre under control of the probasin (Pbsn) promoter. These animal studies will shed light on K2's contribution to PC development and progression in vivo, providing insights into its role in the prostate microenvironment. To sum up, this study emphasizes the crucial role of K2 in the tumorigenic properties of PC cells and tumors. By incorporating CRISPR/Cas9 and innovative mouse models, the study seeks to shed the lights on the mechanisms through which K2 contributes to PC development and progression. Citation Format: Lamyae El Khalki, Wei Wang, Neelum A. Yousaf Zai, Justin Szpendyk, Elzbieta Pluskota, Katarzyna Bialkowska, Edward F. Plow, Lucia R. Languino, Dario C. Altieri, Khalid Sossey-Alaoui. Role of kindlin-2 in prostate cancer progression and metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4380.

Fluoride, a naturally occurring element found in water, soil, food, and atmospheric precipitation, can lead to fluorosis and various health issues when consumed excessively. However, the mechanism of fluorosis is still under investigation. This study utilizes Caenorhabditis elegans as a model organism to investigate the effects of fluoride exposure on biological systems and to explore the mechanisms by which curcumin mitigates fluoride-induced toxicity. Three groups were established: a blank control, a sodium fluoride (NaF) exposure group (concentration 5 mmol/L), and a curcumin intervention group (concentration 25 μmol/L). Physiological parameters, lipofuscin levels, intracellular reactive oxygen species (ROS) levels, mitochondrial membrane potential, and mitochondrial copy numbers were measured to assess the effects of fluoride toxicity and curcumin protection. RNA-seq and qRT-PCR were utilized to investigate the molecular mechanisms underlying fluoride-induced damage and curcumin's mitigating effects. Results indicated that fluoride-exposed nematodes displayed physiological abnormalities, increased ROS production, higher lipofuscin levels, altered mitochondrial membrane potential and mitochondrial copy number, and activated MAPK signaling pathway genes. Curcumin exhibited protective effects on these parameters, suggesting its potential in preventing fluoride-induced harm by modulating oxidative stress and preserving mitochondrial function. This research enhances our understanding of the mechanisms of fluoride toxicity and highlights the potential benefits of curcumin.

Ephrin type-A receptor 2 (EphA2) is a receptor tyrosine kinase that initiates both ligand-dependent tumor-suppressive and ligand-independent oncogenic signaling. We used time-resolved, live-cell fluorescence spectroscopy to show that the ligand-free EphA2 assembles into multimers driven by two types of intermolecular interactions in the ectodomain. The first type entails extended symmetric interactions required for ligand-induced receptor clustering and tumor-suppressive signaling that inhibits activity of the oncogenic extracellular signal-regulated kinase (ERK) and protein kinase B (AKT) protein kinases and suppresses cell migration. The second type is an asymmetric interaction between the amino terminus and the membrane proximal domain of the neighboring receptors, which supports oncogenic signaling and promotes migration in vitro and tumor invasiveness in vivo. Our results identify the molecular interactions that drive the formation of the EphA2 multimeric signaling clusters and reveal the pivotal role of EphA2 assembly in dictating its opposing functions in oncogenesis.

Tongue squamous cell carcinoma (TSCC) is one of the most common malignant tumors of head and neck. Its incidence is on the rise, and the proportion of young patients is gradually increasing, which is prone to tumor recurrence and metastasis. At present, there is no effective method to completely treat TSCC. Studies have shown that brucea javanica oil (BJO) has good antitumor activity against lung cancer and gastrointestinal tumors, but its therapeutic effect on TSCC is not clear. We have previously confirmed that oleic acid, the main component of BJO, can induce apoptosis of TSCC and reduce its invasion and metastasis ability. However, the anticancer effect and mechanism of BJO in TSCC remain unclear. In order to further explore the effects of BJO on the biological characteristics of TSCC cells, we studied the effects of different concentrations of BJO on the migration, invasion ability and epithelial mesenchymal transition (EMT) progression of TSCC cells and the possible mechanisms through in vitro experiments. We found that BJO could inhibit the invasion and metastasis of TSCC and up-regulate miR-138. After BJO treatment, the expression of E-cad was significantly increased, while the expression of EZH2, Slug, p-ERK1/2 and Vimentin was significantly decreased. EZH2 is a miR-138 target gene involved in TSCC. BJO inhibits TSCC invasion and metastasis by regulating the miR-138-EZH2 pathway. In vivo experiments have also well demonstrated the targeting effect of this pathway. This study provides a new therapeutic strategy for the treatment of TSCC.