Search Results (111)

Background: Epithelial-to-mesenchymal transition (EMT) is a cellular reprogramming process characterized by coordinated changes in signaling, membrane organization and metabolism. In a previously established and deeply characterized Huh7 EMT model, it was demonstrated that TGF-β stimulation induces a reproducible shift toward a mesenchymal state accompanied by lipidomic and metabolic remodeling. Building on this framework, the present study evaluates whether extracellular vesicles (EVs)-enriched fractions derived from Capparis spinosa can modulate these EMT-associated alterations. Methods: After detailed physicochemical, molecular, lipidomic and metabolomic characterization, C. spinosa EVs were applied to EMT-induced Huh7 cells. The vesicles were efficiently internalized and, while not inducing a complete epithelial reversion, they attenuated mesenchymal features, indicating a modulatory rather than inhibitory action. Results: Lipidomic profiling showed a partial correction of TGF-β-induced changes including diacylglycerols, phosphoinositides and triglycerides, suggesting interference with lipid signaling and membrane turnover. Metabolomic data further points to reduced mitochondrial and fatty acid oxidation stress, reflected in the re-equilibration of carnitine and acylcarnitine species. Conclusions: Together, these findings indicate that C. spinosa EVs are able to attenuate EMT-associated metabolic and membrane remodeling, positioning them as promising modulators of tumor cell plasticity. Full article

Retained placenta (RP) is a significant postpartum complication in dairy cows. Although abnormal estradiol (E₂) levels are implicated, the underlying cellular mechanisms remain poorly defined. Through RNA-seq analysis of postpartum blood from cows with or without RP, we identified Serum and Glucocorticoid-regulated Kinase 1 (SGK1) as a differentially expressed gene candidate. Analysis of fetal cotyledonary tissues revealed that SGK1 expression was significantly elevated in these tissues, concomitant with markers of suppressed apoptosis, increased levels of tight junction proteins, and an inhibited epithelial–mesenchymal transition (EMT) phenotype. To explore a potential mechanistic link between E₂ and these cellular alterations, we investigated the E₂-SGK1 axis in bovine endometrial epithelial cells in vitro. E₂ treatment upregulated SGK1 expression, reduced apoptosis, increased tight junction protein levels, and suppressed EMT. Conversely, SGK1 knockdown induced apoptosis, disrupted tight junctions, and impaired EMT. Notably, E₂ could not rescue the apoptosis and EMT alterations in SGK1-knockdown cells, indicating that SGK1 is a critical mediator of these E₂ effects in this cellular model. Based on these initial correlative findings in tissues, combined with the subsequent mechanistic experiments in cells, we propose a novel model whereby dysregulation of the E₂- SGK1 axis could contribute to RP pathogenesis by stabilizing the placental interface. Our findings provide the first experimental evidence linking SGK1 to RP and establish a foundation for future in vivo validation. Full article

Objective: This study investigated the anti-prostate cancer mechanism of dictamnine (DIC), focusing on its potential to reverse EMT via DKK1-mediated Wnt/β-catenin inhibition and modulate the tumor microenvironment. Methods: Cell viability, proliferation, migration, and invasion were assessed using CCK-8, colony formation, EdU, wound healing, and Transwell assays. Key targets were identified via transcriptomics and bioinformatics, and validated through molecular docking, co-immunoprecipitation, and cellular thermal shift assay. Protein expression was analyzed by Western blot. Gain/loss-of-function and rescue experiments confirmed target roles. A subcutaneous xenograft model and immunohistochemistry were used for in vivo validation. Results: DIC suppresses prostate cancer malignancy in a concentration-dependent manner. The primary mechanism involves its direct binding to and stabilization of DKK1, which enhances DKK1’s interaction with LRP6. This upregulation of DKK1 inhibits the Wnt/β-catenin signaling pathway, downregulating downstream targets β-catenin/c-Myc/Cyclin D1, and reverses epithelial–mesenchymal transition (EMT) markers. Additionally, DIC modulates key tumor microenvironment factors, including VEGF-A, MMP-9, IL-11, and CXCL-12. Overexpression of DKK1 mimics the antitumor effects of DIC, while knockdown of DKK1 attenuates them. In vivo, DIC inhibits tumor growth, an effect partly mediated through the DKK1/β-catenin axis. Furthermore, DIC potently suppresses angiogenesis (reduced CD31+ staining) independently of DKK1. It also increases tumor-associated macrophage infiltration (elevated F4/80+ cells) in a DKK1-independent manner. Conclusions: DIC exerts its core antitumor effects by targeting DKK1 to inhibit Wnt/β-catenin signaling and EMT. Additionally, it independently suppresses angiogenesis and remodels the immune tumor microenvironment. This multi-level mechanism positions DIC as a promising lead compound for prostate cancer therapy. Full article

Malignant pleural mesothelioma (MPM) is a rare and aggressive tumor associated with asbestos exposure, which represents a current problem. MPM is characterized by a poor prognosis and an unsatisfactory therapeutic approach. Therefore, improving MPM prognosis is the real challenge for research today. Regarding preclinical data, Transforming Growth Factor-β (TGF-β) plays a crucial role in cancer, and its alteration has been associated with tumor progression and invasiveness, in particular through the Epithelial to Mesenchymal Transition (EMT) event. We investigated the role of TGF-β inhibition in proliferation, cell cycle, migration, and invasiveness in human MPM cells. Data obtained clearly highlighted how TGF-β inhibition, through the silencing or treatment of MPM cells with antibody anti-TGF-β (Fresolimumab), significantly reduces cell proliferation (MTT, PCNA) and prevents metastasis, reducing EMT and decreasing the invasiveness and migration of MPM cells. Finally, we also evaluated TGF-β inhibitory effects in 3D MPM cellular models (spheroids), highlighting a significant slowdown in the growth rate of spheroids treated with anti-TGF-β antibody or Fresolimumab, confirming the results previously obtained. Taken as a whole, targeting TGF-β will represent a starting point for future improvements in MPM management. This is particularly important as we foresee a growing increase in MPM in the coming years. Full article

Hepatocellular carcinoma (HCC) remains a leading cause of cancer-related mortality, underscoring the need to elucidate molecular mechanisms that regulate tumor cell state and behavior. Leucine zipper–like post-translational regulator 1 (LZTR1) regulates RAS/mitogen-activated protein kinase (MAPK) signaling, yet LZTR1-dependent transcriptional alterations in HCC cells remain poorly defined. To address this gap and determine how LZTR1 loss reshapes signaling, transcriptional programs, and cellular phenotypes, we established a LZTR1 knockout (KO) Hep3B model and combined pathway profiling with transcriptomic and functional analyses. Immunoblotting revealed increased phosphorylation across the RAF–MEK–ERK–RSK cascade in LZTR1 KO cells. Transcriptome-wide RNA sequencing (RNA-Seq) identified differentially expressed genes, and selected findings were validated by qRT-PCR. Gene set enrichment analysis indicated that the epithelial–mesenchymal transition (EMT) gene set was enriched in control cells. At the protein level, LZTR1 loss remodeled EMT-associated markers in a hybrid epithelial–mesenchymal pattern consistent with epithelial–mesenchymal plasticity (EMP). Vimentin was suppressed at transcript and protein levels. Functionally, LZTR1 KO cells exhibited impaired wound closure and reduced transwell migration and invasion. Collectively, these findings define an EMP-related molecular and phenotypic state associated with LZTR1 deficiency in Hep3B cells, providing insight into how LZTR1 loss reshapes tumor cell behavior in HCC. Full article

Epithelial–mesenchymal plasticity encompasses a spectrum of epithelial and mesenchymal identity states that enable cells to adapt to changing biological contexts. While CD44 isoform usage and epithelial splicing regulators ESRP1/2 are well-characterized in cancer-associated epithelial–mesenchymal transition (EMT), their regulation across physiological, non-transformed identity states remains less well defined. Here, we employed a non-malignant human cellular system comprising primary dermal fibroblasts, induced pluripotent stem (iPS) cells, and iPS-derived mesenchymal stem cells (iPS-MSCs) to define discrete epithelial, intermediate epithelial/mesenchymal, and mesenchymal identity states positioned along an epithelial–mesenchymal identity axis. Morphological assessment, lineage marker profiling, and RT-qPCR analyses revealed reproducible population-level stratification of these states. CD44 expression and alternative splicing followed this hierarchy, with CD44s predominating in fibroblasts, broad variant exon inclusion in iPS cells, and intermediate patterns in iPS-MSCs. ESRP1 expression mirrored CD44 splicing architecture, and ESRP1 silencing in iPS cells induced a shift toward CD44s, confirming its functional contribution to epithelial-associated CD44 splicing. In contrast, Notch-related transcriptional readouts displayed distinct, context-dependent profiles across the examined identity states. Together, this study establishes a tractable non-transformed human model that captures selected molecular features associated with epithelial–mesenchymal plasticity beyond malignant contexts. Full article

Oral squamous cell carcinoma (OSCC) remains a highly prevalent and aggressive malignancy with limited improvements in survival rates. One of the major obstacles to successful treatment is the development of chemoresistance, which contributes to recurrence, metastasis, and treatment failure. This narrative review aims to integrate current evidence on the molecular and cellular mechanisms that drive chemoresistance in OSCC and to delineate how these processes converge under therapeutic pressure. A structured search was performed to identify relevant studies addressing chemoresistance in OSCC, focusing on preclinical and translational evidence. Multiple interconnected mechanisms have been implicated in driving resistance in OSCC, including epigenetic alterations, deregulated signaling pathways, cancer stem cell plasticity, epithelial–mesenchymal transition (EMT), interactions with the tumor microenvironment (TME), drug efflux mediated by ATP-binding cassette (ABC) transporters, and enhanced DNA damage response. In combination, these mechanisms support tumor persistence and limit effective antitumor immunity. Emerging strategies such as epigenetic modulators, signaling pathway inhibitors, immunomodulation, and nanomedicine-based delivery systems have shown promising results in preclinical models. By highlighting convergent resistance networks, this integrative perspective supports the rational design of combination therapies and biomarker-guided strategies aimed at overcoming chemoresistance in OSCC. Full article

Background/Objectives: Neovascular age-related macular degeneration (nAMD) poses a serious threat to the eyesight of older adults, representing a leading cause of irreversible vision loss. Anti-vascular endothelial growth factor (anti-VEGF) treatments are effective but require repeated intraocular injections and show poor responses in some patients. CGK012 is a novel derivative of decursin that inhibits the Wnt/β-catenin pathway. This study aimed to elucidate the mode of action of CGK012 and examine its therapeutic effects. Methods: We performed in vitro cellular studies in a retinal pigment epithelial (RPE) cell line (ARPE-19) and human umbilical vein endothelial cells (HUVECs). We examined the in vivo efficacy of CGK012-loaded implants in laser-induced choroidal neovascularization (CNV) rabbit models. We also determined the implants’ in vitro dissolution, intraocular release, and disposition characteristics. Results: CGK012 decreased angiogenic/proinflammatory factor expression and suppressed the epithelial–mesenchymal transition (EMT) in RPE cells by promoting intracellular β-catenin degradation. Additionally, it repressed the expression of cyclin D1 and c-myc, downstream target genes of β-catenin, and inhibited HUVEC capillary tube formation. CGK012-loaded poly (lactic-co-glycolic acid) (PLGA) intravitreal implants significantly reduced vascular leakage in a laser-induced CNV rabbit model. Notably, CGK012 released from the implant was highly permeable to retina/choroid tissue and downregulated β-catenin, angiogenic/inflammatory factors, and vimentin in the rabbit model. The CGK012 concentration reached a plateau at 28–42 days in the vitreous humor and decayed with a half-life of 14 days without systemic exposure. Conclusions: Our findings demonstrate that CGK012 implants prevent choroidal neovascularization through the Wnt/β-catenin pathway suppression and produce high concentrations of CGK012 in the posterior eye segment with prolonged release. Thus, these implants provide more therapeutic choices for nAMD treatment. Full article

Tumor metastasis is the leading cause of death in cancer patients, with epithelial–mesenchymal transition (EMT) playing a key role. To systematically elucidate the cellular mechanisms and molecular networks through which EMT drives metastasis across cancers, this study integrated transcriptomic data from over 1.2 million single cells across 265 samples representing 12 primary epithelial cancers, constructing a comprehensive pan-cancer single-cell atlas covering diverse stages and metastatic states. By analyzing the metastatic features and interaction networks of malignant epithelial cells and cancer-associated fibroblasts (CAFs), we identified cancer-specific metastasis-related gene sets. Based on these genes, multiple machine learning algorithms were applied to build cancer-specific and cross-cancer metastasis prediction models, leading to the development of the metastasis prediction score (MPS) and global metastasis prediction score (GMPS). Both scores showed excellent predictive performance in independent test and external validation cohorts. MPS exhibited higher cancer specificity, whereas GMPS showed stronger cross-cancer generalization. Moreover, elevated MPS and GMPS reflected immunosuppressive tumor microenvironment features and were significantly associated with poor prognosis across multiple cancer types. Finally, through a drug repositioning framework, we identified several potential anti-metastatic compounds targeting the metastasis network, among which Fostamatinib demonstrated broad-spectrum therapeutic potential against metastasis in multiple cancers. Full article

Metastasis remains the leading cause of cancer deaths, heavily influenced by aging-related biological processes. As global life expectancy increases, cancer incidence and progression complexity in older adults also rise, emphasizing the urgent need to understand how the aging tumor microenvironment (TME) promotes metastasis. This review explores the molecular and cellular mechanisms behind metastatic development in the aged TME, focusing on the combined impacts of cellular senescence, chronic inflammation (inflammaging), immune system decline, extracellular matrix (ECM) changes, and abnormal blood vessel growth. Aging causes an accumulation of senescent cells that secrete a range of cytokines, growth factors, and enzymes (SASPs), which remodel the ECM, making it stiffer and more degradable, and activate pro-metastatic pathways like TGF-β, STAT3, and MAPK, aiding processes like EMT and tumor invasion. Meanwhile, persistent low-grade inflammation attracts immunosuppressive cells, and immune decline hampers tumor surveillance, allowing cancer cells to evade immune detection. The aged TME also undergoes significant vascular and metabolic changes, such as abnormal angiogenesis and hypoxia, supporting the growth of more aggressive, treatment-resistant cancer clones and spreading metastases. These changes are driven by hallmarks of molecular aging, including telomere shortening, oxidative DNA damage, and epigenetic alterations, which lead to genetic instability and turn the aged stroma into fertile ground for metastasis. The review also discusses new therapeutic approaches, including senolytics, anti-inflammatory treatments, immune system rejuvenation, and metabolic strategies, highlighting the importance of age-specific models and precision medicine to enhance outcomes for the growing number of elderly cancer patients. Full article

Background: Three-dimensional (3D) culture systems offer a physiologically relevant alternative to monolayers for studying tumor organization, signaling, and drug response. HER2-positive breast cancers (BCa) account for 15–30% of BCa cases and benefit from HER2-targeted therapies, yet predictive in vitro models remain limited. Objective: To generate and compare 3D spheroids from two HER2+ BCa cell lines, SKBR3 and BT474, and investigate how 3D architecture influences HER2 distribution, intracellular signaling, and cellular organization. Methods: Spheroids were reproducibly generated from SKBR3 and BT474 cells and analyzed after 4 days of culture. Cell viability was evaluated using live/dead staining, HER2 distribution was assessed by confocal microscopy and quantified on cryosections, and protein expression/phosphorylation was measured by Western blotting. Epithelial and EMT markers were visualized by immunofluorescence, and ultrastructural features were examined by transmission electron microscopy (TEM). Results: Both cell lines formed viable spheroids with distinct architectures: SKBR3 spheroids were loose and heterogeneous, whereas BT474 spheroids were compact and highly spherical. Confocal and cryosection imaging showed consistent membrane HER2 localization with a progressive signal decrease toward the core of the spheroids, more pronounced in BT474. Western blotting revealed divergent HER2 expression and AKT phosphorylation: SKBR3 spheroids displayed increased HER2 but reduced pAKT, while BT474 spheroids showed reduced HER2 and pAKT levels. EpCAM and E-cadherin staining revealed cell line-specific epithelial organization, and TEM demonstrated differences in intercellular spacing and mitochondrial morphology, reflecting spheroid compactness. Conclusions: 3D architecture profoundly influences HER2 distribution, signaling, and structural organization in HER2+ BCa spheroids. This model provides a robust platform for investigating architecture-dependent molecular processes, with potential applications in drug response, receptor trafficking, and targeted therapy evaluation. Full article

Lung cancer remains the leading cause of cancer-related mortality worldwide, with non-small-cell lung cancer (NSCLC) being the most prevalent subtype. NSCLC is marked by a complex genetic makeup, involving numerous driver mutations and epigenetic changes that drive tumor growth and resistance to treatment. While several approaches, including chemotherapy and targeted therapy, have been used for lung cancer treatment, their overall responses remain dismal, indicating the need to explore alternative targets implicated in cancer growth. Among various candidates, peroxisome proliferator-activated receptor-gamma (PPARγ), which plays critical roles in regulating cellular functions related to tumorigenesis, has been explored as a promising target for NSCLC intervention. To that end, thiazolidinediones, including pioglitazone, that target PPARγ have shown promise in multiple cellular and preclinical models of NSCLC. Mechanistically, pioglitazone inhibits cancer growth and induces apoptosis via downregulating key signaling pathways, including mitogen-activated protein kinase (MAPK), which play critical roles in regulating cellular activities such as epithelial-to-mesenchymal transition (EMT), cellular bioenergetics, and glucose metabolism. This review highlights the recent updates on the mechanistic insights and the efficacy of PPARγ agonist-based approaches, with an emphasis on pioglitazone, for the treatment of NSCLC. We logically discuss the experimental evidence from the in vitro and in vivo studies exploring pioglitazone’s effect on metabolic pathways, chemical-carcinogen-induced tumorigenesis, the targeting of cell signaling pathways, and then its combination with other therapeutic agents. We also present clinical studies that support pioglitazone’s potential in chemoprevention and underscore its further exploration in large cohorts of NSCLC patients. Full article

Background/Objectives: Cutaneous squamous cell carcinoma (cSCC) is a very common skin malignancy of the head and neck area, with a propensity to spread to local lymph nodes. Epithelial-to-mesenchymal transition (EMT) and cancer-associated fibroblasts (CAFs) play a well-documented role in the progression of the disease. In this study, we developed and characterised multicellular tumour spheroids (MCTS) composed of patient-derived metastatic cSCC cell lines—each exhibiting distinct phenotypes—combined with either dermal- or lymph node-derived fibroblasts. We aimed to investigate how these cellular combinations influence MCTS formation, spatial architecture, and invasive behaviour. We hypothesised that the interplay between different cSCC and fibroblast cell combinations would differentially influence spheroid formation and invasion. Methods: Using live-cell microscopy we assessed the spatial architectures specific to each cell line-fibroblast combination and evaluated the expression of EMT and CAF markers. Furthermore, we utilised MCTS in invasion models to investigate associations between the mode of invasion and the EMT phenotype of the cancer cell line. Results: We show that metastatic cSCC/fibroblast MCTS self-organise into distinct spatial architectures. They also invade through collagen in a manner influenced by fibroblasts but dominated by the EMT status of the originating cancer cells. Conclusions: These findings highlight the physiological relevance and utility of MCTS as models for investigating tumour–stroma interactions and invasion dynamics in metastatic cSCC. Full article

Helicobacter pylori (H. pylori) is a well-known pathogen associated with chronic gastric infection, progressing from gastritis to gastric adenocarcinoma, but the dynamic phenotypic and molecular characteristics of gastric epithelial cells during sustained infection remain unclear. We established a chronic infection model using the human gastric epithelial cell line GES-1, exposed to H. pylori or its lysate across 30 generations, dynamically assessing cell proliferation, migration, invasion, apoptosis, autophagy, and epithelial–mesenchymal transition (EMT) markers, with RNA sequencing for transcriptomic changes and a Mongolian gerbil model to validate chronic pathological progression. Acute H. pylori exposure induced pronounced morphological changes; suppressed proliferation, migration, and invasion; triggered apoptosis; and blocked autophagic flux, while long-term stimulation reversed these effects. EMT markers showed progressive loss of epithelial characteristics with chronic infection. RNA sequencing revealed a dynamic shift from inflammation-driven apoptosis to adaptive survival mechanisms. In vivo, prolonged infection induced dynamic TLR expression alongside progressive gastric pathology, including atrophy and dysplasia. Our study provides new molecular evidence for dynamic cellular and immunological adaptations of gastric epithelial cells under chronic H. pylori infection, highlighting critical intervention windows for preventing gastric carcinogenesis. Full article

Breast cancer is the most prevalent solid tumor diagnosed in women worldwide, remaining a leading cause of cancer-related mortality. Among its subtypes, triple-negative breast cancer (TNBC) is characterized by high aggressiveness and heterogeneity, accounting for approximately 90% of breast cancer-related deaths. Receptor tyrosine kinases (RTKs), such as epidermal growth factor receptor (EGFR) and the insulin-like growth factor I receptor (IGF-IR), are critical cell growth and survival regulators, with their dysregulation closely related to therapy resistance in breast cancer. Studies on RTK targeting have shown promise, and recently attention has shifted toward developing more physiologically relevant preclinical models. Unlike traditional two-dimensional (2D) cell cultures, 3D models such as spheroids better mimic the complex nature of the tumor microenvironment (TME), offering a more accurate representation of tumor behavior and progression. This study utilized both 2D and 3D culture models to assess the effects of EGFR and IGF-IR inhibition, individually and in combination, in two TNBC cell lines with distinct metastatic potential. The results demonstrate that both receptors play critical roles in regulating key cellular functions, including migration, expression of epithelial-to-mesenchymal transition (EMT) markers and matrix metalloproteinases (MMPs). The use of 3D spheroid models enabled the evaluation of additional functional properties, such as spheroid growth and dissemination, revealing treatment-dependent responses to combined receptor inhibition. Overall, this dual-model approach underscores the importance of incorporating 3D culture systems in preclinical cancer research and provides new insights into the regulatory roles of EGFR and IGF-IR in TNBC progression. Full article