Epithelial-mesenchymal Transition Markers Research Articles

3D Scaffold-Based Culture System Enhances Preclinical Evaluation of Natural Killer Cell Therapy in A549 Lung Cancer Cells.

Cell-based immunotherapies have emerged as promising cancer treatment modalities, demonstrating remarkable clinical efficacy. As interest in applying immune cell-based therapies to solid tumors has gained momentum, experimental models that enable long-term monitoring and mimic clinical administration are increasingly necessary. This study explores the potential of scaffold-based cell culture technologies, specifically three-dimensional (3D) extracellular matrix (ECM)-like frameworks, as promising solutions. These frameworks facilitate unhindered immune cell growth and enable continuous cancer cell culture. The three-dimensional (3D) cell culture model was developed using tailored scaffolds for natural killer (NK) cell culture. Within this framework, A549 lung cancer cells were cocultured with NK cells, allowing real-time monitoring for up to 28 days. The expression of critical markers associated with anticancer drug resistance and epithelial-mesenchymal transition (EMT) was evaluated in cancer cells within this 3D culture context. Compared to conventional 2D monolayer cultures, this 3D scaffold-based culture revealed that solid tumor cells, specifically A549 cells, exhibited heightened resistance to anticancer drugs. Additionally, the 3D culture environment upregulated the expression of EMT markers namely vimentin, N-cadherin, and fibronectin, while NK and zEGFR-CAR-NK cells displayed anticancer effects. In the two-dimensional (2D) coculture, only zEGFR-CAR-NK cells exhibited such effects in the 3D coculture system, highlighting an intriguing inconsistency with the 2D culture model, further confirmed by in vivo experiments. This in vitro 3D cell culture model reliably predicts outcomes in NK immunotherapy experiments. Thus, it represents a valuable tool for investigating drug resistance mechanisms and assessing the efficacy of immune cell-based therapies. By bridging the gap between in vitro and in vivo investigations, this model effectively translates potential treatments into animal models and facilitates rigorous preclinical evaluations.

Transglutaminase 2 promotes epithelial-to-mesenchymal transition by regulating the expression of matrix metalloproteinase 7 in colorectal cancer cells via the MEK/ERK signaling pathway

Tissue transglutaminase 2 (TGM2) and matrix metalloproteinase 7 (MMP7) are suggested to be involved in cancer development and progression, however, their specific role in colon cancer remains elusive. The present study investigated whether TGM2 and MMP7 influence epithelial-mesenchymal-transition (EMT) processes of colon cancer cells.TGM2 was either overexpressed or knocked down in SW480 and HCT-116 cells, and MMP7 expression and activity analyzed. Conversely, MMP7 was silenced and its correlation with TGM2 expression and activity examined. Co-immunoprecipitation served to evaluate TGM2-MMP7-interaction. TGM2 and MMP7 expression were correlated with invasion, migration, EMT marker expression (E-cadherin, N-cadherin, Slug, Snail), and ERK/MEK signaling.TGM2 overexpression enhanced MMP7 expression and activity, promoted cell invasion, migration and EMT, characterized by increased N-cadherin and Snail/Slug expression. TGM2 knockdown resulted in the opposite effects. Knocking down MMP7 was associated with reduced TGM2 protein expression, cell invasion and migration. Down-regulation of MMP7 diminished ERK/MEK signaling, whereas its up-regulation activated this pathway. The ERK-inhibitor GDC-0994 blocked phosphorylation of MEK/ERK and suppressed TGM2 and MMP7.TGM2 communicates with MMP7 in colon cancer cells forces cell migration and invasion by the MEK/ERK signaling pathway and triggers EMT. Inhibiting TGM2 could thus offer new therapeutic options to treat patients with colon cancer, particularly to prevent metastatic progression.

AMIGO2 enhances the invasive potential of colorectal cancer by inducing EMT.

In our previous studies, we identified amphoterin-inducible gene and open reading frame 2 (AMIGO2) as a driver gene for liver metastasis and found that AMIGO2 expression in cancer cells worsens the prognosis of patients with colorectal cancer (CRC). Epithelial-mesenchymal transition (EMT) is a trigger for CRC to acquire a malignant phenotype, such as invasive potential, leading to metastasis. However, the role of AMIGO2 expression in the invasive potential of CRC cells remains unclear. Thus, this study aimed to examine AMIGO2 expression and elucidate the mechanisms by which it induces EMT and promotes CRC invasion. Activation of the TGFβ/Smad signaling pathway was found involved in AMIGO2-induced EMT, and treatment with the TGFβ receptor inhibitor LY2109761 suppressed AMIGO2-induced EMT. Studies using CRC samples showed that AMIGO2 expression was highly upregulated in the invasive front, where AMIGO2 expression was localized to the nucleus and associated with EMT marker expression. These results suggest that the nuclear translocation of AMIGO2 induces EMT to promote CRC invasion by activating the TGFβ/Smad signaling pathway. Thus, AMIGO2 is an attractive therapeutic target for inhibiting EMT and metastatic CRC progression.

Inhibiting SNX10 induces autophagy to suppress invasion and EMT and inhibits the PI3K/AKT pathway in cervical cancer.

Cervical cancer (CC) is a prevalent malignancy among women with high morbidity and poor prognosis. Sorting nexin 10 (SNX10) is a newly recognized cancer regulatory factor, while its action on CC progression remains elusive. Hence, this study studied the effect of SNX10 on CC development and investigated the mechanism. The SNX10 level in CC and the overall survival of CC cases with different SNX10 expressions were determined by bioinformatics analysis in GEPIA. The SNX10 expression in tumor tissues and clinical significance were studied in 64 CC cases. The overall survival was assessed using Kaplan-Meier analysis. The formation of LC3 was evaluated using immunofluorescence. Cell invasion was measured using the Transwell assay. Epithelial-to-mesenchymal transition (EMT) was determined by observing cell morphology and assessing EMT marker levels. A xenograft tumor was constructed to evaluate tumor growth. SNX10 was elevated in CC tissues and cells, and the CC cases with high SNX10 levels exhibited poor overall survival. Besides, SNX10 correlated with the FIGO stage, lymph node invasion, and stromal invasion of CC. SNX10 silencing induced CC cell autophagy and suppressed CC cell invasion and EMT. Meanwhile, silenced SNX10 could suppress invasion and EMT via inducing autophagy. Furthermore, SNX10 inhibition suppressed the PI3K/AKT pathway. Moreover, silenced SNX10 restrained the tumor growth, autophagy, and EMT of CC in vivo. SNX10 was enhanced in CC and correlated with poor prognosis. Silenced SNX10 induced autophagy to suppress invasion and EMT and inhibited the PI3K/AKT pathway in CC, making SNX10 a valuable molecule for CC therapy.

UBE2Q1 as a novel cancer biomarker for lung adenocarcinoma: Short Title: Oncogenic function of UBE2Q1 in lung adenocarcinoma

PurposeUbiquitin-conjugating enzymes (E2s) participate in various tumor-promoting processes. UBE2Q1 is a member of the E2 family. This research aimed to detect the expression level of UBE2Q1 in human lung adenocarcinoma and to study its malignant biological function. MethodsWestern blot, qRT-PCR and immunohistochemistry was used to measure the expression of UBE2Q1 in human lung adenocarcinoma tissues. The association between UBE2Q1 expression and clinic-pathological variables in 99 lung adenocarcinoma samples was analyzed by immunohistochemistry. In vitro experiment, establishing UBE2Q1 knockdown pattern, the markers of apoptosis, cell cycle and epithelial-mesenchymal transition (EMT) were analyzed by Western blot. CCK8, colony formation, Transwell and invasion assay analyzed the effect of UBE2Q1 knockdown on the proliferation, metastasis and invasion of lung cancer cells. ResultsUBE2Q1 was overexpressed in lung adenocarcinoma, and the expression level of UBE2Q1 was related with TNM stage, tumor size, and lymph node metastasis. The high level of UBE2Q1 expression was also associated with poor survival and was an independent risk factor. In vitro, It was also confirmed that steady downregulation of UBE2Q1 could promote apoptosis, induce G2/M cell cycle arrest and regulate EMT. UBE2Q1 silencing dramatically reduce lung tumor cells proliferation, migration and invasion capacities. ConclusionsUBE2Q1 may serve as a prognostic biomarker and a new therapeutic target of lung adenocarcinoma.

Xihuang pill suppresses breast cancer malignancy by inhibiting TGF-β signaling and acquires chemotherapy benefits

Ethnopharmacological relevanceBreast cancer (BC) has an extremely high global incidence rate. The Xihuang pill (XHP), a traditional Chinese formula, originates from Hongxu Wang's “Life-Saving Manual of Diagnosis and Treatment of External Diseases” written during the Qing Dynasty. In this book, XHP, was first suggested as an anticancer treatment for BC. However, the regulatory mechanism of XHP on BC requires further investigated. Aim of the studyTo assess the effects of XHP on BC and elucidate the underlying associated signaling network. Materials and methodsThe influence of XHP on cellular viability, proliferation, and apoptosis of MDA-MB-231 and BT-549 cells were examined. The ability to metastasize was evaluated using Transwell invasion and wound healing tests. Western blotting was used to examine the epithelial-mesenchymal transition (EMT) markers expression. RNA sequencing and bioinformatic analysis were utilized to investigate the regulation mechanism of XHP. A subcutaneous tumor model was developed to study the tumor-inhibitory effects of XHP or/and Doxorubicin (Dox) on BALB/c nude mice, and the EMT marker levels in tumor tissues were determined using immunohistochemical labeling. ResultsXHP demonstrated anticancer effects on BC cells by suppressing cell proliferation, inducing cellular apoptosis, and inhibiting EMT progression. XHP may regulate the EMT via the TGF-β axis, as shown by RNA sequencing and Western blotting analysis. Furthermore, the combination of XHP and Dox had a stronger therapeutic effect on BC cell proliferation, apoptosis, and metastasis in both cellular and animal models. ConclusionsWe were the first to reveal that XHP abrogated EMT progression via modulating the TGF-β axis. Furthermore, the combination therapy of XHP and Dox presents a promising novel therapeutic candidate for BC patients.

Oleanolic acid inhibits aldo-keto reductase family 1 member B10-induced cancer stemness and avoids cisplatin-based chemotherapy resistance via the Snail signaling pathway in oral squamous cell carcinoma cell lines

Background/purposeOral squamous cell carcinoma (OSCC) is a common malignancy often associated with poor prognosis due to chemoresistance. In this study, we investigated whether arecoline, a major alkaloid in betel nuts, can stimulate aldo-keto reductase family 1 member B10 (AKR1B10) levels in OSCC, promoting cancer stemness and leading to resistance to cisplatin (CDDP)-based chemotherapy. Materials and methodsGain- and Loss- of AKR1B10 functions were analyzed using WB and q-PCR of OSCC cells. Stemness, epithelial mesenchymal transition (EMT) markers, and CDDP drug resistance in overexpressed AKR1B10 were also identified. ResultsUpregulated AKR1B10 in OSCC significantly increased cell motility and aggregation. The results also showed that the canonical TGF-β1-Smad3 pathway was involved in arecoline-induced AKR1B10 expression, further increasing cancer stemness with CDDP resistance via the Snail-dependent EMT pathway. Moreover, oleanolic acid (OA) and ROS/RNS (reactive oxygen/nitrogen species) inhibitors effectively reversed AKR1B10-induced CDDP-resistance. ConclusionArecoline-induced ROS/RNS to hyper-activate AKR1B10 in tumor sphere cells via the TGF-β1-Smad3 pathway. Furthermore, AKR1B10 enhanced CDDP resistance in OSCC cells via EMT-inducing markers. Finally, Finally, OA may efficiently target CDDP resistance, reverse stemness in OSCC cells, and have the potential as a novel anticancer drug.

Inhibition of Epithelial-Mesenchymal Transition and Migration in Cisplatin-resistant Cancer Through Combined Treatment With Cisplatin and SH003.

This study investigated the synergistic effects of combining cisplatin and SH003 treatment on the viability, apoptosis, cytotoxicity, migration and epithelial-mesenchymal transition (EMT) in cisplatin-resistant cancer cell lines YD-8/CIS, YD-9/CIS and YD-38/CIS. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to assess cell viability, while trypan blue exclusion assay was used to evaluate cytotoxicity. Flow cytometry and western blot analysis measured apoptotic cell death. Wound-healing assays evaluated cell migration and EMT markers. Combination index (CI) plots were used to evaluate the combinatory effects of the treatment. Combination therapy significantly reduced cell viability more effectively than each agent alone, as demonstrated by the MTT assay, with CI plots confirming notable synergism. Trypan blue exclusion assays indicated increased cell death and cytotoxicity in combination treatment than in both monotherapies, although the increase was not significant. Flow cytometry and western blot analysis revealed no significant synergistic effect on apoptotic cell death. However, wound-healing assays revealed that the combination of cisplatin and SH003 significantly inhibited cell migration and regulated EMT markers, indicating the potential reversal of EMT. Combining cisplatin and SH003 therapy may potentially be a more effective strategy for treating cisplatin-resistant cancer by increasing cytotoxicity and inhibiting metastasis. Further research is required to elucidate the underlying mechanisms and evaluate the in vivo efficacy of this combination therapy.

Hsa_circ_0043533 modulates apoptosis and viability of granulosa cells via miR-409-3p/BCL2 and EMT signalling in PCOS: Providing novel perspective of metformin

Polycystic ovary syndrome (PCOS) represents a significant cause of infertility among women of reproductive age. Studies have established a close association between granulosa cells (GCs) and the abnormal follicle formation and ovulation processes characteristic of PCOS. The interactions among hsa_circ_0043533, miR-409–3p, and BCL2 were verified through luciferase activity assays. In PCOS patients, granulosa cells exhibit notably reduced apoptosis but enhanced growth, leading to their accumulation and the development of polycystic ovaries. The involvement of non-coding RNAs in PCOS has been documented, with elevated levels of hsa_circ_0043533 observed in this condition. A comprehensive series of experiments were conducted to explore the role of hsa_circ_0043533 in PCOS and elucidate its underlying mechanisms. Silencing hsa_circ_0043533 was found to promote apoptosis and hinder the migration, proliferation, and viability of KGN cells. Furthermore, we uncovered the regulatory effects of hsa_circ_0043533 on the miR-409–3p/BCL2 axis and key markers of Epithelial-Mesenchymal Transition (EMT). Additionally, it was observed that metformin modulates the hsa_circ_0043533/miR-409–3p/BCL2 axis. Overall, this study provides novel insights into the molecular mechanisms regulating granulosa cell proliferation and apoptosis in PCOS, further elucidating the molecular pathogenesis of this condition.

USP5 promotes tumor progression by stabilizing SLUG in bladder cancer.

Bladder cancer ranks as the second most prevalent urology malignancy globally. Invasive metastasis is a significant contributor to mortality among patients with bladder cancer, yet the underlying mechanisms remain elusive. Deubiquitinases are pivotal in carcinogenesis, with USP5 implicated in the malignant progression of hepatocellular carcinoma, colorectal cancer and non-small cell lung cancer. The present study assessed the role and mechanism of ubiquitin-specific proteinase 5 (USP5) in the malignant progression of bladder cancer. The association between USP5 expression and bladder cancer prognosis and stage was analyzed using The Cancer Genome Atlas database. Moreover, to elucidate the role of USP5 in bladder cancer, USP5 overexpression and knockdown cell lines were established using T24 cells. Cell viability, proliferation and migration were assessed using Cell Counting Kit-8, Transwell and scratch assays, respectively. Cyclohexanamide was used to evaluate the effect of USP5 expression on Snail family zinc finger 2 (SLUG) stability. Immunoprecipitation and immunofluorescence co-localization were utilized to probe the interaction between USP5 and SLUG. Changes in mRNA and protein levels were assessed using reverse transcription-quantitative PCR and western blotting, respectively. The results revealed that patients with bladder cancer with high USP5 expression had significantly shorter survival (P<0.05) and a higher clinicopathologic stage (P<0.05) than those with low USP5 expression. T24 cells overexpressing USP5 demonstrated significantly increased proliferation (P<0.05), invasion (P<0.05) and expression of epithelial-mesenchymal transition markers (P<0.05); whereas T24 cells with knocked-down USP5 expression revealed significantly reduced proliferation (P<0.05), invasion (P<0.05) and epithelial-mesenchymal transition markers (P<0.05). Immunoprecipitation experiments demonstrated the binding of USP5 to SLUG in bladder cancer cells, with further analysis revealing that USP5 upregulated protein levels of SLUG by inhibiting its ubiquitination. Furthermore, the treatment of bladder cancer cells with Degrasyn, a USP5 inhibitor, was associated with a significant inhibition of the proliferation (P<0.05) and invasion (P<0.05) of T24 cells. In conclusion, the findings of the present study underscore the role of USP5 in promoting the malignant progression of bladder cancer through the stabilization of SLUG. Targeting USP5 holds promise as a strategy for inhibiting bladder cancer progression.

Methanolic Extract of Cimicifuga foetida Induces G1 Cell Cycle Arrest and Apoptosis and Inhibits Metastasis of Glioma Cells.

Glioblastoma multiforme (GBM) is among the most aggressive and challenging brain tumors, with limited treatment options. Cimicifuga foetida, a traditional Chinese medicine, has shown promise due to its bioactive components. This study investigates the anti-glioma effects of a methanolic extract of C. foetida (CF-ME) in GBM cell lines. The effects of CF-ME and its index compounds (caffeic acid, cimifugin, ferulic acid, and isoferulic acid) on GBM cell viability were assessed using MTT assays on U87 MG, A172, and T98G cell lines. The ability of CF-ME to induce cell cycle arrest, apoptosis, and autophagy and inhibit metastasis was evaluated using flow cytometry, Western blotting, and functional assays. Additionally, the synergistic potential of CF-ME with temozolomide (TMZ) was explored. CF-ME significantly reduced GBM cell viability in a dose- and time-dependent manner, induced G1 phase cell cycle arrest, promoted apoptosis via caspase activation, and triggered autophagy. CF-ME also inhibited GBM cell invasion, migration, and adhesion, likely by modulating epithelial-mesenchymal transition (EMT) markers. Combined with TMZ, CF-ME further enhanced reduced GBM cell viability, suggesting a potential synergistic effect. However, the individual index compounds of CF-ME exhibited only modest inhibitory effects, indicating that the full anti-glioma activity may result from the synergistic interactions among its components. CF-ME exhibited potent anti-glioma activity through multiple mechanisms, including cell cycle arrest, apoptosis, autophagy, and the inhibition of metastasis. Combining CF-ME with TMZ further enhanced its therapeutic potential, making it a promising candidate for adjuvant therapy in glioblastoma treatment.

Parishin A Inhibits Oral Squamous Cell Carcinoma via the AKT/mTOR Signaling Pathway.

Oral squamous cell carcinoma (OSCC) is an aggressive cancer with limited treatment options. Parishin A, a natural compound derived from Gastrodia elata, possesses multiple therapeutic properties. However, its effects on OSCC remain unexplored. This study explores the anti-cancer potential of Parishin A on OSCC and its mechanisms. OSCC cell lines YD-10B and Ca9-22 were treated with varying Parishin A concentrations. Cell viability was detected using the CCK-8 assay, and colony formation was evaluated in agarose gel. Migration and invasion ability were assessed through wound healing and Matrigel invasion assays. The protein expression levels involved in the PI3K/AKT/mTOR signaling pathway and epithelial-mesenchymal transition (EMT) markers were examined via Western blotting. Parishin A inhibited OSCC cell viability in both dose- and time-dependent manners, with significant reductions at 20, 40, 60, and 80 μM, without affecting normal human gingival fibroblasts. Colony formation decreased substantially at ≥40 μM higher Parishin A concentrations in a dose-dependent manner. Also, migration and invasion assays showed significant suppression by Parishin A treatment concentration ≥40 μM in a dose-dependent manner, as evidenced by decreased wound closure and invasion. Western blot analyses revealed increased E-cadherin levels and decreased N-cadherin and vimentin levels, suggesting EMT inhibition. Parishin A also decreased the phosphorylation levels of PI3K, AKT, and mTOR. Collectively, these findings support the potential of Parishin A as an anti-OSCC agent.

Anlotinib reverses osimertinib resistance via inhibiting epithelial-to-mesenchymal transition and angiogenesis in non-small cell lung cancer.

In the present, we aimed to investigate the effect of anlotinib on the potential reversal of osimertinib resistance by inhibiting the formation of epithelial-to-mesenchymal transition (EMT) and angiogenesis. In a clinical case, anlotinib reversed osimertinib resistance in Non-small cell lung cancer (NSCLC). We performed an immunohistochemical experiment on tumor tissues from three non-small cell lung cancer patients exhibiting osimertinib resistance to analyze alterations in the expression levels of EMT markers and vascular endothelial growth factor A (VEGFA) before and after osimertinib resistance. The results revealed the downregulation of E-cadherin, coupled with the upregulation of vimentin and VEGFA in tumor tissues of patients exhibiting osimertinib resistance, compared with the expression in tissues of patients before taking osimertinib. Subsequently, we established osimertinib-resistant cell lines and found that the osimertinib-resistant cells acquired the EMT features. Then, we analyzed the synergistic effects of the combination therapy to verify whether anlotinib could reverse osimertinib resistance by inhibiting EMT. The expression levels of VEGFA and micro-vessels were analyzed in the combination group in vitro. Finally, we explored the reversal of osimertinib resistance in combination with anlotinib in vivo with 20 nude mice. The combined treatment of osimertinib and anlotinib effectively prevented the metastasis of resistant cells, which also inhibited tumor growth, exerted anti-tumor activity, and ultimately reversed osimertinib resistance in mice. The co-administration of osimertinib and anlotinib demonstrated their synergistic efficacy in inhibiting EMT and angiogenesis in three NSCLC patients, ultimately reversing osimertinib resistance.

The Role of Breastmilk in Macrophage-Tumour Cell Interactions in Postpartum Breast Cancer

Background: Lactation is associated with long-term reduced risk of breast cancer. However, there is a transient increased risk of breast cancer in the 5 to 10 years postpartum and this is associated with a high incidence of metastasis and mortality. Breastmilk is a physiological fluid secreted by the mammary glands intimately connected with breast cells and the microenvironment that may affect postpartum breast cancer development and progression. This study aims to investigate the effect of breastmilk on interactions between breast cancer cells and macrophages in vitro. Methods: Human breastmilk from healthy donors (n = 7) was pooled and incubated with breast cancer (MCF-7 and MDA-MB-231) and macrophage (RAW264.7) cell lines to assess cell proliferation, viability, migration, and expression of key genes associated with epithelial-mesenchymal transition (EMT) and macrophage phenotype. Indirect co-culture studies assessed the effect of breastmilk on interactions between breast cancer cells and macrophages. Results: Breastmilk increased the proliferation and viability of breast cancer cells, reduced EMT markers, and reduced cell migration in MDA-MB-231 cells. Breastmilk decreased mRNA expression of interleukin 1B (IL1B) and interleukin 10 (IL10) in macrophages. Reduced EMT marker expression was observed in breast cancer cells co-cultured with macrophages pre-treated with breastmilk. Macrophages co-cultured with breast cancer cells pre-treated with breastmilk exhibited increased expression of a pro-inflammatory cytokine tumor necrosis factor A (TNFA) and pro-inflammatory nitric oxide synthase 2 (NOS2), and reduced expression of cytokines IL10 and transforming growth factor B1 (TGFB1) which are associated with the alternatively-activated macrophage phenotype. Conclusions: Breastmilk has the potential to promote breast cancer proliferation, however, it can also reduce breast cancer progression through inhibition of breast cancer cell migration and regulation of macrophage polarisation. These findings suggest that breastmilk has potential to shape the tumour microenvironment in postpartum breast cancer.

DDX5-Targeting Fragile X Mental Retardation Protein Regulates the Wnt/β-catenin Signaling Pathway to Promote Epithelial Mesenchymal Transition in Breast Cancer

To investigate the role of fragile X mental retardation protein (FMRP) in promoting cell migration and epithelial-mesenchymal transition (EMT) in breast cancer (BC) and the potential mechanisms involved. The mRNA and protein expressions of FMRP in MCF-10A, a normal human breast epithelial cell line, and four breast cancer cell lines, including MCF-7, BT474, MDA-MB-231, and HCC1937, were analyzed by RT-PCR and Western blot. The expression of FMRP in BC tissues was measured by immunohistochemistry (IHC). FMRP expression in BC and its relationship with clinical prognosis were analyzed using GEO database. Lentiviral infection and siRNA interference were used to construct FMRP overexpression and interference vectors, respectively, and the human breast cancer cell line MCF-7 was subsequently transfected. A Control group, an interference empty vector group (the NC group), a knockdown vector group (the siFMRP group), an overexpression empty vector group (the Lv-NC group), and an overexpression vector group (the Lv-FMRP group) were set up. The migration and invasion abilities of the cells were assessed by scratch assay and Transwell assay. The expression of EMT markers, including E-cadherin, an epithelial marker, N-cadherin, an mesenchymal markers, vimentin, zinc finger E-box binding homeobox 1 (ZEB1), and snail family zinc finger 2 (Slug), in the cells of each group was determined by Western blot. The interaction between FMRP and DEAD-box RNA helicase-5 (DDX5) protein was analyzed by immunocoprecipitation combined with mass spectrometry (IP-MS). The regulatory effect of FMRP on DDX5 protein expression was assessed using the protein synthesis inhibitor cycloheximide (CHX) and proteasome inhibitor MG132. In addition, transfection with siDDX5 vector was conducted to observe whether DDX5 could reverse the effects of FMRP overexpression on cell migration and EMT. The localization and expression of β-catenin were determined by immunofluorescence staining, and the expression of core markers of Wnt/β-catenin signaling pathway was examined by Western blot. FMRP was highly expressed in BC tissues and cells (P<0.05), and overall survival (OS) and recurrence-free survival (RFS) of the FMRP high expression group were significantly lower than those of the FMRP low expression group (P<0.05). The migration ability of MCF-7 cells was weakened after FMRP knockdown, while overexpression of FMRP promoted cell migration (P<0.05). After FMRP knockdown, the expression of E-cadherin was increased, while the expression levels of N-cadherin, vimentin, ZEB1, and Slug were decreased, which inhibited the occurrence of EMT. In contrast, the overexpression of FMRP promoted the EMT process (P<0.05). FMRP interacted with DDX5 protein and promoted DDX5 protein stability by blocking the ubiquitin-proteasome pathway. DDX5 knockdown reversed the effect of FMRP overexpression to promote cell migration and EMT (P<0.05), effectively inhibited β-catenin nuclear translocation, and decreased β-catenin nuclear distribution. Furthermore, it was found that the expression of p-β-catenin, GSK3β and Axin2 protein was increased and the expression of C-myc protein was decreased after DDX5 downregulation (P<0.05). On the other hand, the expression of these proteins was reversed by combined FMRP overexpression (P<0.05). FMRP targets DDX5 and promotes BC cell migration and EMT via the activation of the Wnt/β-catenin signaling pathway.

Tumour stemness and poor clinical outcomes in haemochromatosis patients with hepatocellular carcinoma

AimsPatients with haemochromatosis (HFE) are known to have an increased risk of developing hepatocellular carcinoma (HCC). Available data are conflicting on whether such patients have poorer prognosis, and there is...

UBE2T promotes stage I lung adenocarcinoma progression through PBX1 ubiquitination and PBX1/RORA regulation

BackgroundPost-translational modification pathway of protein ubiquitination is intricately associated with tumorigenesis. We previously reported elevated ubiquitin-conjugating enzyme 2T (UBE2T) as an independent risk factor in stage I lung adenocarcinoma and promoting cellular proliferation. However, its underlying mechanisms needed further investigation.MethodsImmunohistochemistry was used to assess the expression of UBE2T and retinoic acid receptor-related orphan receptor α (RORA) in stage I LUAD. Cell proliferation, migration, and invasion of LUAD cell lines were measured by Cell Counting Kit-8 assay (CCK-8), Colony-forming assay and Transwell assay, respectively. Western blot analysis was performed to determine the expression of epithelial-mesenchymal transition (EMT) markers. A xenograft model was established to evaluate the proliferative capacity of UBE2T and its interaction with RORA in promoting LUAD. Mechanistic insights into the promotion of early-stage LUAD by UBE2T were obtained through luciferase reporter assay, chromatin immunoprecipitation and co-immunoprecipitation.ResultsUBE2T and RORA expression was significantly up- and down-regulated in early-stage LUAD patients which’s proved to be associated with unfavorable outcomes, strengthened cell proliferation, migration, EMT and invasion through its interaction with RORA both in vivo and in vitro. The growth NSCLC xenografts was reduced by down-expression of UBE2T but was suppressed by RORA knockout. Mechanistically, UBE2T mediated the ubiquitination of the intermediate transcription factor PBX1, which played a transcriptional role in downstream regulation of RORA.ConclusionThe oncogenic role of UBE2T and the UBE2T-PBX1-RORA axis in driving malignant progression in Stage I LUAD had been established. UBE2T might be a novel and promising therapeutic target for LUAD treatment.

Plectalibertellenone A suppresses colorectal cancer cell motility and glucose metabolism by targeting TGF-β/Smad and Wnt pathways.

Colorectal cancer (CRC) is the second most common cause of cancer-related death and represents a serious worldwide health problem. CRC metastasis decreases the survival rate of cancer patients, underscoring the need to identify novel anticancer agents and therapeutic targets. Here, we introduce Plectalibertellenone A (B) as a promising agent for the inhibition of CRC cell motility and glucose metabolism and explore its mechanism of action in CRC cells. Plectalibertellenone A suppressed TGF-β gene expression and the activation of the TGF-β/Smad signaling pathway, leading to reverse epithelial to mesenchymal transition (EMT) by modulating the expressions of EMT markers and transcriptional factors such as E-cadherin, N-cadherin, vimentin, Slug, Snail, Twist, and ZEB1/2. Furthermore, disruption of Wnt signaling inhibited CRC motility and glucose metabolism including glycolysis and oxidative phosphorylation, primarily affecting glycolytic enzymes, GLUT1, HK2, PKM2, LDHA, and HIF-1α under hypoxic condition. Therefore, Plectalibertellenone A is a potential drug candidate that can be developed into a promising anticancer treatment to prevent CRC metastasis and inhibit glucose metabolism.

Pregnane X receptor inhibits the transdifferentiation of hepatic stellate cells by down-regulating periostin expression.

Pregnane X receptor (PXR) is a xenobiotic-sensing nuclear receptor that plays a key role in drug metabolism. Recently, PXR was found to attenuate the development of liver cancer by suppressing epithelial-mesenchymal transition (EMT) in liver cancer cells in a mouse model of two-stage chemical carcinogenesis. To elucidate the role of PXR in the EMT of liver cancer cells, we focused on its role in hepatic stellate cells (HSCs), which are components of the tumor microenvironment in hepatocellular carcinoma (HCC). Human HSC-derived LX-2 cells stably expressed destabilization domain (DD)-fused human PXR (hPXR-LX2 cells). Human HCC-derived HepG2 cells were transfected with the EMT marker VIM promoter-regulated reporter plasmid and co-cultured with hPXR-LX2 cells or treated with hPXR-LX2-derived conditioned medium (CM). Co-culture or CM treatment increased reporter activity in HepG2 cells. This induction was attenuated upon PXR activation in hPXR-LX2 cells by treatment with the DD-stabilizing chemical Shield-1 and the human PXR ligand rifampicin. PXR activation in hPXR-LX2 cells exhibited inhibition of TGF-β1-induced transdifferentiation, supported by observations of morphological changes and protein or mRNA levels of the transdifferentiation markers COL1A1 and FN1. PXR activation in hPXR-LX2 cells also attenuated the mRNA levels of the key transdifferentiation factor, POSTN. Treatment of hPXR-LX2 cells with recombinant POSTN restored the PXR-mediated suppression of transdifferentiation. Reporter assays with the POSTN promoter showed that PXR inhibited the NF-κB-mediated transcription of POSTN. Consequently, PXR activation in HSCs is expected to inhibit transdifferentiation by down-regulating POSTN expression, thereby suppressing EMT of liver cancer cells.

Dihydroartemisinin Inhibits Epithelial-Mesenchymal Transition Progression in Medullary Thyroid Carcinoma Through the Hippo Signaling Pathway Regulated by Interleukin-6.

Dihydroartemisinin (DHA), an artemisinin derivative, can influence certain malignancies' inflammatory response and growth. This study used Cell Counting Kit-8 and Transwell assays to show that DHA suppressed the growth, migration, and invasion of medullary thyroid cancer cells. Furthermore, the authors used enzyme-linked immunosorbent assay, Western blotting, and immunofluorescence to confirm the expression of the transcriptional coactivators Yes-associated protein (YAP)/transcriptional coactivator with a PDZ-binding domain (TAZ) downstream of the Hippo pathway and changes in the expression of the epithelial-mesenchymal transition (EMT) process markers E-cadherin and N-cadherin. These results demonstrate that DHA effectively reduced the expression of interleukin (IL)-6 in medullary thyroid carcinoma (MTC) cells and hindered the EMT process by regulating the Hippo pathway. This regulation was achieved by promoting YAP phosphorylation and inhibiting YAP/TAZ protein expression. Additional activation of the Hippo pathway by GA-017 alleviated the inhibitory effect of DHA on IL-6. Hippo pathway activation led to an increase in the expression of E-cadherin, a marker of EMT. In conclusion, DHA was demonstrated to regulate the Hippo pathway by inhibiting IL-6 secretion, leading to the inhibition of EMT in MTC. These findings provide a theoretical foundation for further exploration of the anticancer mechanisms of DHA and offer valuable insights into its potential clinical application as a combinatorial drug.