Hepatocellular Carcinoma Progression Research Articles

Aberrant activation of a miR-101–UBE2D1 axis contributes to the advanced progression and chemotherapy sensitivity in human hepatocellular carcinoma

AbstractChemotherapeutic drugs, such as cisplatin (cis-dichlorodiamineplatinum [II], cDDP) and 5-fluorouracil (5Fu), are widely used in transarterial chemoembolization (TACE), which is a standard therapy for patients with hepatocellular carcinoma (HCC). Chemoresistance is a major cause of TACE treatment failure in HCC patients. Our previous studies have identified the expression levels of miR-101 responsive genes, such as EED, EZH2, STMN1 and JUNB, exhibit significant correlation with the occurrence and progression of HCC, while the role of miR-101 responsive gene signatures in the chemoresistance of HCC treatment remains unclear. In this study, we identified ubiquitin-coupled enzyme E2D1 (UBE2D1) as a crucial regulatory factor in the chemoresistance of HCC, which is a direct target of miR-101 and exhibits significant correlation with miR-101-responsive gene signatures. The bioinformatics analysis showed the expression of UBE2D1 was significantly increased in HCC tissues and was closely correlated with the poor prognosis. In addition, we analyzed the role of miR-101/UBE2D1 axis in regulating chemo-sensitive of HCC cells. Our results showed that miR-101 increases the DNA damage and apoptosis of HCC cells by inhibiting the expression of UBE2D1, which in turn increases the sensitivity of HCC cells to cDDP and 5Fu both in vitro and in vivo. Therefore, simultaneous assessment of miR-101 and UBE2D1 expression levels might provide an effective approach in preselecting HCC patients with survival benefit from TACE treatment. Moreover, further elucidation of the underlying molecular mechanisms of the miR-101/UBE2D1 axis could provide novel insight for targeted therapy of HCC.

Single-cell and spatial transcriptomics reveal apelin/APJ pathway's role in microvessel formation and tumour progression in hepatocellular carcinoma.

The apelin receptor (APJ) is a key player in tumour angiogenesis, but its role in hepatocellular carcinoma (HCC) remains unclear. This study aims to elucidate the function of the apelin/APJ pathway in HCC using a multi-omics approach and identify potential therapeutic biomarkers. Differentially expressed genes related to the apelin/APJ axis were identified from bulk transcriptomics to reveal HCC-associated disparities. Single-cell and spatial transcriptomics were used to localize and analyse the function of these genes. Machine learning models were constructed to predict outcomes based on apelin/APJ expression, and experimental validation was conducted to explore the pathway's impact on HCC angiogenesis. Single cell analysis revealed an overexpression of APJ/Aplin in the endothelium. The stemness of endothelial cell (EC) with high apelin/APJ was enhanced, as well as the expression of TGFb, oxidative stresses and PI3K/AKT pathway genes. Spatial transcriptomics confirmed that EC populations with high APJ scores were enriched within the tumour. Machine learning models showed high prognostic accuracy. High APJ expression was linked to worse outcomes (p = 0.001), and AUC values were high (1 year, 3 year, 5 year) (0.95, 0.97, 0.98). Immune suppression and non-responsiveness of immune therapy were also seen in high-risk groups. The experimental validation showed that silencing apelin reduced angiogenesis (p < 0.05), endothelial proliferation, decreased expression of ANG2, KLF2, VEGFA and lower ERK1/2 phosphorylation. Apelin may serve as a potential therapeutic target in HCC, given its role in promoting tumour angiogenesis and poor patient outcomes.

Exploring SSR1 as a novel diagnostic and prognostic biomarker in hepatocellular carcinoma, and its relationship with immune infiltration.

Although signal sequence receptor subunit 1 (SSR1) has undergone thorough examination in different cancer types, its importance in hepatocellular carcinoma (HCC) remains largely uncharted and warrants further investigation. The aim of this study is to explore the role of SSR1 in HCC progression and to decipher its underlying molecular mechanisms. We employed the ONCOMINE, Tumor IMmune Estimation Resource (TIMER), and The Cancer Genome Atlas databases to assess SSR1 expression levels within tumor tissues. Logistic and Cox regression analyses, Kaplan-Meier survival plots, nomograms, and forest plots were employed to establish correlation between SSR1 and prognosis. Receiver operating characteristic (ROC) curves demonstrated diagnostic utility of SSR1. Additionally, Gene Ontology (GO) and gene set enrichment analysis (GSEA) analyses were conducted to uncover relevant molecular pathways. TIMER was instrumental in elucidating the connection between SSR1 and immune cell infiltration. Actions of SSR1 in HCC proliferation and migration were investigated through quantitative real-time polymerase chain reaction, Cell Counting Kit-8, 5-ethynyl-2'-deoxyuridine cell proliferation assays, and Transwell migration and wound healing experiments. Elevated SSR1 levels were found to be correlated with clinical parameters such as age and pathologic stage, thereby predicting a reduced overall survival (OS) rate in HCC patients. Multivariate survival analysis underscored SSR1 as an independent prognostic marker for OS. A nomogram underscored SSR1's effectiveness as a predictive tool for HCC outcomes, while ROC analysis indicated its high diagnostic accuracy. GO and GSEA analyses suggested that elevated SSR1 expression may be associated with epithelial-mesenchymal transition (EMT) pathway. SSR1 exhibited a negative correlation with cytotoxic cells and a positive correlation with Th2 cells. Our in vitro experiments provided evidence that heightened SSR1 levels may impact HCC proliferation and migration through EMT pathway. SSR1 surfaces as a new diagnostic and potentially prognostic biomarker, showing an association with immune cell infiltration and cell proliferation in HCC.

Aqueous Extract of Rhubarb Promotes Hepatotoxicity via Facilitating PKM2-Mediated Aerobic Glycolysis in a Rat Model of Diethylnitrosamine-Induced Liver Cancer.

To identify the polar parts in Rhubarb that cause hepatotoxicity and explore the underlying mechanisms. The rat model of liver cancer was established by gavage of diethylnitrosamine (DEN; 0.002 g/rat) for 14weeks. Starting from the 11th week, Rhubarb granule (4 g/kg), aqueous, ethyl acetate and n-butanol extract of Rhubarb or Rhein equivalent to a dose of 4 g/kg Rhubarb granule were administered intragastrically for 4 consecutive weeks. Liver tissues from rats treated with DEN and Rhubarb granules were used for non-targeted metabolomics analysis. The correlation between pyruvate kinase isozyme type M2 (PKM2) expression level and the progress and prognosis of hepatocellular carcinoma (HCC) was evaluated through bioinformatics analysis based on TCGA database. Liver tissues and blood samples from rats treated with DEN and aqueous, ethyl acetate and n-butanol extract of Rhubarb were used for the screening of hepatotoxic polar parts of Rhubarb. The liver injuries were evaluated by the changes in pathology, liver function, and the expression levels of proliferating cell nuclear antigen (PCNA) and transforming growth factor beta1 (TGF-β1). The mechanism studies focus on PKM2 expression, and the metabolic reprogramming via detecting the activities of lactate dehydrogenase A (LDHA) and isocitrate dehydrogenase (ICDH). Furthermore, molecular docking analysis was performed to validate the target interaction between Rhein and PKM2, and the hepatotoxicity of Rhein was evaluated by testing liver function in the DEN-induced liver cancer model. The non-targeted metabolomics analysis revealed that Rhubarb promoted aerobic glycolysis in the rat model of DEN-induced liver cancer. And bioinformatics analysis revealed that high PKM2 expression was closely related to the progression and poor prognosis of HCC. In vivo studies indicated that the aqueous extract of Rhubarb, but not ethyl acetate and n-butanol extract, promoted the liver injuries induced by DEN. The mechanism study showed that the aqueous extract of Rhubarb increased the expression of PKM2 and promoted aerobic glycolysis. Moreover, Rhein had a strong binding affinity for PKM2 and aggravated liver injury in the DEN-induced liver cancer model. Aqueous extract of Rhubarb promoted hepatotoxicity via facilitating PKM2-mediated aerobic glycolysis in the rat model of DEN-induced liver cancer.

Glycerophospholipid-driven lipid metabolic reprogramming as a common key mechanism in the progression of human primary hepatocellular carcinoma and cholangiocarcinoma

Metabolic reprogramming, a key mechanism regulating the growth and recurrence of hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA), still lacks effective clinical strategies for its integration into the precise screening of primary liver cancer. This study utilized ultra-high-performance liquid chromatography with quadrupole time-of-flight mass spectrometry to conduct a comprehensive, non-targeted metabolomics analysis, revealing significant upregulation of lipid metabolites such as phosphatidylcholine and lysophosphatidylcholine in patients with HCC and CCA, particularly within the glycerophospholipid metabolic pathway. Hematoxylin and eosin and immunohistochemical staining demonstrated marked upregulation of phospholipase A2 in tumor tissues, further emphasizing the potential of lipid metabolism as a therapeutic target and its important part in the course of cancer. This work provides a new viewpoint for addressing the clinical challenges associated with HCC and CCA, laying the groundwork for the broad application of early diagnosis and personalized treatment strategies, and ultimately aiming to provide tailored and precise therapeutic options for patients.

Carbenoxolone upregulates TRAIL\\TRAILR2 expression and enhances the anti-neoplastic effect of doxorubicin in experimentally induced hepatocellular carcinoma in rats

AimsThis study investigates the in vivo anticancer activity of carbenoxolone (CBX) and its role in fighting hepatocellular carcinoma (HCC) progression and alleviating resistance against doxorubicin (DOX). Moreover, the molecular mechanism of action of CBX is explored. MethodsHCC was induced in Sprague Dawley rats via biweekly administration of thioacetamide (TAA) (200 mg/kg) intraperitoneally (i.p.) for 16 weeks after administering a single dose of diethylnitrosamine (DEN) (200 mg/kg, i.p.). A prophylactic model was established by treating rats with i.p. CBX (20 mg/kg/day) for 4 weeks starting on week 13 post-TAA injection. A therapeutic model was established by treating rats with CBX, DOX, or their combination for 7 weeks following 16 weeks of TAA administration. Serum Alpha-fetoprotein (AFP) and biochemical markers of hepatic functions were assessed. Histopathological examinations of hepatic tissues were performed. Immunohistochemical and qRT-PCR analyses were applied to assess the differential expressions of TRAIL/TRAILR2, Bcl-2, TGF-β1, and caspases 3,8, and 9. ResultsCBX markedly improved hepatic functions, reduced serum AFP levels, and alleviated TAA-induced hepatic histopathological alterations. CBX triggered apoptosis as evident by upregulating apoptotic markers: TRAIL/TRAILR2, caspases 3,8, and 9, and downregulating the antiapoptotic protein Bcl-2. CBX downregulated TGF-β1. Interestingly, CBX/DOX combination mitigated hepatic damage and induced apoptosis in a way that surpassed DOX-only treatment. ConclusionThe current study proposes that CBX is a promising anti-tumor compound, which can work effectively under prophylactic and therapeutic modes. Interestingly, CBX enhanced the anti-tumor effect of DOX. CBX exerted these effects via, in part, stimulating TRAIL-induced apoptosis along with attenuating fibrosis.

Long-Term Exposure of Fresh and Aged Nano Zinc Oxide Promotes Hepatocellular Carcinoma Malignancy by Up-Regulating Claudin-2.

Tumor development and progression is a long and complex process influenced by a combination of intrinsic (eg, gene mutation) and extrinsic (eg, environmental pollution) factors. As a detoxification organ, the liver plays an important role in human exposure and response to various environmental pollutants including nanomaterials (NMs). Hepatocellular carcinoma (HCC) is one of the most common malignant tumors and remains a serious threat to human health. Whether NMs promote liver cancer progression remains elusive and assessing long-term exposure to subtoxic doses of nanoparticles (NPs) remains a challenge. In this study, we focused on the promotional effects of nano zinc oxide (nZnO) on the malignant progression of human HCC cells HepG2, especially aged nZnO that has undergone physicochemical transformation. In in vitro experiments, we performed colony forming efficiency, soft agar colony formation, and cell migration/invasion assays on HepG2 cells that had been exposed to a low dose of nZnO (1.5μg/mL) for 3 or 4 months. In in vivo experiments, we subcutaneously inoculated HepG2 cells that had undergone long-term exposure to nZnO for 4 months into BALB/c athymic nude mice and observed tumor formation. ZnCl2 was administered to determine the role of zinc ions. Chronic low-dose exposure to nZnO significantly intensified the malignant progression of HCC cells, whereas aged nZnO may exacerbate the severity of malignant progression. Furthermore, through transcriptome sequencing analysis and in vitro cellular rescue experiments, we demonstrated that the mechanism of nZnO-induced malignant progression of HCC could be linked to the activation of Claudin-2 (CLDN2), one of the components of cellular tight junctions, and the dysregulation of its downstream signaling pathways. Long-term exposure of fresh and aged nZnO promotes hepatocellular carcinoma malignancy by up-regulating CLDN2. The implications of this work can be profound for cancer patients, as the use of various nanoproducts and unintentional exposure to environmentally transformed NMs may unknowingly hasten the progression of their cancers.

TRPM8 overexpression suppresses hepatocellular carcinoma progression and improves survival by modulating the RTP3/STAT3 pathway.

Hepatocellular carcinoma (HCC) is a malignant tumour associated with high morbidity and mortality rates worldwide. Recently, TRPM8 was reported to play an important role in tumour progression. However, the precise role of TRPM8 in HCC remains unclear. In this study, we explored the expression levels, molecular functions and underlying mechanisms of TRPM8 in HCC. Tissue samples were used to analyse the expression of TRPM8 to assess its diagnostic value for prognosis. Cell Counting Kit-8, EdU and colony formation assays were performed to evaluate the effects of TRPM8 on cell proliferation, whereas the Transwell assay was used to assess cell migration and invasion. The role of TRPM8 invivo was evaluated using a mouse subcutaneous xenograft tumour model. We performed PPI network analyses to understand the possible mechanisms of TRPM8 action. TRPM8 expression was decreased in HCC tissues and was correlated with histological grade and poor patient prognosis. Functionally, TRPM8 repressed the proliferation and metastasis of HCC cells both invitro and invivo by modulating the RTP3/STAT3 signalling pathway. Our findings underscore the critical role of the TRPM8-RTP3-STAT3 axis in maintaining the malignant progression of HCC. Moreover, our study demonstrates that AD80 is involved in anti-tumour processes by upregulating the expression of TRPM8.

Opposing regulation of the STING pathway in hepatic stellate cells by NBR1 and p62 determines the progression of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) emerges from chronic inflammation, to which activation of hepatic stellate cells (HSCs) contributes by shaping a pro-tumorigenic microenvironment. Key to this process is p62, whose inactivation leads to enhanced hepatocarcinogenesis. Here, we show that p62 activates the interferon (IFN) cascade by promoting STING ubiquitination by tripartite motif protein 32 (TRIM32) in HSCs. p62, binding neighbor of BRCA1 gene 1 (NBR1) and STING, triggers the IFN cascade by displacing NBR1, which normally prevents the interaction of TRIM32 with STING and its subsequent activation. Furthermore, NBR1 also antagonizes STING by promoting its trafficking to the endosome-lysosomal compartment for degradation independent of autophagy. Of functional relevance, NBR1 deletion completely reverts the tumor-promoting function of p62-deficient HSCs by rescuing the inhibited STING-IFN pathway, thus enhancing anti-tumor responses mediated by CD8+ Tcells. Therefore, NBR1 emerges as a synthetic vulnerability of p62 deficiency in HSCs by promoting the STING/IFN pathway, which boosts anti-tumor CD8+ Tcell responses to restrain HCC progression.

Preoperative Serum Glycan Levels Reflect Progression of Patients With Hepatocellular Carcinoma.

Abnormal glycosylation is associated with tumors. The clinical value of serum glycans in assessing progression of hepatocellular carcinoma (HCC) patients remains a challenge. A study dynamically comparing levels of fifteen lectin-specific glycans between preoperative and postoperative serum of 65 HCC patients was conducted via lectin biochip technology. Multivariable logistic regression analysis was used to address associations between serum glycan levels and clinicopathological characteristics. Kaplan-Meier analysis was used to evaluate the impacts of serum glycan levels on overall survival (OS) and progression-free survival (PFS) of the HCC patients. HCC patients presented significantly higher levels of the lectin-specific glycans in preoperative serum than disease-free individuals (p < 0.001 - p = 0.029), except ConA. The glycans in preoperative sera were significantly related to tumor size, pTNM, metastasis, BCLC stage, portal hypertension (PHT), and platelet count (PLT), respectively (p < 0.05). Multivariate logistic analyses indicated that tumor size and pTNM independently impact on glycan-specific lectins either LTL, UEA-I, VVL, NPL, WGA, PNA, MAL-I, SNA, or PHA-L (p = 0.003 - p = 0.044); BCLC stage and PLT were independent factors influencing the serum glycans recognizable DSA (p = 0.024) and SNA (p = 0.050), respectively. Surgical excision of tumor mass significantly reduced glycan levels in sera. Tumor differentiation, albumin, and ABO type significantly revealed independent influence on glycan-specific lectins, such as RCA-I (p = 0.024), VVL (p = 0.024), and Con A (p = 0.026) in the postoperative serum. HCC patients with high levels of VVL-binding glycans significantly benefited from a longer OS time (p = 0.016, HR: 0.460, 95% CI: 0.237-0.892) and a better PFS time (p = 0.004; HR: 0.435, 95% CI: 0.237-0.799), respectively. Serum glycans could reflect surgical outcomes in at-risk patients and become valuable biomarkers in evaluating the progression of HCC patients.

Tim-1-mediated extracellular matrix promotes the development of hepatocellular carcinoma.

Tim-1 (T-cell immunoglobulin and mucin domain 1), also known as Kim-1 (kidney injury molecule 1) or hepatitis A virus cellular receptor 1 (HAVCR1), is a transmembrane protein expressed on various immune and epithelial cells. It plays a role in modulating inflammatory and immune responses. In this study, we find that Tim-1 is overexpressed in hepatocellular carcinoma (HCC) samples and that its expression is significantly correlated with postoperative survival. Bulk RNA sequencing reveals a general upregulation of extracellular matrix-related genes in HCC tissues with Tim-1 overexpression. The results of the cell and in vivo experiments reveal that Tim-1 in HCC not only affects biological processes such as the proliferation, migration, and invasion of HCC cells but also broadly promotes extracellular matrix processes by influencing cytokine secretion. Further studies demonstrate that Tim-1 mediates the activation of hepatic stellate cells and upregulates Th1 and Th2 cytokines, thereby promoting HCC progression. Thus, Tim-1 may represent a novel target for future interventions in HCC and liver fibrosis.

Anti-viral and Apoptotic Induction of m-TOR Inhibitor Drugs against Hepatitis C Virus Activity and Hepatocellular Carcinoma Cell Line: In vitro and in silico.

This study investigated the potential of m-TOR inhibitor drugs (sirolimus, everolimus, and tacrolimus) in combating both hepatocellular carcinoma (HCC) and hepatitis C virus (HCV) replication. After treating HepG2 and PBMCs with the mammalian target of Rapamycin (m-TOR) inhibitors drugs; sirolimus, everolimus, and tacrolimus at different concentrations (1, 5, and 10 µM/µl), cell proliferation was assessed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Antioxidant activities (total antioxidant, glutathione S-transferase, and glutathione reductase), Fas-ligand level, tumor necrosis factor-α (TNF-α) level, caspase-3, -8, and -9 activities, and cell cycle analysis were measured. quantitative Real-time PCR, colony forming assay, molecular docking studies after infection of PBMCs with 1 ml (1.5 × 106 HCV) serum then incubated with m-TOR inhibitor drugs at their respective IC50 concentrations. In HepG2 cells, treatment with these inhibitors resulted in suppressed cell viability, increased dead cell accumulation, and enhanced apoptotic signaling through elevated Fas-ligand and caspase activities. Additionally, cell cycle analysis revealed arrest in G0/G1 and G2/M phases, further hindering HCC progression. Furthermore, m-TOR inhibitor drugs significantly reduced HCV viral load and colony formation in infected PBMCs. This antiviral effect was accompanied by decreased TNF-α activity, suggesting potential modulation of the inflammatory response associated with HCV infection. Molecular docking studies provided theoretical support for these findings, with Sovaldi demonstrating the highest binding affinity towards key HCV targets compared to other m-TOR inhibitors. This suggests its potential as a potent HCV inhibitor, while also highlighting the potential of exploring m-TOR inhibitors for future HCV treatment development. Overall, this study provides encouraging evidence for the potential of m-TOR inhibitor drugs as promising therapeutic agents for both HCC and HCV, warranting further investigation and optimization for clinical applications.

ARL8B promotes hepatocellular carcinoma progression and inhibits antitumor activity of lenvatinib via MAPK/ERK signaling by interacting with RAB2A

Tumor recurrence and metastasis are important factors affecting postoperative survival in hepatocellular carcinoma (HCC) patients. ADP Ribosylation factor-like GTPase 8B (ARL8B) plays a crucial role in many biological processes, including lysosomal function, immune response, and cellular communication, all of which are related to the occurrence and development of tumors. However, its role in HCC remains unclear. Herein, we revealed that ARL8B is consistently elevated in HCC tissues compared to normal liver tissues, suggesting an unfavorable outcome in HCC patients. Increased ARL8B levels promoted the malignant phenotype of HCC in vitro and in vivo. Notably, ARL8B also induced epithelial-to-mesenchymal transition (EMT) in HCC cells. Mechanistically, the results of bioinformatics analysis combined with mass spectrometry revealed the potential downstream target molecule RAB2A of ARL8B. ARL8B directly interacted with RAB2A and increased the levels of GTP-bound RAB2A, thereby contributing to the activation of the extracellular signal-regulated kinase (ERK) signaling pathway. Interestingly, knockout of ARL8B in Hep3B cells enhanced the antitumor activity of lenvatinib in vitro and in vivo. Furthermore, AAV-shARL8B enhanced the inhibition of HCC growth through lenvatinib, providing new insights into its mechanism of action in lenvatinib-insensitive patients. In conclusion, ARL8B promotes the malignant phenotype of HCC and EMT via RAB2A mediated activation of the MAPK/ERK signaling pathway and is expected to be a valuable prognostic indicator and therapeutic target for HCC patients.

Gene Expression Aberrations in Alcohol-Associated Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is the most prevalent primary liver cancer, ranking as the sixth most common cancer worldwide and the fourth leading cause of cancer-related deaths. Most HCC cases originate from cirrhotic livers, typically due to chronic liver diseases, such as hepatitis B (HBV) and hepatitis C (HCV) infections, and alcoholism. HCC cells often harbor numerous somatic mutations that are implicated in HCC development, but epigenetic factors, such as miRNA interference, can also affect HCC initiation and progress. miRNA-221 has been explored as a factor affecting HCC development in HCC of viral etiology, but little is known about its effects on gene expression in alcohol-associated HCC. This study aimed to explore potentially similar gene expression aberrations underlying viral and alcohol-induced HCC. We analyzed available transcriptome data from non-tumor hepatocytes and viral-induced HCC tissues. The most notable differences in gene expression associated with miRNA-221 between non-tumor hepatocytes and viral-induced HCC involved NTF-3 and MYBL1 genes. To assess these data in alcohol-induced HCC, we examined 111 tissue samples: tumor tissue and cirrhotic tissue samples from 37 HCC patients and 37 samples from non-tumor liver tissue using RT-Q PCR. We found no significant difference in NTF-3 expression, but MYBL1 expression was significantly lower in HCC tissue compared to non-tumor hepatocytes and cirrhotic tissue. Our findings highlight the importance of the MYBL1 gene in HCC development and emphasize the need for diverse approaches in evaluating tumor mechanisms.

Ficolin-3 induces apoptosis and suppresses malignant property of hepatocellular carcinoma cells via the complement pathway

AimsFicolin 3 (FCN3) has the highest complement-activating capacity through the lectin pathway and is synthesized mainly in the liver and lung. Yet, its potential molecular mechanism in hepatocarcinogenesis is not fully understood. Materials and methodsThe expression of FCN3 in hepatocellular carcinoma (HCC) tumor and non-tumor tissues was analyzed by RT-qPCR, Western blotting and immunofluorescence staining assays. Lentivector-mediated ectopic overexpression was performed to explore the role of FCN3 in vitro and in vivo. Whether FCN3 inhibited HCC cell growth and survival via complement pathway was determined with immunocytochemical staining for C3b, membrane attack complex (MAC) formation and complement killing assay using recombinant FCN3 (rFCN3) in combination with human serum with or without heat inactivation, and with C6 blocking antibody. Key findingsThe transcript and protein of FCN3 were found to be remarkably down-regulated in HCC tumor tissues. FCN3 expression was found to be associated with better survival of HCC patients. Restoration of FCN3 expression significantly inhibited proliferation, migration and anchorage independent growth of HCC cell lines, and xenograft tumor growth. FCN3 expression induced apoptosis of HCC cells. C3 and MAC formation was stimulated by FCN3 overexpression or rFCN3 treatment. rFCN3 enhanced human serum-induced complement activation and cell death. C6 blocking antibody significantly attenuated complement-mediated cell death and restored the growth of FCN3-overexpressing HCC cells. SignificanceFCN3 has a malignant suppressor role in HCC cells. Our study provides new insights into the molecular mechanisms that drive HCC progression and potential therapeutic targets for treating HCC.

Nuclear translocation of plasma membrane protein ADCY7 potentiates T cell-mediated antitumour immunity in HCC

BackgroundThe potency of T cell-mediated responses is a determinant of immunotherapy effectiveness in treating malignancies; however, the clinical efficacy of T-cell therapies has been limited in hepatocellular carcinoma (HCC) owing...

Matrix stiffness-related extracellular matrix signatures and the DYNLL1 protein promote hepatocellular carcinoma progression through the Wnt/β-catenin pathway

BackgroundIn hepatocellular carcinoma (HCC) treatment, first-line targeted therapy in combination with immune checkpoint inhibitors (ICIs) has improved patient prognosis, but the 5-year survival rate is far from satisfactory. Studies have shown that the extracellular matrix (ECM) is an essential part of the tumour microenvironment (TME) and participates in the progression of malignant tumours. ECM remodelling can enhance matrix stiffness in cirrhosis patients, induce an immunosuppressive microenvironment network, and affect the efficacy of targeted therapies and ICIs for treating HCC. However, the exact mechanism is still unclear.MethodsWe downloaded data from public databases, selected differentially expressed ECM proteins associated with matrix stiffness, constructed and validated a prognostic model of HCC using Lasso Cox regression, and investigated the roles and mechanism of one of the ECM proteins, dynein light chain LC8-type 1 (DYNLL1), in HCC proliferation, migration, and apoptosis via in vitro experiments.ResultsIn this study, the risk score of the matrix stiffness-related ECM protein model effectively predicted the prognosis of HCC patients. The high- and low-risk subgroups of the model also showed differences in immune cells, immune functions, and drug sensitivity. DYNLL1 promoted HCC cell progression and migration and inhibited HCC cell apoptosis through the Wnt/β-catenin pathway in vitro.ConclusionThe expression of matrix stiffness-related ECM proteins could be an independent predictor of HCC prognosis. DYNLL1, an oncogenic gene in HCC, has the potential to be a new target for HCC treatment.

METTL3-mediated m6A modification of ZNF384 promotes hepatocellular carcinoma progression by transcriptionally activating ACSM1.

Hepatocellular carcinoma (HCC) is a lethal disease with a high mortality rate, and its development is influenced by various molecular mechanisms. Zinc finger protein 384 (ZNF384) has been reported to be involved in the progression of several cancers; however, its role in HCC remains elusive. mRNA expression levels were analyzed by quantitative real-time polymerase chain reaction, while western blotting and immunohistochemistry were performed to validate protein expression. Cell proliferation, apoptosis, and metabolic activities were examined using clonogenicity, flow cytometry, and specific assay kits. A xenograft mouse model was employed to assess the impact of acyl-CoA synthetase medium-chain family member 1 (ACSM1) depletion on HCC cell malignancy in vivo. Chromatin immunoprecipitation assay and dual-luciferase reporter assay were conducted to explore the association between ZNF384 and ACSM1. We found that ACSM1 and ZNF384 were significantly upregulated in HCC tissues and cells when compared with normal liver tissues and human liver immortalized cells. Knockdown of ACSM1 inhibited HCC cell proliferation and glucose metabolism and induced cell apoptosis. Furthermore, ACSM1 depletion suppressed the malignant progression of HCC cells in vivo. Our data indicated that ZNF384 transcriptionally activated ACSM1 in HCC cells. Overexpression of ACSM1 reversed the inhibitory effect of ZNF384 depletion on HCC cell malignancy. Further, methyltransferase-like 3 (METTL3) stabilized ZNF384 mRNA through m6A methylation. METTL3-mediated m6A modification of ZNF384 contributed to the progression of HCC by transcriptionally activating ACSM1. This finding suggests potential therapeutic targets for this devastating disease.

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.

NADPH oxidase 4 deficiency promotes hepatocellular carcinoma arising from hepatic fibrosis by inducing M2-macrophages in the tumor microenvironment

Hepatocellular carcinoma (HCC) often arises in the cirrhotic livers, highlighting the intricate link between hepatic fibrosis and carcinogenesis. Reactive oxygen species produced by NADPH oxidase 4 (NOX4) contribute to liver injury leading to hepatic fibrosis. Paradoxically, NOX4 is known to inhibit HCC progression. This study aims to elucidate the role of NOX4 in hepatocarcinogenesis in the background of hepatic fibrosis. We established the mouse model of HCC arising from the fibrotic liver by administering diethylnitrosamine and carbon tetrachloride to wild-type (WT) or NOX4−/− mice. Hepatic fibrogenesis, tumorigenesis, and macrophage polarization were assessed by immunohistochemistry, PCR, and flow cytometry using in vivo and in vitro models. In NOX4−/− mice, hepatic fibrosis was attenuated, while the number of tumors and the proliferation of HCC cells were increased compared to WT mice. Notably, a significant increase in M2-polarized macrophages was observed in NOX4−/− mice through immunohistochemistry and PCR analysis. Subsequent experiments demonstrated that NOX4-silenced HCC cells promote macrophage polarization toward M2. In addition to attenuating hepatic fibrogenesis, NOX4 deficiency triggers macrophage polarization towards the M2 phenotype in the fibrotic liver, thereby promoting hepatocellular carcinogenesis. These findings provide novel insights into the mechanism of NOX4-mediated tumor suppression in HCC arising from fibrotic livers.