Hepatocellular Carcinoma Migration Research Articles

GCNT3 Promotes Hepatocellular Carcinoma Progression and EMT by Activating the PI3K/AKT Pathway.

Primary liver cancer, specifically hepatocellular carcinoma (HCC), is a major global health concern. GCNT3 has been identified as an oncogene in various human malignancies. This investigation aimed to discover the GCNT3 function in HCC. The present study employed integrated bioinformatics analyses to assess the expression pattern, prognostic implications, and putative function of GCNT3 in HCC. Transwell flow cytometry, CCK-8, and wound healing assays were performed to examine HCC cell growth, cell cycle, apoptosis, invasion, and migration. In addition, the epithelial-mesenchymal transition (EMT) markers and PI3K/AKT mechanism markers were examined via western blot analysis to elucidate the underlying mechanisms. In HCC, GCNT3 was significantly overexpressed, which was connected with enhanced tumor aggressiveness and an unfavorable prognosis of individuals. In vitro experiments demonstrated that elevated levels of GCNT3 promoted cell growth, migration, cell cycle development, and invasion, in addition to EMT, while suppressing apoptosis. Conversely, knockdown of GCNT3 exerted the opposite effects. GCNT3 overexpression increased PI3K/AKT phosphorylation in HCC cells, and LY294002 counteracted the impacts of upregulated GCNT3 on cell cycle, migration, invasion, proliferation, and EMT in HCC. The investigation showed that GCNT3 may enhance HCC progression and EMT by stimulating PI3K/AKT mechanism.

Hsa_circ_0079875 functions as a competitive endogenous RNA to promote hepatocellular carcinoma progression

ABSTRACT Circular RNA (circRNA) can influence the development of hepatocellular carcinoma (HCC) as a competitive endogenous RNA (ceRNA). However, there are still many circRNAs whose functions are unknown. Our research explores the role of a novel circRNA, hsa_circ_0079875, in HCC. The expression of hsa_circ_0079875 in HCC was verified by next-generation sequencing, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), and fluorescence in situ hybridization (FISH). The distribution of hsa_circ_0079875 in HCC cells was investigated by RNA subcellular isolation and FISH assays. The functional effects on HCC proliferation, invasion, migration, cell cycle, and apoptosis were verified by overexpression and knockdown of hsa_circ_0079875. Moreover, xenograft mouse models and immunohistochemistry experiments were used to assess the function of hsa_circ_0079875 in vivo. Hsa_circ_0079875 was up-regulated in HCC tissues and mainly distributed in the cytoplasm. Higher hsa_circ_0079875 leads to larger tumor tissue, more microvascular invasion(MVI) and higher AFP levels, which in turn leads to a poor prognosis. Overexpression of hsa_circ_0079875 can promote the proliferation, migration, and invasion of HCC cells and inhibit apoptosis in vitro and in vivo. Knocking down hsa_circ_0079875 has the opposite effect. Sequencing and biological information predicted the target miRNA and mRNA of hsa_circ_0079875. Further bioinformatics and clinical correlation analysis revealed that hsa_circ_0079875 promote the malignant biological behaviors of HCC through hsa_circ_0079875/miR-519d-59/NRAS ceRNA net. Therefore, hsa_circ_0079875 can be a potential prognostic marker and therapeutic target for HCC.

TRIM47-CDO1 axis dictates hepatocellular carcinoma progression by modulating ferroptotic cell death through the ubiquitin‒proteasome system

Hepatocellular carcinoma (HCC) is the predominant form of liver cancer, characterized by high morbidity and mortality rates, as well as unfavorable treatment outcomes. Tripartite motif-containing protein 47 (TRIM47) has been implicated in various diseases including tumor progression with the activity of E3 ubiquitin ligase. However, the precise regulatory mechanisms underlying the involvement of TRIM47 in HCC remain largely unexplored. Here, we provide evidence that TRIM47 exhibits heightened expression in tumor tissues, and its expression is in intimate association with clinical staging and patient prognosis. TRIM47 promotes HCC proliferation, migration, and invasion as an oncogene by in vitro gain- and loss-of-function experiments. TRIM47 knockdown results in HCC ferroptosis induction, primarily through CDO1 involvement to regulate GSH synthesis. Subsequent experiments confirm the interaction between TRIM47 and CDO1 dependent on B30.2 domain, wherein TRIM47 facilitates K48-linked ubiquitination, leading to a decrease in CDO1 protein abundance in HCC. Furthermore, CDO1 is able to counteract the promotional effect of TRIM47 on HCC biological functions. Overall, our research provides novel insight into the mechanism of TRIM47 in CDO1-mediated ferroptosis in HCC cells, highlighting its value as a potential target candidate for HCC therapeutic approaches.

SUMOylation of annexin A6 retards cell migration and tumor growth by suppressing RHOU/AKT1–involved EMT in hepatocellular carcinoma

BackgroundThe protein annexin A6 (AnxA6) is involved in numerous membrane-related biological processes including cell migration and invasion by interacting with other proteins. The dysfunction of AnxA6, including protein expression abundance change and imbalance of post-translational modification, is tightly related to multiple cancers. Herein we focus on the biological function of AnxA6 SUMOylation in hepatocellular carcinoma (HCC) progression.MethodsThe modification sites of AnxA6 SUMOylation were identified by LC-MS/MS and amino acid site mutation. AnxA6 expression was assessed by immunohistochemistry and immunofluorescence. HCC cells were induced into the epithelial-mesenchymal transition (EMT)-featured cells by 100 ng/mL 12-O-tetradecanoylphorbol-13-acetate exposure. The ability of cell migration was evaluated under AnxA6 overexpression by transwell assay. The SUMO1 modified AnxA6 proteins were enriched from total cellular proteins by immunoprecipitation with anti-SUMO1 antibody, then the SUMOylated AnxA6 was detected by Western blot using anti-AnxA6 antibody. The nude mouse xenograft and orthotopic hepatoma models were established to determine HCC growth and tumorigenicity in vivo. The HCC patient’s overall survival versus AnxA6 expression level was evaluated by the Kaplan–Meier method.ResultsLys579 is a major SUMO1 modification site of AnxA6 in HCC cells, and SUMOylation protects AnxA6 from degradation via the ubiquitin-proteasome pathway. Compared to the wild-type AnxA6, its SUMO site mutant AnxA6K579R leads to disassociation of the binding of AnxA6 with RHOU, subsequently RHOU-mediated p-AKT1ser473 is upregulated to facilitate cell migration and EMT progression in HCC. Moreover, the SENP1 deSUMOylates AnxA6, and AnxA6 expression is negatively correlated with SENP1 protein expression level in HCC tissues, and a high gene expression ratio of ANXA6/SENP1 indicates a poor overall survival of patients.ConclusionsAnxA6 deSUMOylation contributes to HCC progression and EMT phenotype, and the combination of AnxA6 and SENP1 is a better tumor biomarker for diagnosis of HCC grade malignancy and prognosis.

Abstract 6982: The roles of NDRG1 in HCC progression and its clinical significance

Abstract Background: Hepatocellular carcinoma (HCC) is prevalent globally and the third leading cause of cancer-related mortality. N-myc downstream regulated 1 (NDRG1) was found to be tumor promoting in some cancers while suppressive in others. The expression of NDRG1 is induced by stress signals such as DNA damage and hypoxia. In HCC, it has been found to promote proliferation and metastasis. Contradictory results, however, were also suggested on the functions of NDRG1. Methods: Our own HCC cohort and multiple publicly available databases were examined for NDRG1 expression and clinicopathologic correlation. NDRG1 expression under hypoxic conditions was investigated. Various functional assays were performed for NDRG1-knockdown HCC cell lines Huh-7 and PLC/PRF/5. Results: With RT-qPCR analysis on our 102 pairs of patients’ HCC tumor and corresponding non-tumorous liver samples, we found NDRG1 to be upregulated in HCC tumors. Similarly, RNA-sequencing analysis comparing 41 pairs of samples confirmed NDRG1 enrichment in HCC tumors. The upregulation was significantly and positively correlated with the absence of tumor encapsulation and cellular differentiation. These were further supported by TCGA and ICGC databases, in which upregulation of NDRG1 was significantly correlated with poor HCC patient overall survival. Interestingly, in the TCGA database, only 17% of the HCC cases were inferred to have copy number amplification of NDRG1 gene while NDRG1 mutations were rarely found. As hypoxia is a major tumor microenvironmental factor in HCC, we investigated NDRG1’s expression level in HCC cells under hypoxic or normoxic conditions and found significant increase in NDRG1 expression under hypoxic environment. We further found that NDRG1 knockdown inhibited cell proliferation, self-renewal ability, and migration. RT-qPCR analysis found lower LGR5 and SOX2 expression upon NDRG1 knockdown. This is in line with the positive correlation between NDRG1 and various cancer stemness-related gene expression, including LGR5, CD24, MYC, SOX2, and NOTCH1, in TCGA database. Lastly, in the resected HCC tumors on which spatial transcriptomics using Visium 10X and single-cell RNA-sequencing using Chromium 10X were performed, we found NDRG1 enrichment in both tumors cells and cancer associated fibroblasts (CAFs). Conclusion: NDRG1 was found to be significantly upregulated in HCC cells in our in-house cohort and publicly available databases. NDRG1 consistently promotes various oncogenic features of HCC cells, including proliferation, self-renewal ability, and migration. In addition, NDRG1 was found to be enriched in both the HCC cells and CAFs. Citation Format: Jingyi Lu, Lu Tian, Yu-Man Tsui, Xia Wong, Qingyang Zhang, Daniel Wai-Hung Ho, Irene Oi-Lin Ng. The roles of NDRG1 in HCC progression and its clinical significance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6982.

Abstract 425: GPD1L participates in glycerol-3-phosphate biogenesis and promotes tumor initiation and invasion in hepatocellular carcinoma

Abstract Background: Metabolic reprogramming in hepatocellular carcinoma (HCC) has received rising interest. However, current studies often focus on fatty acid de novo synthesis or oxidation, but less on glycerolipids metabolism. Interestingly, in HCC, we observed frequent dysregulation in enzymes responsible for glycerol-3-phosphate shuttle, which potentially links glycolysis and glycerolipids metabolism. Hence, we aimed to identify the functional significance of glycerol-3-phosphate dehydrogenase 1 like (GPD1L), an enzyme for the shuttle, and its regulation of expression in HCC. Methods: We utilized transcriptomic analysis of clinical HCC samples to validate the frequent overexpression of GPD1L in tumor cells and examined its association with clinicopathological features. In vitro and in vivo assays were performed to explore the functional significance of GPD1L in HCC initiation and progression. Furthermore, with targeted metabolomics analysis, we investigated the role of GPD1L in glycerol-3-phosphate shuttle in HCC. We also performed promoter study to understand the transcriptional regulation on GPD1L expression. Results: Among GPD family members, GPD1L was consistently upregulated at mRNA level in clinical HCC samples in The Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) (N=50) and our in-house RNA-sequencing cohorts (N=41), and in a separate independent in-house cohort as examined by RT-qPCR (N=80). GPD1L overexpression was associated with increased tumor invasiveness (p=0.008) and poorer overall survival (p=0.0017) in HCC patients. Functionally, knockdown of GPD1L suppressed HCC migration (fold change ≤ 0.43) and invasion (fold change ≤ 0.40). GPD1L knockdown also inhibited tumor initiation abilities, as shown by reduced colony (fold change ≤ 0.37) and sphere formation (fold change ≤ 0.41) in vitro, and lower stem cell frequency (p=1.96e-9) in vivo. Furthermore, GPD1L knockdown enhanced sensitivity of HCC cells to chemotherapeutic drugs, sorafenib and doxorubicin. Metabolically, GPD1L facilitated synthesis of glycerolipids precursor, glycerol-3-phosphate (G3P) from glycolysis intermediate dihydroxyacetone phosphate (DHAP) in HCC. Lastly, with chromatin-immunoprecipitation and Luciferase reporter assays, we demonstrated ELF1 bound to GPD1L promoter directly to enhance the gene transcription, while knockdown of ELF1 suppressed GPD1L mRNA and protein expression. Conclusion: Our study demonstrated the overexpression of the glycolysis-related gene, GPD1L, in HCC. GPD1L promoted tumor initiation, invasion and chemoresistance, and in turn might lead to worse clinical outcome. Metabolically, it was responsible for DHAP-to-G3P conversion in HCC and might serve as a linkage between glycolysis and glycerolipid metabolism. Our results also identified the novel role of ELF1 as the transcriptional activator of GPD1L in HCC. Citation Format: Tiffany Ching-Yun Yu, Luqing Zhao, Lu Tian, Charles Shing Kam, Joyce Man-Fong Lee, Daniel Wai-Hung Ho, Lo-Kong Chan, Yu-Man Tsui, Karen Man-Fong Sze, Irene Oi-Lin Ng. GPD1L participates in glycerol-3-phosphate biogenesis and promotes tumor initiation and invasion in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 425.

Tubuloside B, a major constituent of Cistanche deserticola, inhibits migration of hepatocellular carcinoma by inhibiting Hippo-YAP pathway

BackgroundStudies have shown that phenylethanoid glycosides (PhGs) have multiple pharmacological effects such as anti-inflammatory, hepatoprotective or neuroprotective functions, whereas their anti-tumor effects are rarely studied. Tubuloside B (Tub B) is a PhG isolated from Cistanche deserticola, a traditional Chinese medicine. To date, there is a lack of comprehensive research regarding the biological activity of Tub B. PurposeThe subject of the current study was to investigate the anti-hepatocellular carcinoma (HCC) cell activity and the underlying mechanism of Tub B. MethodsWe evaluated the in vitro anti-migratory effect of Tub B by scratch and transwell assays. RNA-seq was employed to identify the differential genes by Tub B. Besides, the functional mechanism of Tub B was investigated by distinct molecular biology techniques including immunofluorescent staining, quantitative PCR, as well as western blot analysis. Subsequently, we utilized Hep3B cells for in vivo metastasis assays through spleen injection and evaluated the anti-migratory effect of Tub B in hepatocellular carcinoma (HCC). ResultsTub B exhibited in vitro and in vivo inhibition of HCC cell migration. Tub B decreased the expression of transcriptional target genes downstream of the Hippo pathway, including CTGF, CYR61, and N-cadherin as determined by RNA-seq. Furthermore, mechanistic studies confirmed that Tub B increased phosphorylation of YAP at S127, which contributes to YAP cytoplasmic localization. Additionally, overexpression of YAP abrogated Tub B-induced inhibition of HCC migration and the mRNA levels of CTGF, CYR61, and N-cadherin. ConclusionsTaken together, these results illustrated that Tub B demonstrated great potential in inhibiting migration of HCC, and a portion of its impact can be attributed to the modulation of the Hippo-YAP pathway.

Histone demethylase KDM5D represses the proliferation, migration and invasion of hepatocellular carcinoma through the E2F1/TNNC1 axis.

This study focused on investigating the mechanism in which the KDM5D/E2F1/TNNC1 axis affected hepatocellular carcinoma (HCC) development. At first, we determined HCC cell proliferation, migration, invasion, and apoptosis, as well as SOD activity, MDA content, and ROS level. ChIP assay was subsequently conducted to examine H3K4me3 modification in the E2F1 promoter region and the binding of E2F1 to the TNNC1 promoter region after KDM5D overexpression. Meanwhile, we performed western blot for testing KDM5D, H3K4me3, and E2F1 expression after KDM5D overexpression in Huh-7 cells. The binding of transcription factor E2F1 to the TNNC1 promoter region was assessed by dual luciferase reporter gene assay. We further observed the tumor growth ability in nude mice transplanted tumor models. Overexpressed KDM5D suppressed HCC proliferation, migration, and invasion, promoted the apoptosis, suppressed SOD activity, elevated MDA content and ROS level, and promoted ferroptosis. KDM5D suppressed H3K4me3 modification in the E2F1 promoter region and suppressed E2F1 expression in HCC cells. Reduced KDM5D, H3K4me3, and E2F1 expression was found after KDM5D overexpression in Huh-7 cells. Overexpressing E2F1 reversed the inhibitory effects of KDM5D on HCC cell proliferative, migratory, and invasive behaviors. KDM5D repressed TNNC1 transcription by inhibiting E2F1 binding to the TNNC1 promoter. In vivo KDM5D overexpression inhibited HCC development via the E2F1/TNNC1 axis. KDM5D inhibits E2F1 expression by suppressing H3K4me3 modification in the E2F1 promoter region, which in turn suppresses the binding of E2F1 to the TNNC1 promoter region, thus leading to the inhibition of HCC development.

ALKBH5 promotes hepatocellular carcinoma cell proliferation, migration and invasion by regulating TTI1 expression.

The objective of this research was to investigate the potential mechanisms of AlkB homolog 5, RNA demethylase (ALKBH5) in hepatocellular carcinoma (HCC). We used The Cancer Genome Atlas (TCGA), Kruskal-Wallis method and Kaplan-Meier (KM) survival analysis to study the expression of ALKBH5 and its correlation with clinical factors in HCC. In vitro experiments verified the expression of ALKBH5 and its effect on HCC cell phenotype. We screened differentially expressed genes (DEGs) from HCC patients associated with ALKBH5. Through this screening we identified the downstream gene TTI1 which is associated with ALKBH5 and investigated its function using Gene Expression Profiling Interaction Analysis (GEPIA) along with univariate Cox proportional hazards regression analysis. Finally, we analyzed the functions of ALKBH5 and TTI1 in HCC cells. Across numerous pan-cancer types, we observed significant overexpression of ALKBH5. In vitro experiments confirmed ALKBH5 as an oncogene in HCC, with its knockdown leading to suppressed cell proliferation, migration, and invasion. Bioinformatics analyses also demonstrated a significant positive correlation between ALKBH5 and TTI1. TTI1, highly expressed in cells, showed promising prognostic ability for patients. Further experiments confirmed that suppressing TTI1 impeded cell growth and movement, with this effect partially offset by increased ALKBH5 expression. Conversely, promoting these cellular processes was observed with TTI1 overexpression, but was dampened by decreased ALKBH5 expression. In conclusion, our findings suggest that ALKBH5 may influence proliferation, migration and invasion of HCC by modulating TTI1 expression, providing a new direction for treating HCC.

HOXD3 promotes the migration and angiogenesis of hepatocellular carcinoma via modifying hepatocellular carcinoma cells exosome-delivered CCR6 and regulating chromatin conformation of CCL20

Angiogenesis plays an essential role in the microenvironment of hepatocellular carcinoma (HCC). HOXD3 is involved in the metastasis and invasion of HCC cells; Whereas the underlying molecular mechanisms in the microenvironment of HCC remain unknown. Wound healing, transwell invasion, tube formation and spheroid sprouting assays were carried out to identify the effects of HCC-HOXD3-exosomes and genes on the migration of HCC cells. ChIP–PCR was applied to test the binding region of HOXD3 on CCR6, Med15, and CREBBP promoter. Exosome isolation and mRNA-seq were applied to examine the morphological characteristics of exosomes and the contained mRNA in exosomes. Co-IP and Immunofluorescence assays were used to demonstrate the role of CREBBP in the chromatin conformation of CCL20. The nude mice were used to identify the function of genes in regulating migration of HCC in vivo. In this study, integrated cellular and bioinformatic analyses revealed that HOXD3 targeted the promoter region of CCR6 and induced its transcription. CCR6 was delivered by exosomes to endothelial cells and promoted tumour migration. Overexpression of CCR6 promoted metastasis, invasion in HCCs and angiogenesis in endothelial cells (ECs), whereas its downregulation suppressed these functions. The role of HOXD3 in the metastasis and invasion of HCC cells was reversed after the suppression of CCR6. Furthermore, CCL20 was demonstrated as the ligand of CCR6, and its high expression was found in HCC tissues and cells, which was clinically associated with the poor prognosis of HCC. Mechanistically, HOXD3 targets the promoter regions of CREBBP and Med15, which affect CCL20 chromatin conformation by regulating histone acetylation and expression of Pol II to enhance the migration of HCCs. This study demonstrated the function of the HOXD3–CREBBP/Med15–CCL20–CCR6 axis in regulating invasion and migration in HCC, thus providing new therapeutic targets for HCC.

Knockdown of ribosome RNA processing protein 15 suppresses migration of hepatocellular carcinoma through inhibiting PATZ1-associated LAMC2/FAK pathway

BackgroundRibosomal RNA processing protein 15 (RRP15) has been found to regulate the progression of hepatocellular carcinoma (HCC). Nevertheless, the extent to which it contributes to the spread of HCC cells remains uncertain. Thus, the objective of this research was to assess the biological function of RRP15 in the migration of HCC.MethodsThe expression of RRP15 in HCC tissue microarray (TMA), tumor tissues and cell lines were determined. In vitro, the effects of RRP15 knockdown on the migration, invasion and adhesion ability of HCC cells were assessed by wound healing assay, transwell and adhesion assay, respectively. The effect of RRP15 knockdown on HCC migration was also evaluated in vivo in a mouse model.ResultsBioinformatics analysis showed that high expression of RRP15 was significantly associated with low survival rate of HCC. The expression level of RRP15 was strikingly upregulated in HCC tissues and cell lines compared with the corresponding controls, and TMA data also indicated that RRP15 was a pivotal prognostic factor for HCC. RRP15 knockdown in HCC cells reduced epithelial-to-mesenchymal transition (EMT) and inhibited migration in vitro and in vivo, independent of P53 expression. Mechanistically, blockade of RRP15 reduced the protein level of the transcription factor POZ/BTB and AT hook containing zinc finger 1 (PATZ1), resulting in decreased expression of the downstream genes encoding laminin 5 subunits, LAMC2 and LAMB3, eventually suppressing the integrin β4 (ITGB4)/focal adhesion kinase (FAK)/nuclear factor κB kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway.ConclusionsRRP15 promotes HCC migration by activating the LAMC2/ITGB4/FAK pathway, providing a new target for future HCC treatment.

Angiopoietin-Related Protein 4-Transcript 3 Increases the Proliferation, Invasion, and Migration of Hepatocellular Carcinoma Cells and Inhibits Apoptosis.

To investigate the functional differences of angiopoietin-related protein 4 (ANGPTL4) transcripts in hepatocellular carcinoma (HCC) cells. By transfecting ANGPTL4-Transcript 1 and ANGPTL4-Transcript 3 overexpression vectors into HepG2 and Huh7 cell lines with ANGPTL4 knockdown, the effects of overexpression of two transcripts on cell viability, invasion, migration, and apoptosis were analyzed. The expression of two transcripts was compared in human liver cancer tissue, and their effects on tumor development were validated in vivo experiments in mice. Compared with control, the overexpression of ANGPTL4-Transcript 1 had no significant effect on viability, invasion, healing, and apoptosis of HepG2 and Huh7 cells. However, these two cell lines overexpressing ANGPTL4-Transcript 3 showed remarkably enhanced cell viability, invasive and healing ability, and decreased apoptosis ability. Furthermore, the mRNA level of ANGPTL4-Transcript 3 was significantly increased in human HCC tissues and promoted tumor growth compared with Transcript 1. Different transcripts of gene ANGPTL4 have distinct effects on HCC. The abnormally elevated Transcript 3 with the specific ability of promoting HCC proliferation, infiltration, and migration is expected to become a new biological marker and more precise intervention target for HCC.

ASIC1 promotes migration and invasion of hepatocellular carcinoma via the PRKACA/AP-1 signaling pathway.

Hepatocellular carcinoma (HCC) exhibits a high mortality rate due to its high invasion and metastatic nature, and the acidic microenvironment plays a pivotal role. Acid-sensing ion channel 1 (ASIC1) is upregulated in HCC tissues and facilitates tumor progression in a pH-dependent manner, while the specific mechanisms therein remain currently unclear. Herein, we aimed to investigate the underlying mechanisms by which ASIC1 contributes to the development of HCC. Using bioinformatics analysis, we found a significant association between ASIC1 expression and malignant transformation of HCC, such as poor prognosis, metastasis and recurrence. Specifically, ASIC1 enhanced the migration and invasion capabilities of Li-7 cells in the in vivo experiment using an HCC lung metastasis mouse model, as well as in the in vitro experiments such as wound healing assay and Transwell assay. Furthermore, our comprehensive gene-chip and molecular biology experiments revealed that ASIC1 promoted HCC migration and invasion by activating the PRKACA/AP-1 signaling pathway. Our findings indicate that targeting ASIC1 could have therapeutic potential for inhibiting HCC progression.

Hsa_circ_0010882 facilitates hepatocellular carcinoma progression by modulating M1/M2 macrophage polarization.

Hepatocellular carcinoma (HCC) is one common malignant tumour with a high immunosuppressive tumour microenvironment and poor outcomes. This study investigated the influence of hsa_circ_0010882 on M1/M2 macrophage polarization in the progression of HCC. A total of 125 paired tissue specimens from HCC patients who underwent hepatectomy were collected. M1 and M2 phenotypes macrophages were induced using THP-1. After co-cultured with macrophages and transfected HCC cells, the viability, migration and invasion of HCC cells were detected by cellular experiments. Bioinformatic databases and dual-luciferase reporter assays were used to predict and validate the interaction between circ_0010882 and miR-382. Expression of circ_0010882 was increased in HCC tissues and associated with shorter overall survival outcomes. The mRNA expression of M2 macrophage markers Arg-1, CD163 and CD206 were elevated in HCC tissues. Interfering with circ_0010882 increased M1-type macrophage markers (TNF-α and iNOS) while decreasing M2-type macrophage markers (Arg-1 and CD206). Silencing of circ_0010882 strengthened the capacity of M1 macrophages to suppress HCC cell viability, migration capacities and invasion potential while reducing the ability of M2 macrophages to promote above cellular abilities. MiR-382 was a direct target miRNA of circ_0010882. The circ_0010882 expression was increased in HCC tissues and associated with poor prognosis of HCC patients. Silencing of circ_0010882 inhibits macrophage M2 polarization in HCC progression by regulating miR-382 expression. Circ_0010882 may serve as a biomarker to provide novel strategies for the treatment of HCC and patient rehabilitation, thereby improving the prognosis of patients.

A hsa_circ_001726 axis regulated by E2F6 contributes to metastasis of hepatocellular carcinoma

BackgroundCircRNAs participate in the development of hepatocellular carcinoma (HCC). This work aims to explore the key tumor promoting circRNA as a gene therapy target.MethodsThe differentially expressed gene circRNAs in HCC tumor tissues was identified by mining GSE121714 dataset. EdU staining, wound healing, transwell invasion assay, TUNEL staining and western blotting examined proliferation, migration, invasion, apoptosis and epithelial mesenchymal transition (EMT). Xenograft mouse model and orthotopic transplantation tumor mouse model were constructed to verify the role of hsa_circ_001726 in growth and metastasis of HCC. The relationship among CCT2, E2F6, hsa_circ_001726, miR-671-5p and PRMT9 was identified by RNA-fluorescence in situ hybridization, luciferase reporter assay and RNA Immunoprecipitation.ResultsEleven differentially expressed circRNAs were found in HCC tumors. Among them, hsa_circ_001726 was highly expressed in HCC tumors and cells, which was transcribed from CCT2. As a transcription factor of CCT2, E2F6 knockdown inactivated CCT2 promoter and reduced hsa_circ_001726 expression. Moreover, hsa_circ_001726 elevated PRMT9 expression by sponging miR-671-5p, and then activated Notch signaling pathway. Additionally, hsa_circ_001726 deficiency repressed malignant phenotypes of HCC cells, including proliferation, migration, invasion, EMT and apoptosis. In vivo, hsa_circ_001726 deficiency reduced tumor growth and lung metastasis of HCC in xenograft mouse models and orthotopic transplantation tumor mouse models.ConclusionHsa_circ_001726 functioned as an oncogene in HCC, which was derived from CCT2 and regulated by E2F6. Hsa_circ_001726 elevated PRMT9 expression by sponging miR-671-5p, and then activated Notch signaling pathway, thereby accelerating malignant phenotypes of HCC. Therefore, targeting hsa_circ_001726 may be a new avenue for HCC treatment.

Angiopoietin-2 and the Vascular Endothelial Growth Factor Promote Migration and Invasion in Hepatocellular Carcinoma- and Intrahepatic Cholangiocarcinoma-Derived Spheroids.

Aggressive hepatocellular carcinoma (HCC) overexpressing Angiopoietin-2 (ANG-2) (a protein linked with angiogenesis, proliferation, and epithelial-mesenchymal transition (EMT)), shares 95% of up-regulated genes and a similar poor prognosis with the proliferative subgroup of intrahepatic cholangiocarcinoma (iCCA). We analyzed the pro-invasive effect of ANG-2 and its regulator vascular endothelial growth factor (VEGF) on HCC and CCA spheroids to uncover posUsible common ways of response. Four cell lines were used: Hep3B and HepG2 (HCC), HuCC-T1 (iCCA), and EGI-1 (extrahepatic CCA). We treated the spheroids with recombinant human (rh) ANG-2 and/or VEGF and then observed the changes at the baseline, after 24 h, and again after 48 h. Proangiogenic stimuli increased migration and invasion capability in HCC- and iCCA-derived spheroids and were associated with a modification in EMT phenotypic markers (a decrease in E-cadherin and an increase in N-cadherin and Vimentin), especially at the migration front. Inhibitors targeting ANG-2 (Trebananib) and the VEGF (Bevacizumab) effectively blocked the migration ability of spheroids that had been stimulated with rh-ANG-2 and rh-VEGF. Overall, our findings highlight the critical role played by ANG-2 and the VEGF in enhancing the ability of HCC- and iCCA-derived spheroids to migrate and invade, which are key processes in cancer progression.

ASTAXANTHIN INHIBITS CELL PROLIFERATION, MIGRATION, INVASION AND INDUCED APOPTOSIS VIA AMPK-mTOR SIGNALING PATHWAY IN HEPATOCELLULAR CARCINOMA HEP 3B CELLS

"Hepatocellular carcinoma (HCC) is a fatal malignancy with a poor prognosis. There is an urgent need to study the molecular mechanisms of HCC development and explore potential drugs to improve survival. This study aims to investigate the antitumor effects of astaxanthin on HCC proliferation, migration, invasion and apoptosis through regulation of adenosine-activated protein kinase (AMPK). CCK8, wound healing, transwell and flow cytometry assays were used to evaluate Hep 3B cell viability, migration, invasion and apoptosis after astaxanthin treatment. Protein expression was determined by Western blot. CCK8 assays showed that all concentrations (200 μM, 400 μM, 500 μM) of astaxanthin used in this study significantly inhibited the proliferation of Hep 3B cells (P < 0.05). Wound healing, transwell and flow cytometry showed that astaxanthin inhibited Hep 3B cell migration and invasion and induced apoptosis. Western blot showed that astaxanthin increased the expression of p-AMPK (P <0.05) and inhibited its downstream p-mTOR protein expression (P < 0.05). The effects of astaxanthin on cell proliferation, migration, invasion and apoptosis was attenuated after addition of the AMPK blocker Compound C. To conclude, astaxanthin inhibits the proliferation, migration and invasion of HCC by regulating AMPK, and promotes tumor cell apoptosis in a dose-dependent manner."

Heat shock protein HSPA13 promotes hepatocellular carcinoma progression by stabilizing TANK

HSPA13, an important member of the heat shock protein family, plays an essential role in the oncogenesis of many organs, but the mechanism and function in hepatocellular carcinoma (HCC) is still unclear. In the present study, we found that HSPA13 was highly expressed in HCC and predicted poor clinical prognosis. Upregulation of HSPA13 was significantly associated with vascular invasion in HCC patients. Functionally, knockdown experiments demonstrated that HSPA13 promoted HCC proliferation, migration, and invasion. Mechanistic investigation showed that HSPA13 could interact with TANK to inhibit its ubiquitination and degradation. In addition, the expression of HSPA13 and TANK were positively correlated in HCC tissues. To conclude, the present study uncovers the oncogenic function of HSPA13 in the progression of HCC by regulating the stability of TANK. These findings suggest that HSPA13 and TANK may serve as promising targets for the diagnosis and treatment of HCC.

M6A modification of AC026356.1 facilitates hepatocellular carcinoma progression by regulating the IGF2BP1-IL11 axis

N6-methyladenosine (m6A) is the most common RNA modification in eukaryotic RNAs. Although the important roles of m6A in RNA fate have been revealed, the potential contribution of m6A to RNA function in various diseases, including hepatocellular carcinoma (HCC), is still unclear. In this study, we identified a novel m6A-modified RNA AC026356.1. We found that AC026356.1 was increased in HCC tissues and cell lines. High expression of AC026356.1 was correlated with poor survival of HCC patients. m6A modification level of AC026356.1 was also increased in HCC and more significantly correlated with poor survival of HCC patients. Functional assays showed that m6A-modified AC026356.1 promoted HCC cellular proliferation, migration, and liver metastasis. Gene set enrichment analysis showed that AC026356.1 activated IL11/STAT3 signaling. Mechanistic investigation showed that m6A-modified AC026356.1 bound to IGF2BP1. The interaction between m6A-modified AC026356.1 and IGF2BP1 promoted the binding of IL11 mRNA to IGF2BP1, leading to increased IL11 mRNA stability and IL11 secretion. Functional rescue assays showed that depletion of IL11 reversed the oncogenic roles of AC026356.1. These findings revealed the potential influences of m6A modification on RNA biological functions and suggested that targeting m6A modification may be a novel strategy for HCC treatment.

ETV4 facilitates angiogenesis in hepatocellular carcinoma by upregulating MMP14 expression

Abnormal vascularization plays a crucial role in cell proliferation, tumor invasion and metastasis of hepatocellular carcinoma (HCC). It has been reported that ETV4 functions as an oncogenic gene in driving the carcinogenesis and progression, and promoting invasion and metastasis of HCC. However, the function of ETV4 on angiogenesis in HCC remains unclear. In the current study, immunohistochemistry showed that knockdown of ETV4 reduced angiogenesis in HCC xenograft tumor tissues. In vitro, tube formation assay verified that ETV4 expression promoted angiogenesis through simulating the angiogenic environment in HCC cells. Transcriptome sequencing indicated that MMP14 was one of the differentially expressed genes enriched in angiogenesis process. Subsequently, it was confirmed that MMP14 was regulated by ETV4 at the transcription level in HCC cells, clinical tissue samples and online databases. Further, we demonstrated that MMP14 induced angiogenesis in ETV4-mediated HCC microenvironment. Collectively, this research further reveals the biological mechanism of ETV4 in promoting the migration and invasion of HCC, and provides novel mechanistic insights and strategic guidance for anti-angiogenic therapy in HCC.