Levels In Hepatocellular Carcinoma Tissues Research Articles

Downregulation of GLYAT correlates with tumour progression and poor prognosis in hepatocellular carcinoma.

Glycine N-acyltransferase (GLYAT), known to influence glycine metabolism, has been implicated in the progression of various malignant tumours. However, its clinical relevance in hepatocellular carcinoma (HCC) remains unexplored. Here, GLYAT expression levels in HCC tissues were significantly reduced compared to normal liver tissues. Similarly, GLYAT expression levels in Huh 7, HepG2, PLC and SK-HEP1 were lower than those in LO2. Receiver operating characteristic curve analysis demonstrated that GLYAT exhibited good diagnostic performance for HCC. Kaplan-Meier analyses suggested that decreased GLYAT expression was correlated with poorer progress in HCC. Low GLYAT expression was significantly associated with gender and histologic grade. Multivariate Cox regression analysis identified low GLYAT expression and T stage as independent prognostic factors. Nomograms based on GLYAT mRNA expression and T stage showed good concordance with actual survival rates at 1, 2, 3 and 5 years. Moreover, GLYAT downregulation in the Huh 7 cell line enhanced cell proliferation, invasion and migration abilities, while GLYAT overexpression in the HepG2 cell line inhibited these abilities. HCC patients with low GLYAT expression exhibited a predisposition to immune escape and poor response to immunotherapy. This research revealed that GLYAT holds promise as both a prognostic biomarker and a potential therapeutic target in HCC.

Nlrp6 overexpression inhibits lipid synthesis to suppress proliferation of hepatocellular carcinoma cells by regulating the AMPK-Srebp1c axis

To investigate the mechanism of Nlrp6 for regulating hepatocellular carcinoma (HCC) progression in light of lipid synthesis regulation. Nlrp6 expression level in HCC tissues of different pathological grades was investigated using RNA-seq data from The Cancer Genome Atlas (TCGA) database, and its correlation with the patients' survival was analyzed with Kaplan-Meier survival analysis. HepG2 cells with adenovirus-mediated Nlrp6 overexpression or knockdown were treated with palmitic acid (PA), and the changes in lipid deposition and cell proliferation were evaluated using Oil Red O staining, CCK-8 assay, EdU staining, and colony formation assay. RT-qPCR and Western blotting were used to detect the changes in expression of lipid synthesis-related genes and the proteins in the AMPK-Srebp1c axis. In a mouse model of hepatic steatosis established in liver-specific Nlrp6 knockout mice by high-fat diet feeding for 24 weeks, liver fibrosis was examined with histological staining, and the changes in expressions of HCC markers and the AMPK-Srebp1c signaling pathway were detected. Nlrp6 expression was significantly reduced in HCC tissues with negative correlations with the pathological grades and the patients' survival (P < 0.0001). In HepG2 cells, Nlrp6 overexpression significantly inhibited lipid deposition and cell proliferation, whereas Nlrp6 knockdown produced the opposite effects. Nlrp6 overexpression strongly suppressed the expression of lipid synthesis-related genes, promoted AMPK phosphorylation, and inhibited Srebp1c expression. The mice with liver-specific Nlrp6 knockout and high-fat feeding showed increased hepatic steatosis, collagen deposition, and AFP expression with reduced AMPK phosphorylation and increased Srebp1c expression. Nlrp6 overexpression inhibits lipid synthesis in HCC cells by regulating the AMPK-Srebp1c axis, which might be a key pathway for suppressing HCC cell proliferation.

Hepatic Bone Morphogenetic Protein and Activin Membrane-Bound Inhibitor Levels Decline in Hepatitis C but Are Not Associated with Progression of Hepatocellular Carcinoma.

Bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI) is an antagonist of transforming growth factor (TGF)-β type 1 signaling. BAMBI functions as an anti-fibrotic protein and exerts pro- as well as anti-cancerogenic activities. Our study aimed to correlate hepatocyte BAMBI protein levels in hepatocellular carcinoma (HCC) with T stage, lymph node invasion, vessel invasion, grading, tumor size and Union for International Cancer Control (UICC) stage, as well as with liver inflammation and fibrosis stages. Hepatocyte BAMBI protein expression was assessed by immunohistochemistry in HCC tissues of 320 patients and non-tumor tissues of 51 patients. In the HCC tissues of the whole cohort and sex-specific analysis, BAMBI protein was not related to T stage, vessel invasion, lymph node invasion, histologic grade, UICC stage and tumor size. Accordingly, BAMBI was not associated with overall survival, recurrence-free and metastasis-free survival. BAMBI protein levels in tumor and non-tumor tissues were not related to inflammation and fibrosis grade. BAMBI protein levels in HCC tissues and non-tumor tissues from HCC patients, which were analyzed by immunoblot in a small cohort and by immunohistochemistry in the tissues of patients described above, were similar. Notably, BAMBI protein was low-abundant in HCC tissues of hepatitis C virus (HCV) compared to hepatitis B virus (HBV)-infected patients with comparable disease severity. Immunoblot analysis revealed reduced BAMBI protein in non-tumor tissues of patients with HCV in comparison to patients with HBV and normal human liver tissues. In summary, this analysis showed that hepatocyte BAMBI protein levels of patients with HCC are related to HCV infection rather than the severity of the underlying liver disease and cancer staging.

Histone lactylation facilitates hepatocellular carcinoma progression by upregulating endothelial cell-specific molecule 1 expression.

Hepatocellular carcinoma (HCC) is a common malignant tumor. Histone lactylation, a novel epigenetic modification, plays a crucial role in various cancers. However, the functional role and underlying mechanism of histone lactylation in HCC progression have not yet been investigated. Histone lactylation levels in HCC tissues and cells were assessed using a densitometric kit and western blot analysis. The role of histone lactylation in cell malignant phenotypes was determined through functional assays in vitro, and a xenograft tumor model was established to verify the function of histone lactylation in vivo. ChIP assay was performed to explore the interaction between histone lactylation and endothelial cell-specific molecule 1 (ESM1). Additionally, gain-and-loss-of-function assays were conducted to investigate the regulatory role of ESM1 in HCC pathogenesis. Histone lactylation levels were increased in HCC tissues and cells, and H3K9 lactylation (H3K9la) and H3K56 lactylation (H3K56la) were identified as the histone modification sites. We observed that H3K9la and H3K56la caused abnormal histone lactylation and were associated with poor prognosis. Functionally, histone lactylation was found to promote HCC cell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) process in vitro. However, histone lactylation inhibition with 2-deoxy-d-glucose (2-DG) reduced the malignant phenotypes of HCC cells. In vivo, 2-DG treatment reduced tumor growth and metastasis in the HCC mouse model. Mechanistically, it was revealed that histone lactylation activated ESM1 transcription in HCC cells. ESM1 was expressed at a high level in HCC and exerted a carcinogenic role. Histone lactylation facilitates cell malignant phenotypes, tumor growth, and metastasis by upregulating ESM1 expression in HCC, which reveals the downstream molecular mechanism of histone lactylation and might provide a novel therapeutic target for HCC therapy.

FAT10 induces immune suppression by upregulating PD-L1 expression in hepatocellular carcinoma.

The upregulation of programmed death ligand 1 (PD-L1) plays a crucial role in facilitating cancer cells to evade immune surveillance through immunosuppression. However, the precise regulatory mechanisms of PD-L1 in hepatocellular carcinoma (HCC) remain undefined. The correlation between PD-L1 and ubiquitin-like molecules (UBLs) was studied using sequencing data from 20 HCC patients in our center, combined with TCGA data. Specifically, the association between FAT10 and PD-L1 was further validated at both the protein and mRNA levels in HCC tissues from our center. Subsequently, the effect of FAT10 on tumor progression and immune suppression was examined through both in vivo and in vitro experiments. Utilizing sequencing data, qPCR, and Western blotting assays, we confirmed that FAT10 was highly expressed in HCC tissues and positively correlated with PD-L1 expression. Additionally, in vitro experiments demonstrated that the overexpression of FAT10 fostered the proliferation, migration, and invasion of HCC cells. Furthermore, the overexpression of FAT10 in HCC cells led to an increase in PD-L1 expression, resulting in the inhibition of T cell proliferation and the enhancement of HCC cell resistance to T cell-mediated cytotoxicity. Moreover, in vivo experiments utilizing the C57BL/6 mouse model revealed that overexpression of FAT10 effectively suppressed the infiltration of CD8 + GZMB + and CD8 + Ki67 + T cells, as well as reduced serum levels of TNF-α and IFN-γ. Mechanistically, we further identified that FAT10 upregulates PD-L1 expression via activating the PI3K/AKT/mTOR pathway, but not in a ubiquitin-like modification. In conclusion, our findings indicate that FAT10 promotes immune evasion of HCC via upregulating PD-L1 expression, suggesting its potential as a novel target to enhance the efficiency of immunotherapy in HCC.

Deficiency of betaine-homocysteine methyltransferase activates glucose-6-phosphate dehydrogenase (G6PD) by decreasing arginine methylation of G6PD in hepatocellular carcinogenesis.

Betaine-homocysteine methyltransferase (BHMT) regulates protein methylation and is correlated with tumorigenesis; however, the effects and regulation of BHMT in hepatocarcinogenesis remain largely unexplored. Here, we determined the clinical significance of BHMT in the occurrence and progression of hepatocellular carcinoma (HCC) using tissue samples from 198 patients. BHMT was to be frequently found (86.6%) expressed at relatively low levels in HCC tissues and was positively correlated with the overall survival of patients with HCC. Bhmt overexpression effectively suppressed several malignant phenotypes in hepatoma cells in vitro and in vivo, whereas complete knockout of Bhmt (Bhmt-/-) produced the opposite effect. We combined proteomics, metabolomics, and molecular biological strategies and detected that Bhmt-/- promoted hepatocarcinogenesis and tumor progression by enhancing the activity of glucose-6-phosphate dehydrogenase (G6PD) and PPP metabolism in DEN-induced HCC mouse and subcutaneous tumor-bearing models. In contrast, restoration of Bhmt with an AAV8-Bhmt injection or pharmacological inhibition of G6PD attenuated hepatocarcinogenesis. Additionally, coimmunoprecipitation identified monomethylated modifications of the G6PD, and BHMT regulated the methylation of G6PD. Protein sequence analysis, generation and application of specific antibodies, and site-directed mutagenesis indicated G6PD methylation at the arginine residue 246. Furthermore, we established bidirectionally regulated BHMT cellular models combined with methylation-deficient G6PD mutants to demonstrate that BHMT potentiated arginine methylation of G6PD, thereby inhibiting G6PD activity, which in turn suppressed hepatocarcinogenesis. Taken together, this study reveals a new methylation-regulatory mechanism in hepatocarcinogenesis owing to BHMT deficiency, suggesting a potential therapeutic strategy for HCC treatment.

SIRT3 suppression resulting from the enhanced β-catenin signaling drives glycolysis and promotes hypoxia-induced cell growth in hepatocellular carcinoma cells

ABSTRACT The precise mechanisms underlying the inhibitory effects of SIRT3, a mitochondrial sirtuin protein, on hepatocellular carcinoma (HCC) development, as well as its impact on mitochondrial respiration, remain poorly understood. We assessed sirtuins 3 (SIRT3) levels in HCC tissues and Huh7 cells cultured under hypoxic condition. We investigated the effects of SIRT3 on cell proliferation, glycolytic metabolism, mitochondrial respiration, mitophagy, and mitochondrial biogenesis in Huh7 cells. Besides, we explored the potential mechanisms regulating SIRT3 expression in hypoxically cultured Huh7 cells. Gradual reduction in SIRT3 expressions were observed in both adjacent tumor tissues and tumor tissues. Similarly, SIRT3 expressions were diminished in Huh7 cells cultured under hypoxic condition. Forced expression of SIRT3 attenuated the growth of hypoxically cultured Huh7 cells. SIRT3 overexpression led to a decrease in extracellular acidification rate while increasing oxygen consumption rate. SIRT3 downregulated the levels of hexokinase 2 and pyruvate kinase M2. Moreover, SIRT3 enhanced mitophagy signaling, as indicated by mtKeima, and upregulated key proteins involved in various mitophagic pathways while reducing intracellular reactive oxygen species levels. Furthermore, SIRT3 increased proxisome proliferator-activated receptor-gamma coactivator 1α levels and the amount of mitochondrial DNA in Huh7 cells. Notably, β-catenin expressions were elevated in Huh7 cells cultured under hypoxic condition. Antagonists and agonists of β-catenin respectively upregulated and downregulated SIRT3 expressions in hypoxically cultured Huh7 cells. The modulationsof glycolysis and mitochondrial respiration represent the primary mechanism through which SIRT3, suppressed by β-catenin, inhibits HCC cell proliferation.

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.

Low SAA4 gene expression is associated with advanced HCC stage and a poor prognosis

At present, although there are tumor markers for hepatocellular carcinoma (HCC), markers with better predictive efficiency are needed. SAA4 gene expression in liver tumor and paracancerous tissues was analyzed using The Cancer Genome Atlas database. The differentially expressed genes (DEGs) were analyzed and visualized by heatmap and volcano plot. Survival analysis was performed based on SAA4 expression. SAA4 expression was compared in patients grouped based on clinicopathological features, and gene set enrichment analysis (GSEA) was conducted. Immunohistochemical staining was used to verify the SAA4 protein staining intensity from The Human Protein Atlas database and our center’s samples. The diagnostic value of SAA4 for HCC was evaluated by receiver operating characteristic curves. SAA4 was expressed at low levels in HCC tissues, and low SAA4 expression was associated with a poor prognosis in HCC. In addition, SAA4 expression decreased with HCC progression. There were 188 upregulated DEGs and 1551 downregulated DEGs between the high and low SAA4 expression groups. Complement and coagulation cascades, fatty acid metabolism, and ECM receptor interaction were significantly enriched in the GSEA. SAA4 had good predictive efficacy for HCC and even early HCC and was superior to AFP. In general, low SAA4 expression was associated with advanced HCC stage and a poor prognosis. In addition, SAA4 may be helpful for the diagnosis of early HCC and may become a novel tumor marker with good predictive power for HCC.

Circ_0007386 Promotes the Progression of Hepatocellular Carcinoma Through the miR-507/ CCNT2 Axis.

Circular RNAs (circRNAs) have been shown to play a crucial role in the initiation and development of Hepatocellular carcinoma (HCC). However, the mechanism and function of circ_0007386 in HCC are still unknown. Circ_0007386 expression level in HCC tissues, and HCC cell lines was further analyzed by qRT-PCR. Agarose gel electrophoresis and Sanger sequencing were used to figure out the structure of circ_0007386. The involvement of circ_0007386 in HCC development was evaluated by experimental investigations conducted in both laboratory settings (in vitro) and living organisms (in vivo). RNA immunoprecipitation, Western blotting, luciferase reporter assay and fluorescence in situ hybridization (FISH) were applied for finding out the interaction among circ_0007386, miR-507 and CCNT2. To assess the connection between circ_0007386 and lenvatinib resistance, lenvatinib-resistant HCC cell lines were employed. The expression of circ_0007386 was found to increase in HCC tissues, and it was observed to be associated with a worse prognosis. Overexpression of circ_0007386 stimulated HCC cells proliferation, invasion, migration and the epithelial-mesenchymal transition (EMT) while silencing of circ_0007386 resulted in the opposite effect. Mechanistic investigations revealed that circ_0007386 acted as a competing endogenous RNA of miR-507 to prevent CCNT2 downregulation. Downregulating miR-507 or overexpressing CCNT2 could reverse phenotypic alterations that originated from inhibiting of circ_0007386. Importantly, circ_0007386 determines the resistance of hepatoma cells to lenvatinib treatment. Circ_0007386 advanced HCC progression and lenvatinib resistance through the miR-507/ CCNT2 axis. Meanwhile, circ_0007386 served as a potential biomarker and therapeutic target in HCC patients.

Circ-AKT3 Inhibits Malignant Progression of Hepatocellular Carcinoma by Negatively Regulating PDK1

This study investigated the clinical and functional significance of circ-AKT3 in hepatocellular carcinoma (HCC) and its impact on tumor progression. Using qRT-PCR, we found reduced circ-AKT3 levels in HCC tissues compared to adjacent liver samples. Clinical data revealed that low circ-AKT3 expression correlated with increased lymphatic and distant metastasis. Functional experiments in MHCC-97H and Hep3B cells, chosen due to their low circ-AKT3 levels, demonstrated that circ-AKT3 overexpression suppressed HCC cell proliferation, invasion, and migration. Bioinformatics analysis and luciferase assays indicated that circ-AKT3 potentially targets PDK1. PDK1 expression was elevated in HCC tissues and cells, inversely correlated with circ-AKT3, and reduced upon circ-AKT3 overexpression. Rescue experiments illustrated that PDK1 overexpression reversed circ-AKT3-induced inhibition of HCC cell proliferation and migration. In conclusion, downregulated circ-AKT3 in HCC, associated with aggressive clinical features, hinders tumor progression by modulating PDK1. This suggests circ-AKT3′s potential as a diagnostic and prognostic biomarker for HCC.

Effects of Circ_0132269 on the Progression of Hepatocellular Carcinoma via Targeting miR-1248/MTO1

We investigated the role of circ_0132269 in hepatocellular carcinoma (HCC). We examined circ_0132269 levels in HCC tissues and cell lines using qRT-PCR. Survival analysis was performed to assess the correlation between circ_0132269 expression and HCC patient survival rates. Knockdown of circ_0132269 was performed to evaluate its impact on HCC cell proliferation, migration, and invasion. Bioinformatics analysis predicted that circ_0132269 could interact with miR-1248. This interaction was confirmed using dual luciferase assays, and the correlation between circ_0132269 and miR-1248 was analyzed. Further functional experiments investigated the effect of miR-1248 on circ_0132269-mediated malignant phenotypes. circ_0132269 was significantly upregulated in HCC tissues and cell lines. Higher circ_0132269 expression correlated with poorer overall and disease-free survival in HCC patients. Silencing circ_0132269 suppressed HCC cell proliferation, migration, and invasion. Bioinformatics analysis revealed a binding site between circ_0132269 and miR-1248. miR-1248 expression was reduced in HCC, while its target MTO1 was highly expressed. miR-1248 levels showed a negative correlation with circ_0132269 and MTO1 levels, while circ_0132269 and MTO1 exhibited a positive correlation. Overexpression of miR-1248 partially reversed the promotive effect of highly expressed circ_0132269 on HCC cell behaviors. circ_0132269 was significantly upregulated in HCC and associated with poor prognosis. It interacts with miR-1248 to regulate HCC malignancy, highlighting its potential as a diagnostic and therapeutic target.

Hsa_circ_0001687 Function as a ceRNA to Facilitate Hepatocellular Carcinoma Progression via miR-140- 3p/FOXQ1 Axis.

Increasingly convincing evidence has revealed that circular RNAs (circRNAs) are critical regulatory components of hepatocellular carcinoma (HCC) genesis. However, the expression of circRNAs in HCC and the relevance of circRNAs to HCC progression remain largely unexplained. qRT-PCR or western blotting was utilized to confirm circ_0001687, miR-140-3p, and Forkhead Box q1 (FOXQ1) levels in HCC tissues or cells. Cell proliferation ability was evaluated via CCK-8 and colony formation assay. The correlation of circ_0001687 or FOXQ1 and miR-140- 3p was determined using dual luciferase reporter assay. Nude mice xenograft tumor model was constructed to verify the effect of circ_0001687 on tumor growth. Circ_0001687 was elevated in HCC. Function assays and the nude mice xenograft tumor model indicated that circ_0001687 acts as a promoting gene in HCC to regulate the proliferation of the tumor cell and foster tumor growth. Further mechanistic exploration revealed that the tumor growth-promoting mechanism of circ_0001687 relied on blocking the inhibitory effect of miR-140- 3p on FOXQ1 and activating FOXQ1 expression. This research indicated the role of circ_0001687/miR-140-3p/FOXQ1 network in regulating HCC development. These may provide new insights into the treatment of HCC.

Curcumin Induces Apoptosis by Suppressing XRCC4 Expression in Hepatocellular Carcinoma

Curcumin is a chemical with various pharmacological activities used for cancer treatment. It inhibits hepatocellular carcinoma (HCC) by inducing apoptosis. Here, the mechanism underlying the effect of curcumin on the apoptosis of HCC cells was studied. Cell counting kit-8 and plate cloning assays were used to assess the proliferation of HCC cells, and acridine orange/ethidium bromide and Annexin V/PI staining were used to analyze their apoptosis. HCC xenograft tumor models were established to validate anti-cancer effects of curcumin. Expression levels of XRCC4 protein in tumor tissues were assessed by immunohistochemistry. Correlation between XRCC4 expression and the prognosis of patients with HCC was analyzed by integrating publicly available gene expression data. Curcumin inhibited HCC cells proliferation in a dose-dependent manner. Compared with the control group, curcumin significantly promoted the apoptosis of HCC cells in vitro and in vivo. Immunohistochemical analysis revealed that curcumin downregulated XRCC4 expression levels in HCC tissues. Prognosis of HCC patients with high XRCC4 expression was poorer than that of patients with low XRCC4 expression. Therefore, curcumin exerts anti-cancer effects by inhibiting cell proliferation and promoting cell apoptosis in HCC. This may be due to curcumin interference in the repair process of the nonhomologous DNA terminal link of HCC cells by downregulating XRCC4 expression.

Arachidonic acid metabolism CYP450 pathway is deregulated in hepatocellular carcinoma and associated with microvascular invasion.

Arachidonic acid metabolism plays a crucial role in the development and progression of inflammatory and metabolic liver diseases. However, its role in hepatocellular carcinoma (HCC) remains unclear. In this study, we investigated the expression of key genes involved in the arachidonic acid metabolism pathway in HCC using a combination of bioinformatics, proteomics and immunohistochemistry analyses. Through a comprehensive analysis of publicly available datasets, clinical HCC tissues, and tissue microarrays, we compared the expression of hepatic arachidonic acid metabolic genes. We observed significant downregulation of cytochrome P450 (CYP450) pathway genes at both the messenger RNA and protein levels in HCC tissues compared to normal liver tissues. Furthermore, we observed a strong correlation between the deregulation of the arachidonic acid metabolism CYP450 pathway and the pathological features and prognosis of HCC. Specifically, the expression of CYP2C8/9/18/19 was significantly correlated with pathological grade (r = -.484, p < .0001), vascular invasion (r = -.402, p < .0001), aspartate transaminase (r = -.246, p = .025), gamma-glutamyl transpeptidase (r = -.252, p = .022), alkaline phosphatase (r = -.342, p = .002), alpha-fetoprotein (r = -.311, p = .004) and carbohydrate antigen 19-9 (r = -.227, p = .047). Moreover, we discovered a significant association between CYP450 pathway activity and vascular invasion in HCC. Collectively, these data indicate that arachidonic acid CYP450 metabolic pathway deregulation is implicated in HCC progression and may be a potential predictive factor for early recurrence in patients with HCC.

Overexpression of TRIM28 predicts an unfavorable prognosis and promotes the proliferation and migration of hepatocellular carcinoma

Abstract Objectives Previous studies have shown that tripartite motif-containing 28 (TRIM28) might be a latent target for cancer therapy. However, the detailed roles and mechanisms of TRIM28 in hepatocellular carcinoma (HCC) remain ambiguous. Methods We systematically analyzed TRIM28 mRNA expression and protein levels in HCC tissues based on large-scale data and publicly available immunohistochemistry images. We estimated the prognostic capacity of TRIM28 in HCC. Additionally, we performed gene enrichment, immune infiltration, and drug sensitivity analyses to further explore the roles of TRIM28 in HCC. To determine the effect of TRIM28 expression on HCC cell proliferation and migration, successful transfection of siRNAs was conducted in MHCC97-L and Huh7 cells, followed by cell functional assays. Results We verified the overexpression of TRIM28 in HCC at the mRNA and protein levels. The summary receiver operating characteristics curve with the area under curve of 0.84 (95 % CI: 0.81–0.87) indicated the high accuracy of increasing TRIM28 expression for discriminating HCC from non-HCC tissues. According to The Cancer Genome Atlas datasets, TRIM28 mRNA expression was significantly related to age, grade, stage, and pathologic T (p&lt;0.05). Increased TRIM28 expression levels were significant correlated to poor survival in HCC patients. An enrichment analysis suggested that TRIM28-reated genes primarily participated in the spliceosome signaling pathway, with hub genes including SNRPA1, SNRPF, SNRPD1, SF3B2, SNRPB, SNRPE, and EFTUD2. TRIM28 expression was correlated with the infiltration of five immune cells. Higher TRIM28 expression was linked to better sensitivity of tumor cells to pluripotin. Molecular docking showed that pluripotin could bind to TRIM28. Further, knockdown of TRIM28 inhibited the proliferation and migration of HCC cells. Conclusions TRIM28 is highly expressed in HCC and contribute to the proliferation and migration of HCC cells, leading to unfavorable outcomes. These findings indicate TRIM28 promise as a novel prognostic indicator.

SNRPD1 inhibition suppresses the proliferation of hepatocellular carcinoma and promotes autophagy through the PI3K/AKT/mTOR/4EBP1 pathway

BackgroundSmall nuclear ribonucleoprotein Sm D1 (SNRPD1) has been reported as an oncogene in some solid cancers. Our previous study suggested that SNRPD1 has diagnostic and prognostic value in hepatocellular carcinoma (HCC), but its role in tumor growth and biological behavior remains unknown. In this study, we aimed to unravel the role and mechanism of SNRPD1 in HCC. MethodsWe investigated the SNRPD1 mRNA level in adjacent normal liver tissues and HCC tissues with different tumor stages in the UALCAN database. The associations between SNRPD1 mRNA expression and HCC prognosis were investigated in TCGA database. Then, 52 pairs of frozen HCC tissues and corresponding adjacent normal liver tissues were collected to perform qPCR and immunohistochemistry assay. Next, we carried out a series of experiments in vitro and in vivo to investigate the effects of SNRPD1 expression on cell invasion, migration, proliferation, autophagy, and the PI3K/AKT/mTOR pathway. ResultsThe bioinformatics analysis and qPCR in our patient cohort demonstrated that the SNRPD1 mRNA level in HCC tissues was higher than in adjacent normal tissues. In addition, the immunohistochemistry assay exhibited an increased SNRPD1 protein level with the tumor stage increase. Survival analysis suggested that higher expression of SNRPD1 was significantly associated with unfavorable prognosis of patients with HCC. The functional experiments in vitro indicated that SNRPD1 knockdown suppressed the cellular proliferation, migration, and invasion capacities. Furthermore, SNRPD1 inhibition induced cellular apoptosis and arrested the HCC cells at the G0/G1 phase of the cell cycle. Mechanistic analyses demonstrated that SNRPD1 knockdown induced the increase of autophagic vacuoles and the expression of autophagy-related genes (ATG5, ATG7, and ATG12) and blocked the PI3K/AKT/mTOR/4EBP1 pathway in vitro. Moreover, SNRPD1 inhibition suppressed tumor growth and expression of the Ki67 protein in vivo. ConclusionsSNRPD1 may serve as an oncogene in HCC and promote tumor proliferation via inhibiting autophagy induced through the PI3K/Akt/mTOR/4EBP1 pathway.

ASAP2 interrupts c-MET-CIN85 interaction to sustain HGF/c-MET-induced malignant potentials in hepatocellular carcinoma

BackgroundSustained activation of hepatocyte growth factor (HGF)/c-MET signaling is a major driver of hepatocellular carcinoma (HCC) progression, but underlying mechanism is unclear. ArfGAP With SH3 Domain, Ankyrin Repeat And PH Domain 2 (ASAP2) can reportedly activate GTPases and promote receptor tyrosine kinase signaling. However, the exact role of ASAP2 in HCC, especially for c-MET activation, also remains elusive.MethodsASAP2 expression levels in HCC tissues and cells were quantified using qRT-PCR, western blot (WB) analysis, and immunohistochemistry staining. Cell counting kit-8 (CCK-8) and colony formation assays were performed to evaluate cell proliferation rates. Flow cytometry assays were conducted to assess apoptosis rates. Wound healing and Transwell assays were performed to determine cell migration and invasion capacities. Epithelial-mesenchymal transition (EMT)-related marker expression levels were also examined. Subcutaneous implantation and tail vein injection models were applied for in vivo growth and metastasis evaluations, respectively. Bioinformatics analyses of The Cancer Genome Atlas and STRING datasets were performed to explore ASAP2 downstream signaling. Co-immunoprecipitation and Cycloheximide chasing experiments were performed to assess protein–protein interactions and protein half-life, respectively.ResultsASAP2 had higher expression levels in HCC tissues than in normal liver, and also predicted poor prognosis. Knocking down ASAP2 significantly impaired cell proliferation, migration, and invasion capacities, but promoted apoptosis in HCC cells in vitro. However, overexpression of ASAP2 achieved the opposite effects. In vivo experiments confirmed that ASAP2 could promote HCC cell growth and facilitate lung metastasis. Interestingly, ASAP2 was essential for triggering EMT. Gene Set Enrichment Analysis demonstrated that c-MET signaling was greatly enriched in ASAP2-high HCC cases. Additionally, c-MET signaling activity was significantly decreased following ASAP knockdown, evidenced by reduced c-MET, p-AKT, and p-ERK1/2 protein levels. Importantly, ASAP2 knockdown effectively attenuated HGF/c-MET signaling-induced malignant phenotypes. c-MET and ASAP2 expression levels were positively correlated in our cohort. Mechanistically, ASAP2 can directly bind to CIN85, thereby disrupting its interaction with c-MET, and can thus antagonize CIN85-induced c-MET internalization and lysosome-mediated degradation. Notably, knocking down CIN85 can rescue the observed inhibitory effects caused by ASAP2 knockdown.ConclusionsThis study highlights the importance of ASAP2 in sustaining c-MET signaling, which can facilitate HCC progression.

CISD2 protects against Erastin induced hepatocellular carcinoma ferroptosis by upregulating FSP1

Abstract Objectives CDGSH iron sulfur domain 2 (CISD2) is essential to maintain iron (Fe) and reactive oxygen species (ROS) homeostasis, and ferroptosis suppressor protein 1 (FSP1) can protect cells from ferroptosis by inhibiting lipid peroxidation. Here, we investigate the role and potential mechanism of CISD2 and FSP1 in ferroptosis of hepatocellular carcinoma (HCC). Methods Human HCC cells were exposed to ferroptosis inducer Erastin, and the expression changes of CISD2 and FSP1 during ferroptosis were detected. Subsequently, we investigated the effect of overexpression of CISD2 on ferroptosis and FSP1 expression in HCC cells. Finally, we also investigated the effect of overexpression of FSP1 on ferroptosis in HCC cells. Results Erastin induced ferroptosis in hepatoma cells, and HepG2 cells were sensitive to Erastin. In addition, it was found that the expression of CISD2 was significantly upregulated and the expression of FSP1 was significantly downregulated in Erastin treated HepG2 cells. Subsequently, CISD2 was found to be highly expressed in HCC tissues, and overexpression of CISD2 reversed ferroptosis induced by Erastin in HepG2 cells and upregulated the expression of FSP1. Meanwhile, FSP1 showed a low expression level in HCC tissues and cells, and overexpression of FSP1 could reverse the ferroptosis induced by Erastin in HepG2 cells. Conclusion CISD2 and FSP1 are involved in the ferroptosis process of HCC induced by Erastin. CISD2 protects against the ferroptosis of HCC induced by Erastin by upregulating the expression of FSP1.

Six immune-related promising biomarkers may promote hepatocellular carcinoma prognosis: a bioinformatics analysis and experimental validation

BackgroundAbnormal miRNA and mRNA expression and dysregulated immune microenvironment have been found to frequently induce the progression of hepatocellular carcinoma (HCC) in recent reports. In particular, the immune-related competing endogenous RNAs (ceRNA) mechanism plays a crucial role in HCC progression. However, the underlying mechanisms remain unclear.MethodsDifferentially expressed immune-related genes were obtained from the Immport, GEO, and TCGA databases. The mRNA and protein expression levels in HCC tissues and adjacent normal tissues were confirmed, and we further investigated the methylation levels of these biomarkers to explore their function. Then, the TIMER and TISCH databases were used to assess the relationship between immune infiltration and hub genes. Survival analysis and univariate and multivariate Cox models were used to evaluate the association between hub genes and HCC diagnosis. Hub gene expression was experimentally validated in six HCC cell lines and 15 HCC samples using qRT-PCR and immunohistochemistry. The hub genes were uploaded to DSigDB for drug prediction enrichment analysis.ResultsWe identified that patients with abnormal miRNAs (hsa-miR-125b-5p and hsa-miR-21-5p) and their targeted genes (NTF3, PSMD14, CD320, and SORT1) had a worse prognosis. Methylation analysis of miRNA-targeted genes suggested that alteration of methylation levels is also a factor in the induction of tumorigenesis. We also found that the development of HCC progression caused by miRNA-mRNA interactions may be closely correlated with the infiltration of immunocytes. Moreover, the GSEA, GO, and KEGG analysis suggested that several common immune-related biological processes and pathways were related to miRNA-targeted genes. The results of qRT-PCR, immunohistochemistry, and western blotting were consistent with our bioinformatics results, suggesting that abnormal miRNAs and their targeted genes may affect HCC progression.ConclusionsBriefly, our study systematically describes the mechanisms of miRNA-mRNA interactions in HCC and predicts promising biomarkers that are associated with immune filtration for HCC progression.