Hepatocellular Carcinoma Migration Research Articles

PCDH17 is regulated by methylation of DNMT3B and affects the malignant biological behavior of HCC through EMT

Hepatocellular carcinoma (HCC) is one of the malignant tumors most frequently encountered in the clinic. Studies have shown that the abnormal expression of various genes leads to the malignant progression of tumors, and the modification in DNA methylation can cause a change in gene expression. Increasing evidence has shown that abnormal expression of PCDH17 is found in many human cancers. However, its functional role in HCC remains unexplored. Herein, we found that PCDH17 was expressed at low levels in HCC tissues and cell lines. There is a significant correlation between low expression of PCDH17 and poor prognosis of HCC patients. Increased expression of PCDH17 significantly suppressed cell proliferation, migration and invasion in HCC. The low expression of PCDH17 was due to its high DNA methylation level, and changing the expression of DNMT3B significantly affect the DNA methylation level of PCDH17 and increase its protein expression. Furthermore, methylation of PCDH17 regulated by DNMT3B affects the malignant biological behavior of HCC through EMT. In conclusion, PCDH17 participates in malignant biological behavior of HCC and that DNMT3B plays an important role in the regulation of PCDH17 methylation, which affects the malignant progression of HCC.

ARPC2: A Pan-Cancer Prognostic and Immunological Biomarker That Promotes Hepatocellular Carcinoma Cell Proliferation and Invasion.

Background: Actin-related protein 2/3 complex subunit 2 (ARPC2) plays a fundamental role in actin filament nucleation and is critical for tumor cell migration and invasion. However, its abnormal expression, clinical significance, and biological function in human pan-cancer have been poorly explored. Thus, we focused on ARPC2 as an entry point for identifying novel pan-cancer prognostic biomarkers. Methods: The Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) databases were used to assess the differential expression of ARPC2 in pan-cancer. The Human Protein Atlas was used for the tissue/cell-specific expression analysis of ARPC2. The genetic alteration information of ARPC2 was obtained from the cBioPortal database and the GSCALite platform. The prognostic value of ARPC2 was explored in pan-cancer using Cox regression and Kaplan–Meier analyses. Spearman correlation analysis was performed to investigate the relationship between ARPC2 expression and tumor mutational burden (TMB), DNA methyltransferases, microsatellite instability (MSI), immune-related genes, and mismatch repairs (MMRs). The ESTIMATE and CIBERSORT algorithms were used to evaluate the association between ARPC2 expression and the tumor microenvironment (TME) and immune infiltrating cells. We also conducted differential expression analysis of ARPC2 in hepatocellular carcinoma (HCC) tissues and cell lines using qPCR, western blotting, and immunohistochemistry and explored its role in tumor proliferation, migration, and invasion of HCC cells. Results: ARPC2 expression was significantly upregulated in multiple tumor types and significantly correlated with worse prognosis and higher clinicopathological stage. Genetic alterations and DNA methylation in tumor tissues may contribute to the aberrant expression of ARPC2. ARPC2 expression was significantly correlated with the tumor microenvironment (TME), infiltrating immune cells, TMB, microsatellite instability (MSI), and immune checkpoint-related genes in certain cancer types. In this experimental study, we found that the expression of ARPC2 was dramatically upregulated in HCC tissues and cell lines compared to adjacent liver tissues and normal liver cell lines. Functionally, ARPC2 silencing in HCC cells significantly inhibited cell proliferation, migration, and invasion, while the overexpression of ARPC2 promotes tumor proliferation, migration, and invasion. Conclusion: ARPC2 is a promising prognostic and immunological biomarker for multiple tumor types and is likely to play an important role in HCC progression and metastasis.

METTL16 promotes hepatocellular carcinoma progression through downregulating RAB11B-AS1 in an m6A-dependent manner

BackgroundThe molecular mechanisms driving hepatocellular carcinoma (HCC) remain largely unclear. As one of the major epitranscriptomic modifications, N6-methyladenosine (m6A) plays key roles in HCC. The aim of this study was to investigate the expression, roles, and mechanisms of action of the RNA methyltransferase methyltransferase-like protein 16 (METTL16) in HCC.MethodsThe expression of METTL16 and RAB11B-AS1 was determined by RT-qPCR. The regulation of RAB11B-AS1 by METTL16 was investigated by RNA immunoprecipitation (RIP), methylated RIP (MeRIP), and RNA stability assays. In vitro and in vivo gain- and loss-of-function assays were performed to investigate the roles of METTL16 and RAB11B-AS1.ResultsMETTL16 was upregulated in HCC, and its increased expression was correlated with poor prognosis of HCC patients. METTL16 promoted HCC cellular proliferation, migration, and invasion, repressed HCC cellular apoptosis, and promoted HCC tumoral growth in vivo. METTL16 directly bound long noncoding RNA (lncRNA) RAB11B-AS1, induced m6A modification of RAB11B-AS1, and decreased the stability of RAB11B-AS1 transcript, leading to the downregulation of RAB11B-AS1. Conversely to METTL16, RAB11B-AS1 is downregulated in HCC, and its decreased expression was correlated with poor prognosis of patients with HCC. Furthermore, the expression of RAB11B-AS1 was negatively correlated with METTL16 in HCC tissues. RAB11B-AS1 repressed HCC cellular proliferation, migration, and invasion, promoted HCC cellular apoptosis, and inhibited HCC tumoral growth in vivo. Functional rescue assays revealed that overexpression of RAB11B-AS1 reversed the oncogenic roles of METTL16 in HCC.ConclusionsThis study identified the METTL16/RAB11B-AS1 regulatory axis in HCC, which represented novel targets for HCC prognosis and treatment.

Upregulation of miR-520c-3p via hepatitis B virus drives hepatocellular migration and invasion by the PTEN/AKT/NF-κB axis

Hepatitis B virus (HBV) is a major risk factor for the development and progression of hepatocellular carcinoma (HCC). It has been reported that viral infection can interfere with the expression of cellular microRNA (miRNA) to affect oncogenesis. In this study, we showed that miR-520c-3p was upregulated in liver tumor specimens, and we revealed that HBV infection enhanced the expression of miR-520c-3p through the interaction of viral protein HBV X protein (HBx) with transcription factor CREB1. We further showed that miR-520c-3p induced by HBV transfection/infection caused epithelial-mesenchymal transition (EMT). Using the miRNA target prediction database miRBase and luciferase reporter assays, we identified PTEN as a novel target gene of miR-520c-3p and miR-520c-3p directly targeted PTEN’s 3′-untranslated region. Moreover, we discovered that HBV promoted EMT via the miR-520c-3p-PTEN to activate AKT-NFκB signaling pathway, leading to increased HCC migration and invasion. Importantly, miR-520c-3p antagomir significantly represses invasiveness in HBx-induced hepatocellular xenograft models. Our findings indicate that miR-520c-3p is a novel regulator of HBV and plays an important role in HCC progression. It may serve as a new biomarker and molecular therapeutic target for HBV patients.

MiR-129-5p exerts Wnt signaling-dependent tumor-suppressive functions in hepatocellular carcinoma by directly targeting hepatoma-derived growth factor HDGF

BackgroundIn hepatocellular carcinoma (HCC), histone deacetylases (HDACs) are frequently overexpressed. This results in chromatin compaction and silencing of tumor-relevant genes and microRNAs. Modulation of microRNA expression is a potential treatment option for HCC. Therefore, we aimed to characterize the epigenetically regulated miR-129-5p regarding its functional effects and target genes to understand its relevance for HCC tumorigenesis.MethodsGlobal miRNA expression of HCC cell lines (HLE, HLF, Huh7, HepG2, Hep3B) and normal liver cell lines (THLE-2, THLE-3) was analyzed after HDAC inhibition by miRNA sequencing. An in vivo xenograft mouse model and in vitro assays were used to investigate tumor-relevant functional effects following miR-129-5p transfection of HCC cells. To validate hepatoma-derived growth factor (HDGF) as a direct target gene of miR-129-5p, luciferase reporter assays were performed. Survival data and HDGF expression were analyzed in public HCC datasets. After siRNA-mediated knockdown of HDGF, its cancer-related functions were examined.ResultsHDAC inhibition induced the expression of miR-129-5p. Transfection of miR-129-5p increased the apoptosis of HCC cells, decreased proliferation, migration and ERK signaling in vitro and inhibited tumor growth in vivo. Direct binding of miR-129-5p to the 3′UTR of HDGF via a noncanonical binding site was validated by luciferase reporter assays. HDGF knockdown reduced cell viability and migration and increased apoptosis in Wnt-inactive HCC cells. These in vitro results were in line with the analysis of public HCC datasets showing that HDGF overexpression correlated with a worse survival prognosis, primarily in Wnt-inactive HCCs.ConclusionsThis study provides detailed insights into the regulatory network of the tumor-suppressive, epigenetically regulated miR-129-5p in HCC. Our results reveal for the first time that the therapeutic application of mir-129-5p may have significant implications for the personalized treatment of patients with Wnt-inactive, advanced HCC by directly regulating HDGF. Therefore, miR-129-5p is a promising candidate for a microRNA replacement therapy to prevent HCC progression and tumor metastasis.

Nascent Proteome and Glycoproteome Reveal the Inhibition Role of ALG1 in Hepatocellular Carcinoma Cell Migration.

The online version contains supplementary material available at 10.1007/s43657-022-00050-5.

A Novel and Robust Prognostic Model for Hepatocellular Carcinoma Based on Enhancer RNAs-Regulated Genes.

Evidence has demonstrated that enhancer RNAs (eRNAs) play a vital role in the progression and prognosis of cancers, but few studies have focused on the prognostic ability of eRNA-regulated genes (eRGs) for hepatocellular carcinoma (HCC). Using gene expression profiles of HCC patients from the TCGA-LIHC and eRNA expression profiles from the enhancer RNA in cancers (eRic) data portal, we developed a novel and robust prognostic signature composed of 10 eRGs based on Lasso-penalized Cox regression analysis. According to the signature, HCC patients were stratified into high- and low-risk groups, which have been shown to have significant differences in tumor immune microenvironment, immune checkpoints, HLA-related genes, DNA damage repair-related genes, Gene-set variation analysis (GSVA), and the lower half-maximal inhibitory concentration (IC50) of Sorafenib. The prognostic nomogram combining the signature, age, and TNM stage had good predictive ability in the training set (TCGA-LIHC) with the concordance index (C-index) of 0.73 and the AUCs for 1-, 3-, and 5-year OS of 0.82, 0.77, 0.74, respectively. In external validation set (GSE14520), the nomogram also performed well with the C-index of 0.71 and the AUCs for 1-, 3-, and 5-year OS of 0.74, 0.77, 0.74, respectively. In addition, an important eRG (AKR1C3) was validated using two HCC cell lines (Huh7 and MHCC-LM3) in vitro, and the results demonstrated the overexpression of AKR1C3 is related to cell proliferation, migration, and invasion in HCC. Altogether, our eRGs signature and nomogram can predict prognosis accurately and conveniently, facilitate individualized treatment, and improve prognosis for HCC patients.

Epigenetic regulation of the DNMT1/MT1G/KLF4/CA9 axis synergises the anticancer effects of sorafenib in hepatocellular carcinoma

Sorafenib, a multikinase inhibitor, has been widely used as a first-line anticancer drug for advanced hepatocellular carcinoma (HCC). However, the development of drug resistance to sorafenib is frequently observed in clinical applications. Potential nonkinase targets of sorafenib have not been well documented and may provide insights into reversing drug resistance and enhancing drug efficacy. Herein, we report that sorafenib exerts its anticancer effects by activating metallothionein 1 G (MT1G) expression. MT1G is a novel marker in HCC that correlates well with patient survival. MT1G overexpression suppressed the cellular proliferation, migration, invasion, and tumour formation of HCC and sensitised cells to sorafenib treatment. However, the disruption of MT1G attenuated the anticancer effects of sorafenib. Mechanistically, sorafenib upregulated MT1G expression via hypomethylation of its promoter region by binding and inhibiting DNA methyltransferase 1 (DNMT1) and increasing its promoter accessibility in HCC cells. Activation of MT1G also inhibited CA9 transcription through the suppression of HIF1A as mediated by KLF4. Our collective data revealed that sorafenib exerts its anticancer effects through epigenetic regulation of the DNMT1/MT1G/KLF4/CA9 axis in HCC and the activation of MT1G might constitute a strategy for enhancing the effect of sorafenib to suppress HCC cells.

Omega-3 Polyunsaturated Fatty Acids Provoke Apoptosis in Hepatocellular Carcinoma through Knocking Down the STAT3 Activated Signaling Pathway: In Vivo and In Vitro Study.

Hepatocellular carcinoma (HCC) is a common type of liver cancer and is a leading cause of death worldwide. Signal transducer and activator of transcription 3 (STAT3) is involved in HCC progression, migration, and suppression of apoptosis. This study investigates the apoptotic effect of the dietary antioxidant (n-3 PUFAs) on HepG2 cells and analyzes the underlying molecular mechanisms of this effect both in vivo and in vitro. In vivo study: Seventy-five adult male albino rats were divided into three groups (n = 25): Group I (control): 0.9% normal saline, intraperitoneal. Group II: N-Nitrosodiethylamine (200 mg/kg b.wt) intraperitoneal, followed by phenobarbital 0.05% in drinking water. Group III: as group II followed by n-3 PUFAs intubation (400 mg/kg/day). In vivo study: liver specimens for biochemical, histopathological, and immunohistochemical examination. In vitro study: MTT assay, cell morphology, PCR, Western blot, and immunohistochemical analysis. n-3 PUFAs significantly improved the histopathologic features of HCC and decreased the expression of anti-apoptotic proteins. Further, HepG2 cells proliferation was suppressed through inhibition of the STAT3 signaling pathway, cyclin D1, and Bcl-2 activity. Here we report that n-3 PUFAs may be an ideal cancer chemo-preventive candidate by targeting STAT3 signaling, which is involved in cell proliferation and apoptosis.

Exosomal miR-142-3p secreted by hepatitis B virus (HBV)-hepatocellular carcinoma (HCC) cells promotes ferroptosis of M1-type macrophages through SLC3A2 and the mechanism of HCC progression.

Most patients with hepatitis B virus (HBV) infection will develop hepatocellular carcinoma (HCC). This study aimed to explore the potential mechanism of miR-142-3p in HCC caused by HBV infection. HepG2 cells and M1 macrophages were cocultured and then infected with HBV to establish an in vitro model. MicroRNA (miRNA) and messenger RNA (mRNA) expression was analyzed by quantitative reverse transcription polymerase chain reaction (RT-qPCR) and Western blot. The protein expressions of COX2, ACSL4, PTGS2, GPX4, and NOX1 were analyzed by Western blot. Flow cytometry and TUNEL assays were used to assess cell reactive oxygen species (ROS) and ferroptosis, respectively. Cell invasion and migration were measured by Transwell assay. To evaluate the ferroptosis of M1-type macrophages, glutathione (GSH), malondialdehyde (MDA), and Fe2+ content was detected by corresponding kits. Dual luciferase reporter gene detection verified the targeting relationship between miR-142-3p and SLC3A2. MiR-142-3p was highly expressed in HBV-infected HCC patients and HBV-infected M1-type macrophages. Inhibition of miR-142-3p or overexpression of SLC3A2 reversed ferroptosis and inhibited the proliferation, migration, and invasion of HCC cells. Our findings indicated that miR-142-3p promoted HBV-infected M1-type macrophage ferroptosis through SLC3A2, affecting the production of GSH, MDA, and Fe2+ and accelerating the development of HCC. The regulation of miR-142-3p and its target genes will help to clarify the pathogenesis of HCC induced by HBV infection and provide new theoretical foundations and therapeutic targets.

LncRNA NEAT1 sponges miR-214 to promoted tumor growth in hepatocellular carcinoma.

Live cancer is the sixth most prevalent diagnosed malignant tumor and the fourth leading cause of cancer-related deaths worldwide. Hepatocellular carcinoma (HCC) is the main histological type of liver cancer. Here, we attempt to evaluate the role of long non coding RNA NEAT1 in HCC, and explore its potential mechanism in this disease. Initially, we detected the expression of NEAT1 in HCC cell lines (SMMC-7721 and Huh7 cells) using qRT-PCR. Then we transfected si-NC or si-NEAT1 into SMMC-7721 and Huh7 cells by RNA interference. CCK-8 assay, transwell assay, flow cytometry, qRT-PCR and western blotting were used to evaluate the role of NEAT1 in the biological behavior of SMMC-7721 and Huh7 cells. The rescue experiment, RIP assay and MeRIP were devoted to the underlying mechanism. NEAT1 expression level was significantly elevated in SMMC-7721 and Huh7 cells. Knockdown of NEAT1 inhibited proliferation and migration, induced apoptosis of HCC cell lines. NEAT1 serves as a sponge for miR-214. Besides, PSMB8 was a direct target of miR-214. Furthermore, ALKBH5 could up-regulate NEAT1 expression by inhibiting m6A enrichment. ALKBH5-induced NEAT1 promoted cell proliferation and migration of HCC by sponging miR-214 in vitro, which may provide a potential therapeutic target for HCC.

Cinobufotalin inhibits the epithelial-mesenchymal transition of hepatocellular carcinoma cells through down-regulate β-catenin in vitro and in vivo

Hepatocellular carcinoma (HCC) is one of the malignant tumors with high incidence and mortality. The prognosis of HCC is poor due to the high postoperative recurrence rate and metastasis rate. Epithelial-mesenchymal transition (EMT) plays a key role in the metastasis of HCC, which is closely related to the invasion, intrahepatic metastasis and low survival rate. Here we demonstrated that cinobufotalin can upregulate epithelial markers (E-cadherin) and downregulate mesenchymal markers (N-cadherin, snail, slug and ZEB1) in HepG2, SMMC-7721 and SNU-368 cells. We further found that the mRNA and protein expression of β-catenin and its target genes (i.e. MMP7 and DKK1), which are related to tumor invasion and metastasis, were decreased after cinobufotalin treatment. Overexpression of β-catenin promoted EMT of HepG2 and SMMC-7721 cells, and cinobufotalin could antagonize this induction. While Knockdown of β-catenin could inhibit EMT and cinobufotalin enhanced this inhibition. In addition, cinobufotalin significantly suppressed the tumor EMT, as demonstrated by increased E-cadherin expression and decreased N-cadherin and vimentin expression, and inhibited formation and metastasis of lung metastases in vivo. In conclusion, our study has revealed a novel anticancer mechanism of cinobufotalin, which inhibits EMT progress by downregulating β-catenin, and then prevents the migration and invasion of HCC. These results provide convincing evidence for the development of cinobufotalin as a potential HCC metastasis inhibitor.

Roles of lncRNAs Mediating Wnt/β-Catenin Signaling in HCC.

Hepatocellular carcinoma (HCC) is considered the second most deadly cancer worldwide. Due to the absence of early diagnostic markers and effective therapeutic approaches, distant metastasis and increasing recurrence rates are major difficulties in the clinical treatment of HCC. Further understanding of its pathogenesis has become an urgent goal in HCC research. Recently, abnormal expression of long noncoding RNAs (lncRNAs) was identified as a vital regulator involved in the initiation and development of HCC. Activation of the Wnt/β-catenin pathway has been reported to obviously impact cell proliferation, invasion, and migration of HCC. This article reviews specific interactions, significant mechanisms and molecules related to HCC initiation and progression to provide promising strategies for treatment.

C-Met up-regulates the expression of PD-L1 through MAPK/NF-κBp65 pathway.

Sorafenib acquired drug resistance during the treatment of hepatocellular carcinoma (HCC) reduces the efficacy of the drug. The immune escape effect induced by PD-L1 is largely associated with drug resistance of HCC. However, the regulated mechanism of PD-L1 is unclear. This research aimed to clarify the control mechanism of PD-L1. c-Met was found abnormally highly expressed in Huh-7SR with high PD-L1 expression. In addition, c-Met, as the upstream target molecule of PD-L1, promoted the proliferation and migration of HCC in vitro and in vivo. We also found that c-Met activated the MAPK signaling pathway and the downstream NF-κBp65 transcription factor, which interacts with the proximal region of the PD-L1 promoter to promote PD-L1 expression. In conclusion, c-Met regulates the transcription of PD-L1 through the MAPK/NF-κBp65 pathway, thereby promoting the progress of HCC. The role of c-Met and PD-L1 in HCC needs to be further studied, but it is a potential target for the treatment of HCC. KEY MESSAGES: In the study, it was found that c-Met is also abnormally highly expressed in Huh-7SR with high PD-L1 expression and can promote the development of HCC in vitro and in vivo. PD-L1 and c-Met expression levels are positively correlated. In the follow-up mechanism study, we found that c-Met activated the MAPK signaling pathway and subsequently activated the downstream NF-κBp65 transcription factor, which interacts with the proximal region of the PD-L1 promoter to promote PD-L1 expression. Our study found that c-Met regulates the transcription of PD-L1 through the MAPK/NF-κBp65 pathway, thereby promoting the progress of HCC.

Long non-coding RNA PAARH promotes hepatocellular carcinoma progression and angiogenesis via upregulating HOTTIP and activating HIF-1\u03b1/VEGF signaling

Hepatocellular carcinoma (HCC) is one of the leading lethal malignancies and a hypervascular tumor. Although some long non-coding RNAs (lncRNAs) have been revealed to be involved in HCC. The contributions of lncRNAs to HCC progression and angiogenesis are still largely unknown. In this study, we identified a HCC-related lncRNA, CMB9-22P13.1, which was highly expressed and correlated with advanced stage, vascular invasion, and poor survival in HCC. We named this lncRNA Progression and Angiogenesis Associated RNA in HCC (PAARH). Gain- and loss-of function assays revealed that PAARH facilitated HCC cellular growth, migration, and invasion, repressed HCC cellular apoptosis, and promoted HCC tumor growth and angiogenesis in vivo. PAARH functioned as a competing endogenous RNA to upregulate HOTTIP via sponging miR-6760-5p, miR-6512-3p, miR-1298-5p, miR-6720-5p, miR-4516, and miR-6782-5p. The expression of PAARH was significantly positively associated with HOTTIP in HCC tissues. Functional rescue assays verified that HOTTIP was a critical mediator of the roles of PAARH in modulating HCC cellular growth, apoptosis, migration, and invasion. Furthermore, PAARH was found to physically bind hypoxia inducible factor-1 subunit alpha (HIF-1α), facilitate the recruitment of HIF-1α to VEGF promoter, and activate VEGF expression under hypoxia, which was responsible for the roles of PAARH in promoting angiogenesis. The expression of PAARH was positively associated with VEGF expression and microvessel density in HCC tissues. In conclusion, these findings demonstrated that PAARH promoted HCC progression and angiogenesis via upregulating HOTTIP and activating HIF-1α/VEGF signaling. PAARH represents a potential prognostic biomarker and therapeutic target for HCC.

Prevention and Treatment of Hepatocellular Carcinoma Using miRNAs.

Hepatocellular carcinoma (HCC) is the second leading cause of death due to cancer. Liver transplantation, surgical liver resection, chemotherapy, and radiotherapy are the main options for the treatment of HCC. However, these methods are unable to limit the growth, survival, and metastasis of HCC cells. Several signaling pathways control propagation, metastasis, and recurrence of HCC. Recent studies have established new approaches for the prevention and treatment of HCC using miRNA technology. MicroRNAs are a class of non-coding RNAs with an average of 22 nucleotides that play critical roles in controlling gene expression in a variety of biological processes. miRNAs can induce or suppress HCC proliferation, migration, metastasis, and tumorigenesis. The anti-cancer effects of molecular agents can be evaluated directly in animal models or indirectly through the injection of HCC cell lines treated with anti-cancer agents. Targeting cancer-specific signaling pathways with miRNAs can be novel therapeutic strategies against HCC. This study provides the latest findings on using miRNAs in the control of HCC in both in vitro and in vivo models.

ACADL Functions as a Tumor Suppressor in Hepatocellular Carcinoma Metastasis by Inhibiting Matrix Metalloproteinase 14.

High aggressiveness is the main reason for the poor prognosis of hepatocellular carcinoma (HCC) patients. However, its molecular mechanisms still remain largely unexplored. ACADL, a mitochondrial enzyme that facilitates the primary regulated step in mitochondrial fatty acid oxidation, plays a role in HCC growth inhibition. However, the function of ACADL in tumor metastasis is not well elucidated. We found that the reduced expression of ACADL is closely associated with the loss of tumor encapsulation, extrahepatic metastasis, and poor prognosis in HCC patients. Upregulation of ACADL significantly inhibited HCC migration and invasion ability. Whereas knockdown of ACADL markedly enhanced cell invasive capability. Expression of matrix metalloproteinase-14 (MMP14) was negatively associated with the content of ACADL in HCC specimens. MMP14-positive patients with a low expression of ACADL showed worse outcome. Treatment with MMP14 agonist reversed the inhibitory effect of ACADL on HCC metastasis. In addition, ACADL negatively regulated MMP14 expression by inhibiting the STAT3 signaling pathway, as the sustained activation of STAT3 effectively restored the level of MMP14 in ACADL-overexpressed cells. Collectively, these findings disclose that ACADL represses HCC metastasis via STAT3-MMP14 pathway. This study may propose a promising strategy for the precise treatment of metastatic HCC patients.

MicroRNA-377-3p inhibits hepatocellular carcinoma growth and metastasis through negative regulation of CPT1C-mediated fatty acid oxidation

BackgroundAltered lipid metabolism is closely related to the occurrence and development of hepatocellular carcinoma (HCC). Carnitine palmitoyltransferase 1C (CPT1C) is a member of CPT1 family and plays a key role in cancer development and progression. However, how microRNAs (miRNAs) regulate CPT1C-mediated fatty acid transport and oxidation remains to be elucidated.MethodsOil Red O staining, mitochondrial, and lipid droplets immunofluorescence staining were used to detect the functions of miR-377-3p and CPT1C in fatty acid oxidation. Colocalization of palmitate and mitochondria was performed to investigate the function of miR-377-3p and CPT1C in fatty acid transport into mitochondria. Fatty acid oxidation (FAO) assay was used to detect the function of miR-377-3p and CPT1C in FAO. Cell proliferation, migration and invasion assays and animal experiments were used to evaluate the role of miR-377-3p/CPT1C axis in HCC progression in vitro and in vivo. Immunofluorescence staining was used to identify the clinical significance of miR-377-3p and CPT1C in HCC patients.ResultsMiR-377-3p inhibits CPT1C expression by targeting its 3’-untranslated region. Through repression of CPT1C, miR-377-3p suppresses fatty acid oxidation by preventing fatty acid from entering into mitochondria and decreasing ATP production in HCC cells. Inhibiting fatty acid oxidation abolishes the ability of miR-377-3p/CPT1C axis to regulate HCC proliferation, migration, invasion and metastasis in vitro and in vivo. In HCC patients, CPT1C is significantly upregulated, and miR-377-3p expression and lipid droplets are negatively correlated with CPT1C expression. High expression of miR-377-3p and CPT1C predict better and worse clinical outcomes, respectively.ConclusionsWe uncover the key function and the relevant mechanisms of the miR-377-3p/CPT1C axis in HCC, which might provide a potential target for the treatment of HCC.

Circular RNA circ_0008934 promotes hepatocellular carcinoma growth and metastasis through modulating miR-1305/TMTC3 axis.

Circular RNAs (circRNAs) play important roles in the progression of hepatocellular carcinoma (HCC). However, the exact function of circ_0008934 in HCC is unknown. Our study aimed to investigate the expression characteristics of circ_0008934 in HCC and its effects on the proliferation and metastasis of HCC, and to explore the potential mechanism. In this study, circ_0008934 expression was found to be significantly upregulated in HCC tissues and cell lines by qRT-PCR. High level of circ_0008934 is closely associated with higher serum AFP (P < 0.001), larger tumor diameter (P = 0.012), microvascular invasion (P = 0.008) and poorer prognosis (P = 0.007) of HCC patients. Functionally, knockdown of circ_0008934 inhibited HCC cell proliferation, invasion and migration in vitro and vivo. Mechanically, circ_0008934 was a sponge of miR-1305 to facilitate the TMTC3 expression, and the TMTC3 expression in HCC tissues was negatively associated with the survival of HCC patients. Furthermore, rescued assays revealed that the circ_0008934 facilitated HCC proliferation, invasion and migration by regulating miR-1305/ TMTC3 signaling pathways. Overall, these results demonstrate that downregulation of circ_0008934 repress HCC growth and metastasis by upregulating miR-1305 to inhibit TMTC3, suggesting circ_0008934/ miR-1305/ TMTC3 regulatory axis may be a possible novel therapeutic target for HCC.

Circular BANP knockdown inhibits the malignant progression of residual hepatocellular carcinoma after insufficient radiofrequency ablation.

Background:Circular RNAs (circRNAs) are endogenous non-coding RNAs, some of which have pathological roles. The current study aimed to explore the role of circRNA BTG3-associated nuclear protein (circ-BANP) binding with let-7f-5p and its regulation of the toll-like receptor 4 (TLR4)/signal transducer and activator of transcription 3 (STAT3) signaling pathway in residual hepatocellular carcinoma (HCC) after insufficient radiofrequency ablation (RFA).Methods:Circ-BANP, let-7f-5p, and TLR4 expressions in HCC samples were assessed using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blotting. Bioinformatics prediction, RNA pull-down assay, and dual luciferase reporter gene assay were used to analyze the relationships among circ-BANP, let-7f-5p, and TLR4. Huh7 cells were used to generate an in vitro model of residual HCC, defined as Huh7-H cells, which were transfected with either a plasmid or the sequence of circ-BANP, let-7f-5p, or TLR4. Expression of circ-BANP, let-7f-5p, and TLR4 mRNA was determined by RT-qPCR. TLR4, STAT3, p-STAT3, vascular endothelial growth factor A, vascular endothelial growth factor receptor-2, and epithelial-mesenchymal transformation (EMT)-related factors proteins were determined by Western blotting. Cell proliferation was determined by cell counting kit-8 and 5-Ethynyl-2’-deoxyuridine (EdU) assay and cell migration and invasion by Transwell assay. Animal studies were performed by inducing xenograft tumors in nude mice.Results:Circ-BANP and TLR4 mRNAs were upregulated in HCC tissues (the fold change for circ-BANP was 1.958 and that for TLR4 was 1.736 relative to para-tumors) and expression further increased following insufficient RFA (fold change for circ- BANP was 2.407 and that of TLR4 was 2.224 relative to para-tumors). Expression of let-7f-5p showed an opposite tendency (fold change for let-7f-5p in HCC tissues was 0.491 and that in tumors after insufficient RFA was 0.300 relative to para-tumors). Competitive binding of circ-BANP to let-7f-5p was demonstrated and TLR4 was identified as a target of let-7f-5p (P < 0.01). Knockdown of circ-BANP or elevation of let-7f-5p expression inhibited the TLR4/STAT3 signaling pathway, proliferation, invasion, migration, angiogenesis, and EMT in Huh7 and Huh7-H cells (P < 0.01). The effects induced by circ-BANP knockdown were reversed by let-7f-5p inhibition. Overexpression of TLR4 reversed the impact of let-7f-5p upregulation on the cells (P < 0.01). Silencing of circ-BANP inhibited the in vivo growth of residual HCC cells after insufficient RFA (P < 0.01).Conclusions:Knockdown of circ-BANP upregulated let-7f-5p to inhibit proliferation, migration, and EMT formation in residual HCC remaining after insufficient RFA. Effects occur via regulation of the TLR4/STAT3 signaling pathway.