Levels In Hepatocellular Carcinoma Tissues Research Articles

MicroRNA-1251-5p promotes tumor growth and metastasis of hepatocellular carcinoma by targeting AKAP12

MicroRNAs (miRNA) are small RNA molecules that have emerged as important regulators of gene expression in hepatocellular carcinoma (HCC). However, the expression, function and mechanism of miR-1251-5p in HCC remain poorly understood. In the present study, it was observed that miR-1251-5p expression was upregulated in HCC. Furthermore, higher miR-1251-5p level was correlated with poor prognosis, large tumor size, vascular invasion and high tumor-node-metastasis (TNM) stages of HCC patients. Functionally, miR-1251-5p drove HCC cell proliferation, migration and invasion in vitro, and promoted growth and metastasis of HCC cells in vivo. A-kinase anchor protein 12 (AKAP12) was screened as a direct target of miR-1251-5p by using the starBase V3.0 online platform. The AKAP12 mRNA expression was downregulated and negatively correlated with miR-1251-5p level in HCC tissues. Furthermore, in vitro experiments confirmed that AKAP12 was targeted and negatively regulated by miR-1251-5p. Importantly, AKAP12 overexpression decreased HCC cell proliferation, migration and invasion, whereas inhibition of AKAP12 rescued the miR-1251-5p knockdown-attenuated HCC cell proliferation, migration and invasion. Overall, the present study indicates that miR-1251-5p plays an oncogenic role in HCC by targeting AKAP12, and may be a potential therapeutic target for HCC treatment.

MicroRNA-627-5p inhibits the proliferation of hepatocellular carcinoma cells by targeting BCL3 transcription coactivator.

Hepatocellular carcinoma (HCC) is a common malignant tumour. An increasing number of studies indicate that microRNAs (miRNAs) are critical regulators in the carcinogenesis and progression of HCC. MiR-627-5p has been identified as a tumour suppressor in colorectal cancer and glioblastoma multiforme. However, the function of miR-627-5p in HCC progression remains unclear yet. In our present study, miR-627-5p was determined to be low-expressed in HCC tissues and cell lines. Furthermore, miR-627-5p was expressed at significantly lower levels in HCC tissues with tumour size >5cm or advanced tumour stages (III+IV). Additionally, HCC patients with low miR-627-5p level had a significantly poorer overall survival. Functionally, ectopic expression of miR-627-5p obviously inhibited the proliferation, and induced G1 phase arrest and apoptosis of Hep3B and SMMC-7721 cells. Conversely, miR-627-5p silencing facilitated HCC cell proliferation, cell cycle progression and apoptosis resistance. In vivo experiments further confirmed that miR-627-5p overexpression repressed the growth of Hep3B cells in mice. Mechanistically, BCL3 transcription coactivator was predicted as a direct target of miR-627-5p. MiR-627-5p overexpression reduced, whereas miR-627-5p knockdown enhanced the expression of BCL3 protein in HCC cells. Luciferase reporter assay confirmed the direct binding between miR-627-5p and 3'UTR of BCL3. The expression of BCL3 protein was negatively correlated with miR-627-5p level in HCC tissues. More importantly, re-expression of BCL3 partially reversed miR-627-5p induced inhibitory effects on Hep3B cells. In conclusion, these results demonstrated that miR-627-5p functioned as a tumour suppressor in HCC possibly by attenuating BCL3. This finding might offer a new therapeutic target for HCC treatment.

An indel polymorphism in the 3' untranslated region of HMGB1 confers risk for hepatocellular carcinoma by regulating HMGB1 transcriptional activity in a Chinese population.

The present study aimed to assess the association of rs34000982 polymorphism located in the 3'untranslated region (3'UTR) of high mobility group box 1 (HMGB1) gene and the risk of hepatocellular carcinoma (HCC), and further to explore the underlying mechanism. Genomic DNA was extracted from peripheral blood of 320 patients with HCC and 360 matched controls. Rs34000982 polymorphism was genotyped by a polymerase chain reaction-polyacrylamide gel electrophoresis assay. The genotype-phenotype association of HMGB1 mRNA and protein expression in HCC tissues with different genotypes was detected by quantitative (q) PCR assay and western blot. Vectors containing the insertion (ins)/ins or deletion (del)/del genotype of the rs34000982 polymorphism were constructed and the HMGB1 transcriptional activity affected by the rs34000982 polymorphism was detected by the luciferase assay. It was identified that the ins/ins genotype of rs34000982 significantly increased the risk of HCC compared with the del/del genotype. Further the qPCR results demonstrated that the HMGB1 mRNA expression level in HCC tissues with ins/ins genotype was 2.24 times that of HCC tissues with ins/del and del/del genotypes and there was a similar trend at protein level. In addition, the insertion allele of rs34000982 disturbed the binding of miR-636 with the 3'UTR of HMGB1, thereby increasing HMGB1 transcriptional activity in vitro. These data suggest that the rs34000982 polymorphism may contribute to HCC susceptibility, in full or at least partially through the effect on HMGB1 transcriptional activity by disturbing the binding of miR-636 with the 3'UTR of HMGB1.

High interleukin-8 and/or extracellular signal-regulated kinase 2 expression predicts poor prognosis in patients with hepatocellular carcinoma.

Interleukin (IL)-8 and extracellular signal-regulated kinase (ERK) 2 play key roles in tumor progression, but the relationship between IL-8 and/or ERK2 expression in hepatocellular carcinoma (HCC) tissues and postoperative recurrence or survival is unclear. The expression levels of IL-8 and ERK2 in both HCC tissues and non-tumor liver tissues were analyzed using the Oncomine™ database and immunohistochemistry assay. Reverse transcription-quantitative PCR was then used to evaluate the expression levels of IL-8 and ERK2 in the tumor tissues of 67 patients with HCC undergoing radical hepatectomy. Pearson's correlation, Kaplan-Meier, Cox univariate and multivariate survival analyses were utilized to determine the correlation between IL-8 and ERK2 expression in HCC tissues, and their potential prognostic significance. As indicated by the data from the Oncomine™ database, and the patient samples, IL-8 and ERK2 were expressed at significantly higher levels in HCC tissues than in non-tumor liver tissues (P<0.05). The rates of high IL-8 and ERK2 expression in HCC tissues were 43.28 (29/67) and 34.33% (23/67), respectively, and the IL-8 and ERK2 expression levels were positively correlated (r=0.764; P<0.001). Both ERK2 expression and IL-8/ERK2 co-expression were significantly associated with tumor size and differentiation (P<0.05). Additionally, high expression levels of IL-8, ERK2 and IL-8/ERK2 co-expression were all significantly associated with poor overall survival (OS; P<0.05) and disease-free survival (DFS; P<0.05). Multivariate Cox regression analysis also showed that high expression levels of IL-8, ERK2, and IL-8 and ERK2 were independent prognostic factors for OS and DFS (P<0.05). The results of the present study indicate a significant increase in the risk of recurrence and mortality in HCC patients with high expression levels of IL-8 and/or ERK2, compared with patients with low expression. Therefore, IL-8 and ERK2 may be predictors of postoperative prognosis in patients with HCC.

Retracted:Propofol exerts anticancer activity on hepatocellular carcinoma cells by raising lncRNA DGCR5

Hepatocellular carcinoma is one of the most fatal cancers worldwide. Propofol is an intravenous anesthetic extensively used in clinical. Herein, we tested the anticancer activity of propofol on hepatocellular carcinoma, along with the internal molecular mechanism related to lncRNA DiGeorge syndrome critical region gene 5 (DGCR5). Followed by propofol stimulation, hepatocellular carcinoma Huh-7 and HepG2 cell viability, proliferation, migration, invasion, and apoptosis were tested, respectively. Then, DGCR5 expression levels in hepatocellular carcinoma tissues and cells were measured. sh-DGCR5 was transfected to silence DGCR5 expression. Subsequently, the influence of DGCR5 silence on propofol caused Huh-7 and HepG2 cell viability loss, proliferation inhibition, migration and invasion suppression, apoptosis induction, as well as Raf1/ERK1/2 and Wnt/β-catenin pathways inactivation were assessed, respectively. We discovered that propofol declined Huh-7 and HepG2 cell viability, proliferation, migration and invasion, but increased cell apoptosis. DGCR5 had a relatively lower expression level in hepatocellular carcinoma tissues and cells. Propofol elevated DGCR5 expression in Huh-7 and HepG2 cells. Increased expression of DGCR5 was connected with the anticancer activity of propofol on Huh-7 and HepG2 cells. Besides, propofol repressed Raf1/ERK1/2 and Wnt/β-catenin pathways through elevating DGCR5 expression. In conclusion, the anticancer activity of propofol on hepatocellular carcinoma was verified in this study. Propofol repressed hepatocellular carcinoma Huh-7 and HepG2 cell growth and metastasis at least by elevating DGCR5 and hereafter inactivating Raf1/ERK1/2 and Wnt/β-catenin pathways.

Targeting Ceramide Metabolism in Hepatocellular Carcinoma: New Points for Therapeutic Intervention.

Hepatocellular Carcinoma (HCC) is one of the greatest global health burdens because of its uncontrolled cell growth and proliferation, aggressive nature as well as inherited chemoresistance. In spite of different treatment options currently available for HCC, the 5-year relative survival rates for HCC patients with regional and distant stages of the disease are still low, which highlights the urgent need for novel therapeutic strategies for HCC. Recent findings strongly suggest that specific lipid species, such as sphingolipids, play a prominent role in tumorigenesis. We will give an overview of recent literature findings on the role of ceramide metabolism in the pathogenesis and treatment of HCC. HCC is characterised by dysregulation of ceramide metabolism, which could be ascribed to altered activity and expression of ceramide synthases 2, 4 and 6, and acid and alkaline ceramidases 2 and 3, as well as to deregulation of Sphingosine kinases (SphK) 1 and 2 and sphingosine-1- phosphate receptors, in particular, S1PR1. Among them, SphK2 has emerged as a clinically relevant drug target in HCC whose inhibition by ABC294640 is currently being investigated in a clinical trial in patients with advanced HCC. Another promising strategy includes restoration of ceramide levels in HCC tissues, whereby nanoliposomal ceramides, in particular C6-ceramide, has emerged as an effective therapeutic agent against HCC whose safety and recommended dosing is currently being clinically investigated. Development of novel drugs specifically targeting ceramide metabolism could provide an enhanced therapeutic response and improved survival outcome in HCC patients.

STAT gene family mRNA expression and prognostic value in hepatocellular carcinoma.

BackgroundSignal transducer and activator of transcription (STAT) proteins are well-known transcription factors that play an important role in the progression of cancer. However, the association between STAT family genes and hepatocellular carcinoma (HCC) remains unclear. This study investigates the expression level, the prognostic value and the potential mechanism of STAT family genes in HCC.MethodsData from 365 HCC patients in The Cancer Genome Atlas (TCGA) database and 241 HCC patients in the Gene Expression Omnibus (GEO) database were used to investigate the diagnostic and prognostic values of STAT genes by survival analysis and nomogram. Gene set enrichment analysis (GSEA) was used to investigate the potential mechanism of the STAT genes in the development of HCC.ResultsOur results showed that STAT4/5B mRNA expression levels in HCC tissues were lower than those in normal tissues. Importantly, our results indicated that high expression of STAT5A, STAT5B and STAT6 was associated with better overall survival in HCC patients. Joint effects analysis of STAT5A, STAT5B and STAT6 suggested that the prognosis difference for any combination of genes was more significant than that for any individual gene. Then, we developed a risk score model could predict HCC prognosis and the nomogram visualized gene expression and clinical factors of probability for HCC prognosis. The ROC and calibration curves showed good performance in survival prediction in both the TCGA and the GEO databases. GSEA suggested that high expression of STAT5A, STAT5B and STAT6 were involved in immune-related biological processes, drug metabolism cytochrome P450, JAK-STAT signalling pathway, and PPAR signalling pathways.ConclusionOur data suggest that STAT5A, STAT5B and STAT6 expression may be potential prognostic markers of HCC and, in combination, have a better predictive value for HCC prognosis.

MiR-450b-5p loss mediated KIF26B activation promoted hepatocellular carcinoma progression by activating PI3K/AKT pathway

BackgroundKinesin family member 26B (KIF26B) is unveiled acted as important role in many solid tumors, however, the function of KIF26B in hepatocellular carcinoma (HCC) is unclear.MethodsThe expression of KIF26B in HCC tissues and cell lines were measured with immunochemistry, real-time PCR and western blotting. The correlation between KIF26B expression and clinicopathological characteristics were analyzed by SPSS19.0. Functional experiments of KIF26B was conducted by CCK-8, transwell, EDU, colony formation in vitro and tumorigenesis in vivo. The gene set enrichment analysis was used to search the downstream pathway, luciferase reporter experiment was used to find the upstream regulatory factor of KIF26B.ResultsIn this study, we found that KIF26B was overexpressed both in HCC tissues and cell lines. High expression of KIF26B was associated with poor overall survival (OS), late TNM stage and poor differentiation. Loss of function experiments showed that suppression of KIF26B could inhibit cell viability, proliferation rate and invasion ability of HCC cells. KEGG and GO analysis showed that expression of KIF26B was highly relevant with PI3K/AKT signal pathway, and suppression of KIF26B could decrease the expression of m-TOR, p-PI3K and p-AKT. Further study demonstrated that expression of KIF26B was negative correlated with miR-450b-5p level in HCC tissues, and miR-450b-5p could inhibit cell viability, proliferation rate and invasion ability of HCC cells via targeted inhibiting KIF26B.ConclusionOur study demonstrated that miR-450-5p/KIF26B/AKT axis is critical for progression of HCC, and might provide novel prognostic biomarker and therapeutic target for HCC.

MicroRNA-548b inhibits proliferation and invasion of hepatocellular carcinoma cells by directly targeting specificity protein 1.

Emerging studies have revealed that microRNAs (miRNAs) are aberrantly expressed in hepatocellular carcinoma (HCC), and the dysregulation of miRNAs exerts crucial roles in the carcinogenesis and development of HCC. Therefore, elucidating the relationship between miRNAs and HCC progression is of great importance to develop novel therapeutic techniques and to improve the prognosis of patients with this malignancy. Recently, miR-548b-3p (miR-548b) has been demonstrated to be a cancer-associated miRNA in tongue squamous cell carcinoma and glioma. However, the expression and function of miR-548b in HCC remain poorly understood. In the present study, it was found that miR-548b is expressed at low levels in HCC tissues and cell lines. Decreased miR-548b expression was found to be positively associated with the clinical features of HCC, including the TNM stage and lymph node metastasis. Functional experiments revealed that upregulation of miR-548b expression decreased proliferation and invasion of HCC cells. Specificity protein 1 (SP1) was verified to be a direct target of miR-548b in HCC cells; as Spearman's correlation analysis identified miR-548b expression to be negatively correlated with that of SP1 expression in HCC tissue specimens. In addition, SP1 inhibition exhibited similar effects as miR-548b overexpression in HCC cells. SP1 reintroduction significantly reversed the suppressive effects of miR-548b upregulation on the proliferation and invasion of HCC cells. In conclusion, the results presented in the present study demonstrated that miR-548b may serve as a tumor suppressive miRNA that inhibits the proliferation and invasion of HCC cells by directly targeting SP1. Consequently, miR-548b can be exploited as a novel therapeutic target for treating patients with HCC in the future, but this needs to be investigated further.

Up-regulation of SNHG6 activates SERPINH1 expression by competitive binding to miR-139-5p to promote hepatocellular carcinoma progression

ABSTRACT We aimed to assess the roles of small nucleolar RNA host gene 6 (SNHG6) in hepatocellular carcinoma (HCC) progression, and establish the lncRNA-miRNA-mRNA regulation mechanism for HCC therapy. SNHG6 is one of the host genes in small nucleolar RNAs (snoRNAs), which make a difference in the development of human cancers. SERPINH1 is a gene encoding a member of the serpin superfamily of serine proteinase inhibitors with miRNA predicted by TargetScan and DIANA Tools. SNHG6, serpin family H member 1 (SERPINH1) and miR-139-5p expression levels in HCC tissues and cells were determined by quantitative real-time PCR (qRT-PCR). Migration and invasion of HCC cells were measured by transwell assay. Cell cycle analysis was determined by using flow cytometry. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay and colony formation assay were performed for cell viability analysis. The expression of SERPINH1 was detected by qRT-PCR and western blot. Dual-luciferase reporter gene assay was conducted to identify the targeted relationship between miR-139-5p and SNHG6, as well as SERPINH1 and miR-139-5p. The positive regulation between SNHG6 and SERPINH1 was demonstrated in this study. In contrast, miR-139-5p was significantly down-regulated in HCC cells, the inhibition of miR-139-5p promotes the proliferation of HCC cells, and accelerated the cell cycle of HCC cells. Our study demonstrated the co-expression of SNHG6 and SERPINH1 in HCC cells for the first time, which revealed that SNHG6 could serve as a novel oncogene for the HCC therapy by its regulation.

MicroRNA-106b-5p promotes hepatocellular carcinoma development via modulating FOG2.

Background: A recent study has revealed that miR-106b-5p might promote hepatocellular carcinoma (HCC) stemness maintenance and metastasis by targeting PTEN via PI3K/Akt pathway based on HCC cell lines and animal models. Its clinical relevance remains unknown.Purpose: Herein, we aimed to evaluate associations of miR-106b-5p dysregulation with various clinicopathological features of HCC patients and investigate its functions during HCC progression.Patients and methods: At first, miR-106b-5p expression in 130 pairs of HCC and adjacent normal liver tissues was detected by quantitative PCR. Chi-square test was then performed to determine clinical significance. Further investigations on its functions were performed by miRNA target prediction and validation, as well as cellular experiments.Results: miR-106b-5p levels in HCC tissues were significantly higher than those in the adjacent normal liver tissues (P<0.001). High miR-106b-5p expression was significantly associated with advanced tumor stage (P=0.02) and high tumor grade (P=0.03). In addition, Friend of GATA 2 (FOG2) was identified as a direct target of miR-106b-5p in HCC cells. Moreover, the clinical relevance to HCC progression of the combined high miR-106b-5p and low FOG2 expression was more significant than high miR-106b-5p alone. Functionally, enforced expression of miR-106b-5p reduced FOG2 expression and promoted the proliferation and invasion of HCC cells. Furthermore, co-transfection of FOG2 restored the oncogenic roles of miR-106b-5p over-expression.Conclusion: Our data offer the convincing evidence that miR-106b-5p upregulation may promote the aggressive progression of HCC. miR-106b-5p overexpression may promote HCC cell proliferation and invasion by suppressing FOG2, implying its potentials as a promising therapeutic target for HCC patients.

LncRNA PLAC2 upregulates p53 to induce hepatocellular carcinoma cell apoptosis

It is known that lncRNA PLAC2 can inhibit glioma. This study explored the function of PLAC2 in hepatocellular carcinoma (HCC). Our data showed that PLAC2 expression in HCC was not affect by HCV and HBV infection, while PLAC2 levels were significantly lower in HCC tissues comparing to non-cancer tissues. Low PLAC2 levels in HCC tissues were associated with low overall 5-year survival rate. P53 mRNA was also downregulated in HCC and positively correlated with PLAC2. PLAC2 overexpression caused upregulated p53 and increase cancer cell apoptosis. P53 overexpression failed to affect PLAC2. In addition, p53 silencing reduced the effects of PLAC2 overexpression. Therefore, PLAC2 upregulated p53 to mediate cancer cell apoptosis.

MiR-1307-3p promotes tumor growth and metastasis of hepatocellular carcinoma by repressing DAB2 interacting protein

Increasing studies provide evidence to support that microRNAs (miRNAs) play important roles in regulating hepatocellular carcinoma (HCC) initiation and progression. However, whether miR-1307-3p is aberrantly expressed in HCC and affects malignant behaviors of cancer cells remain unknown. In this study, we found that miR-1307-3p expression was obviously up-regulated in HCC compared to adjacent nontumor tissues. Moreover, miR-1307-3p expression was prominently higher in HCC cells compared with the normal hepatic cell line LO2. Patients with venous infiltration, tumor size ≥5 cm and advanced tumor stages (III + IV) had significant higher levels of miR-1307-3p in HCC tissues. Notably, the high level of miR-1307-3p predicted poor clinical outcomes of HCC patients. Functionally, miR-1307-3p knockdown inhibited the proliferation, migration and invasion of MHCC97H and HCCLM3 cells, and suppressed the in vivo growth and metastasis of HCCLM3 cells. Conversely, overexpression of miR-1307-3p facilitated Hep3B cell proliferation, migration and invasion. Mechanistically, DAB2 interacting protein (DAB2IP) was screened as a direct target of miR-1307-3p. The expression of DAB2IP mRNA was down-regulated and inversely correlated with miR-1307-3p level in HCC tissues. miR-1307-3p knockdown increased the level of DAB2IP in HCC cells. Luciferase reporter assay confirmed the direct interaction between miR-1307-3p and 3′UTR of DAB2IP. Importantly, DAB2IP overexpression significantly suppressed the proliferation, migration and invasion of HCCLM3 cells. DAB2IP knockdown rescued miR-1307-3p silencing-attenuated HCC cell proliferation, migration and invasion. Taken together, our findings suggest that miR-1307-3p plays a driving role in HCC progression by targeting DAB2IP. Our study may provide new therapeutic targets for HCC treatment.

Endoplasmic Reticulum Stress Causes Liver Cancer Cells to Release Exosomal miR-23a-3p and Up-regulate Programmed Death Ligand 1 Expression in Macrophages.

Endoplasmic reticulum (ER) stress promotes tumor cell escape from immunosurveillance. However, the underlying mechanisms remain unknown. We hypothesized that ER stress induces hepatocellular carcinoma (HCC) cells to release exosomes, which attenuate antitumor immunity by modulating the expression of programmed death ligand 1 (PD-L1) in macrophages. In this study, we demonstrated that expression of several ER stress markers (glucose-regulated protein 78, activating transcription factor 6, protein kinase R-like ER kinase, and inositol-requiring enzyme 1α) was up-regulated in HCC tissues and negatively correlated with the overall survival and clinicopathological scores in patients with HCC. Expression of ER stress-related proteins positively correlated with CD68+ macrophage recruitment and PD-L1 expression in HCC tissues. High-throughput sequencing analysis identified miR-23a-3p as one of the most abundant microRNAs in exosomes derived from tunicamycin (TM)-treated HCC cells (Exo-TMs). miR-23a-3p levels in HCC tissues negatively correlated with overall survival. Treatment with Exo-TMs up-regulated the expression of PD-L1 in macrophages in vitro and in vivo. Bioinformatics analysis suggests that miR-23a-3p regulates PD-L1 expression through the phosphatase and tensin homolog (PTEN)-phosphatidylinositol 3-kinase-protein kinase B (AKT) pathway. This notion was confirmed by in vitro transfection and coculture experiments, which revealed that miR-23a-3p inhibited PTEN expression and subsequently elevated phosphorylated AKT and PD-L1 expression in macrophages. Finally, coculture of T cells with Exo-TM-stimulated macrophages decreased CD8+ T-cell ratio and interleukin-2 production but increased T-cell apoptosis in vitro. Conclusion: ER-stressed HCC cells release exosomes to up-regulate PD-L1 expression in macrophages, which subsequently inhibits T-cell function through an exosome miR-23a-PTEN-AKT pathway. Our findings provide insight into the mechanism how tumor cells escape from antitumor immunity.

Downregulation of nucleolar and spindle-associated protein 1 expression suppresses liver cancer cell function.

The aim of the present study was to determine the role of nucleolar and spindle-associated protein 1 (NuSAP1) in human liver cancer. NuSAP1 expression was determined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), western blotting and immunohistochemistry in hepatocellular carcinoma (HCC) and adjacent tissues. The expression of NuSAP1 gene was detected by RT-qPCR in liver cancer cell lines. Expression information for NuSAP1 was determined using the UALCAN and Oncomine databases. The Kaplan-Meier plotter and The Cancer Genome Atlas databases were used to obtain overall survival data for liver cancer. Liver cancer cell lines HepG2 and Huh-7 were transfected with lentiviral particles to silence the endogenous NuSAP1 gene expression. RT-qPCR and western blotting were performed to confirm the silencing efficiency. Cell Counting Kit-8 was used to estimate the effects of NuSAP1 silencing on HepG2 and Huh-7 cell proliferation. Cell cycle and apoptosis analyses were performed using flow cytometry. Cell invasion was assessed using the Transwell assay with microscopy imaging. The results revealed that the NuSAP1 expression levels in HCC tissues were significantly increased compared with the adjacent tissues. The survival time of patients with HCC with a high NuSAP1 expression was markedly decreased compared with that of patients with HCC with a low expression level of NuSAP1. Functional studies revealed that NuSAP1 silencing significantly reduced HepG2 and Huh-7 cell proliferation and invasion. Increased apoptosis and cell cycle arrest at the G1 phase were observed following NuSAP1 knockdown. NuSAP1 silencing significantly inhibited the mRNA expression of DNA methyltransferase but not glioma-associated oncogene. NuSAP1 contributed to liver cancer development by reducing apoptosis and promoting cell cycle progression. The abnormal expression level of NuSAP1 may serve a role in promoting liver cancer progression.

LncRNA SNHG16 aggravates tumorigenesis and development of hepatocellular carcinoma by sponging miR-4500 and targeting STAT3.

Hepatocellular carcinoma (HCC) is the most common type of primary liver tumor and becomes a lethal malignancy on account of high mortality and increasing incidence. A growing body of studies has proved that long noncoding RNAs (lncRNAs) participate in the development of diverse cancers. Although it has been commonly accepted that SNHG16 is a procancer gene in numerous cancers, the regulatory mechanism of SNHG16 in HCC still needs more explorations. In this study, our results delineated that SNHG16 presented much higher expression levels in HCC tissues and cells, particularly in advanced stages of HCC. Enhanced SNHG16 expression was strongly related to poor prognosis. SNHG16 facilitated HCC progression by promoting cell proliferation, migration, invasion, and epithelial-mesenchymal transition process as well as inhibiting cell apoptosis. SNHG16 served as a sponge for miR-4500 in HCC and miR-4500 neutralized the influences of SNHG16 knockdown on HCC. SNHG16 was confirmed to compete with signal transducer and activator of transcription 3 (STAT3) to bind with miR-4500. SNHG16 aggravated the development of HCC via sponging miR-4500 so as to upregulate STAT3. In other words, this study was the first to investigate the potential mechanism of SNHG16 in HCC and verified SNHG16 exerted its carcinogenesis by miR-4500/STAT3 axis, suggesting SNHG16 may be a new underlying therapeutic target for HCC treatment.

MicroRNA‑663b targets GAB2 to restrict cell proliferation and invasion in hepatocellular carcinoma.

Previous studies have demonstrated that numerous tumor‑specific microRNAs (miRNAs) are upregulated or downregulated in hepatocellular carcinoma (HCC), and that their dysregulation is implicated in HCC occurrence and development. Therefore, investigation of crucial miRNAs involved in HCC oncogenesis and progression may provide novel insights into the therapy of patients with this malignant tumor. In the present study, reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) assays were performed to detect tissue and cellular expression levels of miRNA‑663b (miR‑663b) in HCC. The effects of miR‑663b overexpression on the proliferation and invasion of HCC cells were examined using Cell Counting Kit‑8 and Transwell invasion assays, respectively. The direct target of miR‑663b in HCC cells was determined by bioinformatics analysis, luciferase reporter assay, RT‑qPCR and western blot analysis. It was observed that miR‑663b was expressed at low levels in HCC tissues and cell lines. miR‑663b upregulation suppressed the proliferative and invasive abilities of HCC cells. Additionally, Grb2‑associated binding 2 (GAB2) was regarded as a direct target gene of miR‑663b in HCC cells. Furthermore, GAB2 was overexpressed in HCC tissues, and overexpression of GAB2 was inversely correlated with levels of miR‑663b. GAB2 overexpression was able to rescue the suppressive effects of miR‑663b on HCC cells. These results demonstrated that this newly‑identified miR‑663b/GAB2 axis may be implicated in HCC occurrence and development.

Targeting genes and signaling pathways of transcriptional suppressor ZHX2 in hepatocellular carcinoma: a Chromatin Immunoprecipitation-sequencing (ChIP-seq) investigation

Hepatocellular carcinoma (HCC) is one of the most common malignancies in the world. The unclear molecular mechanisms underlying could provide important theoretical basis for the prevention and control of HCC. This study performed chromatin immunoprecipitation-sequencing (ChIP-seq) to analyze the binding sites between zinc fingers and homeoboxes 2 (ZHX2) and its genome-wide target genes, and bioinformatics was used to analyze their gene transcription regulation network. Immunohistochemistry was used to detect the ZHX2 expression in HCC, and its association with the clinicopathological characteristics of HCC. Results of RT-PCR and western blot showed the expression of ZHX2 in HepG2 cells was obviously lower compared with normal liver cells. ZHX2 could be amplified in ChIP products, then ChIP-seq reveals there were 232 genes binding in promoter regions. GO analysis of functions revealed these genes were mainly associated with biological processes (BP), cellular components (CC), and molecular functions (MF). In addition, PTEN was found enriched in certain biological functions in BP analysis. Then, four pathways of these genes based on Kyoto Encyclopedia of Genes and Genomes (KEGG) were found P<0.05. Last analysis of immunohistochemistry showed the rates of ZHX2 expression and PTEN expression in paracancerous tissues both were significantly higher than that in HCC tissues (P=0.042; P<0.001), with negative correlations with AFP values (r=-0.246, P=0.040; r=-0.263, P=0.028). Further, PTEN expression was positively correlated with the differentiation level in HCC tissues (r=0.267, P=0.025). Spearman correlation analysis revealed that the expression profiles of ZHX2 and PTEN were positively correlated in HCC tissues (r=0.258, P=0.031). This study is the first to use ChIP-seq technology to analyze the specific regulatory mechanisms of the transcription suppressor ZHX2 in the context of HCC at the genome level.

Endoplasmic reticulum resident oxidase ERO1-Lalpha promotes hepatocellular carcinoma metastasis and angiogenesis through the S1PR1/STAT3/VEGF-A pathway

Mounting evidence demonstrates that expression of ERO1α, an endoplasmic reticulum (ER)-resident oxidase, is a poor prognosis factor in a variety of human cancers. However, the clinical relevance of ERO1α and its molecular mechanisms underlying tumor progression have not been determined for hepatocellular carcinoma (HCC). ERO1α expression levels in HCC tissues and cells were detected by quantitative real-time PCR and western blotting. ERO1α shRNAs and overexpression vector were transfected into HCC cells to downregulate or upregulate ERO1α expression. In vitro and in vivo assays were performed to investigate the function of ERO1α in invasion, metastasis, and angiogenesis of HCC. We found high ERO1α expression in HCC tissues and cells that was significantly associated with metastasis and poor clinicopathologic features of vascular invasion, advanced Edmondson Grade, and TNM stage. Loss-of-function and gain-of-function studies showed that ERO1α prompted migration, invasion, epithelial–mesenchymal transition (EMT), and angiogenesis of HCC cells both in vitro and in vivo. Further studies verified a positive correlation between ERO1α and S1PR1, upregulated in metastatic HCC tissues compared with HCC tissues without metastasis. S1PR1 knockdown markedly diminished the effects of ERO1α on HCC cell migration, invasion and vascular endothelial growth factor (VEGF) expression. Most importantly, ERO1α knockdown significantly repressed the death of HCC xenograft mouse models by reducing tumor distant metastasis, and host angiogenesis by suppressing the expression of S1PR1, p-STAT3, and VEGF-A in HCC cells. Our findings suggest that ERO1α is significantly correlated with reduced survival and poor prognosis, and promotes HCC metastasis and angiogenesis by triggering the S1PR1/STAT3/VEGF-A signaling pathway. ERO1α might be a novel candidate in HCC prognosis and therapy.

Microarray-based measurement of microRNA-449c-5p levels in hepatocellular carcinoma and bioinformatic analysis of potential signaling pathways

The clinical role and potential molecular mechanisms of microRNA-449c-5p (miR-449c-5p) in hepatocellular carcinoma (HCC) tissues remains unclear. Combining multiple bioinformatic tools, we studied the miR-449c-5p expression levels in HCC tissues and explored possible target genes and related signaling pathways. First, miR-449c-5p expression data from microarrays provided by publicly available sources were mined and analyzed using various meta-analysis methods. Next, genes that were downregulated after miR-449c-5p mimic transfection into HCC cells were identified, and in silico methods were used to predict potential target genes. Several bioinformatic assessments were also performed to evaluate the possible signaling pathways of miR-449c-5p in HCC. Five microarrays were included in the current study, including GSE98269, GSE64632, GSE74618, GSE40744 and GSE57555. The standard mean difference was 0.44 (0.07–0.80), and the area under the curve was 0.68 (0.63–0.72), as assessed by meta-analyses, which consistently indicated the upregulation of miR-449c-5p in HCC tissues. A total of 2244 genes were downregulated after miR-449c-5p mimic transfection into an HCC cell line, while 5217 target genes were predicted by in silico methods. The overlap of these two gene pools led to a final group of 428 potential target genes of miR-449c-5p. These 428 potential target genes were primarily enriched in the homologous recombination pathway, which includes DNA Polymerase Delta 3 (POLD3). Data mining with Oncomine and the Human Protein Atlas showed a decreasing trend in POLD3 mRNA and protein levels in HCC tissue samples. This evidence suggests that miR-449c-5p could play an essential role in HCC through various pathways and that POLD3 could be a potential miR-449c-5p target. However, these in silico findings should be validated with further experiments.