Invasion Of HCCLM3 Cells Research Articles

Long non-coding RNA LINC01554 overexpression suppresses viability, migration, and invasion of liver cancer cells through regulating miR-148b-3p/EIF4E3

BackgroundLong non-coding RNAs (lncRNAs) can be severed as competing endogenous RNAs (ceRNAs) to regulate target genes or mRNAs via sponging microRNAs (miRNAs). This study explored the effect of LINC01554 on liver cancer cells through the ceRNA mechanism. MethodsFive significantly down-regulated lncRNAs were selected for further verification, and then through bioinformatics, interactive miRNAs and mRNAs of lncRNAs were identified. The relationship between LINC01554, miR-148b-3p and EIF4E3 was detected by the dual luciferase reporter gene assay. Afterwards, HCCLM3 cells were transfected with pCDH-LINC01554, miR-148b-3p inhibitor and miR-148b-3p mimics. Cell viability, apoptosis, migration and invasion were measured by Cell Counting Kit-8, flow cytometer, and Transwell assays. Real-time quantitative PCR (RT-qPCR) and Western blot were used to measure the expressions of related genes and proteins. ResultsLINC01554 was significantly down-regulated in the liver cancer cell lines, and was expressed in the cytoplasm of HCCLM3 cells. LINC01554 overexpression inhibited proliferation, migration, and invasion of HCCLM3 cells, and promote their apoptosis (P < 0.05). Besides, LINC01554 overexpression also significantly increased the levels of BAX, BCL2/BAX, P53, cleaved-Caspase3, TIMP3, E-cadherin and EIF4E3 (P < 0.05). Through bioinformatics and dual-luciferase reporter gene assay, LINC01554, miR-148b-3p and EIF4E3 were proved to interact with each other. Furthermore, the effects of miR-148b-3p knockdown on HCCLM3 cells were similar with those of LINC01554 overexpression, and miR-148b-3p mimics could reverse the changes of cell viability, apoptosis, migration, and invasion induced by LINC01554 overexpression. ConclusionsLINC01554 overexpression could suppress the growth and metastasis of HCCLM3 cells via miR-148b-3p/EIF4E3.

Effect of PI3K/AKT/mTOR Signaling Pathway on Regulating and Controlling the Anti-Invasion and Metastasis of Hepatoma Cells by Bufalin.

Autophagy plays a "double-edged sword" in the process of tumorigenesis, development and metastasis. In this study, we explored the effect of PI3K/AKT/mTOR autophagy-related signaling pathway on regulating and controlling the invasion and metastasis of liver cancer cells by Bufalin. The cell counting, migration, adhesion and invasion assay were used to evaluate the effect of Bufalin on cell proliferation, invasion and metastasis. The protein expression of PI3K/AKT/ mTOR signaling pathway were detected by the Western Blotting technique. After inhibiting autophagy of HCC-LM3 cells, the inhibitory effect of Bufalin on adhesion, migration and invasion of HCC-LM3 cells was significantly enhanced. Synergistic inhibition was strongest when different autophagy inhibitors were combined with 3MA and CQ. After inhibiting autophagy, Bufalin significantly inhibited the protein expression of P-AKT, Cyclin D1, MMP- 2, MMP-9 and VEGF in HCC-LM3 cells. The protein expression of PTEN and E-Cadherin in HCC-LM3 cells was significantly increased. The present study shows that the anti-tumor effect of Bufalin mainly inhibit proliferation, extracellular matrix degradation and angiogenesis of HCC by influencing autophagy. These findings confirm the capability of Bufalin in inhibiting metastasis of HCC and in parallel to current patents, could be applied as a novel therapeutic strategy in the prevention of metastasis of HCC.

Chinese herbal formula QHF inhibits hepatocellular carcinoma metastasis via HGF/c-Met signaling pathway

Hepatocellular Carcinoma (HCC) is one of the most common malignant tumors, and high recurrence and metastasis are the major obstacles to successful treatment of HCC. Traditional Chinese medicine has little known and unique advantages in the treatment of HCC. Previous studies have confirmed that Chinese herbal formula Qingrejiedu (clears away heat and toxins), Huoxuehuayu (promotes blood flow to remove stasis) and Fuzhengguben (strengthens healthy qi and root) (QHF) has a significant effect on patients with advanced HCC, improves the quality of life and prolongs the survival time of patients significantly. In this study, we investigated the effect of QHF on proliferation, migration and invasion of human high metastatic hepatocellular carcinoma cell line HCCLM3 and its underlying mechanism. The results from our in vitro experiments showed that QHF has the ability to inhibit the proliferation by inducing G2/M phase cell cycle arrest and induce apoptosis. Moreover, QHF can also inhibit migration and invasion of HCCLM3 cells and the expression of the p-c-Met protein in HCCLM3 cells was down-regulated. c-Met is closely related to the metastasis of HCC, then we constructed a stable transfected cell line HepG2-met with high expression of c-Met by transfection. Further study in vivo revealed that c-Met gene will promote the growth of tumors and lung metastases in nude mice, and QHF intervention can reduce tumor lung metastases by inhibiting the HGF/c-Met signaling pathway. In conclusion, our study reveals that QHF can inhibit the proliferation, migration and invasion of HCCLM3, and this effect may be related to inhibiting HGF/c-Met signaling pathway.

BICD1 functions as a prognostic biomarker and promotes hepatocellular carcinoma progression

Hepatocellular carcinoma (HCC) is the most predominant type of primary liver cancer and has a high degree of malignancy as well as mortality rate. Many drivers are involved in the development and progression of HCC. A recent study has reported that high BICD cargo adaptor 1 (BICD1) expression indicates poor prognosis of glioblastoma. But, the expression and biological function of BICD1 in HCC remain unclear. In current study, we found that the expression of BICD1 was markedly up-regulated in HCC tissues compared to adjacent nontumor tissues. GEPIA dataset and GSE datasets were consistently indicated the elevated expression of BICD1 in HCC. Furthermore, BICD1 was highly expressed in HCC cell lines including Hep3B, Huh7, MHCC97H and HCCLM3 as compared with that in LO2 cells. High BICD1 expression was positively correlated with malignant clinical features, such as tumor size ≥ 5 cm, venous infiltration and advanced tumor stages. HCC patients highly expressing BICD1 showed a significant shorter overall survival compared to BICD1 low-expression cases. Moreover, TCGA-LIHC data further demonstrated that the up-regulated BICD1 expression predicted poor prognosis of HCC patients. Next, we revealed that BICD1 knockdown prominently suppressed the proliferation, migration and invasion of HCCLM3 cells. Conversely, ectopic expression of BICD1 remarkably facilitated these malignant behaviors of Hep3B cells. Interestingly, BICD1 knockdown abolished hypoxia-induced HCC cell proliferation, migration and invasion. In conclusion, we provide the first evidence to support that BICD1 functions as a predictor for the prognosis of HCC and may serve as a promising therapeutic target for further HCC treatment.

MicroRNA-769-5p contributes to the proliferation, migration and invasion of hepatocellular carcinoma cells by attenuating RYBP

Hepatocellular carcinoma (HCC) is the commonest primary liver cancer with highly aggressive features. MicroRNAs (miRNAs) are demonstrated to play important roles in the tumorigenesis and progression of HCC. miR-769-5p is a recently identified cancer-associated miRNA. But, the expression level of miR-769-5p and its function in HCC are unexplored. In this study, we found that miR-769-5p expression was obviously increased in HCC samples compared to adjacent noncancerous liver tissues. Additionally, we revealed that miR-769-5p was over-expressed in HCC cells as compared with LO2 cells. Notably, HCC tissues from patients with tumor size ≥5 cm, venous infiltration and advanced tumor stages showed higher levels of miR-769-5p compared to those from corresponding controls. Interestingly, our data indicated that HCC patients highly expressing miR-769-5p had significant shorter survivals. Next, functional experiments verified that miR-769-5p knockdown markedly suppressed HCC cell proliferation, migration and invasion. Conversely, ectopic expression of miR-769-5p promoted these biological behaviors of Hep3B cells. Furthermore, depletion of miR-769-5p repressed the growth and metastasis of HCCLM3 cells in vivo. Importantly, miR-769-5p inversely modulated RING1 and YY1 binding protein (RYBP) by directly binding to 3’ untranslated region (UTR) in HCC cells. The expression of RYBP mRNA was down-regulated in HCC tissues and negatively correlated with miR-769-5p level. RYBP overexpression remarkably inhibited the proliferation, migration and invasion of HCCLM3 cells. Accordingly, knockdown of RYBP partially abolished miR-769-5p silencing-induced tumor suppressive effects on HCCLM3 cells. In summary, our study revealed the up-regulated expression of miR-769-5p, which contributed to HCC progression possibly by targeting RYBP.

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.

SHMT1 inhibits the metastasis of HCC by repressing NOX1-mediated ROS production

BackgroundHepatocellular carcinoma (HCC) is the most major type of primary hepatic cancer. Serine hydroxymethyltransferase 1 (SHMT1) is recently found to play critical roles in human cancers including lung cancer, ovarian cancer and intestinal cancer. However, the expression, function and the underlying mechanisms of SHMT1 in HCC remain uncovered.MethodsqRT-PCR, immunohistochemistry and immunoblotting were performed to detect the expression of SHMT1 in HCC tissues and cell lines. HCC cell migration and invasion were determined by Boyden chamber and Transwell assay in vitro, and tumor metastasis was assessed via lung metastasis model in mice. The expression of key factors involved in epithelial-to-mesenchymal transition (EMT) process was evaluated by western blotting.ResultsIn this study, data mining of public databases and analysis of clinical specimens demonstrated that SHMT1 expression was decreased in HCC. Reduced SHMT1 level was correlated with unfavorable clinicopathological features and poor prognosis of HCC patients. Gain- and loss-of-function experiments showed that SHMT1 overexpression inhibited the migration and invasion of HCCLM3 cells while SHMT1 knockdown enhanced the metastatic ability of Hep3B cells. Furthermore, qRT-PCR and western blotting showed that SHMT1 inhibited EMT and matrix metallopeptidase 2 (MMP2) expression. In vivo experiments showed that SHMT1 suppressed the lung metastasis of HCC cells in mice. Mechanistically, SHMT1 knockdown enhanced reactive oxygen species (ROS) production, and thus promoted the motility, EMT and MMP2 expression in Hep3B cells. Furthermore, NADPH oxidase 1 (NOX1) was identified to be the downstream target of SHMT1 in HCC. NOX1 expression was negatively correlated with SHMT1 expression in HCC. Rescue experiments revealed that NOX1 mediated the functional influence of SHMT1 on HCC cells.ConclusionsThese data indicate that SHMT1 inhibits the metastasis of HCC by repressing NOX1 mediated ROS production.

LncRNA A1BG-AS1 suppresses proliferation and invasion of hepatocellular carcinoma cells by targeting miR-216a-5p.

Extensive evidence indicate that long noncoding RNAs (lncRNAs) regulates the tumorigenesis and progression of hepatocellular carcinoma (HCC). However, the expression and biological function of lncRNA A1BG antisense RNA 1 (A1BG-AS1) were poorly known in HCC. Here, we found the underexpression of A1BG-AS1 in HCC via analysis of The Cancer Genome Atlas database. Further analyses confirmed that A1BG-AS1 expression in HCC was markedly lower than that in noncancerous tissues based on our HCC cohort. Clinical association analysis revealed that low A1BG-AS1 expression correlated with poor prognostic features, such as microvascular invasion, high tumor grade, and advanced tumor stage. Follow-up data indicated that low A1BG-AS1 level evidently correlated with poor clinical outcomes of HCC patients. Moreover, forced expression of A1BG-AS1 repressed proliferation, migration, and invasion of HCC cells in vitro. Conversely, A1BG-AS1 knockdown promoted these malignant behaviors in HepG2 cells. Mechanistically, A1BG-AS1 functioned as a competing endogenous RNA by directly sponging miR-216a-5p in HCC cells. Notably, miR-216a-5p restoration rescued A1BG-AS1 attenuated proliferation, migration and invasion of HCCLM3 cells. A1BG-AS1 positively regulated the levels of phosphatase and tensin homolog and SMAD family member 7, which were reduced by miR-216a-5p in HCC cells. Altogether, we conclude that A1BG-AS1 exerts a tumor suppressive role in HCC progression.

MEG2 inhibits the growth and metastasis of hepatocellular carcinoma by inhibiting AKT pathway

MEG2 was recently found to have important functions in human cancers. However, the expression status and biological functions of MEG2 in hepatocellular carcinoma (HCC) remain unknown. In this study, we demonstrated that MEG2 expression was reduced in HCC tissues and cell lines using qRT-PCR, western blot and immunohistochemical staining. Decreased MEG2 expression predicted unfavorable clinical features and decreased overall survival and disease-free survival of HCC patients. In vitro functional assays showed that overexpression of MEG2 inhibited the cell viability, migration and invasion of HCCLM3 cells while MEG2 knockdown promoted these biological functions of Hep3B cells. Subcutaneous injection model and tail vein injection model showed that forced expression of MEG2 in HCCLM3 decreased the growth and lung metastasis of HCCLM3 cells in nude mice. Mechanically, MEG2 inhibited the EMT and AKT phosphorylation of HCC cells. The promoting effects of MEG2 knockdown on EMT, cell viability, proliferation, migration and invasion of Hep3B cells was blocked by AKT phosphorylation inhibition. In all, this study demonstrates that MEG2 inhibits the growth and metastasis of hepatocellular carcinoma by inhibiting AKT pathway.

Quercetin Inhibits the Migration and Invasion of HCCLM3 Cells by Suppressing the Expression of p-Akt1, Matrix Metalloproteinase (MMP) MMP-2, and MMP-9

BackgroundQuercetin is a natural bioactive flavonoid that is present in a wide variety of vegetables and fruits and exhibits a promising anti-metastasis property in various human cancer cells. However, the effect of quercetin on human HCCLM3 cells is unclear.Material/MethodsIn the current study, a wound-healing assay was performed using quercetin-treated HCCLM3 cells to further explore whether quercetin affects the motility of human HCCLM3 cells. Transwell assay was used to explore the potential effect of quercetin in HCCLM3 cells on cell migration and cell invasion. Western blotting analysis was used to explore the expression of p-Akt1, MMP-2, and MMP-9 in quercetin-treated HCCLM3 cells.ResultsThe wound-healing time was delayed in quercetin-treated HCCLM3 cells, and the ability to migrate and invade was inhibited in quercetin-treated human HCCLM3 cells. Moreover, the protein levels of p-Akt1, MMP-2, and MMP-9 were down-regulated in quercetin-treated HCCLM3 cells, as detected by Western blotting.ConclusionsOur data show that quercetin attenuated cell migration and invasion by suppressing the protein levels of p-Akt1, MMP-2, and MMP-9 in HCCLM3 cells.

AGO1 may influence the prognosis of hepatocellular carcinoma through TGF-\u03b2 pathway

AGO1 is a major component of RNA-induced silencing complexes and plays a crucial role in solid tumors. The aim of our study was to investigate AGO1 functions in hepatocellular carcinoma (HCC). Using small interfering RNA, AGO1 functions were investigated in HCCLM3 cell lines. Cell proliferation, immigration, and invasion significantly decreased after AGO1 depletion using MTT, wound-healing, and transwell assay. The associated proteins in the epithelial–mesenchymal transition (EMT) and the activation of its signal pathways were measured using western blot. After AGO1 depleted, increased E-cadherin and decreased N-cadherin, Vimentin, Snail, and Zeb1 were founded. In its upstream pathway, the phosphorylation of ERK1/2(Thr202/Tyr204), Smad2(S425/250/255), and Smad4 were significantly inhibited. Meanwhile, inhibitor of ERK1/2(LY3214996) significantly inhibited the growth and migration of the AGO1 cells. The nuclear importing of Smad4 was blocked and furthermore, the transcription of Snail was also influenced for the decrease of combination between Smad4 and the promotor region of Snail. After Snail was overexpressed, the invasion of HCCLM3 cells was significantly rescued. Immunohistochemistry in tissue microarrays consisting of 200 HCC patients was used to analyze the associations between AGO1 expression and prognosis. Intratumoral AGO1 expression was an independent risk factor for overall survival (P = 0.008) and recurrence-free survival (P < 0.001). In conclusion, AGO1 may promote HCC metastasis through TGF-β pathway, and AGO1 may be a reliable prognostic factor in HCC.