scholarly journals A study of the mechanism of lncRNA-CR594175 in regulating proliferation and invasion of hepatocellular carcinoma cells in vivo and in vitro

2020 ◽  
Vol 15 (1) ◽  
Author(s):  
Quan Liu ◽  
Xuxu Yu ◽  
Minjie Yang ◽  
Xiangke Li ◽  
Xuejia Zhai ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
High Throughput Screening ◽  
Hepatoma Cell ◽  
Screening Method ◽  
Negative Regulation ◽  
Hepatoma Cell Line ◽  
Incidence And Mortality ◽  
Proliferation And Invasion

Abstract Background Hepatocellular carcinoma (HCC) is one of the cancers of highest incidence and mortality worldwide. The proliferation and invasion of tumor cells are the main reason for poor prognosis after HCC surgery. Long non-coding RNA (lncRNA) has been shown to play a key role in the progression of HCC. LncRNA-CR594175 is one of the highly expressed lncRNAs in HCC tumors and their metastatic tumors that we have obtained by the High-throughput screening method. Methods To elucidate the role of lncRNA-CR594175 in regulating the proliferation and invasion of human hepatoma cell line, HepG2, we operated through lncRNA-CR594175 silencing to inhibit the progression of HCC, either through in vitro or in vivo experiments. Results We found that lncRNA-CR594175 was lower in adjacent non-cancerous tissues than in primary HCC, and was lower in primary HCC than in its metastasis. Silencing of lncRNA-CR594175 inhibited the proliferation and invasion of HepG2 cells and growth of subcutaneous tumors. The results revealed that lncRNA-CR594175, as a RNA sponge, broke the negative regulation of hsa-miR-142-3p on Catenin, beta-1 (CTNNB1), and once lncRNA-CR594175 was silenced, the hsa-miR142-3p regained its negative regulation on CTNNB1 which can promote HCC progression by activating the wnt pathway. Conclusions Our present study demonstrated for the first time that lncRNA-CR594175 silencing suppressed proliferation and invasion of HCC cells in vivo and in vitro by restoring the negative regulation of hsa-miR-142-3p on CTNNB1, laying a solid theoretical base for using lncRNA-CR594175 as genetic target therapy for HCC and offering a reasonable explanation for inactivation of miRNA in different tumors or in the tumor at different stages.

scholarly journals A study of the mechanism of lncRNA-CR594175 in regulating proliferation and invasion of hepatocellular carcinoma cells in vivo and in vitro

2020 ◽  
Author(s):  
Quan Liu ◽  
Xuxu Yu ◽  
Minjie Yang ◽  
Xiangke Li ◽  
Xuejia Zhai ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
High Throughput Screening ◽  
Hepatoma Cell ◽  
Screening Method ◽  
Negative Regulation ◽  
Hepatoma Cell Line ◽  
Incidence And Mortality ◽  
Proliferation And Invasion

Abstract Abstract Background Hepatocellular carcinoma (HCC) is one of the cancers of highest incidence and mortality worldwide. The proliferation and invasion of tumor cells are the main reason for poor prognosis after HCC surgery. Long non-coding RNA (lncRNA) has been shown to play a key role in the progression of HCC. LncRNA-CR594175 is one of the highly expressed lncRNAs in HCC tumors and their metastatic tumors that we have obtained by the High-throughput screening method.Methods To elucidate the role of lncRNA-CR594175 in regulating the proliferation and invasion of human hepatoma cell line, HepG2, we operated through lncRNA-CR594175 silencing to inhibit the progression of HCC, either through in vitro or in vivo experiments.Results We found that lncRNA-CR594175 was lower in adjacent non-cancerous tissues than in primary HCC, and was lower in primary HCC than in its metastasis. Silencing of lncRNA-CR594175 inhibited the proliferation and invasion of HepG2 cells and growth of subcutaneous tumors. The results revealed that lncRNA-CR594175, as a RNA sponge, broke the negative regulation of hsa-miR-142-3p on Catenin, beta-1 (CTNNB1), and once lncRNA-CR594175 was silenced, the hsa-miR142-3p regained its negative regulation on CTNNB1 which can promote HCC progression by activating the wnt pathway. Conclusions Our present study demonstrated for the first time that lncRNA-CR594175 silencing suppressed proliferation and invasion of HCC cells in vivo and in vitro by restoring the negative regulation of hsa-miR-142-3p on CTNNB1, laying a solid theoretical base for using lncRNA-CR594175 as genetic target therapy for HCC and offering a reasonable explanation for inactivation of miRNA in different tumors or in the tumor at different stages.

scholarly journals A study of the mechanism of lncRNA-CR594175 in regulating proliferation and invasion of hepatocellular carcinoma cells in vivo and in vitro

2020 ◽  
Author(s):  
Quan Liu ◽  
Xuxu Yu ◽  
Minjie Yang ◽  
Xiangke Li ◽  
Xuejia Zhai ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Hepg2 Cells ◽  
High Throughput Screening ◽  
Target Therapy ◽  
Screening Method ◽  
Negative Regulation ◽  
Incidence And Mortality ◽  
Proliferation And Invasion

Abstract Background Hepatocellular carcinoma (HCC) is one of the cancers of highest incidence and mortality worldwide. The proliferation and invasion of tumor cells are the main reason for poor prognosis after HCC surgery. Long non-coding RNA (lncRNA) has been shown to play a key role in the progression of HCC. LncRNA-CR594175 is one of the highly expressed lncRNAs in HCC tumors and their metastatic tumors that we have obtained by the High-throughput screening method.Methods To elucidate the role of CR594175 in regulating the proliferation and invasion of HepG2 cells, and to initially try to use a genetic engineering operation plan-CR594175 silence to inhibit the HCC progression through functional experiments in vitro and subcutaneous tumor-bearing experiments.Results We found that lncRNA-CR594175 was lower in adjacent noncancerous tissues than in primary HCC, and was lower in primary HCC than in their metastases. SilencingCR594175 inhibited the proliferation and invasion of HepG2 cells and growth of subcutaneous tumors. The mechanism research revealed that CR594175, as a RNA sponge, broke the negative regulation of hsa-miR-142-3p on CTNNB1 (Catenin, beta-1), and once CR594175 was silenced, the hsa-miR142-3p regained its negative regulation on CTNNB1 which can promote HCC progression by activating the wnt pathway. Conclusions Our present study demonstrates for the first time that CR594175 silencing suppressed proliferation and invasion of HCC cells in vivo and in vitro by restoring the negative regulation of hsa-miR-142-3p on CTNNB1, laying a solid theoretic base for using lncRNA-CR594175 as genetic target therapy for HCC and offering a reasonable explanation for inactivation of miRNA in different tumors or tumor at different stages.

scholarly journals Anti-tumor effects of LncRNA-CR594175 silenceing on hepatocellular carcinoma cell line HepG2 in vivo and in vitro

2020 ◽  
Author(s):  
Quan Liu ◽  
Xuxu Yu ◽  
Minjie Yang ◽  
Xiangke Li ◽  
Xuejia Zhai ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
High Throughput Screening ◽  
Wnt Signaling Pathway ◽  
Negative Regulation ◽  
Primary Hepatocellular Carcinoma ◽  
Incidence And Mortality ◽  
Proliferation And Invasion ◽  
Hcc Cells

Abstract Background Hepatocellular carcinoma (HCC) is one of the cancers of highest incidence and mortality worldwide. Methods High-throughput screening was applied to find the most varied expressed LncRNA in primary hepatocellular carcinoma and explore the interaction network of it. Results We found that lncRNA-CR594175 was the lncRNA of most varied expression, which was lower in adjacent noncancerous tissues than in primary HCC, and was lower in primary HCC than in its metastases. Knockdown of CR594175 inhibited the proliferation and invasion of HCC cells HepG2. The mechanism study revealed that CR594175, as a RNA sponge, broke the negative regulation of CTNNB1 (Catenin, beta-1) by hsa-miR142-3p, and once CR594175 was silenced, the highly expressed hsa-miR142-3p regained its regulation on CTNNB1 and inhibited the proliferation and invasion through Wnt signaling pathway. Conclusions Our present study demonstrates for the first time that CR594175 silencing suppressed proliferation and invasion of HCC cells HepG2 in vivo and in vitro by restoring the negative regulation of CTNNB1 by hsa-miR142-3p, laying a solid theoretic base for using lncRNA-CR594175 as genetic therapy target for HCC and offering a reasonable explanation for inactivation of miRNA in difference tumors or tumor of different stages.

scholarly journals Anti-tumor effects of LncRNA-CR594175 silenceing on hepatocellular carcinoma cell line HepG2 in vivo and in vitro

2020 ◽  
Author(s):  
Quan Liu ◽  
Xuxu Yu ◽  
Minjie Yang ◽  
Xiangke Li ◽  
Xuejia Zhai ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
High Throughput Screening ◽  
Wnt Signaling Pathway ◽  
Negative Regulation ◽  
Primary Hepatocellular Carcinoma ◽  
Incidence And Mortality ◽  
Proliferation And Invasion ◽  
Hcc Cells

Abstract Background Hepatocellular carcinoma (HCC) is one of the cancers of highest incidence and mortality worldwide. Methods High-throughput screening was applied to find the most varied expressed LncRNA in primary hepatocellular carcinoma and explore the interaction network of it. Results We found that lncRNA-CR594175 was the lncRNA of most varied expression, which was lower in adjacent noncancerous tissues than in primary HCC, and was lower in primary HCC than in its metastases. Knockdown of CR594175 inhibited the proliferation and invasion of HCC cells HepG2. The mechanism study revealed that CR594175, as a RNA sponge, broke the negative regulation of CTNNB1 (Catenin, beta-1) by hsa-miR142-3p, and once CR594175 was silenced, the highly expressed hsa-miR142-3p regained its regulation on CTNNB1 and inhibited the proliferation and invasion through Wnt signaling pathway. Conclusions Our present study demonstrates for the first time that CR594175 silencing suppressed proliferation and invasion of HCC cells HepG2 in vivo and in vitro by restoring the negative regulation of CTNNB1 by hsa-miR142-3p, laying a solid theoretic base for using lncRNA-CR594175 as genetic therapy target for HCC and offering a reasonable explanation for inactivation of miRNA in difference tumors or tumor of different stages.

scholarly journals KLF7/VPS35 axis contributes to hepatocellular carcinoma progression through CCDC85C-activated β-catenin pathway

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yarong Guo ◽  
Bao Chai ◽  
Junmei Jia ◽  
Mudan Yang ◽  
Yanjun Li ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Proliferation ◽  
Tumor Growth ◽  
Luciferase Reporter ◽  
Aberrant Expression ◽  
Proliferation And Invasion ◽  
Hcc Cells

Abstract Objective Dysregulation of KLF7 participates in the development of various cancers, but it is unclear whether there is a link between HCC and aberrant expression of KLF7. The aim of this study was to investigate the role of KLF7 in proliferation and migration of hepatocellular carcinoma (HCC) cells. Methods CCK8, colony growth, transwell, cell cycle analysis and apoptosis detection were performed to explore the effect of KLF7, VPS35 and Ccdc85c on cell function in vitro. Xenografted tumor growth was used to assess in vivo role of KLF7. Chip-qPCR and luciferase reporter assays were applied to check whether KLF7 regulated VPS35 at transcriptional manner. Co-IP assay was performed to detect the interaction between VPS35 and Ccdc85c. Immunohistochemical staining and qRT-PCR analysis were performed in human HCC sampels to study the clinical significance of KLF7, VPS35 and β-catenin. Results Firstly, KLF7 was highly expressed in human HCC samples and correlated with patients’ differentiation and metastasis status. KLF7 overexpression contributed to cell proliferation and invasion of HCC cells in vitro and in vivo. KLF7 transcriptional activation of VPS35 was necessary for HCC tumor growth and metastasis. Further, co-IP studies revealed that VPS35 could interact with Ccdc85c in HCC cells. Rescue assay confirmed that overexpression of VPS35 and knockdown of Ccdc85c abolished the VPS35-medicated promotion effect on cell proliferation and invasion. Finally, KLF7/VPS35 axis regulated Ccdc85c, which involved in activation of β-catenin signaling pathway, confirmed using β-catenin inhibitor, GK974. Functional studies suggested that downregulation of Ccdc85c partly reversed the capacity of cell proliferation and invasion in HCC cells, which was regulated by VPS35 upregulation. Lastly, there was a positive correlation among KLF7, VPS35 and active-β-catenin in human HCC patients. Conclusion We demonstrated that KLF7/VPS35 axis promoted HCC cell progression by activating Ccdc85c-medicated β-catenin pathway. Targeting this signal axis might be a potential treatment strategy for HCC.

scholarly journals Pyrrolizidine Alkaloids Induce Cell Death in Human HepaRG Cells in a Structure-Dependent Manner

2020 ◽  
Vol 22 (1) ◽  
pp. 202
Author(s):  
Josephin Glück ◽  
Julia Waizenegger ◽  
Albert Braeuning ◽  
Stefanie Hessel-Pras
Keyword(s):  
Cell Death ◽  
Cell Viability ◽  
Pyrrolizidine Alkaloids ◽  
Defense Mechanism ◽  
Hepatoma Cell ◽  
Hepatoma Cell Line ◽  
Dependent Manner ◽  
Human Hepatoma Cell

Pyrrolizidine alkaloids (PAs) are a group of secondary metabolites produced in various plant species as a defense mechanism against herbivores. PAs consist of a necine base, which is esterified with one or two necine acids. Humans are exposed to PAs by consumption of contaminated food. PA intoxication in humans causes acute and chronic hepatotoxicity. It is considered that enzymatic PA toxification in hepatocytes is structure-dependent. In this study, we aimed to elucidate the induction of PA-induced cell death associated with apoptosis activation. Therefore, 22 structurally different PAs were analyzed concerning the disturbance of cell viability in the metabolically competent human hepatoma cell line HepaRG. The chosen PAs represent the main necine base structures and the different esterification types. Open-chained and cyclic heliotridine- and retronecine-type diesters induced strong cytotoxic effects, while treatment of HepaRG with monoesters did not affect cell viability. For more detailed investigation of apoptosis induction, comprising caspase activation and gene expression analysis, 14 PA representatives were selected. The proapoptotic effects were in line with the potency observed in cell viability studies. In vitro data point towards a strong structure–activity relationship whose effectiveness needs to be investigated in vivo and can then be the basis for a structure-associated risk assessment.

scholarly journals Antioxidant supplements promote tumor formation and growth and confer drug resistance in hepatocellular carcinoma by reducing intracellular ROS and induction of TMBIM1

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Vanilla Xin Zhang ◽  
Karen Man-Fong Sze ◽  
Lo-Kong Chan ◽  
Daniel Wai-Hung Ho ◽  
Yu-Man Tsui ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Hepatoma Cell ◽  
Tumor Formation ◽  
In Vitro Assays ◽  
Migration And Invasion ◽  
Intracellular Ros ◽  
Antioxidant Supplements ◽  
Sphere Formation

Abstract Background Controversy over the benefits of antioxidants supplements in cancers persists for long. Using hepatocellular carcinoma (HCC) as a model, we investigated the effects of exogenous antioxidants N-acetylcysteine (NAC) and glutathione (GSH) on tumor formation and growth. Methods Multiple mouse models, including diethylnitrosamine (DEN)-induced and Trp53KO/C-MycOE-induced HCC models, mouse hepatoma cell and human HCC cell xenograft models with subcutaneous or orthotopic injection were used. In vitro assays including ROS assay, colony formation, sphere formation, proliferation, migration and invasion, apoptosis, cell cycle assays were conducted. Western blot was performed for protein expression and RNA-sequencing to identify potential gene targets. Results In these multiple different mouse and cell line models, we observed that NAC and GSH promoted HCC tumor formation and growth, accompanied with significant reduction of intracellular reactive oxygen species (ROS) levels. Moreover, NAC and GSH promoted cancer stemness, and abrogated the tumor-suppressive effects of Sorafenib both in vitro and in vivo. Exogenous supplementation of NAC or GSH reduced the expression of NRF2 and GCLC, suggesting the NRF2/GCLC-related antioxidant production pathway might be desensitized. Using transcriptomic analysis to identify potential gene targets, we found that TMBIM1 was significantly upregulated upon NAC and GSH treatment. Both TCGA and in-house RNA-sequence databases showed that TMBIM1 was overexpressed in HCC tumors. Stable knockdown of TMBIM1 increased the intracellular ROS; it also abolished the promoting effects of the antioxidants in HCC cells. On the other hand, BSO and SSA, inhibitors targeting NAC and GSH metabolism respectively, partially abrogated the pro-oncogenic effects induced by NAC and GSH in vitro and in vivo. Conclusions Our data implicate that exogenous antioxidants NAC and GSH, by reducing the intracellular ROS levels and inducing TMBIM expression, promoted HCC formation and tumor growth, and counteracted the therapeutic effect of Sorafenib. Our study provides scientific insight regarding the use of exogenous antioxidant supplements in cancers.

scholarly journals In vivo and in vitro regulation of erythropoietin mRNA: measurement by competitive polymerase chain reaction

Blood ◽  
1993 ◽  
Vol 81 (3) ◽  
pp. 617-623 ◽  
Author(s):  
J Fandrey ◽  
HF Bunn
Keyword(s):  
Polymerase Chain Reaction ◽  
Hepatoma Cell ◽  
Hepatoma Cell Line ◽  
Mrna Levels ◽  
Chain Reaction ◽  
Human Hepatoma Cell ◽  
Competitive Polymerase Chain Reaction ◽  
Polymerase Chain

Abstract The regulation of erythropoietin (Epo) production was investigated by competitive polymerase chain reaction, a highly sensitive and accurate means of measuring Epo mRNA levels. Co-amplification of the test sample with added mutant Epo cDNA template corrects for variability in the efficiency of amplification. Epo mRNA levels were determined in tissues of normal rats and in animals with varying degrees of anemia. Reduction of the hematocrit level from 0.40 to 0.15–0.20 resulted in a 300-fold increase in kidney Epo mRNA, which comprised 80% of the total Epo mRNA versus 20% from the liver. In contrast, very low levels detected in lung and spleen were not significantly increased by anemia. The human hepatoma cell line, Hep3B, secretes high levels of Epo in response to hypoxia. This regulation is, to a large extent, transcriptional. When Hep3B cells were incubated in the presence of decreasing O2 tension from 160 to 7 mm Hg, there was a monotonic increase in Epo mRNA to 50 to 100 times the normoxic level. Hyperoxia did not suppress basal expression. When cells were incubated at a PO2 of 7 mm Hg, induction of Epo mRNA was first noted at 30 minutes and was maximal at 5 to 6 hours. After Epo mRNA was boosted by a 4-hour hypoxic incubation, cells were then exposed to normoxia, which shut off further transcription of the Epo gene. The decay of Epo mRNA levels closely followed first order kinetics with a half-life of 2 hours, an effective measurement of message stability.

Non-genotoxic hepatocarcinogenesis in vitro: the FaO hepatoma line responds to peroxisome proliferators and retains the ability to undergo apoptosis

1993 ◽  
Vol 104 (2) ◽  
pp. 307-315 ◽  
Author(s):  
A.C. Bayly ◽  
N.J. French ◽  
C. Dive ◽  
R.A. Roberts
Keyword(s):  
Cell Death ◽  
Cell Line ◽  
Cell Lines ◽  
Hepatoma Cell ◽  
Hepatoma Cell Line ◽  
Peroxisome Proliferator ◽  
Peroxisome Proliferators ◽  
Hepatoma Cell Lines

A range of hepatoma cell lines (RH1, HTC, FaO, 7800C1 and MH1C1), has been studied with the aim of establishing an in vitro model to investigate the molecular mechanisms of hepatocarcinogenicity induced by the peroxisome proliferator class of non-genotoxic carcinogens. In view of speculation that peroxisome proliferators suppress hepatocyte apoptosis in vivo, we have placed particular emphasis on evaluating whether hepatoma cell lines retain the ability to undergo apoptotic cell death. Expression of the liver-specific differentiation marker albumin and the peroxisome proliferator-activated receptor (PPAR) was highest in the Reuber hepatoma cell line, FaO. This cell line also demonstrated the most marked response to the peroxisome proliferator nafenopin with a 2.2-fold induction of the microsomal enzyme cytochrome p450IVA1. This response was found to display intercellular heterogeneity by immunocytochemistry. Thus, the FaO cell line maintained characteristics of hepatocytes, both in vivo and in vitro, in terms of expression of constitutive and inducible markers. However, none of the cell lines tested mirrored the hyperplastic response of hepatocytes to nafenopin, since no increase in cell growth kinetics was observed on addition of nafenopin to the growth medium. The mode of cell death in confluent FaO cultures was characterised as apoptosis, by fluorescence microscopy and agarose gel electrophoresis of extracted DNA. Cells detaching from confluent FaO cultures exhibited chromatin condensation and DNA fragmentation patterns characteristic of cels undergoing apoptotic death.Interestingly, no apoptosis was seen in monolayer cells, suggesting that apoptosis in vitro is associated with cell shrinkage and detachment similar to that documented for the liver in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)

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