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 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 ◽  
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 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 Potential Role of microRNA-183 as a Tumor Suppressor in Hepatocellular Carcinoma

2018 ◽  
Vol 51 (5) ◽  
pp. 2065-2072 ◽  
Author(s):  
Wei Bian ◽  
Hongfei Zhang ◽  
Miao Tang ◽  
Shaojun Zhang ◽  
Lichao Wang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Hepg2 Cells ◽  
Molecular Mechanisms ◽  
The Cancer Genome Atlas ◽  
Migration And Invasion ◽  
Metastatic Progression ◽  
Mesenchymal Transition ◽  
Reporter Assays

Background/Aims: Disseminated tumors, known as metastases, are responsible for ninety-percent of mortality due to cancer. Epithelial to mesenchymal transition, a phenomenon required for morphological conversion of non-motile discoid shaped epithelial cells to highly motile spindle-shaped mesenchymal cells, is thought to be a pre-requisite for metastatic progression. Metastasis-associated 1 (MTA1) protein is a prime inducer of EMT and metastatic progression in all solid tumors including hepatocellular carcinoma (HCC). However, the molecular mechanisms that regulate the expression and function of MTA1 in HCC have not been elucidated. Methods: In silico prediction algorithms were used to find microRNAs (miRNAs) that may target MTA1. We examined the relationship between the expression of MTA1 and miR-183 using quantitative real time PCR. We also determined the levels of the MTA1 protein using immunohistochemistry. Reporter assays, in the presence and absence of the miR-183 mimic, were used to confirm MTA1 as a bona fide target of miR183. The effect of miR-183 on HCC pathogenesis was determined using a combination of in vitro migration and invasion assay, together with in vivo xenograft experiments. The correlation between miR-183 and MTA1 expression was also studied in samples from HCC patients, and in The Cancer Genome Atlas dataset. Results: Analysis of the sequence database revealed that MTA1 is a putative target of miR-183. MTA1 protein and RNA expression showed opposite trends to miR-183 expression in breast, renal, prostate, and testicular tissue samples from cancer patients, and in the metastatic HCC cell line HepG2. An inverse correlation was also observed between MTA1 (high) and miR-183 (low) expression within samples from HHC patients and in the TCGA dataset. Reporter assays in HepG2 cells showed that miR-183 could inhibit translation of a reporter harboring the wild-type, but not the mutant miR-183 3’-untranslated region (UTR). In addition, miR-183 significantly inhibited in vitro migration and invasion in HepG2 cells, and in vivo hepatic metastasis. Conclusion: Our results reveal a novel post-transcriptional regulatory mechanism for MTA1 expression via miR-183, which is suppressed during HCC pathogenesis.

scholarly journals High Throughput Screening Method for Identification of New Lipofection Reagents

2001 ◽  
Vol 6 (4) ◽  
pp. 245-254 ◽  
Author(s):  
Anne E. Regelin ◽  
Erhard Fernholz ◽  
Harald F. Krug ◽  
Ulrich Massing
Keyword(s):  
High Throughput Screening ◽  
Screening Method ◽  
Genetic Material ◽  
Cell Types ◽  
Cationic Lipids ◽  
Adherent Cell ◽  
Gene Assay

Lipofection, the transfer of genetic material into cells by means of cationic lipids, is of growing interest for in vitro and in vivo approaches. In order to identify ideal lipofection reagents in a HTS, we have developed an automated lipofection method for the transfer of reporter genes into cells and for determination of the lipofection results. The method has specifically been designed and optimized for 96-well microtiter plates and can successfully be carried out by a pipetting robot with accessory equipment. It consists of two separate parts: (1) pretransfection (preparation of liposomes, formation of lipoplexes, and lipoplex transfer to the cells) and (2) posttransfection (determination of the reporter enzyme activity and the protein content of the transfected cells). Individual steps of the lipofection method were specifically optimized—for example, lipoplex formation and incubation time as well as cell lysis, cell cultivating, and the reporter gene assay. The HTS method facilitates characterization of the transfection properties (efficiency and cytotoxicity) of large numbers of (cationic) lipids in various adherent cell types.

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