Ginsenoside Rg3 Inhibits the Growth of Osteosarcoma and Attenuates Metastasis through the Wnt/β-Catenin and EMT Signaling Pathway

Osteosarcoma (OS) is the most common primary malignant bone cancer. An increasing number of studies have demonstrated that ginsenoside Rg3 (Rg3), which is extracted from the roots of the traditional Chinese herb Panax ginseng, plays a tumor suppression role in various malignant tumors. In the present study, we aimed at investigating the role of Rg3 in the proliferation, migration, and invasion of OS and at exploring the underlying mechanisms. Cell viability and proliferation were observed by MTT assay and crystal violet staining. The migration and invasion of cells were measured by wound-healing assay and Transwell method. Signaling pathway screening was investigated using luciferase reporter gene assay. qRT-PCR and western blot were performed to measure the expression of molecules involved in cell epithelial-mesenchymal transition (EMT), and Wnt/β-catenin pathway. Results suggested that Rg3 could not only inhibit proliferation but also hamper the migration and invasion of OS. qRT-PCR and western blot demonstrated that a reduced level of MMP2/MMP7/MMP9 was induced after Rg3 treatment. In addition, the expression levels of proteins related to EMT and the Wnt/β-catenin pathway were downregulated. In summary, our data revealed that Rg3 could inhibit the proliferation, migration, and invasion of OS cells. This effect of Rg3 might be mediated by downregulating MMP2, MMP7, and MMP9 expression and suppressing EMT as well as the Wnt/β-catenin pathway. Thus, Rg3 might be a potential agent for the treatment of OS.

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Capicua homology protein inhibits the progression of gastric cancer through the PI3K/AKT signaling pathway

10.21203/rs.3.rs-29464/v1 ◽

2020 ◽

Author(s):

LU GE ◽

Chang-long Hu ◽

Zheng-hui Ge ◽

Chun-rong Wang ◽

Li Qian ◽

...

Keyword(s):

Gastric Cancer ◽

Western Blot ◽

Signaling Pathway ◽

Cell Lines ◽

Epithelial Mesenchymal Transition ◽

Akt Signaling ◽

Akt Signaling Pathway ◽

Mesenchymal Transition ◽

Matrigel Invasion ◽

Qrt Pcr

Abstract Purpose Capicua homolog protein (CIC) played a broad role in the development of cancer in humans, however, its role in the progression of gastric cancer (GC) specifically has been unclear. This study aimed to explore the expression of CIC and its potential clinical value in patients with GC. Methods The CIC levels in GC tissues and cell lines were examined by quantitative real-time polymerase chain reaction (qRT-PCR). And the in-vitro effects of CIC expression in MGC-803 cells on their proliferation, invasion, and the progression of epithelial-mesenchymal transition were assessed by CCK-8 assays, Matrigel-invasion analysis, qRT-PCR and Western blot assays, separately. In addition, the effects of downregulation of CIC on the activation of PI3K/AKT signaling pathway were measured using Western-blot analysis. Results The results showed CIC levels were lower in GC tissues and GC cell lines, and these lower CIC levels were correlated with tumor differentiation, Helicobacter pylori infection, TNM stage, and patient survival. In addition, CIC overexpression could promote cell proliferation, invasion, and progression of epithelial-mesenchymal transition in MGC-803 cells. Notably, exotic expression of CIC inactivated the phosphoinositide 3-kinase/protein kinase B signaling pathway. Conclusions In conclusion, our finding suggested CIC could serve as a potential diagnostic and prognostic biomarker and a probable therapy target for GC.

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Downregulation of UBAP2L Inhibits the Epithelial-Mesenchymal Transition via SNAIL1 Regulation in Hepatocellular Carcinoma Cells

Cellular Physiology and Biochemistry ◽

10.1159/000470824 ◽

2017 ◽

Vol 41 (4) ◽

pp. 1584-1595 ◽

Cited By ~ 6

Author(s):

Tao Ye ◽

Jing Xu ◽

Ling Du ◽

Wenhui Mo ◽

Yiming Liang ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Western Blot ◽

Cell Lines ◽

Epithelial Mesenchymal Transition ◽

Critical Role ◽

Migration And Invasion ◽

Biological Traits ◽

Mesenchymal Transition ◽

Qrt Pcr ◽

E Cadherin

Background/Aims: Dysregulation of ubiquitin-associated protein 2-like (UBAP2L) has been reported in tumors, but its role in hepatocellular carcinoma (HCC) progression is unclear. Methods: The expression levels of UBAP2L in HCC tissues and HCC cell lines were detected by western blot and quantitative real-time (qRT) PCR. The effects of UBAP2L expression on HCC cell biological traits, including migration and invasion, were investigated by wound healing assay and matrigel transwell assay. Simultaneously, the expression of epithelial-mesenchymal transition (EMT) markers including E-cadherin, CK-18, N-cadherin, Vimentin, Claudin7 and the promoter activity of E-cadherin were detected by western blot and qRT-PCR. Subsequently, role of SNAIL1 in UBAP2L-mediated EMT and the mechanism underlying UBAP2L-mediated SNAIL1 expression were further investigated. Results: UBAP2L was overexpressed in human HCC tissues compared with peri-tumoral tissues. Downregulation of UBAP2L inhibited migration, invasion and the EMT in highly metastatic HCC cell lines. Furthermore, UBAP2L knockdown inhibited expression of the transcriptional repressor SNAIL1 and its ability to bind to the E-cadherin promoter via SMAD2 signaling pathway, which in turn resulted in increased E-cadherin expression. Additionally, bioinformatics analysis showed that expression of UBAP2L is correlated with poor prognosis in patients with HCC. Conclusions: UBAP2L plays a critical role in maintenance of the metastatic ability of HCC cells via SNAIL1 Regulation and is predictive of a poor clinical outcome.

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miR-4458 inhibits the epithelial–mesenchymal transition of hepatocellular carcinoma cells by suppressing the TGF-β signaling pathway via targeting TGFBR1

Acta Biochimica et Biophysica Sinica ◽

10.1093/abbs/gmaa029 ◽

2020 ◽

Vol 52 (5) ◽

pp. 554-562

Author(s):

Yuke Zhang ◽

Kun Shi ◽

Hang Liu ◽

Wei Chen ◽

Yunhai Luo ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Signaling Pathway ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Epithelial Mesenchymal Transition ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Mesenchymal Transition ◽

Gene Assay ◽

Hcc Cells

Abstract Hepatocellular carcinoma (HCC) is one of the most lethal cancers in the world. MicroRNAs play a pivotal role in the progression of various cancers. To date, very little attention has been paid to miR-4458. Therefore, the aim of our study was to explore the function and underlying molecular mechanism of miR-4458 in HCC. We found that the expression of miR-4458 was reduced in HCC tissues and cell lines. Forced overexpression of miR-4458 inhibited the migration, invasion, and epithelial–mesenchymal transition (EMT) of HCC cells, while downregulation of miR-4458 promoted the aggressive phenotype. Furthermore, transforming growth factor beta receptor 1 (TGFBR1), the modulator of the TGF-β signaling pathway, was verified to be a novel target gene of miR-4458 by dual-luciferase reporter gene assay. Upregulated miR-4458 dramatically abolished TGFBR1 and p-Smad2/3 expression, thus blocking the TGF-β signaling pathway. Moreover, restoration of TGFBR1 partially rescued the miR-4458-mediated suppressive effect on the migration, invasion, and EMT and reactivated the TGF-β signaling pathway in HCC cells. In summary, our findings first demonstrated a mechanism of miR-4458 in HCC cell migration, invasion, and EMT by regulating the TGF-β signaling pathway via directly targeting TGFBR1.

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Long Non-Coding RNA TMEM220-AS1 Suppressed Hepatocellular Carcinoma by Regulating the miR-484/MAGI1 Axis as a Competing Endogenous RNA

10.21203/rs.3.rs-88668/v1 ◽

2020 ◽

Author(s):

Cong Cao ◽

Jun Li ◽

Guangzhi Li ◽

Gaoyu Hu ◽

Zhihua Deng ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Western Blot ◽

Luciferase Reporter ◽

Mesenchymal Transition ◽

Qrt Pcr ◽

Non Coding Rna ◽

Gene Assay ◽

Hcc Cells

Abstract BackgroundLong non-coding RNA has a considerable regulative influence in multiple biological processes. Nevertheless, the role of TMEM220-AS1 in hepatocellular carcinoma (HCC) remains unclear.MethodsWe used the TCGA database to analyze differentially expressed lncRNAs. qRT-PCR was used to verify the results in a large population. Afterwards, in vitro effects of TMEM220-AS1 on HCC cells were determined by CCK-8, EdU, Flow cytometry experiment and transwell assays in HCC cells. We adopted qRT-PCR, western blot to identify epithelial-mesenchymal transition (EMT). Moreover, we adopted bioinformatics analysis, western blot, dual luciferase reporter gene assay and RIP to investigate underlying molecular mechanisms of TMEM220-AS1 function. Finally, the function of TMEM220-AS1 was verified in vivo.ResultsTMEM220-AS1 was remarkably decreasedin HCC. It was demonstrated that malignant phenotypes and EMT of HCC cells were promoted by knocking TMEM220-AS1 down both in vivo and in vitro. TMEM220-AS1, which was detected distributing mainly in the cytoplasm, worked as a miRNA sponge to sponge miR-484 and promote the level of MAGI1, therefore curbed malignant phenotypes of HCC cells.ConclusionsIn conclusion, downregulation of TMEM220-AS1promotes HCC through miR-484/MAGI1 axis.

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Circ_0000043 promotes breast cancer cell proliferation, migration, invasion and epithelial-mesenchymal transition via the miR-136/ Smad3 axis

Biochemistry and Cell Biology ◽

10.1139/bcb-2020-0219 ◽

2020 ◽

Author(s):

Xiaoling Leng ◽

Guofu Huang ◽

Jianbing Ding ◽

Fucheng Ma

Keyword(s):

Breast Cancer ◽

Cell Proliferation ◽

Western Blot ◽

Epithelial Mesenchymal Transition ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Circular Rnas ◽

Tissue Samples ◽

Mesenchymal Transition ◽

Smad3 Expression

Circular RNAs (circRNAs) are a type of tissue-specific RNA with more stable structure than linear RNAs, and its association with breast cancer (BC) is poorly understood. This study aimed at probing the biological effect of circ_0000043 in the progression of BC. In this study, expression of circ_0000043 in BC tissue samples was measured using quantitative real-time polymerase chain reaction (qRT-PCR). Immunohistochemistry (IHC) and Western blot were used to detect the expression of Smad family member 3 (Smad3). CCK-8, wound healing and Transwell assays were used to assess the effect of circ_0000043 in regulating BC cell proliferation, migration and invasion. Moreover, the binding relationships between circ_0000043 and miR-136, and miR-136 and Smad3 were detected by dual-luciferase reporter assay. Additionally, Western blot was used to detect the expressions of markers related to epithelial-mesenchymal transition (EMT), including E-cadherin, N-cadherin and vimentin. Our results showed that circ_0000043 expression was up-regulated in BC tissues and cell lines. Proliferation, migration, invasion and EMT of BC cells were significantly inhibited by circ_0000043 knockdown, and overexpression of circ_0000043 had the opposite effects. Additionally, circ_0000043 could up-regulate Smad3 expression by sponging miR-136. In conclusion, our study demonstrates that circ_0000043 can promote BC progression via regulating the miR-136/Smad3 axis.

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miR-491-3p is Downregulated in Retinoblastoma and Inhibit Tumor Cells Growth and Metastasis by Targeting SNN

Biochemical Genetics ◽

10.1007/s10528-020-10007-w ◽

2020 ◽

Author(s):

Yang Hu ◽

Ming Zhao ◽

Li Li ◽

Jie Ding ◽

Yu-Min Gui ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Lines ◽

Target Genes ◽

Expression Profiles ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Mesenchymal Transition ◽

Qrt Pcr ◽

Gene Assay ◽

E Cadherin

Abstract Retinoblastoma (Rb) is the most common pediatric malignant tumor of the eyes. Previous studies demonstrated that miR-491-3p is downregulated in various cancers. However, its function in Rb remains unknown. A total of 15 pairs of primary Rb tissues and adjacent noncancerous tissues were collected. Quantitative real-time PCR (qRT-PCR) was used to investigate the expression profiles of miR-491-3p. qRT-PCR, western blotting and in situ immunocytochemistry were performed to investigate the expression profiles of epithelial–mesenchymal transition-related proteins (E-cadherin, Vimentin and N-cadherin) in Rb tissues and Rb cell lines as well as cell morphology. Cell proliferation was estimated by MTS and colony formation assays. Apoptosis was determined by FACS, cell migration and invasion were analyzed using transwell chambers. MiR-491-3p’s target genes were predicted using target gene prediction databases. The interplay between miR-491-3p and SNN was evaluated through dual luciferase reporter gene assay. MiR-491-3p was significantly downregulated in mixed collection of 15 pairs of Rb tissues and Rb cell lines. Overexpression of miR-491-3p enhanced apoptosis, and significantly suppressed proliferation, migration and invasion of Rb cells. In contrast, the present of miR-491-3p inhibitor showed reversed results which apoptosis decreased, while cell proliferation of ARPE-19 cells increased. In addition, miR-491-3p increased the expression of E-cadherin, and dramatically decreased the expression of Vimentin and N-cadherin in Rb tissues and Rb cell lines, noticeable changes in morphology, too, as cells became less cohesive and more adhering. We found out that SNN was the pairing target of miR-491-3p and result showed that miR-491-3p and SNN interacted with each other. We also found out that the effects of miR-491-3p were in Rb cells were almost entirely canceled out at the overexpression of SNN. Our findings collectively suggest that miR-491-3p is an important tumor suppressor in Rb, which inhibits tumor growth and metastasis in Rb. These implicate it may be explored as a new therapeutic target in Rb.

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CircRNA_100290 promotes GC cell proliferation and invasion via the miR-29b-3p/ITGA11 axis and is regulated by EIF4A3

Cancer Cell International ◽

10.1186/s12935-021-01964-2 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Gang Wang ◽

Dan Sun ◽

Wenhui Li ◽

Yan Xin

Keyword(s):

Cell Proliferation ◽

Western Blot ◽

Rna Binding ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Mesenchymal Transition ◽

Qrt Pcr ◽

Node Metastasis

Abstract Background Circular RNAs (circRNAs) have been reported to be important regulators of the development and progression of various carcinomas. However, the role of circRNA_100290 in gastric cancer (GC) is still unclear. This study aimed to investigate the role of circRNA_100290 in GC invasion and metastasis and the possible underlying mechanism. Methods The expression of circRNA_100290 in GC cells and tissues was examined using quantitative real-time polymerase chain reaction (qRT-PCR). The role of circRNA_100290 in cell proliferation, migration, and invasion was evaluated in the AGS and HGC-27 cell lines in vitro. Bioinformatics tools, dual-luciferase reporter assays, Western blot assays and qRT-PCR were used to explore the pathways downstream of circRNA_100290. The mechanism underlying the regulation of circRNA_100290 expression was explored using RNA immunoprecipitation, qRT-PCR, and Western blot assays. Results The expression of circRNA_100290 was significantly upregulated in GC cells and 102 GC tissues, and high circRNA_100290 expression in GC was closely related to Borrmann’s type, lymph node metastasis and tumour-node-metastasis stage. In vitro, knockdown of circRNA_100290 in AGS and HGC-27 cells significantly inhibited cell proliferation, migration, and invasion. Mechanistically, a dual-luciferase reporter assay confirmed the direct interaction between circRNA_100290 and miR-29b-3p, which targets ITGA11, an oncogene that is closely related to epithelial–mesenchymal transition (EMT). In addition, EIF4A3, an RNA-binding protein (RBP), could inhibit the formation of circRNA_100290 by binding to the flanking sites of circRNA_100290. Low EIF4A3 expression in GC was related to a poor prognosis. Conclusions Elevated circRNA_100290 expression in GC promotes cell proliferation, invasion and EMT via the miR-29b-3p/ITGA11 axis and might be regulated by EIF4A3. CircRNA_100290 might be a promising biomarker and target for GC therapy. Graphical abstract

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Overexpression of Long Noncoding RNA H19 Downregulates miR-140-5p and Activates PI3K/AKT Signaling Pathway to Promote Invasion, Migration and Epithelial-Mesenchymal Transition of Ovarian Cancer Cells

BioMed Research International ◽

10.1155/2021/6619730 ◽

2021 ◽

Vol 2021 ◽

pp. 1-13

Author(s):

Hao Xu ◽

Yuan Ding ◽

Xiangying Yang

Keyword(s):

Ovarian Cancer ◽

Western Blot ◽

Cancer Cells ◽

Signaling Pathway ◽

Epithelial Mesenchymal Transition ◽

Akt Signaling ◽

Luciferase Reporter ◽

Ovarian Cancer Cells ◽

Akt Signaling Pathway ◽

Mesenchymal Transition

Objective. The abnormal expression of LncRNA H19 and miR-140-5p has been linked to ovarian cancer (OC). Whether H19 directly regulates miR-140-5p in ovarian cancer cells has been unclear. In this study, we deeply explored the relationship between H19 and miR-140-5p in ovarian cancer and the mechanism of action in regulating OC progression. Methods. A total of 66 patients with OC admitted to the hospital from June 2017 to June 2019 were selected as the research group (RG), and meanwhile, 60 cases of healthy subjects were selected as the control group (CG). In addition, OC cells and normal ovarian epithelial cells were used to detect H19 and miR-140-5p expression levels and to analyze the effect of H19 on OC cells. The activation of the PI3K/AKT pathway and downstream proteins were analyzed by western blot. Results. H19 was highly expressed while miR-140-5p was lowly expressed in OC patients and cell lines ( P < 0.050 ). The proliferation, invasion, migration ability, and epithelial-mesenchymal transition (EMT) of OC cells were reduced after inhibiting H19 expression, and the apoptosis rate was increased. Transfection of cells with miR-140-5p mimics brought opposite effects. Online prediction and dual-luciferase reporter (DLR) confirmed that H19 directly binds miR-140-5p. Western blot assay indicated overexpression activated the PI3K/AKT signaling pathway in OC cells. Moreover, overexpression promoted tumor growth in nude mice and was suppressed by PI3K inhibitor. Conclusion. LncRNA H19 downregulation of miR-140-5p to activate the PI3K/AKT signaling pathway and promote the proliferation, invasion, migration and EMT of OC.

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Up-regulation of miR-1825 inhibits the progression of glioblastoma by suppressing CDK14 though Wnt/β-catenin signaling pathway

10.21203/rs.3.rs-18711/v2 ◽

2020 ◽

Author(s):

Fengqin Lu ◽

Chunhong Li ◽

Yuping Sun ◽

Ting Jia ◽

Na Li ◽

...

Keyword(s):

Western Blot ◽

Cell Viability ◽

Signaling Pathway ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Invasion And Migration ◽

Protein Levels ◽

Qrt Pcr ◽

Cell Progression ◽

Tissue Specimens

Abstract Background: Mounting evidences displayed that miRNAs play crucial roles in tumor initiation and development. However, the regulation and relevant mechanism of miR- miR-1825 in glioblastoma (GBM) remain unclear. Methods: qRT-PCR was used to detect miR-1825 and CDK14 mRNA expression. Western blot was applied for testing protein levels (VEGF, E-cadherin, N-cadherin, vimentin, β-catenin, c-myc, p-c-Jun). MTT and transwell assays were used for detecting GBM cell progression, including cell viability, migration, and invasion.Results: The results showed that miR-1825 was decreased in GBM tissue specimens by qRT-PCR and it was confirmed as a prognostic marker of GBM by Kaplan-Meier survival analysis. Moreover, we also found that miR-1825 up-regulation suppressed GBM cell viability, tumor growth, invasion and migration. Furthermore, CDK14 was first identified as the direct target of miR-1825 by Luciferase reporter assay. CDK14 acted as an oncogene in GBM development by Immunohistochemistry. In addition, Western blot analysis demonstrated that miR-1825 regulated Wnt/β-catenin signaling pathway in GBM development. Conclusion: In conclusion, miR-1825 up-regulation suppressed GBM progression by targeting CDK14 through Wnt/β-catenin pathway.

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LncRNA NEAT1 regulated diabetic retinal epithelial-mesenchymal transition through regulating miR-204/SOX4 axis

PeerJ ◽

10.7717/peerj.11817 ◽

2021 ◽

Vol 9 ◽

pp. e11817

Author(s):

Yang Yang ◽

Jing Zhou ◽

Wei hong Li ◽

Zhi xiong Zhou ◽

Xiao bo Xia

Keyword(s):

High Glucose ◽

Epithelial Mesenchymal Transition ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Mesenchymal Transition ◽

High Glucose Condition ◽

Gene Assay ◽

Related Proteins ◽

Lncrna Neat1 ◽

Glucose Condition

Aim Epithelial-mesenchymal transition (EMT) of retinal pigment epithelium (RPE) cells is the key of the development of diabetic retinopathy (DR), and lncRNA NEAT1 could accelerate EMT in diabetic nephropathy. Meanwhile, as a diabetes susceptibility gene, whether sex-determining region Y-related (SRY) high-mobility group box 4 (SOX4) has relationship with lncRNA NEAT1 in DR remains unclear. Methods Firstly, NEAT1, SOX4 and miR-204 were evaluated by qRT-PCR (quantitative reverse-transcriptase PCR) under high glucose condition. Then, cell viability, proliferation, migration and invasion were respectively detected by MTT, BrdU staining, wound healing and transwell assay after NEAT1 knockdown or miR-204 overexpression. Also, the EMT-related proteins were examined by western blot and cell immunofluorescence assay. In order to confirm the relationship between miR-204 and NEAT1 or SOX4, dual luciferase reporter gene assay was conducted. At the same time, the protein levels of SOX4 and EMT-related proteins were investigated by immunohistochemistry in vivo. Results High glucose upregulated NEAT1 and SOX4 and downregulated miR-204 in ARPE19 cells. NEAT1 knockdown or miR-204 overexpression inhibited the proliferation and EMT progression of ARPE19 cells induced by high glucose. NEAT1 was identified as a molecular sponge of miR-204 to increase the level of SOX4. The effect of NEAT1 knockdown on the progression of EMT under high glucose condition in ARPE19 cells could be reversed by miR-204 inhibitor. Also, NEAT1 knockdown inhibited retinal EMT in diabetic mice. Conclusion NEAT1 regulated the development of EMT in DR through miR-204/SOX4 pathway, which could provide reference for clinical prevention and treatment.

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