scholarly journals MicroRNA-518-3p suppresses cell proliferation, invasiveness, and migration in colorectal cancer via targeting TRIP4

2020 ◽  
Vol 98 (5) ◽  
pp. 575-582
Author(s):  
Heng Yang ◽  
Jia Ren ◽  
Yu Bai ◽  
Jielin Jiang ◽  
Shiyao Xiao
Keyword(s):  
Colorectal Cancer ◽  
Cell Viability ◽  
Colony Formation ◽  
Thyroid Hormone Receptor ◽  
Formation Rate ◽  
Negative Control ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Closure Rate ◽  
Normal Tissues

MicroRNA (miR)-518-3p has been shown to function as a tumor suppressor. This study was conducted to investigate the effects of miR-518-3p in colorectal cancer (CRC). The miR-518-3p mimic, mimic negative control (NC), miR-518-3p inhibitor, inhibitor-NC, ShRNA-TRIP4, and ShRNA-NC vectors were transfected into SW480 cells using Lipofectamine 2000. Cell viability was detected using CCK-8. Colony formation, cell invasiveness, and cell migration were assessed by plate colony formation, Transwell assays, and wound healing assays, respectively. Relative mRNA and protein levels were detected using RT–qPCR and Western blot, respectively. The target gene thyroid hormone receptor interactor 4 (TRIP4) of miR-518-3p was identified and further verified using dual-luciferase reporter assay. Compared with normal tissues, levels of miR-518-3p were decreased and TRIP4 was significantly increased in the tissues from patients with CRC. Following transfection with a miR-518-3p mimic or ShRNA-TRIP4, cell viability decreased in a time-dependent manner, and colony formation rate, wound closure rate, and the number of invasive cells were much lower for the transfected cells than in the corresponding NC and control groups. miR-518-3p overexpression or silencing of TRIP4 significantly down-regulated the expression of MMP-2 and MMP-9. Knockdown of miR-518-3p had the opposite effects, and TRIP4 was identified as a target of miR-518-3p. The inhibitory effects of miR-518-3p on the progressions of CRC are associated with TRIP4.

Downregulation of MicroRNA-135b-5p Inhibits Esophageal Cancer Cell Progression via Targeting GNG7

2019 ◽  
Vol 9 (9) ◽  
pp. 1222-1231
Author(s):  
Jun Chen
Keyword(s):  
Cell Viability ◽  
Colony Formation ◽  
Biological Behavior ◽  
Luciferase Reporter ◽  
P53 Expression ◽  
Normal Tissues ◽  
Cell Progression ◽  
Esophageal Cancer Cell ◽  
Ec Cells ◽  
Guanine Nucleotide Binding

Objective: MicroRNAs (miRNAs) have emerged as the critical modulators of the tumorigenesis and tumor progression. Guanine nucleotide binding (G) protein gamma 7 (GNG7) has been shown to be closely associated with the occurrence of tumors. We aimed to investigate the effects of microRNA-135b-5p (miR-135b-5p) mediated GNG7 knockdown on the biological behavior of esophageal carcinoma (EC) cells. Material and Methods: The expression of miR-135b-5p and GNG7 in 33 cases of EC clinical tissues and EC cell lines, including TE-1, ECA109 and EC9706, were examined by PCR and western blotting assays to select the optimum cell line for subsequent experiments. After ECA109 cells were transfected with miR-135b-5p mimics or inhibitor, siRNA GNG7 and NC plasmids, the cell viability, migration invasion, proliferation and apoptosis of ECA109 cells were evaluated by MTT, wound healing, Transwell, colony formation and flow cytometry assays, respectively. The expression of relevant molecules (p53 and VEGF-A) was determined by western blot analysis. Results: The results indicated that the levels of miR-135b-5p were upregulated while GNG7 were downregulated in EC tissues in comparison with the corresponding normal tissues, and miR-135b-5p mimics and knockdown of GNG7 sharply increased cell viability, migration, invasion and colony formation of ECA109 cells, and inhibited cell apoptosis. The luciferase reporter assay identified that miR-135b-5p decreased the level of GNG7 via binding to the 3′-UTR of GNG7 gene. In addition, p53 expression were decreased and VEGFA expression was up-regulated in ECA109 cells after transfected with miR-135b-5p mimics and GNG7 siRNA. However, inhibition of miR-135b-5p suppressed biological behavior abilities and promoted apoptosis of ECA109 cells that was promoted by miR-135b-5p mimics and siRNA GNG7. Conclusions: In conclusion, miR-135b-5p is identified as a potential tumor-promoting miRNA likely through the regulation of GNG7 expression, which could be developed as a new therapeutic target for EC.

scholarly journals Aspirin enhances the sensitivity of hepatocellular carcinoma side population cells to doxorubicin via miR-491/ABCG2

2018 ◽  
Vol 38 (6) ◽  
Author(s):  
Zheng-Yuan Xie ◽  
Mao-Sheng Liu ◽  
Cong Zhang ◽  
Peng-Cheng Cai ◽  
Zhi-Hua Xiao ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Viability ◽  
Colony Formation ◽  
Side Population ◽  
Inhibitory Effect ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Flow Cytometry Analysis ◽  
Side Population Cells ◽  
Concentration Dependent Manner

Objective: To explore whether aspirin (ASA) enhances the sensitivity of hepatocellular carcinoma (HCC) side population (SP) cells to doxorubicin (Doxo) via miR-491/ATP-binding cassette sub-family G member 2 (ABCG2).Methods: Non-SP and SP cells were isolated from MHCC-97L cell line using flow cytometry analysis and fluorescence-activated cell sorting. Colony formation assay was performed to determine the colony-formation ability of cells. Cell viability of SP cells was determined with the MTT assay. Luciferase reporter assay was applied in confirming the binding between miR-491 and ABCG2.Results: Although the Doxo treatment lowered the colony-formation ability of both non-SP and SP cells, the colony-formation ability of SP cells was 2-fold higher than that of non-SP cells (P<0.05). Doxo slightly inhibited the cell viability of SP cells in a concentration-dependent manner; the addition of ASA dramatically enhanced the inhibitory effect of Doxo on SP cell viability in a concentration-dependent manner (P<0.05). Compared with non-SP cells, the miR-491 expression was significantly decreased in SP cells, which was significantly reversed by ASA (P<0.05). miR-491 directly controlled the ABCG2 expression. In the presence of Doxo, miR-491 inhibitor reduced the inhibitory effect of ASA on the cell viability of SP cells, which was significantly reversed by knockdown of ABCG2 (P<0.05).Conclusion: ASA enhanced the sensitivity of SP cells to Doxo via regulating the miR-491/ABCG2 signaling pathway.

scholarly journals Circular RNA circFAT1(e2) Promotes Colorectal Cancer Tumorigenesis via the miR-30e-5p/ITGA6 Axis

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Fei Pan ◽  
Dongqing Zhang ◽  
Na Li ◽  
Mei Liu
Keyword(s):  
Colorectal Cancer ◽  
Circular Rna ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Circular Rnas ◽  
Small Interfering Rnas ◽  
Regulatory Relationship ◽  
Downstream Target ◽  
Normal Tissues

circRNAs (circular RNAs) are a family of noncoding RNAs and have diverse physiological and pathological functions. However, the functions and mechanisms of circRNAs in the development and progression of colorectal cancer (CRC) remain largely unknown. Here, we aimed to explore the functions and roles of circFAT1(e2) in CRC. qRT-PCR revealed that circFAT1(e2) in CRC tumor tissues was upregulated compared with that in adjacent normal tissues and was also upregulated in CRC cell lines. Small interfering RNAs (siRNAs) against circFAT1(e2) were used to decrease the expression of circFAT1(e2) in HCT116 and RKO cells in vitro. The roles of circFAT1(e2) in CRC cell metastasis and proliferation were then determined by transwell and CCK-8 assays. The results showed that circFAT1(e2) silencing markedly suppressed CRC growth. Moreover, we identified circFAT1(e2) as a promoter of CRC metastasis. Knockdown of circFAT1(e2) evidently reduced HCT116 and RKO cell migration and invasion. Furthermore, the regulatory relationship between circFAT1(e2) and its target miRNAs was verified by a luciferase reporter assay. We demonstrated that circFAT1(e2) could sponge miR-30e-5p, which regulated the expression level of integrin α6 (ITGA6), the downstream target gene of miR-30e-5p. Rescue assays demonstrated that knockdown of miR-30e-5p enhanced CRC proliferation and migration via ITGA6. Taken together, our results reveal the novel oncogenic roles of circFAT1(e2) in CRC through the miR-30e-5p/ITGA6 axis.

Long Non-Coding RNA-Colon Cancer Associated Transcript 1 Down-Regulation Inhibits Cell Proliferation and Promotes Apoptosis in Colorectal Cancer Through Enhancing miR-152 Expression

2020 ◽  
Vol 10 (12) ◽  
pp. 1766-1772
Author(s):  
Jindong Li ◽  
Xi Wang ◽  
Xin Huang ◽  
Na Li ◽  
Ya Ling ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Viability ◽  
Cancer Cell ◽  
Cancer Progression ◽  
Biological Effects ◽  
Colorectal Cancer Cell ◽  
Luciferase Reporter ◽  
Pcr Analysis ◽  
Cancer Tissue ◽  
Colorectal Cancer Progression

Colorectal cancer is a common malignant cancer that is characterized by high mortality rate. CCAT1 is a type of newly discovered lncRNA. This research was conducted to study the role of CCAT1 in colorectal cancer. The findings showed that there was significant up-regulation of CCAT1 expression in colorectal cancer. Then, online bioinformatic database and dual-luciferase reporter assay to prove CCAT1 and miR-152 have direct binding sites. Many researches demonstrated that miR-152 played a crucial role in development of colorectal cancer. Therefore, we then explored the relationship between CCAT1 and miR-152 in colorectal cancer. qRT-PCR analysis showed that miR-152 was lowly expressed in cancer tissue and cells. We then explored the effect of CCAT1 and miR-152 on the biological effects of colorectal cancer cells. MiR-152 up-regulation significantly reduced colorectal cancer cell viability and enhanced apoptosis. Furthermore, CCAT1-shRNA inhibited colorectal cancer cell viability and enhanced cell apoptosis were significantly eliminated by miR-152 inhibitor. Combined with all results, CCAT1/miR-152 axis was related to colorectal cancer progression regulation, which might be used as new therapeutic targets for colorectal cancer treatment.

scholarly journals MALAT1 modulates miR-146’s protection of microvascular endothelial cells against LPS-induced NF-κB activation and inflammatory injury

2019 ◽  
Vol 25 (7) ◽  
pp. 433-443
Author(s):  
Lin-Lin Feng ◽  
Wei-Na Xin ◽  
Xiu-Li Tian
Keyword(s):  
Endothelial Cells ◽  
Cell Viability ◽  
Combined Treatment ◽  
Luciferase Assay ◽  
Luciferase Reporter ◽  
Microvascular Endothelial Cells ◽  
Dependent Manner ◽  
Inflammatory Injury ◽  
Tnf Α ◽  
Microvascular Endothelial

To investigate the role of miR-146 and its possible relationship with MALAT1 in LPS-induced inflammation in human microvascular endothelial cells (HMECs), HMEC-1 cells were treated with LPS to construct an inflammatory injury cell model, and the cell viability, TNF-α and IL-6 secretion and the expression levels of VCAM-1, SELE and ICAM-1 were analysed as markers of inflammatory injury. The regulation mechanisms of miR-146 interacted with MALAT1 and the downstream NF-κB signalling were also verified by dual-luciferase assay and knockdown technology. LPS significantly decreased the cell viability, increased levels of VCAM-1, SELE and ICAM-1 and also up-regulated miR-146a/b, TNF-α and IL-6 in a dose-dependent manner. Over-expression of miR-146a resulted in down-regulation of TNF-α and IL-6, as well as VCAM-1, SELE and ICAM-1, while inhibition of miR-146a led to opposite results. The dual-luciferase reporter assay showed both miR-146a and miR-146b directly targeted and negatively regulated the expression of MALAT1. Silencing of MALAT1 suppressed LPS-induced NF-κB activation and TNF-α and IL-6 secretion, reducing the cell inflammatory injury, but these changes were reversed after combined treatment with miR-146a inhibitor. Taken together, we demonstrate that miR-146 protects HMECs against inflammatory injury by inhibiting NF-κB activation. This process is modulated by MALAT1.

Circ_0007031 enhances tumor progression and promotes 5-fluorouracil resistance in colorectal cancer through regulating miR-133b/ABCC5 axis

2020 ◽  
Vol 29 (4) ◽  
pp. 531-542
Author(s):  
Xiaowen He ◽  
Jun Ma ◽  
Mingming Zhang ◽  
Jianhua Cui ◽  
Hao Yang
Keyword(s):  
Colorectal Cancer ◽  
Colony Formation ◽  
Human Cancer ◽  
Animal Experiments ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Circular Rnas ◽  
Cell Progression ◽  
Rna Immunoprecipitation

Colorectal cancer (CRC) remains one of the most commonly diagnosed malignancies worldwide. Circular RNAs (circRNAs) are being found to play crucial roles in human cancer, including CRC. The purpose of this study was to explore the function and mechanism of circ_0007031 on CRC progression and 5-fluorouracil (5-FU) resistance. The levels of circ_0007031, ATP-binding cassette subfamily C member 5 (ABCC5) and miR-133b were assessed by quantitative real-time polymerase chain reaction (qRT-PCR) or western blot. Cell survival and proliferation were detected by the 3-(4,5-dimethylthiazol-2yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay. Cell colony formation was evaluated using a standard colony formation assay. Transwell assays were performed to determine cell migration and invasion. Targeted correlations among circ_0007031, miR-133b and ABCC5 were verified by dual-luciferase reporter, RNA immunoprecipitation (RIP) and RNA pulldown assays. Animal experiments were performed to observe the role of circ_0007031 in vivo. Our data indicated that circ_0007031 up-regulation was associated with CRC resistance to 5-FU. Circ_0007031 knockdown repressed CRC cell proliferation, migration and invasion and enhanced 5-FU sensitivity. Circ_0007031 directly interacted with miR-133b. Moreover, circ_0007031 knockdown regulated CRC cell progression and 5-FU sensitivity by miR-133b. ABCC5 was a direct target of miR-133b, and circ_0007031 mediated ABCC5 expression via acting as a miR-133b sponge. Furthermore, miR-133b overexpression regulated CRC cell progression and sensitivity to 5-FU by down-regulating ABCC5. Additionally, circ_0007031 knockdown suppressed tumor growth in vivo. Our current work had led to the identification of circ_0007031 knockdown that repressed CRC cell malignant progression and enhanced 5-FU sensitivity via regulating ABCC5 expression by sponging miR-133b.

IKKα as a potential novel target for treatment of colorectal cancer.

2020 ◽  
Vol 38 (4_suppl) ◽  
pp. 174-174
Author(s):  
Jean A. Quinn ◽  
Meera Patel ◽  
Kathryn AF Pennel ◽  
Dustin Flanagan ◽  
Paul G. Horgan ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Viability ◽  
Mouse Models ◽  
Dependent Manner ◽  
Cancer Specific Survival ◽  
Similar Reduction ◽  
Novel Target ◽  
Dose Dependent ◽  
Tumour Differentiation

174 Background: Colorectal cancer (CRC) is a heterogeneous disease leading to different survival outcomes for patients with the same stage of disease. The non-canonical NF-κB pathway has been shown to have a key role in tumorigenesis, and the aim of this study was to investigate the role of IKKα, the main catalytic component of this pathway in CRC. Methods: A tissue microarray was retrospectively constructed from a patient cohort (1033) with stage I-III CRC who underwent surgery. IHC was utilised to examine cytoplasmic and punctate IKKα expression and determine any association with clincopathological features and cancer specific survival (CSS). To assess IKKα inhibition, organoids were prepared from wild type (WT) mouse colon, mouse models of CRC (Apc and Apc.KRAS.pT53.TGFbR2 (AKPT)) and patient derived human organoids. These were treated with an IKKα inhibitor, SU1433 and organoid size and cell viability assessed. Results: High cytoplasmic expression of IKKα was associated with increasing T stage (p = 0.012), poor tumour differentiation (p = 0.010), tumour necrosis (p = 0.013) and low proliferation status (p = 0.013) but was not associated with CSS. High punctate IKKα expression associated with tumour differentiation (p = 0.001), necrosis (p = 0.004), proliferation (p = 0.044) and MMR competence (p < 0.001) and was also significantly associated with reduced CSS (HR1.20 95%CI 1.02-1.42, p < 0.001). SU1433 did not significantly impact on WT (C57/B16) organoid viability up to a concentration of 1 uM, however organoid size and cell viability was significantly reduced in a dose dependent manner in organoids from both Apc and AKPT mouse models. A similar reduction was observed in patient derived human organoids. Conclusions: Punctate IKKα expression was associated with poor cancer specific survival in CRC patients, and inhibition with SU1433, impacted on CRC mouse and patient derived human organoid size and cell viability. These results suggest that, following further investigation and confirmation, IKKα may be employed as a novel therapeutic target in CRC.

scholarly journals hsa_circ_0000231 Promotes Colorectal Cancer Cell Growth Through Upregulation of CCND2 by IGF2BP3/miR-375 Dual Pathway

2020 ◽  
Author(s):  
Wei Zhang ◽  
Bo Wang ◽  
Yang Yang ◽  
Zhen Zhang ◽  
Quan Wang ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Proliferation ◽  
In Situ Hybridization ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Cancer Cell Growth ◽  
Normal Tissues

Abstract Background circular RNAs (circRNAs) recently have been emerged as vital regulators for involvement of initiation and progression of diverse kinds of human cancers. This study aimed to investigate the role of circRNAs in colorectal cancer (CRC). Methods The expression profile of circRNAs in 5 pairs of CRC tissues and adjacent normal tissues were analyzed by Microarray. Quantitative real-time PCR and in situ hybridization and Base Scope Assay were used to determine the level and prognostic values of hsa_circ_0000231. Then, functional experiments in vitro and in vivo were performed to investigate the effects of hsa_circ_0000231 on cell proliferation. Mechanistically, fluorescent in situ hybridization, dual luciferase reporter assay, RNA pull-down and RNA immunoprecipitation experiments were performed to confirm the interaction between hsa_circ_0000231 and IGF2BP3 or has_miR-375. Results hsa_circ_0000231 was evidently up-regulated in CRC primary tissues, which was indicated to poor prognosis of CRC patients. The results demonstrated that hsa_circ_0000231 could promote CRC cell proliferation as well as tumorigenesis in vitro and in vivo. Mechanistic analysis showed that hsa_circ_0000231 might on the one hand act as a ceRNA (competing endogenous RNA) of miR-375 to regulate cyclin D2 (CCND2), and on the other hand bind to IGF2BP3 protein to protect CCND2 from being degraded. Conclusion Our findings suggest that hsa_circ_0000231 facilitated CRC progression by sponging miR-375 or binding to IGF2BP3 to modulate CCND2. This discovery implied that has_circ_0000231 may be a potential new diagnostic and therapeutic biomarker for CRC.

scholarly journals Restoring HOXD10 Exhibits Therapeutic Potential for Ameliorating Malignant Progression and 5-Fluorouracil Resistance in Colorectal Cancer

2021 ◽  
Vol 11 ◽  
Author(s):  
Weijie Pan ◽  
Kaijing Wang ◽  
Jiayong Li ◽  
Hanhua Li ◽  
Yuchan Cai ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Lines ◽  
Therapeutic Potential ◽  
Methylation Status ◽  
Suppressor Gene ◽  
Resistant Cell ◽  
The Cancer Genome Atlas ◽  
Migration And Invasion ◽  
Dependent Manner ◽  
Normal Tissues

Emerging evidence suggests that hypermethylation of HOXD10 plays an important role in human cancers. However, the biological and clinical impacts of HOXD10 overmethylation and its downstream targets in colorectal cancer remain unknown. We evaluated the methylation level of HOXD10 in paired cancer and normal tissues (n = 42) by using pyrosequencing, followed by validation of the methylation status of HOXD10 from The Cancer Genome Atlas (TCGA) datasets with 302 cancer tissues and 38 normal tissues. The biological function of HOXD10 was characterized in cell lines. We further evaluated the effects of HOXD10 and its targets on chemoresistance in our established resistant cell lines and clinical cohort (n = 66). HOXD10 was found frequently methylated in colorectal cancer, and its hypermethylation correlates with its low expression level, advanced disease, and lymph node metastasis. Functionally, HOXD10 acts as a tumor suppressor gene, in which HOXD10-expressing cells showed suppressed cell proliferation, colony formation ability, and migration and invasion capacity. Mechanistically, DNMT1, DNMT3B, and MeCP2 were recruited in the HOXD10 promoter, and demethylation by 5-Aza-2′-deoxycytidine (5-Aza-CdR) treatment or MeCP2 knockdown can sufficiently induce HOXD10 expression. HOXD10 regulates the expressions of miR-7 and IGFBP3 in a promoter-dependent manner. Restoration of the expression of HOXD10 in 5-fluorouracil (5-FU)-resistant cells significantly upregulates the expressions of miR-7 and IGFBP3 and enhances chemosensitivity to 5-FU. In conclusion, we provide novel evidence that HOXD10 is frequently methylated, silenced, and contributes to the development of colorectal cancers. Restoration of HOXD10 activates the expressions of miR-7 and IGFBP3 and results in an inhibited phenotype biologically, suggesting its potential therapeutic relevance in colorectal cancer (CRC).

scholarly journals Melatonin Inhibits the Progression of Oral Squamous Cell Carcinoma via Inducing miR-25-5p Expression by Directly Targeting NEDD9

2020 ◽  
Vol 10 ◽  
Author(s):  
Yanling Wang ◽  
Bo Tao ◽  
Jiaying Li ◽  
Xiaoqun Mao ◽  
Wei He ◽  
...  
Keyword(s):  
Squamous Cell Carcinoma ◽  
Wound Healing ◽  
Oral Squamous Cell Carcinoma ◽  
Cell Viability ◽  
Cell Carcinoma ◽  
Squamous Cell ◽  
Colony Formation ◽  
Biological Behavior ◽  
Luciferase Reporter

Melatonin exerts anti-cancer roles in various types of cancers. However, to the best of our knowledge, its role in oral squamous cell carcinoma (OSCC) is unknown. The present study aimed to investigate the role of melatonin and its underlying mechanism in OSCC. MTT, colony formation, wound healing, and transwell invasion assays proved that melatonin played anti-tumor effects in OSCC cells by inhibiting cell viability, proliferation, migration, and invasion in a concentration-dependent manner. The RT-qPCR analysis showed that miR-25-5p was significantly upregulated after melatonin treatment. Further, miR-25-5p might be involved in melatonin-induced inhibitory effects on the biological behavior of OSCC. The expression of miR-25-5p was decreased in tumor tissues and OSCC cells detected by RT-qPCR. MTT assay, colony formation assay, and TUNEL staining indicated miR-25-5p overexpression inhibited OSCC cell viability, proliferation, and induced OSCC cell apoptosis. Furthermore, wound healing, transwell invasion assay, and animal experiments suggested that miR-25-5p might exert suppressive effects on the migration, invasion, and tumor formation of OSCC cells, while miR-25-5p knockdown exhibited the opposite effects in OSCC cells. Bioinformatics analysis, western blot analysis, and luciferase reporter assay suggested that neural precursor cell expressed developmentally downregulated protein 9 (NEDD9) was proved to be a putative target for miR-25-5p. The role of NEDD9 in inhibiting OSCC cell proliferation, invasion, and migration was verified with NEDD9 siRNA transfection. Thus, melatonin exerted anti-proliferative, anti-invasive, and anti-migrative effects on OSCC via miR-25-5p/NEDD9 pathway. Melatonin could be applied as a potential novel drug on treating OSCC.

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