scholarly journals Tumor-derived extracellular vesicles inhibit HGF/c-Met and EGF/EGFR pathways to accelerate the radiosensitivity of nasopharyngeal carcinoma cells via microRNA-142-5p delivery

2022 ◽  
Vol 8 (1) ◽  
Author(s):  
Changyu Zhu ◽  
Xiaolei Jiang ◽  
Hua Xiao ◽  
Jianmei Guan
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Extracellular Vesicles ◽  
Nude Mice ◽  
Peripheral Blood ◽  
Cell Apoptosis ◽  
Expression Patterns ◽  
Carcinoma Cells ◽  
Luciferase Reporter ◽  
Relationship Of ◽  
Dual Luciferase Reporter Assay

AbstractRadioresistance prevails as one of the largest obstacles in the clinical treatment of nasopharyngeal carcinoma (NPC). Meanwhile, tumor-derived extracellular vesicles (TEVs) possess the ability to manipulate radioresistance in NPC. However, its mechanism remains to be further explored. Therefore, the current study set out to explore the mechanism of microRNA (miR)-142-5p delivered by TEVs in regard to the radiosensitivity of NPC. Firstly, peripheral blood samples were collected from patients with radioresistance and radiosensitivity, followed by RT-qPCR detection of miR-142-5p expression. A dual-luciferase reporter assay was carried out to elucidate the targeting relationship of miR-142-5p with HGF and EGF. In addition, radiotherapy-resistant NPC cell models were established by screening NPC cells with gradient increasing radiation exposure, and co-incubated with EVs isolated from miR-142-5p mimic-transfected NPC cells, followed by overexpression of HGF and EGF. Moreover, cell viability was detected by means of MTS, cell proliferation with a colony formation assay, cell apoptosis with flow cytometry, and expression patterns of related genes with the help of Western blot analysis. NPC xenotransplantation models in nude mice were also established by subcutaneous injection of 5-8FR cells to determine apoptosis, tumorigenicity, and radiosensitivity in nude mice. It was found that miR-142-5p was poorly expressed in peripheral blood from NPC patients with radioresistance. Mechanistic experimentation illustrated that miR-142-5p inversely targeted HGF and EGF to inactivate the HGF/c-Met and EGF/EGFR pathways, respectively. NPC cell apoptosis was observed to be augmented, while their radioresistance and proliferation were restricted by EVs-miR-142-5p or HGF silencing, or EGF silencing. Furthermore, EVs-miR-142-5p inhibited growth and radioresistance and accelerated the apoptosis of radiotherapy-resistant NPC cells in nude mice by inhibiting the HGF/c-Met and EGF/EGFR pathways. Collectively, our findings indicated that TEVs might inhibit the HGF/c-Met and EGF/EGFR pathways by delivering miR-142-5p into radiotherapy-resistant NPC cells to enhance radiosensitivity in NPC.

scholarly journals LncRNA FOXD3-AS1/miR-135a-5p function in nasopharyngeal carcinoma cells

Open Medicine ◽  
2020 ◽  
Vol 15 (1) ◽  
pp. 1193-1201
Author(s):  
Zhang E ◽  
Chunli Li ◽  
Yuandi Xiang
Keyword(s):  
Cell Growth ◽  
Nasopharyngeal Carcinoma ◽  
Cell Apoptosis ◽  
Target Gene ◽  
Carcinoma Cells ◽  
Luciferase Reporter ◽  
Western Blot Assay ◽  
Direct Target ◽  
Polymerase Chain ◽  
Dual Luciferase Reporter Assay

AbstractThis research aimed to illustrate the biological function and associated regulatory mechanism of lncRNA FOXD3-AS1 (FOXD3-AS1) in nasopharyngeal carcinoma (NPC). This research initially found that FOXD3-AS1 was obviously upregulated in NPC cell lines by quantitative reverse transcription polymerase chain reaction (qRT-PCR) detection. Next, the direct target of FOXD3-AS1 was predicted by bioinformatics and further verified by dual-luciferase reporter assay. MiroRNA-135a-5p (miR-135a-5p) was identified as the target gene of FOXD3-AS1 and down-expressed in C666-1 cells compared to NP69. In addition, function assays were conducted in C666-1 cells, including methyl tetrazolium assay, flow cytometry, Caspase3 activity detection, and western blot assay. Our results suggested that miR-135a-5p upregulation inhibited NPC cell growth, enhanced cell apoptosis, promoted Caspase3 activity, increased cleaved-Caspase3, and reduced pro-Caspase3 level. Moreover, we found that FOXD3-AS1 knockdown notably inhibited C666-1 cell proliferation, increased cell apoptosis, enhanced Caspase3 activity, enhanced cleaved-Caspase3 expression, and suppressed pro-Caspase3 level in C666-1 cells. However, these findings were reversed in C666-1 cells by miR-135a-5p mimic co-transfection. To sum up, our data showed that FOXD3-AS1 knockdown regulated cell growth and apoptosis in NCP cells via altering miR-135a-5p expression, suggesting that FOXD3-AS1 might be a therapeutic target for NPC diagnosis and treatment.

scholarly journals miR-206 regulates a metabolic switch in nasopharyngeal carcinoma by suppressing HK2 expression

2020 ◽  
Author(s):  
Chunying Luo ◽  
Min Liu ◽  
Jianwei Zhang ◽  
Guoqiang Su ◽  
Zhonghua Wei
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Warburg Effect ◽  
Carcinoma Cells ◽  
Luciferase Reporter ◽  
Metabolic Shift ◽  
Invasion And Metastasis ◽  
Metabolic Switch ◽  
And Function ◽  
The Relationship

Abstract Background: Many studies have shown that microRNAs play key functions in nasopharyngeal carcinoma proliferation, invasion and metastasis. However, whether the dysregulated level of miRNAs contributes to the metabolic shift in nasopharyngeal carcinoma is not completely understood.Objectives: This study was conducted to explore the expression and function of miR-206 in nasopharyngeal carcinoma.Methods: miR-206 expression level was examined by real-time PCR. miR-206 inhibitor, mimics, and scrambled control were transiently transfected into nasopharyngeal carcinoma cells and their effects on colony formation, glucose uptake, and lactate secretion were observed in vitro. Moreover, the relationship between the levels of miR-206 and HK2 was examined by luciferase reporter and assay western blot.Results: In our study, we reported downregulation of miR-206 expression leads to metabolic change in nasopharyngeal carcinoma cells. miR-206 controls this function by enhancing HK2 expression. The enhancement of aerobic metabolism activity induced by miR-206 leads to the rapid proliferation of nasopharyngeal carcinoma cells.Conclusions: Our data demonstrated that miR-206 was involved in the regulation of Warburg effect in nasopharyngeal carcinoma by suppressing HK2 expression.

scholarly journals Loading MicroRNA-376c in Extracellular Vesicles Inhibits Properties of Non-Small Cell Lung Cancer Cells by Targeting YTHDF1

2020 ◽  
Vol 19 ◽  
pp. 153303382097752
Author(s):  
Jianying Zhou ◽  
Dan Xiao ◽  
Tingting Qiu ◽  
Jun Li ◽  
Zhentian Liu
Keyword(s):  
Lung Cancer ◽  
Small Cell Lung Cancer ◽  
Extracellular Vesicles ◽  
Cell Lung Cancer ◽  
Rna Binding ◽  
Biological Activities ◽  
Expression Patterns ◽  
Small Cell ◽  
Luciferase Reporter ◽  
Small Cell Lung

Objective: Extracellular vesicles (Evs) secreted from cells have been revealed to mediate signal transduction between cells. Nevertheless, the mechanisms through which molecules transported by EVs function remain to be elucidated. In the present study, the functional relevance of endothelial cells (ECs)-secreted Evs carrying microRNA-376c (miR-376c) in the biological activities of non-small cell lung cancer (NSCLC) cells was investigated, including the related mechanisms. Methods: Two cell lines with the highest YTH N6-methyladenosine (m6A) RNA binding protein 1 (YTHDF1) expression were selected for subsequent experiments. Cellular proliferation, migration, invasion and apoptosis were measured by EdU, wound healing, Transwell assays and flow cytometry, respectively. The binding relationship between miR-376c and YTHDF1 was analyzed by dual-luciferase reporter assays. The miR-376c, YTHDF1 and β-catenin expression was evaluated by qPCR assays and western blot assays. Results: The expression patterns of YTHDF1 were higher in NSCLC cells, whereas miR-376c was reduced versus the normal bronchial epithelial cells. Silencing of YTHDF1 repressed NSCLC cell proliferation, invasion and migration abilities, whereas enhanced apoptosis. miR-376c negatively modulated YTHDF1 expression. Under co-culture conditions, ECs transmitted miR-376c into NSCLC cells through Evs, and inhibited the intracellular YTHDF1 expression and the Wnt/β-catenin pathway activation. Rescue experiments revealed that YTHDF1 overexpression reversed the inhibitory role of miR-376c released by EC-Evs in NSCLC cells. Conclusion: EC-delivered Evs inhibit YTHDF1 expression and the Wnt/β-catenin pathway induction via miR-376c overexpression, thus inhibiting the malignant phenotypes of NSCLC cells.

miR-141 Regulates Proliferation and Apoptosis of Renal Clear Cell Carcinoma by Targeting Phosphatase and Tensin Homolog (PTEN)

2020 ◽  
Vol 10 (5) ◽  
pp. 676-681
Author(s):  
Jianjun Zhao ◽  
Chao Tan ◽  
Haibin Chen ◽  
Xiaolong Cheng ◽  
Limei Hui
Keyword(s):  
Cell Carcinoma ◽  
Cancer Cells ◽  
Renal Cancer ◽  
Cell Apoptosis ◽  
Akt Signaling ◽  
Pten Expression ◽  
Luciferase Reporter ◽  
Tumor Tissues ◽  
Pten Mrna ◽  
Relationship Of

Objective: PTEN inhibits the activity of PI3K/AKT pathway. Abnormal miR-141 expression is associated with kidney cancer. Bioinformatics analysis revealed a relationship of miR-141 with PTEN. This study assessed miR-141's role in renal cancer cells. Methods: The dual luciferase reporter gene assay validated the relationship of miR-141 with PTEN. The tumor tissues and adjacent tissues of patients with renal cell carcinoma were collected to measure miR-141 and PTEN level. A498 cells were divided into miR-NC group and miR-141 inhibitor group followed by analysis of the expressions of miR-141, PTEN and p-AKT, cell apoptosis and proliferation by flow cytometry. Results : There is a relationship of miR-141 with PTEN. Compared with those in adjacent tissues, miR-141 was upregulated and PTEN mRNA was downregulated in tumor tissues. There was a negative correlation between miR-141 and PTEN mRNA (r = – 0 646,P < 0 001). Compared with that in HK-2 cells, miR-141 expression was increased in RCC4 and A498 cells, with decreased PTEN expression. Transfection of miR-141 inhibitor significantly up-regulated PTEN in A498 cells, reduced PI3K/AKT signaling activity, decreased cell proliferation and colony formation, as well as promoted cell apoptosis. Conclusion: miR-141 participates in reducing PTEN expression and promoting the pathogenesis of renal cancer. Inhibiting miR-141 expression up-regulates PTEN, inhibits PI3K/AKT signaling, and attenuates proliferation and promotes apoptosis of renal cancer cells.

miR-9-5p Increases Sensitivity to Cisplatin in Thyroid Cancer Cells by Down-Regulating BRAF Expression

2019 ◽  
Vol 9 (6) ◽  
pp. 751-759
Author(s):  
Wanzhi Chen ◽  
Jichun Yu ◽  
Rong Xie ◽  
Meijun Zhong
Keyword(s):  
Cell Proliferation ◽  
Wound Healing ◽  
Thyroid Cancer ◽  
Cancer Cells ◽  
Cell Apoptosis ◽  
Cell Number ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Thyroid Cancer Cells ◽  
Dual Luciferase Reporter Assay

Objective: To explore the expression of miR-9-5p and BRAF in cisplatin resistant strain thyroid cancer cells and reversal effect of drug resistance as well as the possible mechanism. Methods: The cisplatin-resistant thyroid cancer cells (FTC-133/DDP and TPC-1/DDP) were respectively divided into 3 groups as NC, DDP and DDP + miRNA groups. Measuring cell proliferation by MTT assay and cell apoptosis by flow cytometry; Evaluating invasion cell number and wound healing rates by transwell and wound healing assay. The relative proteins (BRAF, Mek and Erk1/2) were measured by WB assay. The correlation between miR-9-5p and BRAF by dual-luciferase reporter assay in FTC-133/DDP and TPC-1/DDP cells. Results: In FTC-133/DDP and TPC-1/DDP cells experiment, compared with DDP group, with miR-9-5p supplement, the cell proliferation rats were significantly depressed with cell apoptosis increasing (P < 0.001, respectively); invasion cell number and wound healing rats were significantly down-regulation (P < 0.001, respectively) in DDP + miRNA groups. Meanwhile, the BRAF, Mek and Erk1/2 proteins expressions were significantly depressed in DDP + miRNA groups were significantly suppressed compared with those in DDP groups (P < 0.001, respectively). By dual-luciferase reporter assay, BRAF was the target gene of miR-9-5p in FTC133/DDP and TPC-1/DDP cells. Conclusion: miR-9-5p increases sensitivity to cisplatin in thyroid cancer cells by down-regulating BRAF expression.

scholarly journals NEAT1 Overexpression Indicates a Poor Prognosis and Induces Chemotherapy Resistance via the miR-491-5p/SOX3 Signaling Pathway in Ovarian Cancer

2021 ◽  
Vol 12 ◽  
Author(s):  
Xinzhuan Jia ◽  
Lan Wei ◽  
Zhengmao Zhang
Keyword(s):  
Ovarian Cancer ◽  
Cell Lines ◽  
Cell Apoptosis ◽  
Expression Patterns ◽  
Chemotherapy Resistance ◽  
Direct Interaction ◽  
Luciferase Reporter ◽  
Tissue Samples ◽  
Qrt Pcr

BackgroundAccumulated studies have reported that dysregulated long non-coding RNAs (lncRNAs) are crucial in ovarian cancer (OC) initiation and development. However, detailed biological functions of lncRNA NEAT1 during the progression of OC remains to be uncovered.PurposeOur aim was to identify the role of NEAT1 in cisplatin resistance of ovarian cancer and the underlying mechanisms.MethodsThe expression patterns of NEAT1 in OC cell lines and tissue samples were identified by qRT-PCR. The cisplatin (DDP) sensitivity of OC cells was detected by MTT and CCK8 assay, while OC cell apoptosis and cell cycle were detected using flow cytometer assays. In addition, effects of NEAT1 on tumor growth were determined by xenograft tumor model. Luciferase reporter assay was conducted to prove the regulatory relation of miR-491-5p, NEAT1, and SOX3. Importantly, the expression of NEAT1 in exosomes from cisplatin-resistant patients was also determined by using qRT-PCR.ResultsIn this study, upregulated NEAT1 was detected in OC cell lines and tissues. Meanwhile, NEAT1 was also increased in cisplatin-resistant OC cell lines and tissues. Upregulation of NEAT1 inhibited cisplatin-induced OC cell apoptosis and promoted cell proliferation, while knockdown of NEAT1 played the opposite role. These effects were also observed in vivo. Furthermore, direct interaction was observed between NEAT1 and miR-491-5p. NEAT1 led to the upregulation of miR-491-5p-targeted SOX3 mRNA. Importantly, this study also showed upregulated NEAT1 expression in serum exosomes derived from cisplatin-resistant patients.ConclusionNEAT1 is vital in the chemoresistance of ovarian cancer through regulating miR-491-5p/SOX3 pathway, showing that NEAT1 might be a potential target for OC resistance treatment.

scholarly journals MiR-375 gene therapy attenuates drug resistance of hepatocellular carcinoma cells by inhibiting cell autophagy

2020 ◽  
Author(s):  
Dan Wang ◽  
Jingbo Yang
Keyword(s):  
Hepatocellular Carcinoma ◽  
Drug Resistance ◽  
Western Blotting ◽  
Regulatory Mechanism ◽  
Negative Control ◽  
Carcinoma Cells ◽  
Luciferase Reporter ◽  
Sorafenib Treatment ◽  
Hcc Cells ◽  
Dual Luciferase Reporter Assay

Abstract Objective To probe into the regulatory mechanism of miR-375 in hepatocellular carcinoma (HCC) cells under sorafenib treatment. Methods Western blotting and qRT-PCR were applied to measure the expressions of miR-375 and SIRT5 in parental HCC cells (HepG2 and Huh7) and sorafenib-resistant HCC cells (HepG2/so and Huh7/so). HepG2/so cells were accordingly transfected with miR-375 mimic, miR-375 inhibitor, sh-SIRT5, pcDNA3.1-SIRT5 or negative control. Western blotting measured the expressions of p62, LC3I and LC3II in HCC cells. CCK-8 and flow cytometry assessed the survivability and apoptosis of HCC cells, respectively. Bioinformatics techniques and dual-luciferase reporter assay predicted and verified the targeting relationship between miR-375 and SIRT5. Results MiR-375 was under-expressed and SIRT5 was over-expressed in HCC cells. Autophagy inhibitor impaired the survival of HepG2/so cells transfected with miR-375 inhibitor. Autophagy activator enhanced the drug resistance of HepG2/so cells transfected with miR-375 mimic. MiR-375 suppressed the drug resistance of HepG2/so cells by inhibiting autophagy. SIRT5 enhanced the drug resistance of HepG2/so cells by promoting autophagy and it could be targeted by miR-375. Conclusion MiR-375 suppresses autophagy to attenuate the drug resistance of HCC cells by regulating SIRT5. The findings of this study may provide new therapeutic targets for treating hepatocellular carcinoma.

MiR-30a-5p inhibits proliferation, migration and invasion of nasopharyngeal carcinoma cells by targeting NUCB2

2021 ◽  
pp. 096032712199191
Author(s):  
M Li ◽  
Y Wang ◽  
Q Zhao ◽  
W Ma ◽  
J Liu
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Tumor Growth ◽  
Carcinoma Cells ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Binding Interaction ◽  
Downstream Target ◽  
Downstream Target Gene

Background: Nasopharyngeal carcinoma (NPC) is a malignant head and neck tumor arising in the nasopharynx. MicroRNAs (miRNAs) are elucidated to exert tumor-suppressing function in human cancers. Numerous studies have manifested that miR-30a-5p serves as an anti-oncogene in various cancers. Objective: To research the biological function and molecular mechanism of miR-30a-5p in NPC. Methods: The morphology of NPC tissues was revealed by H&E staining. Transwell and wound healing assays were applied to investigate the effects of miR-30a-5p on NPC cell migration. The binding interaction between miR-30a-5p and nucleobindin 2 (NUCB2) was identified by luciferase reporter assay. Xenograft nude mice were used to detect the influence of miR-30a-5p on NPC tumor growth. Results: MiR-30a-5p was downregulated in NPC tissues and cells. The overexpression ofmiR-30a-5p inhibited proliferation, migration and invasion abilities of NPC cells. Moreover, NUCB2 was revealed to be a downstream target gene of miR-30a-5p, and knockdown of NUCB2 repressed the malignant behaviors of NPC cells and tumor growth. Additionally, rescue experiments revealed that miR-30a-5p suppressed the proliferation, migration and invasion of NPC cells via targeting NUCB2 in vitro. Meanwhile, in vivo assays depicted that NUCB2 overexpression rescued the effects induced by miR-30a-5p upregulation on tumor growth. Conclusion: MiR-30a-5p modulates NPC progression by targeting NUCB2. These findings lay a foundation for exploring the clinical treatment of NPC.

Sign in / Sign up

Export Citation Format

Share Document