scholarly journals IL-1β-Mediated Up-Regulation of WT1D via miR-144-3p and Their Synergistic Effect with NF-κB/COX-2/HIF-1α Pathway on Cell Proliferation in LUAD

2018 ◽  
Vol 48 (6) ◽  
pp. 2493-2502 ◽  
Author(s):  
Chen Wu ◽  
Xiaodong Li ◽  
Dachuan Zhang ◽  
Bin Xu ◽  
Wenwei Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
De Novo ◽  
Lentiviral Vector ◽  
Mrna Level ◽  
A549 Cells ◽  
Xenograft Model ◽  
Suppressive Effect ◽  
Luciferase Reporter ◽  
Transcriptional Level ◽  
Cox 2

Background/Aims: IL-1β is an important mediator of “inflammation-cancer" transformation through IL-1β/NF-κB/COX-2/HIF-1α signaling pathway, whereas certain portion of patients with lung adenocarcinoma (LUAD) still suffer from rapid tumor progression in clinical practice, indicating the occurrence of potential bypass. Methods: Real-time polymerase chain reaction was applied to examine the expressions of mir-144-3p, WT1, NF-κB, COX2 and HIF-1α at the mRNA level in 127 LUAD samples and corresponding adjacent tissues. miR-144-3p mimic and antagormiR were used to trigger activation and suppression of miR-144-3p in A549 cells, respectively. MTT assay and Western blotting analysis were carried out to evaluate the cell proliferation. Stable clones with over-expression or knockdown of WT1 were generated with plasmid or shRNA by lentiviral vector technology in H1568 and H1650 NSCLC cell lines, respectively. Dual luciferase reporter assay was performed to validate the effect of miR-144-3p on WT1D. Xenograft model was established for in vivo experiment, and TCGA data were extracted for validation. Results: miR-144-3p could suppress the WT1D expression at the post-transcriptional level, hence regulating cell proliferation in LUAD. WT1 and COX-2 were independent prognostic factors of LUAD patients. In addition, inhibition of IL-1β/miR-144-3p/WT1D and IL-1β/NF-κB/COX-2/HIF-1α pathways using miR-144-3p mimic and Celecoxib, respectively, displayed synergistic suppressive effect on cell proliferation in LUAD. Conclusion: A de novo IL-1β/miR-144-3p/WT1D axis was involved in proliferative regulation of LUAD. Moreover, simultaneous blockade of both IL-1β/miR-144-3p/WT1D and IL-1β/NF-κB/COX-2/ HIF-1α pathways might have synergistic suppressive effect on cell proliferation in LUAD.

scholarly journals CircFOXM1 promotes the proliferation, migration, invasion, and glutaminolysis of glioblastoma by regulating the miR-577/E2F5 axis

2021 ◽  
Author(s):  
Xuhui Fan ◽  
Meng Liu ◽  
Li Fei ◽  
Zhihui Huang ◽  
Yufeng Yan
Keyword(s):  
Cell Proliferation ◽  
Xenograft Model ◽  
Circular Rna ◽  
Suppressive Effect ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
In Vivo Experiments

Circular RNA (circRNA) is a key regulator of tumor progression. However, the role of circFOXM1 in glioblastoma (GBM) progression is unclear. The aim of this study was to investigate the role of circFOXM1 in GBM progression. The expression levels of circFOXM1, miR-577 and E2F transcription factor 5 (E2F5) were examined by real-time quantitative PCR. Cell counting kit 8 assay, EdU staining and transwell assay were used to detect cell proliferation, migration, and invasion. The levels of glutamine, glutamate and α-ketoglutarate were determined to evaluate the glutaminolysis ability of cells. Protein expression was tested by western blot analysis. Dual-luciferase reporter assay, RNA pull-down assay and RNA immunoprecipitation assay were employed to verify the interaction between miR-577 and circFOXM1 or E2F5. Mice xenograft model for GBM was constructed to perform in vivo experiments. Our results showed that circFOXM1 was highly expressed in GBM tumor tissues and cells. Silencing of circFOXM1 inhibited GBM cell proliferation, migration, invasion, glutaminolysis, as well as tumor growth. MiR-577 could be sponged by circFOXM1, and its inhibitor could reverse the suppressive effect of circFOXM1 downregulation on GBM progression. E2F5 was a target of miR-577, and the effect of its knockdown on GBM progression was consistent with that of circFOXM1 silencing. CircFOXM1 positively regulated E2F5 expression, while miR-577 negatively regulated E2F5 expression. In conclusion, our data confirmed that circFOXM1 could serve as a sponge of miR-577 to enhance the progression of GBM by targeting E2F5, which revealed that circFOXM1 might be a biomarker for GBM treatment.

scholarly journals MiR-188-3p and miR-133b Suppress Cell Proliferation in Human Hepatocellular Carcinoma via Post-Transcriptional Suppression of NDRG1

2021 ◽  
Vol 20 ◽  
pp. 153303382110330
Author(s):  
Zhenzhao Luo ◽  
Yue Fan ◽  
Xianchang Liu ◽  
Shuiyi Liu ◽  
Xiaoyu Kong ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Proliferation ◽  
Cell Migration ◽  
Cell Growth ◽  
Suppressive Effect ◽  
Luciferase Reporter ◽  
Invasion And Migration ◽  
And Migration ◽  
Cancer Tissues ◽  
Suppress Cell Proliferation

Background: Previous studies reported that N-myc downstream-regulated gene 1 (NDRG1) was upregulated in various cancer tissues and decreased expression of miR-188-3p and miR-133b could suppress cell proliferation, metastasis, and invasion and induce apoptosis of cancer cells. However, the molecular mechanism of NRDG1 involved in hepatocellular carcinoma (HCC) tumorigenesis is still unknown. Methods: The expressions of miR-188-3p, miR-133b, and NRDG1 in HCC tissues and cells were quantified by qRT-PCR and Western blot. MTT assay and transwell invasion assay were performed to evaluate cell growth and cell migration, respectively. Luciferase reporter assay were performed to determine whether miR-188-3p and miR-133b could directly bind to NRDG1 in HCC cells. Results: The results showed that NRDG1 was upregulated and these 2 microRNAs were downregulated in HCC tissues. NRDG1 was negatively correlated with miR-188-3p and miR-133b in HCC tissues. MiR-188-3p and miR-133b were demonstrated to directly bind to 3′UTR of NRDG1 and inhibit its expression. Upregulation of miR-188-3p and miR-133b reduced NRDG1 expression in hepatocellular carcinoma cell lines, which consequently inhibited cell growth and cell migration. Conclusions: Our finding suggested that miR-188-3p and miR-133b exert a suppressive effect on hepatocellular carcinoma proliferation, invasion, and migration through downregulation of NDRG1.

scholarly journals METTL14 promotes tumorigenesis by regulating lncRNA OIP5-AS1/miR-98/ADAMTS8 signaling in papillary thyroid cancer

2021 ◽  
Vol 12 (6) ◽  
Author(s):  
Xiaoping Zhang ◽  
Dan Li ◽  
Chengyou Jia ◽  
Haidong Cai ◽  
Zhongwei Lv ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Thyroid Cancer ◽  
Papillary Thyroid Cancer ◽  
Xenograft Model ◽  
Luciferase Reporter ◽  
Papillary Thyroid ◽  
Qrt Pcr ◽  
The Relationship

Abstract Background Papillary thyroid cancer (PTC) is the most common type of cancer of the endocrine system. Long noncoding RNAs (lncRNAs) are emerging as a novel class of gene expression regulators associated with tumorigenesis. Through preexisting databases available for differentially expressed lncRNAs in PTC, we uncovered that lncRNA OIP5-AS1 was significantly upregulated in PTC tissues. However, the function and the underlying mechanism of OIP5-AS1 in PTC are poorly understood. Methods Expression of lncRNA OIP5-AS1 and miR-98 in PTC tissue and cells were measured by quantitative real-time PCR (qRT-PCR). And expression of METTL14 and ADAMTS8 in PTC tissue and cells were measured by qRT-PCR and western blot. The biological functions of METTL14, OIP5-AS1, and ADAMTS8 were examined using MTT, colony formation, transwell, and wound healing assays in PTC cells. The relationship between METTL14 and OIP5-AS1 were evaluated using RNA immunoprecipitation (RIP) and RNA pull down assay. And the relationship between miR-98 and ADAMTS8 were examined by luciferase reporter assay. For in vivo experiments, a xenograft model was used to investigate the effects of OIP5-AS1 and ADAMTS8 in PTC. Results Functional validation revealed that OIP5-AS1 overexpression promotes PTC cell proliferation, migration/invasion in vitro and in vivo, while OIP5-AS1 knockdown shows an opposite effect. Mechanistically, OIP5-AS1 acts as a target of miR-98, which activates ADAMTS8. OIP5-AS1 promotes PTC cell progression through miR-98/ADAMTS8 and EGFR, MEK/ERK pathways. Furthermore, RIP and RNA pull down assays identified OIP5-AS1 as the downstream target of METTL14. Overexpression of METTL14 suppresses PTC cell proliferation and migration/invasion through inhibiting OIP5-AS1 expression and regulating EGFR, MEK/ERK pathways. Conclusions Collectively, our findings demonstrate that OIP5-AS1 is a METTL14-regulated lncRNA that plays an important role in PTC progression and offers new insights into the regulatory mechanisms underlying PTC development.

scholarly journals Circ-BANP Contributes to Cell Carcinogenesis and Aerobic Glycolysis by Regulating MAPK1 Through miR-874-3p in Colorectal Cancer

2021 ◽  
Author(s):  
Yunxin Zhang ◽  
Kexin Shen ◽  
Hanyi Zha ◽  
Wentao Zhang ◽  
Haishan Zhang
Keyword(s):  
Colorectal Cancer ◽  
Cell Proliferation ◽  
Aerobic Glycolysis ◽  
Lactate Production ◽  
Xenograft Model ◽  
Mitogen Activated Protein Kinase ◽  
Cell Counting ◽  
Nuclear Antigen ◽  
Luciferase Reporter

Abstract BackgroundCircular RNA-BTG3 associated nuclear protein (circ-BANP) was identifified to involve in cell proliferation of colorectal cancer (CRC). The aerobic glycolysis is a key metabolism mediating cancer progression. However, the role of circ-BANP on aerobic glycolysis in CRC remains unknown. MethodsThe expression of circ-BANP, microRNA (miR)-874-3p, and mitogen-activated protein kinase 1 (MAPK1) mNRA was detected using quantitative real-time polymerase chain reaction. Cell viability and invasion were measured by cell counting kit-8 assay or transwell assay. Glucose consumption and lactate production were assessed by a glucose and lactate assay kit. XF Extracellular Flux Analyzer was used to determine extracellular acidifification rate (ECAR). Western blot was used to analyze the levels of hexokinase-2 (HK2), pyruvate kinase M2 (PKM2), MAPK1, proliferating cell nuclear antigen (PCNA), Cyclin D1, N-cadherin, E-cadherin, hypoxia inducible factor-1α (HIF-1α), glucose transport protein 1(GLUT1), and c-Myc. The interaction between miR-874-3p and circ-BANP or MAPK1 was confifirmed by dual luciferase reporter assay. In vivo experiments were conducted through the murine xenograft model. ResultsCirc-BANP was up-regulated in CRC tissues and cell lines. Circ-BANP knockdown suppressed CRC cell proliferation, invasion and aerobic glycolysis in vitro as well as inhibited tumor growth in vivo. Circ-BANP was a sponge of miR-874-3p and performed anti-tumor effffects by binding to miR-874-3p in CRC cells. Subsequently, we confifirmed MAPK1 was a target of miR-874-3p and circ-BANP indirectly regulated MAPK1 expression by sponging miR-874-3p. After that, we found MAPK1 overexpression partially reversed circ-BANP deletion-mediated inhibition on cell carcinogenesis and aerobic glycolysis in CRC. ConclusionCirc-BANP accelerated cell carcinogenesis and aerobic glycolysis by regulating MAPK1 through miR- 874-3p in CRC, suggesting a promising therapeutic strategy for CRC treatment.

MiR-4458 inhibits breast cancer cell growth, migration, and invasiveness by targeting CPSF4

2019 ◽  
Vol 97 (6) ◽  
pp. 722-730 ◽  
Author(s):  
Jianrong Wu ◽  
Juan Miao ◽  
Ye Ding ◽  
Yayun Zhang ◽  
Xiaohao Huang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Proliferation ◽  
Cell Growth ◽  
Target Genes ◽  
Human Lung ◽  
Lung Tumor ◽  
Luciferase Reporter ◽  
Bioinformatics Analyses ◽  
Cox 2

Numerous studies have reported that CPSF4 is over-expressed in a large percentage of human lung cancers, and CPSF4 has been identified as a potential oncogene of human lung tumor. Downregulation of CPSF4 inhibits the proliferation and promotes the apoptosis of lung adenocarcinoma cells. A previous study by our group also found overexpression of CPSF4 in breast cancer (BC), and was closely associated with a poor prognosis for the patient. This study investigates microRNAs (miRNAs) that target CPSF4 to modulate BC cell proliferation. We found that miR-4458 was noticeably reduced in BC tissues and cells. Using a miR-4458 mimic, we found that cell proliferation, migration, and invasiveness were suppressed by miR-4458 overexpression, and were enhanced by reducing the expression of miR-4458. Moreover, the results from bioinformatics analyses suggest a putative target site in the CPSF4 3′-UTR. Furthermore, using luciferase reporter assays and Western blotting, we verified that miR-4458 directly targets the 3′-UTR of CPSF4 and downregulates COX-2 and h-TERT, which are downstream target genes of CPSF4. Additionally, PI3K/AKT and ERK were shown to be inhibited by miR-4458 overexpression in BC cells. Moreover, miR-4458 suppresses BC cell growth in vivo. Consequently, these results suggest that the miR-4458–CPSF4–COX-2–hTERT axis might serve as a potential target for the treatment of BC patients.

scholarly journals MicroRNA-381 Negatively Regulates TLR4 Signaling in A549 Cells in Response to LPS Stimulation

2015 ◽  
Vol 2015 ◽  
pp. 1-8 ◽  
Author(s):  
Zhihao Xu ◽  
Dapeng Dong ◽  
Xiaofei Chen ◽  
Huaqiong Huang ◽  
Shenglan Wen
Keyword(s):  
Target Gene ◽  
Target Genes ◽  
Bioinformatics Analysis ◽  
Respiratory Infections ◽  
A549 Cells ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Elisa Kit ◽  
Cox 2

It is widely reported that miR-381 is dysregulated in various tumors. However, the specific role of miR-381 in respiratory infections has not been reported. To probe this role, A549 cells were pretreated with 1 μg/mL LPS for 24 h. The level of miR-381 was detected using RT-qPCR. The expression of proinflammatory cytokines was determined using an ELISA kit and western blotting. Bioinformatics analysis was used to predict the target genes of miR-381, and a luciferase reporter assay was used to validate the expression of the target genes. miR-381 expression was increased in A549 cells treated with LPS, which is a ligand of TLRs. Further study revealed that the overexpression of miR-381 increased the activity of NF-κB signaling, thereby increasing the expression of IL-6, TNFα, and COX-2. Further study revealed that IκBα was a target gene of miR-381. The upregulation of miR-381 under LPS stimulation contributes to respiratory infections mainly by targeting IκBα.

Effect of 5-Fluorouracil on the Regulation of Zebrafish Thymidylate Synthase Gene Expression

2013 ◽  
Vol 641-642 ◽  
pp. 732-735
Author(s):  
Chun Xia Song ◽  
Qian Zhang ◽  
Yu Liang Xiao
Keyword(s):  
Gene Expression ◽  
Thymidylate Synthase ◽  
De Novo ◽  
Mrna Level ◽  
Translation System ◽  
Transcriptional Level ◽  
Important Target ◽  
Reticulocyte Lysate ◽  
First Time ◽  
Translational Level

Thymidylate synthase (TS) is an important target in cancer therapy, which is a folate-dependent enyme, catalyzing the de novo synthesis of dUMP. In this report, the effect of 5-flurorouracil (5-FU) on the regulation of TS gene expression was estimated in zebrafish. The results showed 5-FU could significantly increase the TS expression in zebrafish embryos. However, TS mRNA level were remained unchanged. To determine the effect of 5-FU and 5-FdUMP on translation of TS mRNA, a rabbit reticulocyte lysate translation system was used. Addition of 5-FU, not inhibited the translation of TS mRNA. While addition of 5-FdUMP, completely repressed the translation of TS mRNA. Therefore, induced expression of thymidylate synthase by 5-FU in zebrafish occurred in translational level, not in transcriptional level. The findings demonstrated that zebrafish TS protein was able to bind to its own cogate mRNA and the 5-FU regulated TS in the translational level. This is the first time to confirm that the regulation of TS is affected by TS and its cognant mRNA interaction in the whole animal level.

scholarly journals MicroRNA-224 Promotes Pancreatic Cancer Cell Proliferation and Migration by Targeting the TXNIP-Mediated HIF1α Pathway

2018 ◽  
Vol 48 (4) ◽  
pp. 1735-1746 ◽  
Author(s):  
Guanghui Zhu ◽  
Lianming Zhou ◽  
Haijun Liu ◽  
Yuanzhou Shan ◽  
Xueli Zhang
Keyword(s):  
Cell Proliferation ◽  
Nuclear Export ◽  
Xenograft Model ◽  
Ductal Adenocarcinoma ◽  
Luciferase Reporter ◽  
Mouse Xenograft Model ◽  
And Migration ◽  
Proliferation And Migration

Background/Aims: MicroRNAs (miRNAs) have been shown to participate in the development of pancreatic ductal adenocarcinoma (PDAC) by modulating multiple cellular processes. Increased miR-224 expression enhances proliferation and metastasis in human cancers. This study aimed to investigate the role of miR-224 and its underlying mechanism of action in PDAC. Methods: BrdU, MTT, and cell migration assays were performed to determine cell proliferation, viability, and migration, respectively. The binding sites of miR-224 were identified using a luciferase reporter system, whereas protein expression of target genes was determined by immunoblotting and immunofluorescence analyses. A BALB/c nude mouse xenograft model was used to evaluate the role of miR-224 in vivo. Results: We demonstrated that miR-224 expression was enhanced in PDAC cells and tissues, and was related to migration and proliferation. Noticeably, miR-224 overexpression promoted the proliferation, migration, and metastasis of Panc1 cells, while miR-224 inhibition had the reverse effect on PDAC cells. Moreover, we found that thioredoxin-interacting protein (TXNIP) is a target of miR-224. The results also indicated that miR-224 inversely regulated TXNIP by binding directly to its 3′-untranslated region, which resulted in the activation of hypoxia-inducible factor 1α (HIF1α). Further, either TXNIP re-expression or HIF1α depletion abolished the effects of miR-224 on the proliferation and migration of PDAC cells in vitro and in vivo. Regarding the relationship of TXNIP and HIF1α, we found that TXNIP mediated the nuclear export of HIF1α and its degradation by forming a complex with HIF1α. Conclusion: The miR-224-TXNIP-HIF1α axis may be useful in developing novel therapies for PDAC.

scholarly journals Identification of a Previously Unrecognized Promoter That Drives Expression of the UXP Transcription Unit in the Human Adenovirus Type 5 Genome

2010 ◽  
Vol 84 (21) ◽  
pp. 11470-11478 ◽  
Author(s):  
Baoling Ying ◽  
Ann E. Tollefson ◽  
William S. M. Wold
Keyword(s):  
Transcription Initiation ◽  
Promoter Activity ◽  
Mrna Level ◽  
A549 Cells ◽  
Initiation Site ◽  
Transcription Unit ◽  
Transcription Initiation Site ◽  
Luciferase Reporter ◽  
Rnase Protection ◽  
Ccaat Box

ABSTRACT We previously identified an adenovirus (Ad) protein named U exon protein (UXP) encoded by a leftward-strand (l-strand) transcription unit. Here we identify and characterize the UXP promoter. Primer extension and RNase protection assays mapped the transcription initiation site at 32 nucleotides upstream of the UXP gene initiation codon. A series of viral mutants with mutations at two putative inverted CCAAT (I-CCAAT) boxes and two E2F sites were generated. With mutants lacking the proximal I-CCAAT box, the UXP mRNA level decreased significantly to 30% of the Ad type 5 (Ad5) mRNA level as measured by quantitative reverse transcription-PCR. Decreased UXP was also observed by immunoblotting and immunofluorescence. UXP mRNA and protein levels were similar to those of Ad5 for mutants lacking the distal I-CCAAT box or both putative E2F sites. Ad DNA levels were similar in mutant- and wild-type Ad5-infected cells during the late stage of infection, strongly suggesting that the decreased UXP mRNA and protein from mutants lacking the proximal I-CCAAT box was due to decreased promoter activity. Electrophoretic mobility shift assays (EMSA) indicated that a cellular factor binds specifically to the proximal I-CCAAT box of the UXP promoter. An in vitro luciferase reporter assay demonstrated that basal promoter activity lies between bp −158 and +30 of the transcription initiation site. No E1A-mediated promoter transactivation was observed in 293 cells compared with A549 cells. Thus, we propose that there is a previously unidentified Ad5 promoter that drives expression of the UXP transcription unit. This promoter is embedded within the gene for fiber, and it contains a proximal I-CCAAT box critical for UXP mRNA transcription.

scholarly journals Downregulation of circ-YES1 Suppresses NSCLC Migration and Proliferation Through The miR-142-3p–HMGB1 Axis

2020 ◽  
Author(s):  
Yongbin Chi ◽  
Yan Wang ◽  
Dawei Zhou ◽  
Wanchao Liu ◽  
Ruodong Han ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Tumor Development ◽  
Xenograft Model ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Downstream Targets ◽  
Cell Proliferation And Migration ◽  
Mouse Xenograft Model ◽  
And Migration ◽  
Proliferation And Migration

Abstract Background: Circular RNAs (circRNAs) are a new family of abundant regulatory RNAs with roles in various types of cancer. While the hsa_circ_0046701 (circ-YES1) function in non-small cell lung cancer (NSCLC) is unclear. Methods: Circ-YES1 expression in normal pulmonary epithelial and NSCLC cells was examined. The small interfering RNA for circ-YES1 was prepared, cell proliferation and migration were assessed. Tumorigenesis in nude mice was assayed to validate the role of circ-YES1. Bioinformatics analyses and luciferase reporter assays were utilized to identify downstream targets of circ-YES1. Results: Compared to normal pulmonary epithelial cells, the circ-YES1 expression increased in NSCLC cells, and cell proliferation and migration were suppressed after circ-YES1 knockdown. Both high mobility group protein B1 (HMGB1) and miR-142-3p were found to be downstream targets of circ-YES1, and miR-142-3p inhibition and HMGB1 overexpression reversed the effects of circ-YES1 knockdown on cell proliferation and migration. Similarly, HMGB1 overexpression reversed the miR-142-3p overexpression effects on these two processes. The imaging experiment results revealed that circ-YES1 knockdown impeded tumor development and metastasis in a nude mouse xenograft model. Conclusion: Taken together, our results show that circ-YES1 promotes tumor development through the miR-142-3p–HMGB1 axis and support the development of circ-YES1 as a new therapeutic NSCLC target.

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