scholarly journals GATA1-induced upregulation of LINC01503 promotes carboplatin resistance in ovarian carcinoma by upregulating PD-L1 via sponging miR-766-5p

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Yao Li ◽  
Yan Zhai ◽  
Yuxuan Chen
Keyword(s):  
Ovarian Carcinoma ◽  
Inhibitory Concentration ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Invasive Ability ◽  
Protein Levels ◽  
Reverse Transcription Quantitative Pcr ◽  
Non Coding Rnas ◽  
Dual Luciferase Reporter Assay

Abstract Background Ovarian Carcinoma (OCa) is a high-mortality malignancy derived from female reproductive system. Increasing evidence has identified long non-coding RNAs (lncRNAs) as important regulators in OCa chemoresistance. In this study, we intended to explore the role of LINC01503 in OCa resistance to carboplatin (CBP). Methods Gene expression was measured by reverse transcription-quantitative PCR (RT-qPCR) in OCa cells. Western blot was adopted to detect protein levels of GATA1, PD-L1, E-cadherin, N-cadherin, Vimentin, Bcl-2, Bax, cleaved caspase-3. To assess the effects of LINC01503 on the resistance of OCa cells to CBP, Cell Counting Kit-8 (CCK-8), colony formation, Transwell, and flow cytometry experiments were performed to evaluate half-maximal inhibitory concentration (IC50), cell viability, migrative and invasive ability, as well as cell apoptosis. Dual-luciferase reporter assay was employed to assess the associations between the genes. Results LINC01503 was upregulated in CBP-resistant OCa cells. LINC01503 knockdown reduced CBP resistance in OCa cells. Besides, GATA-binding protein 1 (GATA1) activated LINC01503 transcription in CBP-resistant OCa cells. MiR-766-5p was lowly expressed in CBP-resistant cells and confirmed as a target for LINC01503. In addition, miR-766-5p overexpression increased CBP sensitivity in OCa cells. PD-L1 was verified as the target of miR-766-5p. Besides, LINC01503 upregulated PD-L1 level by regulating miR-766-5p. Furthermore, rescue experiments showed that PD-L1 overexpression abrogated the inhibited impacts of blocking LINC01503 on CBP resistance in OCa cells. Conclusion GATA1-induced LINC01503 expedited CBP resistance in OCa cells via the miR-766-5p/PD-L1 axis, providing a new target for improving the efficacy of OCa chemotherapy. Graphical Abstract

scholarly journals LncRNA SNHG14 regulates the DDP-resistance of non-small cell lung cancer cell through miR-133a/HOXB13 pathway

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Li Xu ◽  
Yan Xu ◽  
Min Yang ◽  
Jia Li ◽  
Fang Xu ◽  
...  
Keyword(s):  
Cell Line ◽  
A549 Cells ◽  
Parental Cell ◽  
Luciferase Reporter ◽  
Inhibitory Effects ◽  
Through Flow ◽  
Non Coding Rnas ◽  
Apoptosis Related Protein ◽  
Dual Luciferase Reporter Assay

Abstract Background Recently, long non-coding RNAs (lncRNAs) have been reported to be involved in regulating chemo-resistance of NSCLC, however, the role of lncRNA SNHG14 in the DDP-resistance of NSCLC remains unexplored. Methods Relative expression of SNHG14, HOXB13 and miR-133a in DDP-resistant A549 (A549/DDP) cell and its parental cell A549 were measured using qRT-PCR. Cell proliferation viability of indicated A549/DDP cell was estimated via CCK-8 and colony formation experiments. Cell cycle and apoptosis were analyzed through flow cytometry. Expression of apoptosis-related protein and HOXB13 were detected via western blot. The interaction among SNHG14, HOXB13 and miR-133a was predicted by bioinformatics and validated by dual-luciferase reporter assay. Results LncRNA SNHG14 and HOXB13 were upregulated while miR-133a was downregulated in A549/DDP cell line compared to A549 cell line. SNHG14 knockdown or miR-133a overexpression was demonstrated to increase the DDP-sensitivity of A549/DDP cells. SNHG14 was revealed to compete with HOXB13 for miR-133a binding in A549/DDP cells. Inhibition of miR-133a in A549 cells could reverse the promotive effects of SNHG14 knockdown on DDP-sensitivity, as well as the inhibitory effects on HOXB13 expression. HOXB13 overexpression was revealed to abolish the enhanced effects of miR-133a on the sensitivity of A549/DDP cell to DDP. Conclusion Our findings demonstrated that SNHG14 was involved in the development of DDP-resistance of A549/DDP cells through miR-133a/HOXB13 axis, which may present a path to novel therapeutic stratagems for DDP resistance of NSCLC.

scholarly journals Long noncoding RNA LBX2-AS1-modulated miR-4766-5p regulates gastric cancer development through targeting CXCL5

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
LiPan Peng ◽  
ZeZhong Chen ◽  
GuangChuan Wang ◽  
ShuBo Tian ◽  
Shuai Kong ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Proliferation ◽  
Noncoding Rna ◽  
Mrna Level ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Polymerase Chain ◽  
Dual Luciferase Reporter Assay

Abstract Background Long noncoding RNAs (LncRNAs) have been reported to critically regulate gastric cancer (GC). Recently, it was reported that LBX2 antisense RNA 1 (LBX2-AS1) is abnormally expressed in GC. However, the role of LBX2-AS1 in the malignancy of GC is worth further discussion. Methods Quantitative real-time polymerase chain reaction (qRT-PCR) was used to determine the LBX2-AS1, miR-4766-5p and C-X-C motif chemokine (CXCL5) expression in GC tissues and cells. Dual-luciferase reporter assay was applied to examine the target relationship between LBX2-AS1 and miR-4766-5p or miR-4766-5p and CXCL5. Cell counting kit-8 (CCK-8) and Transwell assays were used to detect cell proliferation, migration and invasion rates. The protein expression of CXCL5 was confirmed using western blot. The RNA pull down experiment was used to verify the specificity of LBX2-AS1 and miR-4766-5p on BGC-823 and SGC-7901 cells. Results LBX2-AS1 was up-regulated in GC tissues and cells, and its knockdown suppressed proliferation, migration and invasion of GC cells. While, overexpression of LBX2-AS1 increased proliferation and increased CXCL5 mRNA level. CXCL5 improved cell proliferation, migration and invasion of GC cells. LBX2-AS1 could bind to miR-4766-5p to regulate CXCL5 expression. Overexpression of CXCL5 overturned those effects of miR-4766-5p in GC cells. RNA Pull down shown that BGC-823 and SGC-7901 cells, miR-4766-5p specifically binds to LBX2-AS1. Conclusions In short, this study demonstrated that LBX2-AS1 promoted proliferation, migration and invasion through up-regulation CXCL5 mediated by miR-4766-5p in GC. The LBX2-AS1/miR-4766-5p/CXCL5 regulatory axis provides a theoretical basis for the research on lncRNA-directed therapeutics in GC.

scholarly journals The Effect of lncRNA SNHG3 Overexpression on Lung Adenocarcinoma by Regulating the Expression of miR-890

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Baojie Kang ◽  
Caihong Qiu ◽  
Ying Zhang
Keyword(s):  
Lung Adenocarcinoma ◽  
Cell Lines ◽  
Bioinformatics Analysis ◽  
Underlying Mechanism ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Expression Levels ◽  
Reverse Transcription Quantitative Pcr ◽  
Dual Luciferase Reporter Assay

The lncRNA small nucleolar host gene 3 (SNHG3) was discovered to play an important role in the occurrence and development of lung adenocarcinoma (LUAD). However, the underlying molecular mechanism of SNHG3 in LUAD remains unclear. In the present study, SNHG3 expression levels in LUAD tissues and cell lines were analyzed using reverse transcription-quantitative PCR. The effects of SNHG3 on the proliferation, apoptosis, migration, and invasion of LUAD cells were determined using Cell Counting Kit-8, colony formation, flow cytometry, wound healing, and Transwell chamber assays, respectively. The specific underlying mechanism of SNHG3 in LUAD was investigated using bioinformatics analysis and a dual luciferase reporter assay. The results revealed that SNHG3 expression levels were downregulated in LUAD tissues and cell lines. Functionally, SNHG3 overexpression suppressed the proliferation, migration, and invasion of LUAD cells, while promoting apoptosis. Mechanistically, microRNA- (miR-) 890 was identified as a potential target of SNHG3, and its expression was negatively regulated by SNHG3. Notably, SNHG3 was found to promote LUAD progression by targeting miR-890. In conclusion, the findings of the present study revealed that lncRNA SNHG3 promoted the occurrence and progression of LUAD by regulating miR-890 expression.

scholarly journals Murine miR-483-3p promotes proliferation and suppresses differentiation of C2C12 cells by targeting SRF

2017 ◽  
Author(s):  
Jing Huang ◽  
Ying Liu ◽  
Liangliang Fu ◽  
Hegang Li ◽  
Bingkun Xie ◽  
...  
Keyword(s):  
Muscle Development ◽  
Myogenic Differentiation ◽  
C2c12 Cells ◽  
Luciferase Reporter ◽  
Back Muscle ◽  
Mouse Development ◽  
Protein Levels ◽  
Complicated Process ◽  
Non Coding Rnas ◽  
Dual Luciferase Reporter Assay

Myogenesis is a complicated process, which is regulated by numerous regulators. MicroRNAs (miRNAs) are conserved non-coding RNAs of ~22 nucleotides, which regulate post-transcriptional gene expression in many physiological and pathophysiological processes. Recent studies have indicated that microRNAs are critical regulators of muscle development. Here, we report miR-483-3p as a new essential regulator of muscle development, mediating myoblast proliferation and myogenic differentiation. miR-483-3p is strongly and almost exclusively expressed in muscle-related tissues such as leg muscle, back muscle, and heart. Its expression is downregulated during mouse development. Overexpression of miR-483-3p in C2C12 cells promotes proliferation and suppresses myogenic differentiation. A dual-luciferase reporter assay demonstrated miR-483-3p direct targets to the 3′-UTR of the SRF gene. Overexpression of miR-483-3p reduced SRF protein levels in C2C12 myoblasts. These results reveal a novel function of miR-483-3p as a positive regulator of C2C12 proliferation and inhibitor of myogenic differentiation via SRF downregulation.

scholarly journals CircCDC45 promotes the malignant progression of glioblastoma by modulating the miR-485-5p/CSF-1 axis

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Rongcai Liu ◽  
Weimin Dai ◽  
An Wu ◽  
Yunping Li
Keyword(s):  
Cell Proliferation ◽  
Cancer Progression ◽  
Biological Effects ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Invasion And Migration ◽  
Protein Levels

Abstract Background Glioblastoma (GBM) is characterized by progressive growth and metastasis. Numerous studies claim that the deregulation of circular RNAs (circRNAs) is associated with cancer progression. However, the role of circRNAs in GBM is largely limited. The purpose of this study was to investigate the functions of circCDC45 in GBM and provide a feasible functional mechanism to support its role. Methods The expression of circCDC45, miR-485-5p and colony-stimulating factor 1 (CSF-1) mRNA was examined using quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation was assessed using cell counting kit − 8 (CCK-8) assay and colony formation assay. Cell migration and cell invasion were monitored using transwell assay. The protein levels of proliferation-related markers and CSF-1 were determined using western blot. The target relationship was predicted using bioinformatics tools and validated using dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. Animal models were constructed to verify the role of circCDC45 in vivo. Results The expression of circCDC45 and CSF-1 was elevated in GBM tissues and cells, while the expression of miR-485-5p was declined. Downregulation of circCDC45 or CSF-1 blocked GBM cell proliferation, invasion and migration as well as tumor growth in vivo. In mechanism, circCDC45 positively regulated the expression of CSF-1 by targeting miR-485-5p. Inhibition of miR-485-5p reversed the biological effects caused by circCDC45 downregulation in GBM cells. Conclusion CircCDC45 promoted the progression of GBM by mediating the miR-485-5p/CSF-1 axis, and circCDC45 might be a promising plasmatic biomarker for GBM diagnosis and treatment.

scholarly journals Sirt-1 Regulates Physiological Process and Exerts Protective Effects against Oxidative Stress

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Lei Liu ◽  
Guangyuan Xia ◽  
Peifan Li ◽  
Yiming Wang ◽  
Qian Zhao
Keyword(s):  
Oxidative Stress ◽  
Transcriptional Activity ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Therapeutic Drug ◽  
Mrna Levels ◽  
P53 Expression ◽  
Protein Levels ◽  
Gene Assay

Background. Recent studies suggest a correlation between the reduced Sirt-1 expression with Alzheimer’s diseases (AD) and depression, respectively, suggesting a possible pathogenic role of the altered Sirt-1 expression in neuronal degenerative diseases, such as AD and depression. However, the molecular mechanisms underlying how Sirt-1 reduction impairs neuronal functions remain unknown. Methods. We used the SK-N-SH neuroblastoma cells to study the role of Sirt-1 expression on physiological roles in neuronal cells. Gain of Sirt-1 was achieved by transiently transfecting Sirt-1 expression plasmid. Sirt-1-specific shRNA was used to elucidate the role of Sirt-1 loss of function. CCK-8 (Cell Counting Kit-8) assay and flow cytometry were used to evaluate cell proliferation. Semiquantitative western blotting was used to detect relative protein levels. A further luciferase reporter gene assay was employed to examine the effect of Sirt-1 expression on the transcriptional activity of p53. RT-qPCR was used to determine the mRNA levels of p21, Bax, and Bcl-2, which were the downstream target genes of p53. Results. Sirt-1 suppressed the p53 downstream gene p21 transcription, while shRNA-mediated Sirt-1 knockdown resulted in a significant increase in p21 expression, implying a possibility that Sirt-1 promotes neuron proliferation through suppressing p53 transcriptional activity. The mRNA and protein levels of p53 were not affected by the altered Sirt-1 expression, suggesting that Sirt-1 regulates the transcriptional regulatory activity of p53 rather than p53 expression. Indeed, we further confirmed that Sirt-1 appeared to inhibit p53 transcriptional activity by attenuating its acetylation and resulted in a decrease of p53’s binding to the p21 promoter. Overexpressed Sirt-1 scavenged reactive oxygen species (ROS) production in SK-N-SH with H2O2. Knockdown of Sirt-1 presented opposite effect; the addition of EX527 (Sirt-1 inhibitor) increased ROS accumulation. Conclusions. Oxidative stress induces Sirt-1 in neuron cells, and Sirt-1 promotes proliferation in SK-N-SH cells, which protects them from oxidative stress-induced cell death, potentially via suppressing the transcriptional activity of p53. These results provide a molecular explanation underlying how the reduced Sirt-1 potentially causes the AD and depression-related diseases, supporting the idea that Sirt-1 can possibly be used as a diagnostic biomarker and/or therapeutic drug target for the AD and depression-related diseases.

scholarly journals Murine miR-483-3p promotes proliferation and suppresses differentiation of C2C12 cells by targeting SRF

2017 ◽  
Author(s):  
Jing Huang ◽  
Ying Liu ◽  
Liangliang Fu ◽  
Hegang Li ◽  
Bingkun Xie ◽  
...  
Keyword(s):  
Muscle Development ◽  
Myogenic Differentiation ◽  
C2c12 Cells ◽  
Luciferase Reporter ◽  
Back Muscle ◽  
Mouse Development ◽  
Protein Levels ◽  
Complicated Process ◽  
Non Coding Rnas ◽  
Dual Luciferase Reporter Assay

Myogenesis is a complicated process, which is regulated by numerous regulators. MicroRNAs (miRNAs) are conserved non-coding RNAs of ~22 nucleotides, which regulate post-transcriptional gene expression in many physiological and pathophysiological processes. Recent studies have indicated that microRNAs are critical regulators of muscle development. Here, we report miR-483-3p as a new essential regulator of muscle development, mediating myoblast proliferation and myogenic differentiation. miR-483-3p is strongly and almost exclusively expressed in muscle-related tissues such as leg muscle, back muscle, and heart. Its expression is downregulated during mouse development. Overexpression of miR-483-3p in C2C12 cells promotes proliferation and suppresses myogenic differentiation. A dual-luciferase reporter assay demonstrated miR-483-3p direct targets to the 3′-UTR of the SRF gene. Overexpression of miR-483-3p reduced SRF protein levels in C2C12 myoblasts. These results reveal a novel function of miR-483-3p as a positive regulator of C2C12 proliferation and inhibitor of myogenic differentiation via SRF downregulation.

scholarly journals Downregulation of lncRNA ANRIL Inhibits Osteogenic Differentiation of Periodontal Ligament Cells via Sponging miR-7 through NF-κB Pathway

2021 ◽  
Author(s):  
Xinwei Liu ◽  
Yue Zhou
Keyword(s):  
Western Blot ◽  
Osteogenic Differentiation ◽  
Periodontal Ligament ◽  
Molecular Mechanisms ◽  
Luciferase Reporter ◽  
Periodontal Ligament Cells ◽  
Protein Levels ◽  
Alp Activity ◽  
Non Coding Rnas ◽  
Dual Luciferase Reporter Assay

Abstract Background Long non-coding RNAs (lncRNAs) are dysregulation in periodontitis development and involved in osteogenesis. The current study aimed was to investigate the function of lncRNA ANRIL in periodontal ligament cells (PDLCs) and potential molecular mechanisms.Methods Firstly, the level of ANRIL was tested by qPCR. Then PDLCs were treated with a mineralizing solution to induce the osteogenic differentiation. ALP activity was measured and protein levels of BMP2, Osterix, and OCN were measured by western blot. A target of ANRIL was verified using dual-luciferase reporter assay. MiR-7 level was measured by qPCR and the signalings of NF-κB pathway were tested by western blot.Results ANRIL expression was downregulated in PDL tissues. Next, ALP activity and protein levels of BMP2, Osterix, and OCN were reduced to show PDLCs were differentiated. ANRIL level was increased in differential PDLCs, and which knockdown inhibited osteogenic differentiation. Then, miR-7 was found as a target of ANRIL. The miR-7 level was upregulated in PDL tissues and reduced in differential PDLCs. Inhibition of miR-7 suppressed ALP activity and BMP2, Osterix, and OCN expression. Moreover, inhibition of miR-7 reversed the effects on the osteogenic differentiation induced by knockdown of ANRIL. Besides, the levels of p-P65 and p-IκBα were elevated by ANRIL downregulation and were rescued by suppressing miR-7.Conclusions Knockdown of ANRIL inhibited osteogenic differentiation via sponging miR-7 through the NF-κB pathway, suggesting that ANRIL might be a therapeutic target for periodontitis.

scholarly journals LINC_00355 promotes gastric cancer progression by upregulating PHF19 expression through sponging miR-15a-5p

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Jishui Zhang ◽  
Wenhao Lv ◽  
Yagang Liu ◽  
Weihua Fu ◽  
Baosheng Chen ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cancer Progression ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Phd Finger ◽  
Protein Levels ◽  
The Relationship ◽  
Gastric Cancer Progression ◽  
Dual Luciferase Reporter Assay

Abstract Background Long non-coding RNAs exert vital roles in several types of cancer. The objective of this study was to explore the role of LINC_00355 in gastric cancer (GC) progression and its potential mechanism. Methods The expression levels of LINC_00355 in GC tissues and cells were detected by quantitative real-time PCR, followed by assessing the effects of LINC_00355 knockdown or overexpression on cell properties. Dual-luciferase reporter assay was utilized to identify the relationship between LINC_00355 and microRNA (miR)-15a-5p and miR-15a-5p and PHD finger protein 19 (PHF19), followed by the rescue experiments. Results The results showed that LINC_00355 was highly expressed in GC tissues and cells compared with the corresponding control. LINC_00355 knockdown decreased the viability, migration, and invasion and increased the accumulation of GC cells in G1 phase and apoptosis. Meanwhile, LINC_00355 downregulation markedly increased cleaved caspase 3 and cleaved poly (ADP-ribose) polymerase protein levels, whereas decreased cyclin D1, cyclin E, matrix metalloproteinase (MMP) 9, MMP2, and N-cadherin protein levels in GC cells. However, LINC_00355 overexpression had the opposite effects. It was verified that LINC_00355 upregulated the expression of PHF19 through sponging miR-15a-5p. Furthermore, PHF19 overexpression reversed the effect of LINC_00355 knockdown on GC cell properties, including cell viability, migration, invasion, and apoptosis. Conclusions Collectively, these results suggest that LINC_00355 promotes GC progression by up-regulating PHF19 through sponging miR-15a-5p. Our findings may provide an important clinical basis for reversing the malignant phenotype of GC.

scholarly journals Let-7a-5p regulates the inflammatory response in chronic rhinosinusitis with nasal polyps

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Jianwei Zhang ◽  
Lei Han ◽  
Feng Chen
Keyword(s):  
Inflammatory Response ◽  
Chronic Rhinosinusitis ◽  
Nasal Polyps ◽  
Mapk Pathway ◽  
Inhibitory Effect ◽  
Luciferase Reporter ◽  
Western Blot Assay ◽  
Protein Levels ◽  
Tnf Α

Abstract Background Let-7a-5p is demonstrated to be a tumor inhibitor in nasopharyngeal carcinoma. However, the role of let-7a-5p in chronic rhinosinusitis with nasal polyps (CRSwNP) has not been reported. This study is designed to determine the pattern of expression and role of let-7a-5p in CRSwNP. Methods The expression level of let-7a-5p, TNF-α, IL-1β, and IL-6 in CRSwNP tissues and cells were detected by RT-qPCR. Western blot assay was carried out to measure the protein expression of the Ras-MAPK pathway. Dual luciferase reporter assay and RNA pull-down assay were used to explore the relationship between let-7a-5p and IL-6. Results Let-7a-5p was significantly downregulated in CRSwNP tissues and cells. Moreover, the mRNA expression of TNF-α, IL-1β and IL-6 was increased in CRSwNP tissues, while let-7a-5p mimic inhibited the expression of TNF-α, IL-1β and IL-6. Besides that, let-7a-5p was negatively correlated with TNF-α, IL-1β and IL-6 in CRSwNP tissues. In our study, IL-6 was found to be a target gene of let-7a-5p. Additionally, let-7-5p mimic obviously reduced the protein levels of Ras, p-Raf1, p-MEK1 and p-ERK1/2, while IL-6 overexpression destroyed the inhibitory effect of let-7a-5p on the Ras-MAPK pathway in CRSwNP. Conclusion We demonstrated that let-7a-5p/IL-6 interaction regulated the inflammatory response through the Ras-MAPK pathway in CRSwNP.

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