LncRNA ZFAS1 contributed to irradiation resistance in nasopharyngeal carcinoma by inhibiting hsa-miR-7-5p/ENO2

2020 ◽  
Author(s):  
Jiaojiao Peng ◽  
Feng Liu ◽  
Hong Zheng ◽  
Qi Wu ◽  
Shixi Liu
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Luciferase Reporter ◽  
Downstream Target ◽  
Flow Cytometric ◽  
Proliferation And Apoptosis ◽  
Significant Gene ◽  
Apoptosis Assay ◽  
Cell Fractions ◽  
Irradiation Resistance

Abstract Background: In our previous research, we found that lncRNA ZFAS1 could promote nasopharyngeal carcinoma by inhibiting its downstream target axis. But we hadn’t shown whether ZFAS1 had association with irradiation resistance of NPC. In this study, we aimed to study the role of ZFAS1 in irradiation resistance of nasopharyngeal carcinoma. Methods: Bioinformatics analysis was first conducted to identify the potentially significant gene that participated in radio resistance of nasopharyngeal carcinoma. qRT-PCR was used to detect the expression of ZFAS1, miR-7-5p and ENO2 mRNA. The location of ZFAS1 was measured in cell fractions. The relationship between ZFAS1, miR-7-5p and ENO2 mRNA was validated by luciferase reporter assay and RNA pull down. Edu assay and flow cytometric apoptosis assay were conducted to observe how ZFAS1, miR-7-5p and ENO2 affected nasopharyngeal carcinoma proliferation and apoptosis under irradiation. Results：ZFAS1 was upregulated in nasopharyngeal carcinoma, which was associated with radio resistance. ZFAS1 strengthened the ability of irradiation resistance in nasopharyngeal carcinoma. ZFAS1 acted on miR-7-5p to promote ENO2 upregulation, thereby promoting the irradiation resistance. Conclusion: ZFAS1 sponged miR-7-5p to further affect ENO2 expression to augmenting the irradiation resistance in nasopharyngeal carcinoma.

The Clinical Significance of miR-135b-5p and Its Role in the Proliferation and Apoptosis of Hippocampus Neurons in Children with Temporal Lobe Epilepsy

2020 ◽  
Vol 42 (5-6) ◽  
pp. 187-194
Author(s):  
Ruixiang Li ◽  
Jiahua Hu ◽  
Sue Cao
Keyword(s):  
Temporal Lobe Epilepsy ◽  
Cell Viability ◽  
Temporal Lobe ◽  
Luciferase Reporter ◽  
Diagnostic Biomarker ◽  
Flow Cytometric ◽  
Proliferation And Apoptosis ◽  
Apoptosis Assay ◽  
Polymerase Chain ◽  
Adverse Influence

Temporal lobe epilepsy (TLE) is the most familiar localized epilepsy in children. MicroRNAs (miRNAs) are essential for the inhibition or promotion of numerous diseases. This study aimed to detect the expression of miR-135b-5p and primarily uncover its underlying function and mechanism in children with TLE. Quantitative real-time polymerase chain reaction was used to evaluate the expression of miR-135b-5p in children with TLE and in a rat model of epilepsy. MTT assay and flow cytometric apoptosis assay were conducted to evaluate the effects of miR-135b-5p on cell viability and apoptosis. Additionally, the dual luciferase reporter assay was performed to confirm the direct target of miR-135b-5p. Our data showed that the expression of miR-135b-5p was significantly decreased in children with TLE and in the epileptic rat neuron model. The dysregulation of miR-135b-5p could serve as a promising diagnostic biomarker for children with TLE. The overexpression of miR-135b-5p moderated the adverse influence on cell viability and apoptosis induced by magnesium-free medium. SIRT1 was identified as a target gene of miR-135b-5p. These results proved that miR-135b-5p might serve as a potential diagnostic biomarker in children with TLE. Overexpression of miR-135b-5p alleviates the postepileptic influence on cell viability and apoptosis by targeting SIRT1.

scholarly journals Circ_0000215 Exerts Oncogenic Function in Nasopharyngeal Carcinoma by Targeting miR-512-5p

2021 ◽  
Vol 9 ◽  
Author(s):  
Xinping Chen ◽  
Weihua Xu ◽  
Zhichao Ma ◽  
Juan Zhu ◽  
Junjie Hu ◽  
...  
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Cell Lines ◽  
Cell Counting ◽  
Regulatory Subunit ◽  
Luciferase Reporter ◽  
Regulatory Function ◽  
Migration And Invasion ◽  
Circular Rnas

Background: Increasing circular RNAs (circRNAs) are reported to participate in cancer progression. Nonetheless, the role of circRNAs in nasopharyngeal carcinoma (NPC) has not been fully clarified. This work is aimed to probe the role of circ_0000215 in NPC.Methods: Circ_0000215 expression in NPC tissues and cell lines was examined by quantitative real-time polymerase chain reaction (qRT-PCR). Cell counting kit-8 (CCK-8) assay, 5-bromo-2′-deoxyuridine (BrdU) assay, scratch healing assay and Transwell experiment were executed to investigate the regulatory function of circ_0000215 on the proliferation, migration and invasion of NPC cells. RNA immunoprecipitation (RIP), pull-down and dual-luciferase reporter experiments were utilized to determine the binding relationship between circ_0000215 and miR-512-5p, and between miR-512-5p and phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1) 3′UTR. The effects of circ_0000215 on NPC growth and metastasis in vivo were examined with nude mice model. Western blot was applied to detect the regulatory effects of circ_0000215 and miR-512-5p on PIK3R1 expression.Results: Circ_0000215 was overexpressed in NPC tissues and cell lines. The functional experiments confirmed that knockdown of circ_0000215 impeded the growth and metastasis of NPC cells in vitro and in vivo. Additionally, circ_0000215 could also work as a molecular sponge to repress miR-512-5p expression. PIK3R1 was validated as a target gene of miR-512-5p, and circ_0000215 could increase the expression level of PIK3R1 in NPC cells via suppressing miR-512-5p.Conclusion: Circ_0000215 is overexpressed in NPC and exerts oncogenic effects in NPC through regulating miR-512-5p/PIK3R1 axis.

scholarly journals The lncRNA XIST promotes colorectal cancer cell growth through regulating the miR-497-5p/FOXK1 axis

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Nan Wang ◽  
Jia-Xing He ◽  
Guo-Zhan Jia ◽  
Ke Wang ◽  
Shuai Zhou ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Cancer Cell Growth ◽  
Forkhead Box ◽  
Proliferation And Apoptosis ◽  
Lncrna Xist

Abstract Background Recent studies suggest that long noncoding RNAs (lncRNAs) play an important role in tumorigenesis. As a newly identified lncRNA, the role of XIST in colorectal cancer (CRC) has not been established. Here, we sought to characterize the role of XIST and its associated regulatory network in CRC cells. Methods Expression of XIST mRNA, miR-497-5p, and forkhead box k1 (FOXK1) in CRC cells and tissues were detected using quantitative real-time polymerase chain reaction (qRT-PCR). Proliferation and apoptosis of CRC cells were determined using the CCK-8 cell counting assay and flow cytometry. The rate of cell migration and invasion was determined using a transwell assay. The relationships between XIST, miR-497-5p, and FOXK1 were predicted and confirmed using a dual-luciferase reporter assay. Expression of FOXK1 protein was quantified by Western blot. Results XIST and FOXK1 expression were significantly upregulated in CRC tissues and cell lines, while miR-497-5p expression was downregulated. XIST knockdown significantly suppressed CRC cell proliferation, migration, and invasion. Silencing of XIST also reversed the downregulation of miR-497-5p and upregulation of FOXK1. Moreover, blocking XIST expression was shown to inhibit CRC tumor growth in vivo and the effects were antagonized by the loss of miR-497-5p. miR-497-5p was shown to act as a sponge of XIST and also targeted FOXK1 in CRC cells. Conclusions XIST was shown to promote the malignancy of CRC cells by competitively binding to miR-497-5p, resulting in an increase in FOXK1 expression. These results suggest that targeting of XIST may represent a possible treatment for CRC.

scholarly journals LncRNA MEG3 influences the proliferation and apoptosis of psoriasis epidermal cells by targeting miR-21/caspase-8

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Hai-Yan Jia ◽  
Kai Zhang ◽  
Wen-Jing Lu ◽  
Gui-Wen Xu ◽  
Jian-Fen Zhang ◽  
...  
Keyword(s):  
Binding Site ◽  
Cell Model ◽  
Mrna Level ◽  
Epidermal Cells ◽  
Luciferase Reporter ◽  
Psoriatic Skin ◽  
Caspase 8 ◽  
Proliferation And Apoptosis

Abstract Background It was reported that microRNA-21(miR-21) was differentially expressed in the keratinocytes of psoriasis patients, and it may influence the apoptosis and proliferation of cells. The role of lncRNA maternally expressed gene3 (MEG3), a competing endogenous RNAs of miR-21, in the progression of psoriasis remains unclear. We aimed to unfold the influence of MEG3 and miR-21 on the proliferation and apoptosis of psoriasis epidermal cells. Methods 50μg/L TNF-α was used to treat HaCaTs and NHEKs cells for 24 h, and then different experiments were conducted. qRT-PCR were applied for measuring the mRNA level of MEG3, miR-2, and caspase-8, and the protein expression of caspase-8 was measured with western blotting. Flow cytometry was used for assessing apoptosis. Cell proliferation was detected using MTT and colony formation assays. Dual luciferase reporter assay was applied for confirming the binding site between MEG3 and miR-21, miR-21 and Caspase-8. Results A cell model for in vitro studying the role of MEG3 in psoriasis pathophysiology was established using HaCaT and HHEKs. MEG3 was significantly down-regulated in HaCaT, HHEKs, and psoriatic skin samples. MEG3 inhibits proliferation and promotes apoptosis of Activated-HaCaT (Act-HaCaT) and Activated-HHEKs (Act- HHEK) by regulating miR-21, and the binding site between MEG3 and miR-21 was identified. We also found that miR-21 could inhibit the level of caspase-8 and identified the binding site between caspase-8 and miR-21. Some down-stream proteins of caspase-8, Cleaved caspase-8, cytc, and apaf-1 were regulated by miR-21 and MEG3. Conclusion MEG3/miR-21 axis may regulate the expression of caspase-8, and further influence the proliferation and apoptosis of psoriasis keratinocyte, Act-HaCaT and Act- HHEK. Therefore, our findings may provide a new thought for the study of pathogenesis and treatment of psoriasis.

scholarly journals LncRNA-H19 Drives Cardiomyocyte Senescence by Targeting miR-19a/socs1/p53 Axis

2021 ◽  
Vol 12 ◽  
Author(s):  
Yuting Zhuang ◽  
Tingting Li ◽  
Hongwen Xiao ◽  
Jiaxu Wu ◽  
Shuang Su ◽  
...  
Keyword(s):  
Molecular Mechanism ◽  
Noncoding Rna ◽  
Physiological Function ◽  
Luciferase Reporter ◽  
Aged Mouse ◽  
Progressive Decline ◽  
Downstream Target ◽  
Novel Target ◽  
Related Proteins

Purpose: Cardiomyocyte senescence is associated with a progressive decline in cardiac physiological function and the risk of cardiovascular events. lncRNA H19 (H19), a well-known long noncoding RNA (lncRNA), is involved in the pathophysiological process of multiple cardiovascular disease such as heart failure, cardiac ischemia and fibrosis. However, the role of H19 in cardiomyocyte senescence remains to be further explored.Methods: Senescence-associated β-galactosidases (SA-β-gal) staining was used to detect cardiomyocyte senescence. Western blot, qRT-PCR and luciferase reporter assay were employed to evaluate the role of H19 in cardiomyocyte senescence and its underling molecular mechanism.Results: H19 level was significantly increased in high glucose-induced senescence cardiomyocytes and aged mouse hearts. Overexpression of H19 enhanced the number of SA-β-gal-positive cells, and the expression of senescence-related proteins p53 and p21, whereas H19 knockdown exerted the opposite effects. Mechanistically, H19 was demonstrated as a competing endogenous RNA (ceRNA) for microRNA-19a (miR-19a): H19 overexpression downregulated miR-19a level, while H19 knockdown upregulated miR-19a. The expression of SOSC1 was dramatically increased in senescence cardiomyocytes and aged mouse hearts. Further experiments identified SOCS1 as a downstream target of miR-19a. H19 upregulated SOCS1 expression and activated the p53/p21 pathway by targeting miR-19a, thus promoting the cardiomyocytes senescence.Conclusion: Our results show that H19 is a pro-senescence lncRNA in cardiomyocytes acting as a ceRNA to target the miR-19a/SOCS1/p53/p21 pathway. Our research reveals a molecular mechanism of cardiomyocyte senescence regulation and provides a novel target of the therapy for senescence-associated cardiac diseases.

Expression and role of miR-637 before and after lymphadenectomy for thyroid carcinoma

2021 ◽  
Vol 11 (7) ◽  
pp. 1245-1253
Author(s):  
Yanqing Qu ◽  
Xiaoyu Chu ◽  
Cuihong Dong ◽  
Weijiao Wang ◽  
Xiaojian Zhang
Keyword(s):  
Cell Proliferation ◽  
Thyroid Carcinoma ◽  
Biological Significance ◽  
Negative Control ◽  
Luciferase Reporter ◽  
Proliferation And Apoptosis ◽  
Before And After ◽  
Proliferation And Invasion

Thyroid carcinoma (TC) is a common endocrine malignancy that can be partially relieved by surgery, but its recurrence rate remains high. It is speculated that miR-637 exerts certain influence in its occurrence and development. Accordingly, we included 87 TC patients and 72 concurrent healthy controls as the research participants and purchased human papillary thyroid carcinoma cells with which to study and analyze the biological significance of miR-637. The determination of miR-637 and SH2B1 in peripheral blood and tissues was performed using nanoparticle-assisted polymerase chain reaction assay, and the identification of cell proliferation and apoptosis was made by MTT, Transwell, and flow cytometry. The results indicated that after transfection of miR-637 into TPC-1, the cell proliferation and invasion capacities in the mimics-miR-637 group were significantly reduced as compared to that of the inhibition-miR-637 and negative control (NC)-miR groups (P < 0.05). While transfection of SH2B1 into TPC-1 cells led to significantly enhanced cell proliferation and invasion capacities in sh-SH2B1 group than in si-SH2B1 and NC groups (P < 0.05). Finally, a double luciferase reporter assay identified enormously inhibited fluorescence activity of SH2B1-WT by mimics-miR-637. According to the experimental results, it is concluded that miR-637 expression was low in TC but increased after lymphadenectomy for TC. Moreover, by targeting SH2B1, miR-637 interferes with TC progression, which carries significant implications for future diagnosis and treatment of the disease.

scholarly journals WT1 Enhances Proliferation and Impedes Apoptosis in KRAS Mutant NSCLC via Targeting cMyc

2015 ◽  
Vol 35 (2) ◽  
pp. 647-662 ◽  
Author(s):  
Chen Wu ◽  
Sihan Wang ◽  
Caihua Xu ◽  
Andreas Tyler ◽  
Xingru Li ◽  
...  
Keyword(s):  
Negative Impact ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Wild Type ◽  
Small Cell Lung ◽  
Kras Mutations ◽  
Proliferation And Apoptosis ◽  
Mutant Cells ◽  
Dual Luciferase Reporter Assay

Background: A novel link between oncogenic KRAS signalling and WT1 was recently identified. We sought to investigate the role of WT1 and KRAS in proliferation and apoptosis. Methods: KRAS mutations and WT1 (cMyc) expression were detected using Sanger sequencing and real-time PCR in 77 patients with non-small cell lung cancer (NSCLC). Overexpression and knockdown of WT1 were generated with plasmid and siRNA via transient transfection technology in H1299 and H1568 cells. MTT assay for detection of cell proliferation, and TUNEL assay and proteomic profiler assay for apoptosis evaluation were carried out. Dual luciferase reporter assay and ChIP-PCR were performed to validate the effect of WT1 on the cMyc promoter. Results: KRAS mutations showed a negative impact on overall survival (OS). High expressions of WT1 and cMyc were associated with poor OS in KRAS mutant subgroup. The potential mechanisms that WT1 promotes proliferation and impedes apoptosis through affecting multiple apoptosis-related regulators in KRAS mutant NSCLC cells were identified. WT1 could activate cMyc promoter directly in KRAS mutant cells. Conclusion: The results suggest that WT1 and c-MYC expression is important for survival in KRAS mutant tumors as opposed to KRAS wild-type tumors. For treatment of KRAS mutant NSCLC, targeting WT1 and cMyc may provide alternative therapeutic strategies.

scholarly journals Glucocorticoid inhibits cell proliferation in differentiating osteoblasts by microRNA-199a targeting of WNT signaling

2015 ◽  
Vol 54 (3) ◽  
pp. 325-337 ◽  
Author(s):  
Changgui Shi ◽  
Ping Huang ◽  
Hui Kang ◽  
Bo Hu ◽  
Jin Qi ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Wnt Signaling ◽  
Transcriptional Repression ◽  
Wnt Signaling Pathway ◽  
Luciferase Reporter ◽  
Osteoblast Proliferation ◽  
Cellular Processes ◽  
Proliferation And Apoptosis ◽  
Proliferation Assays

The inhibition of osteoblast proliferation by glucocorticoids (GCs) is very important in the etiology of GC-induced osteoporosis. The mechanisms of this process are still not fully understood. The results of recent studies have indicated an important role for microRNAs in GC-mediated responses in various cellular processes, including cell proliferation and apoptosis. Therefore, we developed the hypothesis that these regulatory molecules might be involved in GC-decreased osteoblast proliferation. Western blotting, quantitative real-time PCR, cell proliferation assays, and luciferase assays were employed to investigate the role of miRNAs in GC-inhibited osteoblast proliferation. microRNA-199a-5p was significantly increased in osteoblasts treated with dexamethasone (Dex). To delineate the role of microRNA-199a-5p, we silenced and overexpressed microRNA-199a-5p in osteoblasts. We found that overexpressing microRNA-199a-5p remarkably increased the inhibition effect of Dex on osteoblast proliferation, and depleting microRNA-199a-5p significantly attenuated Dex-inhibited osteoblast proliferation. Results of mechanistic studies indicated that microRNA-199a-5p inhibited FZD4 and WNT2 expression through a microRNA-199a-5p binding site within the 3′-UTR of FZD4 and WNT2. The post-transcriptional repression of FZD4 and WNT2 were further confirmed by luciferase reporter assay. These results indicated that microRNA-199a-5p may play a significant role in GC-inhibited osteoblast proliferation by regulating the WNT signaling pathway.

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.

scholarly journals TRIM25 contributes to the malignancy of acute myeloid leukemia and is negatively regulated by microRNA-137

Open Medicine ◽  
2020 ◽  
Vol 16 (1) ◽  
pp. 095-103
Author(s):  
Sheng Wang ◽  
Bang Shuo Zhang ◽  
Yi Yang ◽  
Ying Li ◽  
Jing Long Lv ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Luciferase Reporter ◽  
Causative Role ◽  
Treatment Target ◽  
Invasion And Migration ◽  
Downstream Target ◽  
And Migration ◽  
Acute Myeloid

Abstract Background Acute myeloid leukemia (AML) is a ubiquitous malignancy that occurs in the hematological system. Tripartite motif-containing 25 (TRIM25) has been found to be involved in various carcinomas comprising AML. However, the function and underlying causative role of TRIM25 in AML are still obscure. Methods and materials Quantitative real-time polymerase chain reaction (qPCR) was used for assaying TRIM25 and miR-137 expression in AML samples and cells. CCK-8 assay, Calcein-acetoxymethylester/propidium iodide staining, and Transwell assay were adopted to assay cell proliferation, invasion, and migration. Dual-luciferase reporter experiment was used for analyzing the interaction of TRIM25 with miR-137. Western blot was used for assaying protein expression levels. Results This study confirmed that TRIM25 expression was upregulated in AML samples and cell lines, whereas miR-137 expression was downregulated. Overexpression of TRIM25 significantly contributed to AML cell’s proliferation, invasion, and migration, whereas knockdown exerted the opposite effect. In addition, TRIM25 was a downstream target of miR-137 in AML cells and negatively modulated by miR-137. Conclusion TRIM25 was targeted and regulated by miR-137, exerted a carcinogenic function in AML, and could be used as a latent biomarker and a treatment target for AML.

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