scholarly journals miR-590-3p Targets Cyclin G2 and FOXO3 to Promote Ovarian Cancer Cell Proliferation, Invasion, and Spheroid Formation

2019 ◽  
Vol 20 (8) ◽  
pp. 1810 ◽  
Author(s):  
Mohamed Salem ◽  
Yanan Shan ◽  
Stefanie Bernaudo ◽  
Chun Peng
Keyword(s):  
Ovarian Cancer ◽  
Cell Proliferation ◽  
Target Genes ◽  
Cancer Development ◽  
Nuclear Accumulation ◽  
Luciferase Reporter ◽  
Spheroid Formation ◽  
Forkhead Box ◽  
Cyclin G2 ◽  
Reporter Assays

Ovarian cancer is the leading cause of death from gynecological cancers. MicroRNAs (miRNAs) are small, non-coding RNAs that interact with the 3′ untranslated region (3′ UTR) of target genes to repress their expression. We have previously reported that miR-590-3p promoted ovarian cancer growth and metastasis, in part by targeting Forkhead box A (FOXA2). In this study, we further investigated the mechanisms by which miR-590-3p promotes ovarian cancer development. Using luciferase reporter assays, real-time PCR, and Western blot analyses, we demonstrated that miR-590-3p targets cyclin G2 (CCNG2) and Forkhead box class O3 (FOXO3) at their 3′ UTRs. Silencing of CCNG2 or FOXO3 mimicked, while the overexpression of CCNG2 or FOXO3 reversed, the stimulatory effect of miR-590-3p on cell proliferation and invasion. In hanging drop cultures, the overexpression of mir-590 or the transient transfection of miR-590-3p mimics induced the formation of compact spheroids. Transfection of the CCNG2 or FOXO3 plasmid into the mir-590 cells resulted in the partial disruption of the compact spheroid formation. Since we have shown that CCNG2 suppressed β-catenin signaling, we investigated if miR-590-3p regulated β-catenin activity. In the TOPFlash luciferase reporter assays, mir-590 increased β-catenin/TCF transcriptional activity and the nuclear accumulation of β-catenin. Silencing of β-catenin attenuated the effect of mir-590 on the compact spheroid formation. Taken together, these results suggest that miR-590-3p promotes ovarian cancer development, in part by directly targeting CCNG2 and FOXO3.

scholarly journals Mir-573 regulates cell proliferation and apoptosis by targeting Bax in human degenerative disc cells following hyperbaric oxygen treatment

2020 ◽  
Author(s):  
Song-Shu Lin ◽  
Chi-Chien Niu ◽  
Li-Jen Yuan ◽  
Tsung-Ting Tsai ◽  
Po-Liang Lai ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Hyperbaric Oxygen ◽  
Caspase 3 ◽  
Target Genes ◽  
Vital Role ◽  
Luciferase Reporter ◽  
Nucleus Pulposus Cells ◽  
Caspase 9 ◽  
Proliferation And Apoptosis ◽  
Reporter Assays

Abstract Background: MicroRNA (miRNA) plays a vital role in the intervertebral disc (IVD) degeneration. The expression level of miR-573 was downregulated whereas Bax was upregulated notably in human degenerative nucleus pulposus cells (NPCs). In this study, we aimed to investigate the role of miR-573 in human degenerative NPCs following hyperbaric oxygen (HBO) treatment. Methods: NPCs were separated from human degenerated IVD tissues. The control cells were maintained in 5% CO2/95% air and the hyperoxic cells were exposed to 100% O2 at 2.5 atmospheres absolute. MiRNA expression profiling was performed via microarray and confirmed by real-time PCR, and miRNA target genes were identified using bioinformatics and luciferase reporter assays. The mRNA and protein levels of Bax were measured. The proliferation of NPCs were detected using MTT assay. The protein expression levels of Bax, cleaved caspase 9, cleaved caspase 3, pro-caspase 9 and pro-caspase 3 were examined.Results: Bioinformatics analysis indicated that the 3′ untranslated region (UTR) of the Bax mRNA contained the “seed-matched-sequence” for hsa-miR-573, which was validated via reporter assays. MiR-573 was induced by HBO and simultaneous suppression of Bax was observed in NPCs. Knockdown of miR-573 resulted in upregulation of Bax expression in HBO-treated cells. In addition, overexpression of miR-573 by HBO increased cell proliferation and coupled with inhibition of cell apoptosis. The cleavage of pro‑caspase 9 and pro‑caspase 3 was suppressed while the levels of cleaved caspase 9 and caspase 3 were decreased in HBO-treated cells. Transfection with anti-miR-573 partly suppressed the effects of HBO. Conclusion: Mir-573 regulates cell proliferation and apoptosis by targeting Bax in human degenerative NPCs following HBO treatment.

scholarly journals Mir-573 regulates cell proliferation and apoptosis by targeting Bax in human degenerative disc cells following hyperbaric oxygen treatment

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Song-Shu Lin ◽  
Chi-Chien Niu ◽  
Li-Jen Yuan ◽  
Tsung-Ting Tsai ◽  
Po-Liang Lai ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Nucleus Pulposus ◽  
Hyperbaric Oxygen ◽  
Caspase 3 ◽  
Target Genes ◽  
Luciferase Reporter ◽  
Nucleus Pulposus Cells ◽  
Caspase 9 ◽  
Proliferation And Apoptosis ◽  
Reporter Assays

Abstract Background MicroRNA (miRNA) plays a vital role in the intervertebral disc (IVD) degeneration. The expression level of miR-573 was downregulated whereas Bax was upregulated notably in human degenerative nucleus pulposus cells. In this study, we aimed to investigate the role of miR-573 in human degenerative nucleus pulposus (NP) cells following hyperbaric oxygen (HBO) treatment. Methods NP cells were separated from human degenerated IVD tissues. The control cells were maintained in 5% CO2/95% air and the hyperoxic cells were exposed to 100% O2 at 2.5 atmospheres absolute. MiRNA expression profiling was performed via microarray and confirmed by real-time PCR, and miRNA target genes were identified using bioinformatics and luciferase reporter assays. The mRNA and protein levels of Bax were measured. The proliferation of NPCs was detected using MTT assay. The protein expression levels of Bax, cleaved caspase 9, cleaved caspase 3, pro-caspase 9, and pro-caspase 3 were examined. Results Bioinformatics analysis indicated that the 3′ untranslated region (UTR) of the Bax mRNA contained the “seed-matched-sequence” for hsa-miR-573, which was validated via reporter assays. MiR-573 was induced by HBO and simultaneous suppression of Bax was observed in NP cells. Knockdown of miR-573 resulted in upregulation of Bax expression in HBO-treated cells. In addition, overexpression of miR-573 by HBO increased cell proliferation and coupled with inhibition of cell apoptosis. The cleavage of pro-caspase 9 and pro-caspase 3 was suppressed while the levels of cleaved caspase 9 and caspase 3 were decreased in HBO-treated cells. Transfection with anti-miR-573 partly suppressed the effects of HBO. Conclusion Mir-573 regulates cell proliferation and apoptosis by targeting Bax in human degenerative NP cells following HBO treatment.

LncRNA LINC00858 enhances cervical cancer cell growth through miR-3064-5p/ VMA21 axis

2021 ◽  
pp. 1-11
Author(s):  
Min Wei ◽  
Youguo Chen ◽  
Wensheng Du
Keyword(s):  
Cervical Cancer ◽  
Cell Proliferation ◽  
Cell Apoptosis ◽  
Luciferase Reporter ◽  
Cancer Cell Growth ◽  
Protein Coding ◽  
Gynecological Malignancy ◽  
Promising Therapeutic Target ◽  
Proliferation And Apoptosis ◽  
Reporter Assays

BACKGROUND: Cervical cancer (CC) is the most common form of gynecological malignancy. Long intergenic non-protein coding RNA 858 (LINC00858) has been identified to participate in multiple cancers. However, the role and mechanism of LINC00858 in CC cells are still elusive. AIM: The aim of this study is to explore the biological functions and mechanisms of LINC00858 in CC cells. METHODS: RT-qPCR analysis was used to examine the expression of LINC00858 in CC cells. EdU and colony formation assay were utilized to assess cell proliferation. TUNEL assay and flow cytometry assay were conducted to assess cell apoptosis. The mechanism regarding LINC00858 was certified through RNA pull down, RIP and luciferase reporter assays. RESULTS: The up-regulated LINC00858 was detected in CC cells. Reduction of LINC00858 effectively subdued CC cells proliferation and stimulated cell apoptosis. LINC00858 was determined to bind with miR-3064-5p and up-regulate VMA21 in CC cells. In rescue assays, miR-3064-5p down-regulation and VMA21 up-regulation were able to counteract the effect caused by LINC00858 decrease on CC cell proliferation and apoptosis. CONCLUSION: LINC00858 enhances cell proliferation, while restraining cell apoptosis in CC through targeting miR-3064-5p/VMA21 axis, implying that LINC00858 may serve as a promising therapeutic target for CC.

Lysine-specific demethylase 1 (LSD1) promotes ovarian cancer cell progression by Forkhead box O 3a (FOXO3a) inhibition

2020 ◽  
Vol 10 (4) ◽  
pp. 594-602 ◽  
Author(s):  
Li Liu ◽  
Fuxing Hao ◽  
Anping Wang ◽  
Xiaolan Chen ◽  
Bin Zhang ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Cell Proliferation ◽  
Cancer Cell ◽  
Ovarian Cancer Cell ◽  
Soft Agar ◽  
Ovarian Cancer Cells ◽  
Cancer Cell Growth ◽  
Forkhead Box ◽  
Lysine Specific Demethylase ◽  
Cell Progression

Recently, LSD1 is considered as a possible therapeutic mark for ovarian epithelial cancer (OEC). Though, the underlying molecular mechanism by which LSD1 endorses the oncogenesis of OEC has not been fully understood. Here, we revealed that overexpression of LSD1 downregulated Forkhead box O 3a (FOXO3a), while knockdown or pharmacological inhibition of LSD1 upregulated FOXO3a expression. Specifically, LSD1 interacted with demethylated FOXO3a. The LSD1-demethylated FOXO3a degraded via an ubiquitin-proteasome pathway. Biologically, LSD1 destabilized FOXO3a to abrogate its functions in the suppression of soft agar colony and cell proliferation formation in HO8910 ovarian cancer cells. Knockdown of FOXO3a rescued the restricted cell proliferation by LSD1 downregulation. As a whole, our study clarifies a way in ovarian cancer cell growth through the negative regulation of FOXO3a by LSD1.

LncRNA SNHG14 Promotes Progression of Ovarian Cancer by Regulating miR-206 Expression

2021 ◽  
Vol 7 (5) ◽  
pp. 3997-4004
Author(s):  
Zhibo Zou ◽  
Lin Peng
Keyword(s):  
Ovarian Cancer ◽  
Flow Cytometry ◽  
Cell Proliferation ◽  
Tumor Growth ◽  
Cancer Progression ◽  
Nude Mice ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Qrt Pcr ◽  
Research Objects

Objective: This study aimed to probe into the effect of LncRNA SNHG14 on ovarian cancer progression by regulating miR-206.Methods: Fifty-seven ovarian cancer (OC) patients who were treated in our hospital from December 2017 to December 2019 were collected as the research objects. During the operation, OC tissues and paracancerous tissues of patients were collected, and the effect of SNHG14 on OC tumor growth in nude mice was detected, and SNHG14 inhibitor was transfected into OC cells. The relative expression of SNHG14 in tissues and cells was detected by qRT-PCR, cell proliferation was testedvia CCK8, migration and invasion were detected through Transwell, apoptosis was assessedvia flow cytometry, and the targeted relationship between SNHG14 and miR-206 was detected by dual luciferase reporter gene.Results: SNHG14 is highly expressed in OC tissues, cells and nude mice. Down-regulating it can inhibit the biological ability of OC cells and inhibit the growth of nude mice tumors. It can directly target miR-206 to regulate CCND1 expression and promote OC progression.Conclusion: LncRNA SNHG14 can act as miR-206 sponge to regulate CCND1 expression downstream of miR-206 and promote OC progression.

Screening candidate microR-15a- IRAK2 regulatory pairs for predicting the response to Staphylococcus aureus-induced mastitis in dairy cows

2019 ◽  
Vol 86 (4) ◽  
pp. 425-431 ◽  
Author(s):  
Zhi Chen ◽  
Jingpeng Zhou ◽  
Xiaolong Wang ◽  
Yang Zhang ◽  
Xubin Lu ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Differential Expression ◽  
High Throughput Sequencing ◽  
Target Genes ◽  
Interleukin 1 ◽  
Differentially Expressed ◽  
Luciferase Reporter ◽  
Expression Of Genes ◽  
Chinese Holstein Cows ◽  
Reporter Assays

AbstractWe established a mastitis model using exogenous infection of the mammary gland of Chinese Holstein cows with Staphylococcus aureus and extracted total RNA from S. aureus-infected and healthy mammary quarters. Differential expression of genes due to mastitis was evaluated using Affymetrix technology and results revealed a total of 1230 differentially expressed mRNAs. A subset of affected genes was verified via Q-PCR and pathway analysis. In addition, Solexa high-throughput sequencing technology was used to analyze profiles of miRNA in infected and healthy quarters. These analyses revealed a total of 52 differentially expressed miRNAs. A subset of those results was verified via Q-PCR. Bioinformatics techniques were used to predict and analyze the correlations among differentially expressed miRNA and mRNA. Results revealed a total of 329 pairs of negatively associated miRNA/mRNA, with 31 upregulated pairs of mRNA and 298 downregulated pairs of mRNA. Differential expression of miR-15a and interleukin-1 receptor-associated kinase-like 2 (IRAK2), were evaluated by western blot and luciferase reporter assays. We conclude that miR-15a and miR-15a target genes (IRAK2) constitute potential miRNA–mRNA regulatory pairs for use as biomarkers to predict a mastitis response.

scholarly journals Overexpression of KIF2A is Suppressed by miR-206 and Associated with Poor Prognosis in Ovarian Cancer

2018 ◽  
Vol 50 (3) ◽  
pp. 810-822 ◽  
Author(s):  
Nan Sheng ◽  
Yun-Zhao Xu ◽  
Qing-Hua Xi ◽  
Hai-Yan Jiang ◽  
Chen-Yi Wang ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Cell Proliferation ◽  
Cancer Cell ◽  
Poor Prognosis ◽  
Ovarian Cancer Cell ◽  
Expression Profiles ◽  
Annexin V ◽  
Prognostic Indicator ◽  
Luciferase Reporter ◽  
Migration And Invasion

Background/Aims: This study aimed to investigate the expression and prognostic value of kinesin family member 2A (KIF2A) and the suppression effects of microRNA-206 (miR-206) on KIF2A in ovarian cancer. Methods: Ovarian cancer tissues from patients and ovarian cancer cell lines (A2780 and SKOV3) were used in this study. miR-206 mimics and control were transiently transfected into cells. RT-qPCR was performed to detect KIF2A mRNA and miR-206 expression levels, Western blot was performed to detect KIF2A protein levels, Dual-Luciferase Reporter Assay was used to examine the inhibition effects of miR-206 on KIF2A mRNA, immunohistochemical staining was used to examine the expression of KIF2A in tissue sections. CCK-8, transwell and Annexin-V-FITC/Propidium Iodide staining with flow cytometry were used to detect the cell proliferation, migration/invasion, and apoptosis respectively. Results: Our study explored the expression profiles of KIF2A and miR-206 in the patients with ovarian cancer. We found that overexpression of KIF2A was associated with a poor prognosis in ovarian cancer. We also found that KIF2A mRNA contains two target sites for miR-206 binding and confirmed that miR-206 directly suppresses KIF2A; inhibits ovarian cancer cell proliferation, migration, and invasion; and induces apoptosis. Conclusion: The results suggest KIF2A could serve a valuable prognostic indicator in ovarian cancer and provide a rationale for treatment of ovarian cancer by targeting KIF2A via miR-206.

scholarly journals Long non-coding RNA PTPRG-AS1 promotes cell tumorigenicity in epithelial ovarian cancer by decoying microRNA-545-3p and consequently enhancing HDAC4 expression

2020 ◽  
Author(s):  
Juanjuan Shi ◽  
Xijian Xu ◽  
Dan Zhang ◽  
Jiuyan Zhang ◽  
Hui Yang ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Cell Proliferation ◽  
Tumor Growth ◽  
Epithelial Ovarian Cancer ◽  
Cell Lines ◽  
Luciferase Reporter ◽  
Non Coding Rna ◽  
Long Non Coding Rna

Abstract Background: Long non-coding RNA PTPRG antisense RNA 1 (PTPRG-AS1) deregulation has been reported in various human malignancies and identified as an important modulator of cancer development. Few reports have focused on the detailed role of PTPRG-AS1 in epithelial ovarian cancer (EOC) and its underlying mechanism. This study aimed to determine the physiological function of PTPRG-AS1 in EOC. A series of experiments were also performed to identify the mechanisms through which PTPRG-AS1 exerts its function in EOC.Methods: Reverse transcription-quantitative polymerase chain reaction was used to determine PTPRG-AS1 expression in EOC tissues and cell lines. PTPRG-AS1 was silenced in EOC cells and studied with respect to cell proliferation, apoptosis, migration, and invasion in vitro and tumor growth in vivo. The putative miRNAs that target PTPRG-AS1 were predicted using bioinformatics analysis and further confirmed in luciferase reporter and RNA immunoprecipitation assays.Results: Our data verified the upregulation of PTPRG-AS1 in EOC tissues and cell lines. High PTPRG-AS1 expression was associated with shorter overall survival in patients with EOC. Functionally, EOC cell proliferation, migration, invasion in vitro, and tumor growth in vivo were suppressed by PTPRG-AS1 silencing. In contrast, cell apoptosis was promoted by loss of PTPRG-AS1. Regarding the mechanism, PTPRG-AS1 could serve as a competing endogenous RNA in EOC cells by decoying microRNA-545-3p (miR-545-3p), thereby elevating histone deacetylase 4 (HDAC4) expression. Furthermore, rescue experiments revealed that PTPRG-AS1 knockdown-mediated effects on EOC cells were, in part, counteracted by the inhibition of miR-545-3p or restoration of HDAC4.Conclusions: PTPRG-AS1 functioned as an oncogenic lncRNA that aggravated the malignancy of EOC through the miR-545-3p/HDAC4 ceRNA network. Thus, targeting the PTPRG-AS1/miR-545-3p/HDAC4 pathway may be a novel strategy for EOC anticancer therapy.

Silencing of miR-1246 Induces Cell Cycle Arrest and Apoptosis in Cisplatin-Resistant Ovarian Cancer Cells by Promoting ZNF23 Transcription

2021 ◽  
pp. 1-13
Author(s):  
Lu Cai ◽  
Qian Zhang ◽  
Lili Du ◽  
Feiyun Zheng
Keyword(s):  
Cell Cycle ◽  
Ovarian Cancer ◽  
Cell Proliferation ◽  
Cell Cycle Arrest ◽  
Cell Cycle Progression ◽  
Luciferase Reporter ◽  
Cycle Phase ◽  
Ovarian Cancer Cells ◽  
Tunel Staining ◽  
Cycle Arrest

Ovarian cancer (OC) is the most frequent cause of death among patients with gynecologic malignancies. In recent years, the development of cisplatin (DDP) resistance has become an important reason for the poor prognosis of OC patients. Therefore, it is vital to explore the mechanism of DDP resistance in OC. In this study, microRNA-1246 (miR-1246) expression in OC and DDP-resistant OC cells was determined by RT-qPCR, and chemosensitivity to DDP was assessed by the CCK-8 assay. A dual-luciferase reporter assay was performed to confirm the interaction between miR-1246 and zinc finger 23 (<i>ZNF23</i>), while changes in <i>ZNF23</i> expression were monitored by RT-qPCR, immunofluorescence, and western blot assays. Moreover, cell proliferation, cycle phase, and apoptosis were determined by EdU staining, flow cytometry, TUNEL staining, and Hoechst staining. Our data showed that miR-1246 was highly expressed in DDP-resistant OVCAR-3 and TOV-112D cells. Functionally, overexpression of miR-1246 markedly enhanced DDP resistance and cell proliferation, and suppressed cell cycle arrest and apoptosis of OC cells. Inhibition of miR-1246 expression significantly attenuated DDP resistance and cell proliferation, and increased cell cycle arrest and apoptosis in DDP-resistant OC cells. Furthermore, <i>ZNF23</i> was identified as a target gene of miR-1246, and ZNF23 protein expression was notably downregulated in DDP-resistant OC cells. Moreover, overexpression of miR-1246 significantly downregulated the <i>ZNF23</i> levels in OVCAR-3 and TOV-112D cells, and inhibition of miR-1246 upregulated the <i>ZNF23</i> levels in the DDP-resistant OVCAR-3 and TOV-112D cells. In conclusion, miR-1246 might be a novel regulator of DDP-resistant OC that functions by regulating <i>ZNF23</i> expression in DDP-resistant cells, as well as cell proliferation, cell cycle progression, and apoptosis.

scholarly journals EIF4A3-induced circular RNA MMP9 (circMMP9) acts as a sponge of miR-124 and promotes glioblastoma multiforme cell tumorigenesis

2018 ◽  
Vol 17 (1) ◽  
Author(s):  
Renjie Wang ◽  
Sai Zhang ◽  
Xuyi Chen ◽  
Nan Li ◽  
Jianwei Li ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Glioblastoma Multiforme ◽  
Aurora Kinase ◽  
Underlying Mechanism ◽  
Circular Rna ◽  
Initiation Factor ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Circular Rnas ◽  
Reporter Assays

Abstract Background Circular RNAs (circRNAs) have been found to play critical roles in the development and progression of various cancers. However, little is known about the effects of the circular RNA network on glioblastoma multiforme (GBM). Methods A microarray was used to screen circRNA expression in GBM. Quantitative real-time PCR was used to detect the expression of circMMP9. GBM cells were transfected with a circMMP9 overexpression vector or siRNA, and cell proliferation, migration and invasion, as well as tumorigenesis in nude mice, were assessed to examine the effect of circMMP9 in GBM. Biotin-coupled miRNA capture, fluorescence in situ hybridization and luciferase reporter assays were conducted to confirm the relationship between circMMP9 and miR-124. Results In this study, we screened differentially expressed circRNAs and identified circMMP9 in GBM. We found that circMMP9 acted as an oncogene, was upregulated in GBM and promoted the proliferation, migration and invasion abilities of GBM cells. Next, we verified that circMMP9 served as a sponge that directly targeted miR-124; circMMP9 accelerated GBM cell proliferation, migration and invasion by targeting miR-124. Furthermore, we found that cyclin-dependent kinase 4 (CDK4) and aurora kinase A (AURKA) were involved in circMMP9/miR-124 axis-induced GBM tumorigenesis. Finally, we found that eukaryotic initiation factor 4A3 (eIF4A3), which binds to the MMP9 mRNA transcript, induced circMMP9 cyclization and increased circMMP9 expression in GBM. Conclusions Our findings indicate that eIF4A3-induced circMMP9 is an important underlying mechanism in GBM cell proliferation, invasion and metastasis through modulation of the miR-124 signaling pathway, which could provide pivotal potential therapeutic targets for the treatment of GBM. Graphical abstract

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