scholarly journals Inhibition of FOSL2 aggravates the apoptosis of ovarian cancer cells by promoting the formation of inflammasomes

2021 ◽  
Author(s):  
Jie Li ◽  
Li Zhou ◽  
Hongye Jiang ◽  
Lin Lin ◽  
Yinguang Li
Keyword(s):  
Ovarian Cancer ◽  
Western Blot ◽  
Interleukin 1Β ◽  
Migration And Invasion ◽  
Enzyme Linked Immunosorbent Assay ◽  
Ovarian Cancer Cells ◽  
Interleukin 18 ◽  
Western Blot Assay ◽  
Cell Functions ◽  
Caspase 1

Abstract Background Ovarian cancer is a common gynecological malignancy among female patients and poses a serious threat to women’s health. Although it has been established that Fos-like antigen 2 (FOSL2) is linked to ovarian cancer (OC), its exact role in the development of OC remains unknown. Objective This article aims to investigate the role of FOSL2 in ovarian cancer development. Methods FOSL2 expression in ovarian carcinoma and adjacent tissues was assessed using real-time fluorescent quantitative PCR and western blot. We constructed OE/sh-FOSL2 plasmids and Caspase-1 specific inhibitors (Yvad-CMK) and transfected A 2780 cells with them to identify the relevant cell functions. Furthermore, we used western blot assay to determine the changes in expression of apoptosis-associated speck-like protein containing a CARD (ASC), cysteine aspartate-specific proteasezymogen procaspase 1 (pro-caspase-1), cysteinyl aspartate-specific proteinase-1 (caspase-1), interleukin-1β precursor (pro-IL-1β), interleukin-1β (IL-1β), interleukin-18 precursor (pro-IL-18), and interleukin-18 (IL-18). In addition, we measured the concentration of IL-1β and IL-18 using an enzyme-linked immunosorbent assay (ELISA). Moreover, Tthe level of lactate dehydrogenase (LDH) in the cell supernatant was measured by LDH release assay kit. Results The expression of FOSL2 was significantly higher compared with the surrounding tissues. The proliferation, migration, and invasion of A2780 cells were enhanced after transfection with OE-FOSL2 plasmids; however, the cell apoptosis was significantly decreased. When FOSL2 was overexpressed, the inflammasome-associated proteins such as ASC, caspase-1, IL-1β, and IL-18 were downregulated. Furthermore, FOSL2 induced apoptosis and activated the production of inflammasomes in A2780 cells. Co-therapy with Yvad-CMK and substantially inhibited apoptosis and activation of inflammasomes. Conclusions Inhibition of FOSL2 promotes the apoptosis of OC cells by mediating the formation of an inflammasome.

Knockdown of BAG3 synergizes with olaparib to kill ovarian cancer cells via repressing autophagy

2020 ◽  
pp. jim-2020-001602
Author(s):  
Kexin Wang ◽  
Jianhua Zheng
Keyword(s):  
Ovarian Cancer ◽  
Western Blot ◽  
Transfection Efficiency ◽  
Parp Inhibitor ◽  
Beclin 1 ◽  
Cell Counting ◽  
Ovarian Cancer Cells ◽  
Western Blot Assay ◽  
Related Proteins

This study aimed at expounding the synergistic effect of Bcl-2-associated athanogene 3 (BAG3) knockdown and poly ADP-ribose polymerase (PARP) inhibitor on ovarian cancer (OC) cells and the potential mechanism. Short hairpin RNA (shRNA) targeting BAG3 (sh-BAG3) was transfected into SK-OV-3 (SKOV-3 ;SKOV3) and A2780 cells, and western blot assay was used to detect transfection efficiency. Cell proliferation and apoptosis were detected by the cell counting kit-8 method, 5-Bromodeoxyuridine (BrdU) experiment and flow cytometry analysis, respectively. The expressions of apoptosis-related proteins Bax and Bcl-2, as well as the expressions of autophagy-related proteins LC3-I, LC3-II and Beclin-1, were examined by western blot assay. Additionally, the cells were treated with autophagy activator rapamycin to investigate whether the tumor-suppressive function of BAG3 knockdown+PARP inhibitor was dependent on autophagy. In this work, we demonstrated that BAG3 knockdown further sensitized OC cells to olaparib treatment, reducing cellular viability and promoting apoptosis. Both sh-BAG3 and olaparib decreased the expression of Beclin-1 and the LC3-Ⅱ:LC3-I ratio, and their synergism further inhibited the process of autophagy. However, the aforementionede effects were reversed after the cells were treated with rapamycin. Based on these results, we concluded that BAG3 knockdown synergizes with olaparib to kill OC cells in vitro by repressing autophagy.

scholarly journals Circ_0015756 promotes the progression of ovarian cancer by regulating miR-942-5p/CUL4B pathway

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Zhenhua Du ◽  
Lei Wang ◽  
Yu Xia
Keyword(s):  
Ovarian Cancer ◽  
Tumor Growth ◽  
Western Blot ◽  
Gynecologic Cancer ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Circular Rnas ◽  
Western Blot Assay

Abstract Background Ovarian cancer (OC) is the gynecologic cancer with the highest mortality. Circular RNAs (circRNAs) play a vital role in the development and progression of cancer. This study aimed to explore the potential role of circ_0015756 in OC and its molecular mechanism. Methods The levels of circ_0015756, microRNA-942-5p (miR-942-5p) and Cullin 4B (CUL4B) were determined by quantitative real-time PCR (qRT-PCR) or Western blot assay. Cell proliferation, apoptosis, migration and invasion were assessed by Cell Counting Kit-8 (CCK-8), colony formation assay, flow cytometry and transwell assay. The levels of proliferation-related and metastasis-related proteins were measured by Western blot assay. The relationship between miR-942-5p and circ_0015756 or CUL4B was verified by dual-luciferase reporter assay, RNA immunoprecipitation assay and RNA pull-down assay. Xenograft assay was used to analyze tumor growth in vivo. Results Circ_0015756 and CUL4B levels were increased, while miR-942-5p level was decreased in OC tissues and cells. Depletion of circ_0015756 suppressed proliferation, migration and invasion and promoted apoptosis in OC cells. Down-regulation of circ_0015756 hindered OC cell progression via modulating miR-942-5p. Also, up-regulation of miR-942-5p impeded OC cell development by targeting CUL4B. Mechanistically, circ_0015756 up-regulated CUL4B via sponging miR-942-5p. Moreover, circ_0015756 silencing inhibited tumor growth in vivo. Conclusion Knockdown of circ_0015756 suppressed OC progression via regulating miR-942-5p/CUL4B axis, suggesting that circ_0015756 might be a potential therapeutic target for ovarian cancer.

Overexpression of transcriptional co-activator with PDZ-binding motif promotes epithelial mesenchymal transformation of ovarian cancer cells by upregulating Smad3 and Snail1

2020 ◽  
Vol 10 (1) ◽  
pp. 120-126
Author(s):  
Guangyuan Chen ◽  
Ping Huang ◽  
Jiabin Xie ◽  
Rihong Li
Keyword(s):  
Ovarian Cancer ◽  
Western Blot ◽  
Cancer Cells ◽  
Migration And Invasion ◽  
Ovarian Cancer Cells ◽  
Control Group ◽  
Binding Motif ◽  
Qrt Pcr ◽  
Associated Proteins ◽  
Mesenchymal Transformation

This study is intended to explore the effect of transcriptional coactivator with PDZ binding motif (TAZ) expression in ovarian cancer cells as well as investigate the expression of signal proteins Smad3 and Snail1. Ovarian cancer cells (SKOV-3) were divided into two groups: control and TAZ overexpression. The overexpression of TAZ in SKOV-3 cells was determined by immunofluorescence, western blot, and qRT-PCR. The proliferation, invasiveness, and expression of epithelial mesenchymal transformation (EMT)-associated proteins were detected, and the expression of Smad3 and Snail1 proteins was determined by qRT-PCR and western blot, respectively. Small interfering RNA (siRNA) targeting TAZ were synthesized and used to transfect SKOV-3. Cell migration and invasion were observed via a wound healing assay and a transwell assay, respectively. The expressions of representative genes involved in proliferation and migration, EMT-associated proteins and Smad3 and Snail1 proteins were also detected by western blot assays. The results of qRT-PCR, immunofluorescence, and western blot showed that, compared with the control group, the expressions of Smad3 and Snail1 protein were upregulated, and the expression of EMT-related genes-including Actin, N-cadherin, and Vimentin protein-was downregulated in the TAZ overexpression group. After TAZ mRNA was suppressed, the migration and invasion ability of the TAZ siRNA group was weaker than that of the control group. In addition, the expression level of Smad3 and Snail1 decreased when TAZ was silenced, while the expression of EMT-related genes increased. Therefore, TAZ in ovarian cancer cells can promote growth, migration, and invasiveness of cancer cells by regulating genes related to proliferation, migration, and invasion.

scholarly journals CircMAPK9 promotes the progression of fibroblast-like synoviocytes in rheumatoid arthritis via the miR-140-3p/PPM1A axis

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Zhihuan Luo ◽  
Shaojian Chen ◽  
Xiaguang Chen
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Inflammatory Response ◽  
Western Blot ◽  
Joint Disease ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Enzyme Linked Immunosorbent Assay ◽  
Western Blot Assay ◽  
Fibroblast Like Synoviocytes

Abstract Background Rheumatoid arthritis (RA) is a chronic inflammatory joint disease, and fibroblast-like synoviocytes (FLSs) are key effector cells in RA development. Mounting evidence indicates that circular RNAs (circRNAs) participate in the occurrence and development of RA. However, the precise mechanism of circRNA mitogen-activated protein kinase (circMAPK9) in the cell processes of FLSs has not been reported. Methods The expression levels of circMAPK9, microRNA-140-3p (miR-140-3p), and protein phosphatase magnesium-dependent 1A (PPM1A) were determined by quantitative real-time polymerase chain reaction (qRT-PCR) or western blot assay. Cell proliferation was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell apoptosis and cycle distribution were assessed by flow cytometry. Cell migration and invasion were tested by transwell assay. All the proteins were inspected by western blot assay. Inflammatory response was evaluated by enzyme-linked immunosorbent assay (ELISA). The interaction between miR-140-3p and circMAPK9 or PPM1A was verified by dual-luciferase reporter assay. Results CircMAPK9 and PPM1A were upregulated and miR-140-3p was downregulated in RA patients and FLSs from RA patients (RA-FLSs). CircMAPK9 silence suppressed cell proliferation, migration, invasion, inflammatory response, and promoted apoptosis in RA-FLSs. MiR-140-3p was a target of circMAPK9, and miR-140-3p downregulation attenuated the effects of circMAPK9 knockdown on cell progression and inflammatory response in RA-FLSs. PPM1A was targeted by miR-140-3p, and circMAPK9 could regulate PPM1A expression by sponging miR-140-3p. Furthermore, miR-140-3p could impede cell biological behaviors in RA-FLSs via targeting PPM1A. Conclusion CircMAPK9 knockdown might inhibit cell proliferation, migration, invasion, inflammatory response, and facilitate apoptosis in RA-FLSs via regulating miR-140-3p/PPM1A axis, offering a new mechanism for the comprehension of RA development and a new insight into the potential application of circMAPK9 in RA treatment.

The Anti-tumor Activity and Mechanisms of rLj-RGD3 on Human Laryngeal Squamous Carcinoma Hep2 Cells

2020 ◽  
Vol 19 (17) ◽  
pp. 2108-2119
Author(s):  
Yang Jin ◽  
Li Lv ◽  
Shu-Xiang Ning ◽  
Ji-Hong Wang ◽  
Rong Xiao
Keyword(s):  
Wound Healing ◽  
Western Blot ◽  
Morphological Changes ◽  
In Vivo Studies ◽  
Migration And Invasion ◽  
Tunel Staining ◽  
Dna Ladder ◽  
Western Blot Assay

Background: Laryngeal Squamous Cell Carcinoma (LSCC) is a malignant epithelial tumor with poor prognosis and its incidence rate increased recently. rLj-RGD3, a recombinant protein cloned from the buccal gland of Lampetra japonica, contains three RGD motifs that could bind to integrins on the tumor cells. Methods: MTT assay was used to detect the inhibitory rate of viability. Giemsa’s staining assay was used to observe the morphological changes of cells. Hoechst 33258 and TUNEL staining assay, DNA ladder assay were used to examine the apoptotic. Western blot assay was applied to detect the change of the integrin signal pathway. Wound-healing assay, migration, and invasion assay were used to detect the mobility of Hep2 cells. H&E staining assay was used to show the arrangement of the Hep2 cells in the solid tumor tissues. Results: In the present study, rLj-RGD3 was shown to inhibit the viability of LSCC Hep2 cells in vitro by inducing apoptosis with an IC50 of 1.23µM. Western blot showed that the apoptosis of Hep2 cells induced by rLj- RGD3 was dependent on the integrin-FAK-Akt pathway. Wound healing, transwells, and western blot assays in vitro showed that rLj-RGD3 suppressed the migration and invasion of Hep2 cells by integrin-FAKpaxillin/ PLC pathway which could also affect the cytoskeleton arrangement in Hep2 cells. In in vivo studies, rLj-RGD3 inhibited the growth, tumor volume, and weight, as well as disturbed the tissue structure of the solid tumors in xenograft models of BALB/c nude mice without reducing their body weights. Conclusion: hese results suggested that rLj-RGD3 is an effective and safe suppressor on the growth and metastasis of LSCC Hep2 cells from both in vitro and in vivo experiments. rLj-RGD3 might be expected to become a novel anti-tumor drug to treat LSCC patients in the near future.

scholarly journals Propofol suppresses cell viability, cell cycle progression and motility and induces cell apoptosis of ovarian cancer cells through suppressing MEK/ERK signaling via targeting circVPS13C/miR-145 axis

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Huan Lu ◽  
Guanlin Zheng ◽  
Xiang Gao ◽  
Chanjuan Chen ◽  
Min Zhou ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Ovarian Cancer ◽  
Cancer Cells ◽  
Rna Binding ◽  
Erk Signaling ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Ovarian Cancer Cells ◽  
Dependent Manner ◽  
Vacuolar Protein

Abstract Background Propofol is a kind of common intravenous anaesthetic agent that plays an anti-tumor role in a variety of cancers, including ovarian cancer. However, the working mechanism of Propofol in ovarian cancer needs further exploration. Methods The viability and metastasis of ovarian cancer cells were assessed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and transwell assays. Flow cytometry was used to evaluate the cell cycle and apoptosis. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to examine the abundance of circular RNA vacuolar protein sorting 13 homolog C (circVPS13C) and microRNA-145 (miR-145). The target relationship between miR-145 and circVPS13C was predicted by circinteractome database and verified by dual-luciferase reporter assay, RNA-binding protein immunoprecipitation (RIP) assay and RNA-pull down assay. Western blot assay was used to detect the levels of phosphorylated extracellular regulated MAP kinase (p-ERK), ERK, p-MAP kinse-ERK kinase (p-MEK) and MEK, in ovarian cancer cells. Results Propofol treatment suppressed the viability, cell cycle and motility and elevated the apoptosis rate of ovarian cancer cells. Propofol up-regulated miR-145 in a dose-dependent manner. Propofol exerted an anti-tumor role partly through up-regulating miR-145. MiR-145 was a direct target of circVPS13C. Propofol suppressed the progression of ovarian cancer through up-regulating miR-145 via suppressing circVPS13C. Propofol functioned through circVPS13C/miR-145/MEK/ERK signaling in ovarian cancer cells. Conclusion Propofol suppressed the proliferation, cell cycle, migration and invasion and induced the apoptosis of ovarian cancer cells through circVPS13C/miR-145/MEK/ERK signaling in vitro.

scholarly journals Long non-coding RNA XIST regulates ovarian cancer progression via modulating miR-335/BCL2L2 axis

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Qingjuan Meng ◽  
Ningning Wang ◽  
Guanglan Duan
Keyword(s):  
Ovarian Cancer ◽  
Cancer Cells ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Ovarian Cancer Cells ◽  
Human Ovarian Cancer ◽  
Invasion And Migration ◽  
Non Coding Rna ◽  
And Migration ◽  
Long Non Coding Rna

Abstract Background X inactivation-specific transcript (XIST) is the long non-coding RNA (lncRNA) related to cancer, which is involved in the development and progression of various types of tumor. However, up to now, the exact role and molecular mechanism of XIST in the progression of ovarian cancer are not clear. We studied the function of XIST in ovarian cancer cells and clinical tumor specimens. Methods RT-qPCR was performed to detect the expression levels of miR-335 and BCL2L2 in ovarian cancer cells and tissues. MTT and transwell assays were carried out to detect cell proliferation, migration, and invasion abilities. Western blot was performed to analyze the expression level of BCL2L2. The interaction between miR-335 and XIST/BCL2L2 was confirmed using a luciferase reporter assay. Results The inhibition of XIST can inhibit the proliferation invasion and migration of human ovarian cancer cells. In addition, the miR-335/BCL2L2 axis was involved in the functions of XIST in ovarian cancer cells. These results suggested that XIST could regulate tumor proliferation and invasion and migration via modulating miR-335/BCL2L2. Conclusion XIST might be a carcinogenic lncRNA in ovarian cancer by regulating miR-335, and it can serve as a therapeutic target in human ovarian cancer.

scholarly journals NEAT1 aggravates sepsis-induced acute kidney injury by sponging miR-22-3p

Open Medicine ◽  
2020 ◽  
Vol 15 (1) ◽  
pp. 333-342
Author(s):  
Yawei Feng ◽  
Jun Liu ◽  
Ranliang Wu ◽  
Peng Yang ◽  
Zhiqiang Ye ◽  
...  
Keyword(s):  
Acute Kidney Injury ◽  
Western Blot ◽  
Cell Apoptosis ◽  
Noncoding Rna ◽  
Cell Injury ◽  
Kidney Injury ◽  
Inflammatory Factors ◽  
Luciferase Reporter ◽  
Enzyme Linked Immunosorbent Assay ◽  
Western Blot Assay

AbstractBackground and aimAcute kidney injury (AKI) is a common complication of sepsis. Long noncoding RNA nuclear-enriched abundant transcript 1 (NEAT1) plays a vital role in various diseases, including AKI. This study aimed to investigate the function and mechanism of NEAT1 in sepsis-induced AKI.Materials and methodsA septic AKI model was established by treating HK-2 cells with lipopolysaccharide (LPS). The levels of NEAT1 and miR-22-3p were measured by quantitative real-time PCR. Cell apoptosis was assessed by flow cytometry. The levels of apoptosis-related protein and autophagy-related factors were examined by the western blot assay. An enzyme-linked immunosorbent assay was used to calculate the contents of inflammatory factors. The interaction between NEAT1 and miR-22-3p was validated by dual-luciferase reporter assay, RNA immunoprecipitation assay, and RNA pull-down assay. The levels of nuclear factor (NF)-κB pathway-related proteins were evaluated by the western blot assay.ResultsNEAT1 was upregulated, while miR-22-3p was downregulated in patients with sepsis and in LPS-stimulated HK-2 cells. LPS treatment triggered cell apoptosis, autophagy, and inflammatory response in HK-2 cells. NEAT1 knockdown attenuated LPS-induced cell injury. NEAT1 modulated LPS-triggered cell injury by targeting miR-22-3p. Furthermore, NEAT1 regulated the NF-κB pathway by modulating miR-22-3p.ConclusionDepletion of NEAT1 alleviated sepsis-induced AKI via regulating the miR-22-3p/NF-κB pathway.

scholarly journals OC125, M11 and OV197 Epitopes Are Not Uniformly Distributed in the Tandem-Repeat Region of CA125 and Require the Entire SEA Domain

2013 ◽  
Vol 34 (4) ◽  
pp. 257-267 ◽  
Author(s):  
Alessandro Bressan ◽  
Francesca Bozzo ◽  
Carlo Alberto Maggi ◽  
Monica Binaschi
Keyword(s):  
Ovarian Cancer ◽  
Monoclonal Antibodies ◽  
Western Blot ◽  
Tandem Repeat ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Human Cancer ◽  
Ovarian Cancer Cells ◽  
Repeat Region ◽  
Tandem Repeat Region

The human cancer antigen 125 (CA125) is over-expressed in epithelial ovarian cancer cells and it plays a role in the pathogenesis of ovarian cancer. This protein presents a repeat region containing up to sixty tandem repeat units. The anti-CA125 monoclonal antibodies have been previously classified into three groups: two major families, the OC125-like antibodies and M11-like antibodies, and a third group, the OV197-like antibodies. A model in which a single repeat unit contains all the epitopes for these antibodies has been also proposed, even if their exact position is still undetermined. In the present work, the affinities of the monoclonal antibodies, representative of the three families, have been investigated for different CA125-recombinant repeats through Western blot analysis. Different patterns of antibody recognition for the recombinant repeats show that CA125 epitopes are not uniformly distributed in the tandem repeat region of the protein. The minimal region for the recognition of these antibodies has been also individuated in the SEA domain through the subcloning of deleted sequences of the highly recognized repeat-25 (R-25), their expression as recombinant fragments inE. coliand Western blot analysis. Obtained data have been further confirmed by ELISA using the entire R-25 as coating antigen.

scholarly journals FBXO6-mediated RNASET2 ubiquitination and degradation governs the development of ovarian cancer

2021 ◽  
Vol 12 (4) ◽  
Author(s):  
Mei Ji ◽  
Zhao Zhao ◽  
Yue Li ◽  
Penglin Xu ◽  
Jia Shi ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Inverse Correlation ◽  
Degradation Pathway ◽  
Migration And Invasion ◽  
Ovarian Cancer Cells ◽  
Poor Overall Survival ◽  
Protein Levels ◽  
Ubiquitin E3 Ligase ◽  
Ovarian Tumorigenesis ◽  
F Box Protein

AbstractRNASET2 (Ribonuclease T2) functions as a tumor suppressor in preventing ovarian tumorigenesis. However, the mechanisms underlying the regulation of RNASET2 protein are completely unknown. Here we identified the F-box protein FBXO6, a substrate recognition subunit of an SCF (Skp1-Cul1-F-box protein) complex, as the ubiquitin E3 ligase for RNASET2. We found that the interaction between FBXO6 and RNASET2 induced RNASET2 instability through the ubiquitin-mediated proteasome degradation pathway. FBXO6 promoted K48-dependent ubiquitination of RNASET2 via its FBA domain. Through analysis of the TCGA dataset, we found that FBXO6 was significantly increased in ovarian cancer tissues and the high expression of FBXO6 was related to the poor overall survival (OS) of ovarian cancer patients at advanced stages. An inverse correlation between the protein levels of FBXO6 and RNASET2 was observed in clinic ovarian cancer samples. Depletion of FBXO6 promoted ovarian cancer cells proliferation, migration, and invasion, which could be partially reversed by RNASET2 silencing. Thus, our data revealed a novel FBXO6-RNASET2 axis, which might contribute to the development of ovarian cancer. We propose that inhibition of FBXO6 might represent an effective therapeutic strategy for ovarian cancer treatment.

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