MicroRNA-16-5p Promoted Fibroblast-Like Synoviocyte Proliferation and Suppressed Apoptosis via Targeting Suppressor of Cytokine Signaling 6 (SOCS6) in Human Rheumatoid Arthritis

2021 ◽  
Vol 11 (9) ◽  
pp. 1744-1751
Author(s):  
Deqian Meng ◽  
Wenyou Pan ◽  
Ju Li
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Cytokine Signaling ◽  
Human Rheumatoid Arthritis ◽  
Reporter Assay ◽  
Functional Roles ◽  
Proliferation And Apoptosis ◽  
Fibroblast Like Synoviocytes

Accumulating evidence have indicated that MicroRNAs (miRNAs) are key regulators in human rheumatoid arthritis (RA). The aim of this study was to explore the functional roles of miR-16-5p in proliferation, inflammation, and apoptosis of fibroblast-like synoviocytes (FLS). The expression of miR-16-5p and SOCS6 in FLA was detected by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation and apoptosis were measured by CCK-8 assay and flow cytometry, respectively. Luciferase reporter assay was used to verify the direct target of miR-16-5p. Western blot analysis was performed to analysis the levels of SOCS6, Bcl-2, Bax and cleaved caspase 3. miR-16-5p expression was significantly upregulated while SOCS6 level was decreased in RA-FLS compared with normal FLS. In addition, luciferase reporter assay confirmed that SOCS6 was the target of miR-16-5p. Silencing of miR-16-5p inhibited cell proliferation, releases of TNF-α, IL-1β, IL-6 and IL-8, and induced the apoptosis. The effects of miR-16-5p silencing on RA-FLS were reversed by downregulation of SOCS6. In summary, knockdown of miR-16-5p could suppress cell proliferation and accelerate the apoptosis of RA-FLS through targeting SOCS6, which may provide a potential therapeutic target for patients with RA.

scholarly journals MiR-129-5p regulates cell proliferation and apoptosis via IGF-1R/Src/ERK/Egr-1 pathway in RA-fibroblast-like synoviocytes

2019 ◽  
Vol 39 (12) ◽  
Author(s):  
Yan Zhang ◽  
Ni Yan ◽  
Xiaoqing Wang ◽  
Yanhai Chang ◽  
Yu Wang
Keyword(s):  
Cell Proliferation ◽  
Signaling Pathway ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Pcr Assay ◽  
Proliferation And Apoptosis ◽  
Direct Target ◽  
Induced Apoptosis ◽  
Fibroblast Like Synoviocytes

Abstract It is reported that miR-129-5p plays an important role in various diseases, but its effect on rheumatoid arthritis (RA) and the potential mechanism remain to be clarified. In the present research, we aimed to investigate the effect of miR-129-5p on RA and the special molecular mechanism. First, the expression of miR-129-5p was analyzed in RA patients and RA Fibroblast-like synoviocytes (RA-FLSs) by RT-PCR assay. The cell viability, apoptotic rate and the relative expression of caspase-3 and caspase-8 were measured by CCK-8, Annexin-FITC/propidium iodide (PI) and ELISA, respectively. Luciferase reporter assay was performed to investigate the target of miR-129-5p. The results revealed that the expression of miR-129-5p was down-regulated in RA patients and RA-FLSs. In addition, miR-129-5p inhibited cell proliferation and induced apoptosis of RA-FLS. Furthermore, luciferase reporter assay demonstrated that insulin-like growth factor-1 receptor (IGF-1R) was the direct target of miR-129-5p, and IGF-1R promoted cell proliferation and inhibited apoptosis by activating Src/ERK/Egr-1 signaling. Furthermoremore, the Src/ERK/Egr-1 signaling pathway was suppressed by miR-129-5p. Collectively, the results of the present study suggested that miR-129-5p regulated cell proliferation and apoptosis via IGF-1R/Src/ERK/Egr-1 signaling pathway in RA.

MiR-1246 Involves in the Proliferation, Apoptosis and Inflammation of Rheumatoid Arthritis Fibroblast-Like Synoviocytes by Targeting Axis Inhibition Protein 2 and Glycogen Synthetase Kinase-3β via Wnt/β-Catenin Pathway in Rheumatoid Arthritis

2021 ◽  
Vol 11 (11) ◽  
pp. 2246-2253
Author(s):  
Siyin Guo ◽  
Shichang Hu ◽  
Guoyuan Zhao ◽  
Weijian Hu ◽  
Yiling Liao ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Bioinformatic Analysis ◽  
Vital Role ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Regulatory Effects ◽  
Apoptosis And Inflammation

Background and Objective: Accumulating evidence supports that fibroblast-like synovial cells (FLS) plays a vital role in the pathogenesis of rheumatoid arthritis (RA). miR-1246 has been reported to be up-regulated in the sera of RA patients. The purpose of the present research was to determine the potential role of miR-1246 in RA and the underlying mechanisms. Methods: miR-1246, GSK3β and AXIN2 levels were determined by RT-qPCR. By exploiting miR-1246 inhibitor, shRNA-GSK3β and shRNA-AXIN2, we detected the effects of miR-1246, GSK3β and AXIN2 on cell proliferation, apoptosis and inflammation in RA-FLSs. Bioinformatics analysis predicted the binding sites of miR-1246 to AXIN2, miR-1246 to GSK3β. Moreover, luciferase reporter assay verified the binding relationship. Results: In comparison with normal human FLSs, higher levels of miR-1246 existed in RA-FLSs. Downregulation of miR-1246 inhibited cell proliferation, inflammation and β-catenin expression, and promoted cell apoptosis. Furthermore, bioinformatic analysis and dual-luciferase reporter assay identified AXIN2 or GSK3β as a target gene of miR-1246. Downregulation of miR-1246 enhanced AXIN2 and GSK3β expression in RA-FLSs. Besides, co-transfection with shRNA-GSK3β or shRNA-AXIN2 partly reversed the regulatory effects of miR-1246 inhibitor in RA-FLSs. Conclusions: Collectively, our in vitro experiments proved that downregulation of miR-1246 might alleviate RA pathogenesis by targeting AXIN2 and GSK3β via Wnt/β-Catenin axis.

scholarly journals Knockdown of Long Noncoding RNA HOXA-AS2 Suppresses Chemoresistance of Acute Myeloid Leukemia via the miR-520c-3p/S100A4 Axis

2018 ◽  
Vol 51 (2) ◽  
pp. 886-896 ◽  
Author(s):  
Xiaoya Dong ◽  
Zhigang Fang ◽  
Mingxue Yu ◽  
Ling Zhang ◽  
Ruozhi Xiao ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Online Programs ◽  
Acute Myeloid

Background/Aims: Among different molecular candidates, there is growing data to support that long noncoding RNAs (lncRNAs) play a significant role in acute myeloid leukemia (AML). HOXA-AS2 is significantly overexpressed in a variety of tumors and associated with anti-cancer drug resistance, however, little is known regarding the expression and function of HOXA-AS2 in the chemoresistance of AML. In this study, we aimed to determine the role and molecular mechanism of HOXA-AS2 in adriamycin-based chemotherapy resistance in AML cells. Methods: Quantitative real-time PCR was used to detect HOXA-AS2 expression in the BM samples and ADR cell lines, U/A and T/A cells. Furthermore, the effects of HOXA-AS2 silencing on cell proliferation and apoptosis were assessed in vitro by CCK8 and flow cytometry, and on tumor growth in vivo. Furthermore, bioinformatics online programs predicted and luciferase reporter assay were used to validate the association of HOXA-AS2 and miR-520c-3p in AML. Results: In this study, we showed that HOXA-AS2 is significantly upregulated in BM samples from AML patients after treatment with adriamycin-based chemotherapy and in U/A and T/A cells. Knockdown of HOXA-AS2 inhibited ADR cell proliferation in vitro and in vivo and promoted apoptosis. Bioinformatics online programs predicted that HOXA-AS2 sponge miR-520c-3p at 3’-UTR with complementary binding sites, which was validated using luciferase reporter assay and anti-Ago2 RIP assay. HOXA-AS2 could negatively regulate the expression of miR-520c-3p in ADR cells. S100A4 was predicted as a downstream target of miR-520c-3p, which was confirmed by luciferase reporter assay. Conclusion: Our results suggest that HOXA-AS2 plays an important role in the resistance of AML cells to adriamycin. Thus, HOXA-AS2 may represent a therapeutic target for overcoming resistance to adriamycin-based chemotherapy in AML.

scholarly journals FENDRR Sponges miR-424-5p to Inhibit Cell Proliferation, Migration and Invasion in Colorectal Cancer

2020 ◽  
Vol 19 ◽  
pp. 153303382098010
Author(s):  
Chuan Cheng ◽  
Huixia Li ◽  
Jiujian Zheng ◽  
Jie Xu ◽  
Peng Gao ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Proliferation ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Reporter Assay ◽  
Differential Analysis ◽  
Inhibit Cell Proliferation ◽  
Dual Luciferase Reporter Assay

Objective: LncRNAs are non-coding RNAs exerting vital roles in the occurrence and development of various cancer types. This study tended to describe the expression pattern of FENDRR in colorectal cancer (CRC), and further investigate the role of FENDRR in CRC cell biological behaviors. Methods: Gene expression profile of colon cancer was accessed from the TCGA database, and then processed for differential analysis for identification of differentially expressed lncRNAs and miRNAs. Some in vitro experiments like qRT-PCR, MTT, colony formation assay, wound healing assay and Transwell assay were performed to assess the effect of FENDRR on cell biological behaviors. Dual-luciferase reporter assay was conducted to further validate the targeting relationship between FENDRR and miR-424-5p, and rescue experiments were carried out for determining the mechanism of FENDRR/miR-424-5p underlying the proliferation, migration and invasion of CRC cells. Results: Bioinformatics analysis suggested that FENDRR was significantly down-regulated in CRC tissue, and low FENDRR was intimately correlated to poor prognosis. FENDRR overexpression could greatly inhibit cell proliferation, migration and invasion. Besides, there was a negative correlation between FENDRR and miR-424-5p. Dual-luciferase reporter assay indicated that miR-424-5p was a direct target of FENDRR. Rescue experiments discovered that FENDRR exerted its role in cell proliferation, migration and invasion in CRC via targeting miR-424-5p. Conclusion: FENDRR is poorly expressed in CRC tissue and cells, and low FENDRR is responsible for the inhibition of cell proliferation, migration and invasion of CRC by means of targeting miR-424-5p.

scholarly journals MiR-18a-5p Promotes NPC Cell Proliferation, Invasion, Migration, and EMT by Targeting SMAD2

2020 ◽  
Author(s):  
Pengcheng Li ◽  
Junhui Xing ◽  
Jianwu Jiang ◽  
Xinyu Tian ◽  
Xuemeng Liu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cell Lines ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Invasion And Migration ◽  
Metastasis Model ◽  
And Migration ◽  
Dual Luciferase Reporter Assay

Abstract Background: Nasopharyngeal carcinoma (NPC) is the most common malignant tumor in the head and neck that is characterized by high local malignant invasion and distant metastasis. miR-18a-5p reportedly plays an important role in tumorigenesis and development. However, little is known about the mechanism underlying miR-18a-5p’s role in NPC.Methods：Quantitative real-time PCR was used to detect the expression of miR-18a-5p in NPC tissues and cell lines. MTT assay and plate clone formation assay were used to detect the effect of miR-18a-5p on NPC cell proliferation. Woundhealing assays and Transwell assays were used to detect the effect of miR-18a-5p on NPC cell invasion and migration. The expressions of epithelialmesenchymal transition (EMT)-related proteins N-cadherin, Vimentin, and E-cadherin were detected by Westernblot. Bioinformatics and dual-luciferase reporter assay were used to detect the targeting interaction between miR-18a-5p and SMAD2. Xenotransplantation and metastasis model were used to detect the effect of miR-18a-5p on NPC growth and metastasis in vivo.Results:miR-18a-5p was highly expressed in NPC tissues and cell lines. Overexpression of miR-18a-5p promotedNPC cell proliferation, invasion, migration, and EMT process, whereas inhibition of miR-18a-5p expression led to the oppositeresults. Results of dual-luciferase reporter assay showed that SMAD2 was the target gene of miR-18a-5p, and SMAD2 could reverse the effect of miR-18a-5p on NPC cell line. Xenotransplantation and metastasis model experiments in nude mice showed that miR-18a-5p promotesNPC growth and metastasis in vivo.Conclusions:Targeting SMAD2 downregulated miR-18a-5p expression, thereby promoting NPC cell proliferation, invasion, migration, and EMT.

scholarly journals LncRNA ACTA2-AS1 Suppress Colon Adenocarcinoma Progression by Sponging miR-4428 Upregulation BCL2L11

2020 ◽  
Author(s):  
Qingyun Pan ◽  
Ying Huang ◽  
Yirui Wang ◽  
Deke Li ◽  
Changjiang Lei
Keyword(s):  
Cell Proliferation ◽  
Colony Formation ◽  
Colon Adenocarcinoma ◽  
Human Colon ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Over Expression ◽  
Non Coding Rna

Abstract BackgroundLong non-coding RNA is considered to be essential to modulate the development and progression of human malignant cancers. And long non-coding RNA can act as crucial modulators by sponging the corresponding microRNA in tumorigenesis. We aimed to elucidate the function of ACTA2-AS1 and its molecular mechanism in colon adenocarcinoma. Materials and MethodsThe expression of ACTA2-AS1, miR-4428 and BCL2L11 in colon adenocarcinoma tissues were detected via qRT-PCR. SW480 and HT29 cells were transfected with shRNA ACTA2-AS1, OE ACTA2-AS1, miRNA mimics of miR-4428, miR-4428 inhibitor, si-BCL2L11 and over-expression of si-BCL2L11. Cell proliferation, colony formation and apoptosis were respectively assessed using CCK-8 assay, colony assay and flow cytometry. Luciferase reporter assay was performed to verify the targets of ACTA2-AS1 and miR-4428. Tumor subcutaneous xenograft mode was constructed to explore tumor growth in vivo. ResultsACTA2-AS1 was obviously downregulated in human colon adenocarcinoma tissues and colon adenocarcinoma cell lines. Silence or over-expression of ACTA2-AS1 promoted or inhibited cell proliferation and colony formation abilities, and regulated apoptosis. The silence of ACTA2-AS1 resulted in the decrease of Bax and increase of Bal2, while restored in OE ACTA2-AS1 group when compared with the control transfected cells. In addition, luciferase reporter assay revealed that ACTA2-AS1 interacted with miR-4428 and suppressed its expression. miR-4428 could bind to 3ʹ untranslated region of BCL2L11 and modulated the expression of BCL2L11 negatively. Knockdown of ACTA2-AS1 and over-expression of BCL2L11 reversed the biological function that ACTA2-AS1 mediated by knockdown ACTA2-AS1 alone. ConclusionOur data demonstrated that ACTA2-AS1 could suppress colon adenocarcinoma progression via sponging miR-4428 to regulate BCL2L11 expression.

scholarly journals Lnc RNA ZFAS1 regulates the proliferation, apoptosis, inflammatory response and autophagy of fibroblast-like synoviocytes via miR-2682-5p/ADAMTS9 axis in rheumatoid arthritis

2020 ◽  
Vol 40 (8) ◽  
Author(s):  
Shanshan Yang ◽  
Wei Yin ◽  
Yan Ding ◽  
Fan Liu
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Inflammatory Response ◽  
Large Subunit ◽  
Luciferase Reporter ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cell Behaviors ◽  
Synovial Tissues ◽  
Related Proteins ◽  
Fibroblast Like Synoviocytes

Abstract Backgrounds: Rheumatoid arthritis (RA) is a frequent autoimmune disease. Emerging evidence indicated that ZNFX1 antisense RNA1 (ZFAS1) participates in the physiological and pathological processes in RA. However, knowledge of ZFAS1 in RA is limited, the potential work pathway of ZFAS1 needs to be further investigated. Methods: Levels of ZFAS1, microRNA (miR)-2682-5p, and ADAM metallopeptidase with thrombospondin type 1 motif 9 (ADAMTS9) were estimated using quantitative real-time polymerase chain reaction (qRT-PCR) assay. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was conducted to explore the ability of cell proliferation in fibroblast-like synoviocytes (FLS-RA). Cell apoptosis was measured via flow cytometry. Also, levels of ADAMTS9, apoptosis-related proteins, cleaved-caspase-3 (active large subunit), and autophagy-related proteins were identified adopting Western blot. Enzyme-linked immunosorbent assay (ELISA) was performed to determine the productions of inflammatory cytokines. Beside, the interrelation between miR-2682-5p and ZFAS1 or ADAMTS9 was verified utilizing dual-luciferase reporter assay. Results: High levels of ZFAS1 and ADAMTS9, and a low level of miR-2682-5p were observed in RA synovial tissues and FLS-RA. Knockdown of ZFAS1 led to the curbs of cell proliferation, inflammation, autophagy, and boost apoptosis in FLS-RA, while these effects were abolished via regaining miR-2682-5p inhibition. Additionally, the influence of miR-2682-5p on cell phenotypes and inflammatory response were eliminated by ADAMTS9 up-regulation in FLS-RA. Mechanically, ZFAS1 exerted its role through miR-2682-5p/ADAMTS9 axis in RA. Conclusion: ZFAS1/miR-2682-5p/ADAMTS9 axis could modulate the cell behaviors, inflammatory response in FLS-RA, might provide a potential therapeutic target for RA treatment.

scholarly journals MiR-196b-5p regulates the proliferation of drug-resistant hepatocellular carcinoma cell lines by activating NFκB/ABCB1 signaling pathway

2020 ◽  
Vol 19 (1) ◽  
pp. 39-44
Author(s):  
Bangming Pu ◽  
Yong Cao ◽  
Yan Li ◽  
Li Tang ◽  
Jiyi Xia ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Cycle ◽  
Cell Proliferation ◽  
Signaling Pathway ◽  
Cell Lines ◽  
Hepg2 Cells ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Drug Resistant ◽  
Reporter Assay

Purpose: To explore the molecular function of miR-196b-5p in hepatocellular carcinoma (HCC).Methods: MiR-196b-5p expression levels in HCC tissue samples were assessed by qRT-PCR. MiR-196b-5p was knocked-down or over-expressed in HepG2 cells by transfecting the cells with plasmids expressing either a miR-196b-5p inhibitor or mimic, respectively, while cell proliferation was  assessed by MTT assay. The interaction of miR-196b-5p with target molecules was confirmed using luciferase reporter assay. Cell cycle was investigated by flow cytometry, while NFκBIA expression was assessed by western blotting.Results: MiR-196b-5p was over-expressed in HCC, and miR-196b-5p expression levels in patients with HCC were related to tumor grade. MiR-196b-5p over-expression promoted cell proliferation and colony formation and suppressed cell cycle arrest and apoptosis. The results of luciferase reporter assay showed that miR-196b-5p reduced NFκBIA expression in HepG2 cells by binding to a response element in the 3′ UTR of NFκBIA. Further investigation showed that NFκBIA interacts with NFκB1 and reduces the concentration of NFκB1 in HepG2 cells. The promoter of ATP-binding cassette sub-family B member 1 (ABCB1) was also targeted and bound by NFκB1, which altered the expression of ABCB1 in HepG2 cells.Conclusion: MiR-196b-5p regulates cell proliferation in drug-resistant HCC cell lines via activation of the NFκB/ABCB1 signaling pathway. Keywords: Hepatocellular carcinoma, miR-196b-5p, NFκBIA, NFκB1, ABCB1

Sign in / Sign up

Export Citation Format

Share Document