scholarly journals Curcumin induces paraoxonase 1 in cultured hepatocytes in vitro but not in mouse liver in vivo

2010 ◽  
Vol 105 (2) ◽  
pp. 167-170 ◽  
Author(s):  
Charlotte Schrader ◽  
Christina Schiborr ◽  
Jan Frank ◽  
Gerald Rimbach
Keyword(s):  
Cultured Cells ◽  
Paraoxonase 1 ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Protein Levels ◽  
Huh7 Cells ◽  
Gene Assay ◽  
B6c3f1 Mice

Paraoxonase 1 (PON1) is an enzyme that is mainly synthesised in the liver and protects LDL from oxidation, thereby exhibiting antiatherogenic properties. Using a luciferase reporter gene assay, we tested curcumin for its ability to induce PON1 in Huh7 hepatocytes in culture. Curcumin ( ≥ 10 μmol/l) dose-dependently induced PON1 transactivation in Huh7 cells. However, dietary supplementation of female B6C3F1 mice with curcumin (500 mg/kg diet) for 2 weeks did not increase the hepatic PON1 mRNA and protein levels. No curcumin was detectable in the plasma of the 12 h fasted mice. In conclusion, curcumin may be a potent PON1 inducer in cultured cells in vitro, but not in the liver of curcumin-fed mice because of its low concentrations in vivo.

scholarly journals Impact of RARα and miR-138 on retinoblastoma etoposide resistance

Tumor Biology ◽  
2021 ◽  
Vol 43 (1) ◽  
pp. 11-26
Author(s):  
Maike Busch ◽  
Natalia Miroschnikov ◽  
Jaroslaw Thomas Dankert ◽  
Marc Wiesehöfer ◽  
Klaus Metz ◽  
...  
Keyword(s):  
Cell Viability ◽  
Cell Lines ◽  
Cancer Progression ◽  
Treated Patient ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Gene Assay ◽  
The Impact

BACKGROUND: Retinoblastoma (RB) is the most common childhood eye cancer. Chemotherapeutic drugs such as etoposide used in RB treatment often cause massive side effects and acquired drug resistances. Dysregulated genes and miRNAs have a large impact on cancer progression and development of chemotherapy resistances. OBJECTIVE: This study was designed to investigate the involvement of retinoic acid receptor alpha (RARα) in RB progression and chemoresistance as well as the impact of miR-138, a potential RARα regulating miRNA. METHODS: RARα and miR-138 expression in etoposide resistant RB cell lines and chemotherapy treated patient tumors compared to non-treated tumors was revealed by Real-Time PCR. Overexpression approaches were performed to analyze the effects of RARα on RB cell viability, apoptosis, proliferation and tumorigenesis. Besides, we addressed the effect of miR-138 overexpression on RB cell chemotherapy resistance. RESULTS: A binding between miR-138 and RARα was shown by dual luciferase reporter gene assay. The study presented revealed that RARα is downregulated in etoposide resistant RB cells, while miR-138 is endogenously upregulated. Opposing RARα and miR-138 expression levels were detectable in chemotherapy pre-treated compared to non-treated RB tumor specimen. Overexpression of RARα increases apoptosis levels and reduces tumor cell growth of aggressive etoposide resistant RB cells in vitro and in vivo. Overexpression of miR-138 in chemo-sensitive RB cell lines partly enhances cell viability after etoposide treatment. CONCLUSIONS: Our findings show that RARα acts as a tumor suppressor in retinoblastoma and is downregulated upon etoposide resistance in RB cells. Thus, RARα may contribute to the development and progression of RB chemo-resistance.

scholarly journals MiR-203 Determines Poor Outcome and Suppresses Tumor Growth by Targeting TBK1 in Osteosarcoma

2015 ◽  
Vol 37 (5) ◽  
pp. 1956-1966 ◽  
Author(s):  
Shiping Liu ◽  
Peng Feng
Keyword(s):  
Cell Growth ◽  
Cell Lines ◽  
Cancer Progression ◽  
Human Cancer ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Poor Overall Survival ◽  
Gene Assay

Background/Aims: Increasing evidence has shown that miR-203 plays important role in human cancer progression. However, little is known about the function of miR-203 in osteosarcoma (OS). Methods: The expression of miR-203 in OS tissues and cell lines were examined by qRT-PCR. The biological role of miR-20 in OS cell proliferation was examined in vitro and in vivo. The targets of miR-203 were identified by a luciferase reporter gene assay. Results: miR-203 was down regulated in OS tissues and cell lines; decreased miR-203 was associated with a poor overall survival in OS patients. Restoration of miR-203 expression reduced cell growth in vitro and suppressed tumorigenicity in vivo. In contrast, inhibition of miR-203 stimulated OS cell growth both in vitro and in vivo. In addition, TANK binding kinase 1 (TBK1) was identified as a direct target of miR-203; overexpression of TBK1 partly reversed the suppressive effects of miR-203. Furthermore, TBK1 was found up-regulated and inversely correlated with miR-203 in OS tissues. Conclusion: Taken together, these findings suggest that miR-203 acts as a tumor suppressor via regulation of TBK1 expression in OS progression, and miR-203 may be a promising therapeutic target for OS.

scholarly journals PRPF6 Induces the Alternative Splicing of lncRNA SNHG16 To Promote Progression and Paclitaxel Resistance of Ovarian Cancer via Regulating GATA3 Transcription

2021 ◽  
Author(s):  
Han Wang ◽  
Yingying Zhou ◽  
Siyang Zhang ◽  
Ya Qi ◽  
Min Wang
Keyword(s):  
Ovarian Cancer ◽  
Alternative Splicing ◽  
Epithelial To Mesenchymal Transition ◽  
Binding Protein ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Mesenchymal Transition ◽  
Gene Assay

Abstract Background Small nucleolar RNA host gene 16 (SNHG16) and pre-mRNA processing factor 6(PRPF6) play vital roles in regulatory mechanisms of multiple cancers, but the mechanisms in ovarian cancer (OC) remains poorly understood. Methods The expression of SNHG16 transcripts-SNHG16-L/S in OC tissues were analyzed by real-time PCR (RT-PCR). The expression of PRPF6 in OC tissues were detected by Immunohistochemistry (IHC). Tumorigenesis, epithelial-to-mesenchymal transition (EMT) and PTX-resistance were detected by western blot, transwell, CCK-8 assays, colony formation assays and flow cytometry analyses. Molecular interactions were examined by dual-luciferase reporter gene assay, RNA immunoprecipitation (RIP) and chromatin immunoprecipitation (ChIP). Results The results indicated the expression of SNHG16-L/S was opposite in chemo-resistance and chemo-sensitivity tissues of OC. And SNHG16-L/S had different effects on the progression and PTX-resistance of OC cells. SNHG16-L inhibited GATA binding protein 3 (GATA3) transcription through CCAAT/enhancer-binding protein b (CEBPB) to further promote tumorigenesis, EMT and PTX-resistance of OC. Moreover, PRPF6 was upregulated in chemo-resistance tissues of OC. PRPF6 promoted tumorigenesis and PTX-resistance in vitro and in vivo. Mechanistically, PRPF6 induced the alternative splicing of SNHG16 to downregulate SNHG16-L, which further mediated progression and PTX-resistance through upregulating GATA3 in OC. Conclusions Totally, the results demonstrated that PRPF6 promoted progression and PTX-resistance in OC through SNHG16-L/CEBPB/GATA3 axis. Thus, PRPF6 may become a valuable target for OC therapy.

scholarly journals miR-495-3p Depresses Cell Proliferation and Migration By Down-Regulating HMGB1 in Colorectal Cancer

2021 ◽  
Author(s):  
Zhang Jieling ◽  
Li Kai ◽  
Zheng Huifen ◽  
Zhu Yiping
Keyword(s):  
Colorectal Cancer ◽  
Cell Proliferation ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Gene Assay ◽  
And Migration ◽  
Cancer Tissues

Abstract Background: MicroRNAs play an important role in the genesis and progression of tumors, including colorectal cancer (CRC), which has a high morbidity and mortality rate. In this research, the role of miR-495-3p and HMGB1 in CRC was investigated.Methods: We performed qRT-PCR to detect the expression of miR-495-3p in colorectal cancer tissues and cell lines. Functional experiments such as CCK-8 assay, EDU assay, Transwell assay and apoptosis assay were conducted to explore the effects of miR-495-3p on the proliferation, migration and apoptosis of CRC cells in vitro. Then, the use of database prediction, dual-luciferase reporter gene assay and functional experiments verified the role of miR-495-3p target gene HMGB1 in CRC. Finally, rescue experiments was performed to investigate whether overexpression of HMGB1 could reverse the inhibitory effect of miR-495-3p on CRC cell proliferation in vivo and in vitro.Results: miR-495-3p was down-regulated in colorectal cancer tissues and cell lines, and could inhibit the proliferation and migration of colorectal cancer cells, and promote cell apoptosis. The database prediction and dual-luciferase reporter gene assay showed that HMGB1 was the downstream target gene of miR-495-3p. We finally demonstrated that miR-495-3p inhibited CRC cell proliferation by targeting HMGB1 in vitro and in vivo.Conclusion: Our research shows that miR-495-3p inhibits the progression of colorectal cancer by down-regulating the expression of HMGB1, which indicates that miR-495-3p may become a potential therapeutic target for colorectal cancer.

scholarly journals IFN-γ Enhances the Efficacy of Exosomes Derived from Mesenchymal Stem Cells in Myocardial Infarction Rats via miR-21 Stimulated by STAT1

2021 ◽  
Author(s):  
Jian Zhang ◽  
Yao Lu ◽  
Yangming Mao ◽  
Yue Yu ◽  
Tianyu Wu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Umbilical Vein ◽  
Luciferase Reporter ◽  
H9c2 Cells ◽  
Luciferase Reporter Gene ◽  
Gene Assay ◽  
Ifn Γ

Abstract Background: Mesenchymal stem cells (MSCs) activated with IFN-γ elicit more powerful physical effects. Exosomes (Exos) secreted from MSCs have protective against myocardial injury. The aim of this study was to investigate whether Exsos derived from IFN-γ-pretreated MSCs exhibit more potent cardioprotective function and the underlying mechanisms. Methods: Exos were isolated from MSCs (Ctrl-Exo) and IFN-γ-primed MSCs (IFN-γ-Exo) and were then delivered to H9c2 cells or human umbilical vein endothelial cells (HUVECs) in vitro under oxygen and glucose deprivation (OGD) condition or in vivo in an infarcted rat heart. RNA sequencing was to identify the different expressed functional transcription factor (TF). Quantitative reverse transcription-PCR (qPCR) was to confirm the upregulated TF and miRNA in IFN-γ-primed MSCs. Dual-luciferase reporter gene assay were to analyze the transcriptional regulation of miRNAs by STAT1. The target of miR-21-5p (miR-21) was disclosed by luciferase reporter assays and qPCR. The function of BTG2 was verified in vitro under OGD condition.Result: IFN-γ-Exo accelerated migration, tube-like structure formation, and prevented H9c2 from OGD-induced apoptosis. Similarly, IFN-γ-Exo leaded to further reduction in fibrosis size, reduced cardiomyocyte apoptosis and improved cardiac function compared to Ctrl-Exo. miR-21 was significantly upregulated in both IFN-γ-primed MSCs and IFN-γ-Exo. STAT1 transcriptionally induced miR-21 expression. Up-regulated miR-21 can inhibit the expression of BTG2. BTG2 promoted H9c2 cells apoptosis and reversed the protective effect of miR-21 under OGD environment.Conclusion: IFN-γ-Exo have enhanced therapeutic efficacy against acute MI possibly through promoting angiogenesis and anti-apoptotic effect through increasing the level of miR-21, which directly targeted on BTG2.

Bioengineered microRNA-1291 effects on pancreatic cancer cell proliferation and xenograft tumor growth.

2018 ◽  
Vol 36 (4_suppl) ◽  
pp. 307-307
Author(s):  
Mei-Juan Tu ◽  
Zhijian Duan ◽  
Qianyu Zhang ◽  
Jing-Xin Qiu ◽  
Frank J Gonzalez ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Tumor Growth ◽  
Therapeutic Potential ◽  
Xenograft Model ◽  
Luciferase Reporter ◽  
Mrna Levels ◽  
Luciferase Reporter Gene ◽  
Protein Levels ◽  
Gene Assay

307 Background: MicroRNAs (miR) have proved to be vital regulators in the control of tumor progression. Our recent studies have revealed miR-1291 is downregulated in patient pancreatic cancer (PC) specimens and re-introduction of miR-1291 suppresses tumorigenesis of PC cells. We have developed a novel ncRNA bioengineering technology to produce a miR-1291 prodrug. In this study, we aimed to assess the effectiveness of this miR-1291 prodrug as a monotherapy, as well as in combination with chemotherapy, for treatment of PC. Methods: Sensitivity of PC cells to miR-1291 prodrug alone, gemcitabine plus nab-paclitaxel (Gem-nP) alone, and their combination was evaluated by CellTiter-Glo assay. Mature miR-1291 and ARID3B mRNA levels were determined by quantitative real-time PCR (q-PCR) assay. A luciferase reporter gene assay was used to validate interaction between miR-1291 and ARID3B 3’UTR. Target protein expression was examined by Western blot and immunofluorescence analyses. PANC-1 and PC patient-derived xenograft (PDX) mouse models were established and used to assess anti-tumor effects of miR-1291 monotherapy and combination therapy with Gem-nP. Results: Cytotoxicity assays showed that miR-1291 prodrug enhanced the sensitivity of PANC-1 and AsPC-1 cells to Gem-nP. Luciferase assays confirmed ARID3B as a target for miR-1291 as predicted by computational analysis. qPCR analysis demonstrated that miR-1291 prodrug was readily processed to mature miR-1291 and subsequently upregulated ARID3B mRNA levels. miR-1291 prodrug also elevated the protein levels of ARID3B. Co-administration of miR-1291 prodrug and Gem-nP increased caspase-3/7 and γH2AX levels in PC cells, compared to miR-1291 or Gem-nP treatment alone. In addition, systemic administration of in vivo-jet PEI formulated miR-1291 prodrug suppressed tumor growth in both a PANC-1 xenograft model and three PDX models, and largely enhanced the efficacy of Gem-nP. All treatments were well tolerated in mice in vivo. Conclusions: Our bioengineered miR-1291 prodrug has therapeutic potential as a monotherapy but also can act as a sensitizing agent to chemotherapy. This novel treatment approach should be further explored for PC.

scholarly journals CircRNA1615 Inhibits Ferroptosis via Modulation of Autophagy by the miRNA152-3p/LRP6 Axis in Cardiomyocytes of Myocardial Infarction

2021 ◽  
Author(s):  
Rui-lin Li ◽  
Cheng-hui Fan ◽  
Shi-yu Gong ◽  
Sheng Kang
Keyword(s):  
Myocardial Infarction ◽  
Molecular Mechanism ◽  
Biological Function ◽  
Myocardial Cell ◽  
Pathological Process ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Gene Assay

Abstract Background Searching for new molecular targets of ferroptosis is gradually becoming the focus in the field of cardiovascular disease research. This study was aimed to explore the biological function and molecular mechanism of ferroptosis of circRNA modulation in cardiomyocytes of myocardial infarction (MI).Method We explored the regulatory effect and molecular mechanism of LPR6 on myocardial cell ferroptosis by establishing a model of MI in vivo and in vitro, constructed the regulatory network of circRNA-miRNA-LRP6 by the bioinformatics analysis, and focused on the biological function and molecular mechanism of circRNA1615 regulating ferroptosis in MI by the overexpression or knockdown of circRNA1615, the RIP experiments, and double luciferase reporter gene assay.Results Ferrostatin-1(ferroptosis inhibitor) can improve the pathological process of MI; LRP6 was involved in the process of ferroptosis in cardiomyocytes; LRP6 deletion regulates ferroptosis in cardiomyocytes through autophagy; Screening and identification of circRNA1615 targets LRP6; circRNA1615 inhibits ferroptosis in cardiomyocytes; circRNA1615 regulates the expression of LRP6 through sponge adsorption of miR-152-3p, and then prevent LRP6-mediated autophagy-related ferroptosis in cardiomyocytes, finally regulate the pathological process of MI.Conclusions CircRNA1615 inhibits ferroptosis via modulation of autophagy by the miRNA152-3p/LRP6 molecular axis in cardiomyocytes of myocardial infarction.

scholarly journals Identification of the ASPM-miR-26b-5p network associated with the aggressive traits of HCC cells

2020 ◽  
Author(s):  
Hong-Guang Li ◽  
Heng-Jun Gao ◽  
Fang-Feng Liu ◽  
Jun Liu
Keyword(s):  
Tumor Model ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Loss Of Function ◽  
Luciferase Reporter Gene ◽  
Luciferase Reporter Gene Assay ◽  
Gene Assay ◽  
Hcc Cells

Abstract Background: Even though earlier reports have revealed that abnormal spindle-like microcephaly associated (ASPM) exert essential roles in diverse malignancies, its relationship between specific microRNAs (miRNAs) in regulation of hepatocellular carcinoma (HCC) progression has never been elaborated. Methods: Bioinformatics analysis detected differentially expressed genes in HCC and normal. qRT-PCR was performed to detect expression of miR-26b-5p in HCC tissues and cells. HCC cells were transfected with plasmids and their proliferative ability and colony formation were detected with loss-of-function assay. The invasion of HCC cells was determined using Transwell assay. The expression of ASPM was detected by western blotting. Luciferase reporter gene assay was performed to detect the interaction between miR-26b-5p and ASPM. ASMP silencing cells were injected into mice to establish xenograft tumor model.Results: Herein, we proved that ASPM was upregulated in HCC and higher level of ASPM was significantly associated with worse survival in HCC patients. ASPM silencing restrained HCC cell proliferation, migration and invasion capacities in vitro. In vivo, downregulation of ASPM also suppressed HCC cells growth. Mechanistic analyses illustrated that ASPM was a directly target of miR-26b-5p. The expression of ASPM was negatively modulated by miR-26b-5p. Rescues assays displayed that miR-26b-5p inhibited HCC cells growth and invasion via modulating the expression of ASPM. Conclusions: Our work validated that miR-26b-5p restrained the aggressiveness of HCC cells through targeting ASPM.

scholarly journals NR2F2-AS1 Suppressed Trastuzumab Effects on Esophageal Cancer By Inhibiting miR-4429 and miR-425-5p Expression Through Targeting IGF1R

2021 ◽  
Author(s):  
Zhibin Zhang ◽  
Xu Zhao ◽  
Rui Zhu ◽  
Hong Ren
Keyword(s):  
Esophageal Cancer ◽  
Cell Viability ◽  
Rna Sequencing ◽  
Inhibitory Effect ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Luciferase Reporter Gene Assay ◽  
Gene Assay

Abstract Aim:In this manuscript, we aimed to investigate the involvement of non-coding RNAs in mediating trastuzumab effects in EAC. Background: Scarce evidences supported that targeted drugs, like Trastuzumab, can be applied to esophageal adenocarcinoma patients (EAC). Objective: Evaluating the role and mechanism of NR2F2-AS1 in regulating Trastuzumab effects in EAC patients. Method: RNA sequencing to screen IGF1R related lncRNAs. qRT-PCR and western blot were used to evaluate the expression level of genes. CCK-8 was used to test the cell proliferation ability. Dual luciferase reporter gene assay and RNA pull-down were used for crosstalk evaluation. Results: NR2F2-AS1 was identified to be associated with HER2 expression by RNA sequencing and its expression related to worse prognosis and advanced T and N stage.NR2F2-AS1 expression induced by Trastuzumab through mediating H3K27ac. Furtherly, miR-4429 and miR-425-5p, which were predicted and proved to interact with NR2F2-AS1 and IGF1R, expressed lowly in esophageal cancer both in vivo and in vitro and suppressed cell viability. Most importantly, miR-4429 and miR-425-5p overexpression could increase trastuzumab’s inhibitory effect on cell viability. Conclusion: Trastuzumab has the potential to suppress EAC progression mainly in the presence of miR-4429 and miR-425-5p overexpression targeting HER2. However, Trastuzumab induces exosomal NR2F2-AS1 expression, which binds to miR-4429 and miR-425-5p to suppress their expression, resulting in the failure of trastuzumab treatment. Therefore, targeting exosomes might be a novel way to develop auxiliary drugs for trastuzumab in EAC.

Linc01559 Served as a Potential Oncogene and Promoted Resistance of Hepatocellular Carcinoma to Oxaliplatin by Directly Sponging miR-6783-3p

2020 ◽  
Vol 21 (2) ◽  
pp. 278-286 ◽  
Author(s):  
Shunbin Dong ◽  
Ying Fu ◽  
Kaibo Yang ◽  
Xing Zhang ◽  
Runchen Miao ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Resistant Cell ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
In Vitro Tests ◽  
Luciferase Reporter Gene ◽  
Migration Ability ◽  
Gene Assay

Background: Oxaliplatin (L-OHP)-based chemotherapy, such as FOLFOX4 (5-fluorouracil, leucovorin, and LOHP), improves the prognosis of patients with late-stage Hepatocellular Carcinoma (HCC). However, the development of resistance to L-OHP leads to failure of chemotherapy. The aim of this study was to investigate the role of linc01559 and miR-6783-3p in regulating resistance to L-OHP. Methods: Quantitative reverse transcription-polymerase chain reaction was used to determine the expression profile. The Cell Counting Kit-8 test and wound healing assay were also used. Dual-luciferase reporter gene assay, RNA pull-down assay, and RNA immunoprecipitation were used to evaluate the interaction between linc01559 and miR-6783-3p. Result: linc01559 expression was associated with response to FOLFOX4, as well as miR-1343-3p and miR-6783-3p expression in vivo. A nomogram, including linc01559 and miR-1343-3p, precisely and accurately predicted the overall survival of patients with HCC. Regarding the in vitro tests, linc01559 showed higher expression in L-OHP-resistant cell lines, whereas miR-6783-3p was downregulated. Knockdown of linc01559 led to decreased proliferation and migration ability, and increased expression of miR-6783-3p; however, it did not influence the expression of miR-1343-3p. We also found that linc01559 directly interacted with miR-6783-3p. Furthermore, linc01559 and miR-6783-3p regulated the viability of L-OHP-resistant cells following treatment with L-OHP. Conclusion: linc01559 promoted the proliferation of HCC by sponging miR-6783-3p. This suggests that linc01559/miR6783-3p may be key factors in regulating resistance and response to L-OHP. Moreover, they may be potential therapeutic targets for improving sensitivity to L-OHP in patients with HCC.

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