MicroRNA-24 alleviates isoflurane-induced neurotoxicity in rat hippocampus via attenuation of oxidative stress

2020 ◽  
Vol 98 (2) ◽  
pp. 208-218 ◽  
Author(s):  
Na Li ◽  
Linli Yue ◽  
Jun Wang ◽  
Zhenzhen Wan ◽  
Wenhao Bu
Keyword(s):  
Oxidative Stress ◽  
Hippocampal Neurons ◽  
Rat Hippocampus ◽  
Luciferase Reporter ◽  
Loss Of Function ◽  
Luciferase Reporter Gene ◽  
Related Factors ◽  
Proliferation And Apoptosis

Several miRNAs have been recently suggested as potential therapeutic targets for anesthesia-related diseases. This study was carried out to explore the biological roles of miR-24 in isoflurane-treated rat hippocampal neurons. Isoflurane was used to induce neurotoxicity in a rat model. Gain- and loss-of-function of miR-24 was performed, and the size and Ca2+ permeability of mitochondria, as well as cell proliferation and apoptosis, and levels of oxidative-stress-related factors were measured both in vivo and in vitro. Dual luciferase reporter gene assays were used to identify the target relationship between miR-24 and p27kip1. In this study, isoflurane treatment decreased miR-24 expression, after which, levels of neuron apoptosis and oxidative-stress-related factors were elevated and neuron viability was reduced. Over-expression of miR-24 inhibited oxidative damage and neuronal apoptosis in hippocampal tissues, and suppressed the size and Ca2+ permeability of mitochondria of hippocampal neurons. miR-24 enhanced the viability of rat hippocampal neurons by targeting p27kip1. To conclude, this study demonstrated that miR-24 attenuates isoflurane-induced neurotoxicity in rat hippocampus via its antioxidative properties and inhibiting p27kip1 expression.

scholarly journals miR-30e targets GLIPR-2 to modulate diabetic nephropathy: in vitro and in vivo experiments

2017 ◽  
Vol 59 (2) ◽  
pp. 181-190 ◽  
Author(s):  
Dong Zhao ◽  
Jinhua Jia ◽  
Hong Shao
Keyword(s):  
High Glucose ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Luciferase Reporter Gene ◽  
Related Factors ◽  
Qrt Pcr ◽  
In Vivo Experiments ◽  
Renal Tubular

The objectives of this study are to investigate the effect of miR-30e targeting GLIPR-2 on the pathological mechanism of DN. The renal tissues of db/db and db/m mice at different age of weeks were stained with PAS. qRT-PCR was applied to detect the expression of miR-30e and GLIPR-2, not only in the renal tissues of mice but also in the renal tubular epithelial cells (RTECs). By luciferase reporter gene assays, we found the 3′-UTR of the GLIPR-2 mRNA as a direct target of miR-30e. The RTECs cultured in high glucose were divided into blank control, NC, miR-30e mimics, miR-30e inhibitors, miR-30e inhibitor + si-GLIPR-2 and si-GLIPR-2 groups. MTT and flow cytometry were utilized to measure the proliferation and apoptosis of RTECs, while qRT-PCR and Western blot to detect the expression of GLIPR-2- and EMT-related factors. The following results were obtained: In the renal tissues of over 8-week-old db/db mice and the RTECs cultured for 6 h in high glucose, miR-30e was downexpressed while GLIPR-2 was upregulated in a time-dependent manner. Besides, overexpression of miR-30e and si-GLIPR-2 can not only greatly improve the proliferation of RTECs cultured in high glucose, but also downregulate the apoptosis rate of RTECs and the expressions of GLIPR-2, vimentin, α-SMA, Col-I and FN and upregulate E-cadherin. Moreover, si-GLIPR-2 can reverse the proliferation reduction, GLIPR-2 and EMT occurrence caused by the downexpression of miR-30e in RTECs. In conclusion, miR-30e is downregulated in DN, and the overexpression of miR-30e can inhibit GLIPR-2, promote the proliferation of RTECs and inhibit EMT, ultimately avoid leading to renal fibrosis in DN.

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 LncRNA MYLK-AS1 facilitates tumor progression and angiogenesis by targeting miR-424-5p/E2F7 axis and activating VEGFR-2 signaling pathway in hepatocellular carcinoma

2020 ◽  
Author(s):  
Fei Teng ◽  
Ju-Xiang Zhang ◽  
Qi-Meng Chang ◽  
Xu-Bo Wu ◽  
Wei-Guo Tang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Tumor Progression ◽  
Underlying Mechanism ◽  
Luciferase Reporter ◽  
Loss Of Function ◽  
Luciferase Reporter Gene ◽  
Tumor Tissues ◽  
Vegfr2 Signaling

Abstract BackgroundLong non-coding RNA is essential for the metastasis, invasion, angiogenesis and progression of hepatocellular carcinoma (HCC). However, their specific mechanisms are still controversial. Here, we found that Lnc-MYLK-AS1 is a potential oncogene. We systematically analyzed the clinical significance and mechanism of Lnc-MYLK-AS1 in HCC metastasis, invasion and angiogenesis.MethodsDetermine research goals through bioinformatics analysis. The expression of MYLK-AS1 in matched tumor and non-tumor tissues of 156 HCC patients was detected by quantitative reverse transcription PCR. The in vitro and in vivo biological functions of MYLK-AS1 were examined through the loss of function and function gain experiments. The use of dual luciferase reporter gene analysis, quantitative PCR, Western blotting, and fluorescence in situ hybridization (FISH) clarified the underlying mechanism of this competitive endogenous RNA (ceRNA). ResultsMYLK-AS1 is up-regulated in HCC cell lines and tumor tissues, which is related to tumor progression and enhancement of angiogenesis. Overexpression of MYLK-AS1 promotes the proliferation, metastasis, invasion, and angiogenesis of HCC cells, while down-regulation of MYLK-AS1 can reverse these effects in vivo and in vitro. Dual analysis of luciferase and RNA immunoprecipitation showed that microRNA miR-424-5p is the direct target of MYLK-AS1, and MYLK-AS1 acts as ceRNA, which can regulate angiogenesis in HCC. Mechanism studies have shown that miR-424-5p specifically targets E2F transcription factor 7 (E2F7), while the complex MYLK-AS1/miR-424-5p activates VEGFR2 signaling through E2F7, thereby promoting tumor proliferation and angiogenesis.ConclusionThe up-regulation of MYLK-AS1 is related to tumor cell proliferation, increased angiogenesis and poor prognosis in HCC patients. MYLK-AS1 regulates E2F7 expression and VEGFR2 signaling by acting as a ceRNA of miR-424-5p.

scholarly journals Circular RNA FLNA acts as a sponge of miR-486-3p in promoting lung cancer progression via regulating XRCC1 and CYP1A1

2021 ◽  
Author(s):  
Jiongwei Pan ◽  
Gang Huang ◽  
Zhangyong Yin ◽  
Xiaoping Cai ◽  
Enhui Gong ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Cancer Cells ◽  
Cancer Cell ◽  
Cancer Progression ◽  
Lung Cancer Cells ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Related Factors

AbstractSignificantly high-expressed circFLNA has been found in various cancer cell lines, but not in lung cancer. Therefore, this study aimed to explore the role of circFLNA in the progression of lung cancer. The target gene of circFLNA was determined by bioinformatics and luciferase reporter assay. Viability, proliferation, migration, and invasion of the transfected cells were detected by CCK-8, colony formation, wound-healing, and transwell assays, respectively. A mouse subcutaneous xenotransplanted tumor model was established, and the expressions of circFLNA, miR-486-3p, XRCC1, CYP1A1, and related genes in the cancer cells and tissues were detected by RT-qPCR, Western blot, or immunohistochemistry. The current study found that miR-486-3p was low-expressed in lung cancer. MiR-486-3p, which has been found to target XRCC1 and CYP1A1, was regulated by circFLNA. CircFLNA was located in the cytoplasm and had a high expression in lung cancer cells. Cancer cell viability, proliferation, migration, and invasion were promoted by overexpressed circFLNA, XRCC1, and CYP1A1 but inhibited by miR-486-3p mimic and circFLNA knockdown. The weight of the xenotransplanted tumor was increased by circFLNA overexpression yet reduced by miR-486-3p mimic. Furthermore, miR-486-3p mimic reversed the effect of circFLNA overexpression on promoting lung cancer cells and tumors and regulating the expressions of miR-486-3p, XRCC1, CYP1A1, and metastasis/apoptosis/proliferation-related factors. However, overexpressed XRCC1 and CYP1A1 reversed the inhibitory effect of miR-486-3p mimic on cancer cells and tumors. In conclusion, circFLNA acted as a sponge of miR-486-3p to promote the proliferation, migration, and invasion of lung cancer cells in vitro and in vivo by regulating XRCC1 and CYP1A1.

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 LINC00680 enhances hepatocellular carcinoma stemness behavior and chemoresistance by sponging miR-568 to upregulate AKT3

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Gege Shu ◽  
Huizhao Su ◽  
Zhiqian Wang ◽  
Shihui Lai ◽  
Yan Wang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Xenograft Model ◽  
Luciferase Reporter ◽  
Loss Of Function ◽  
Mechanism Study ◽  
Downstream Signaling ◽  
Mouse Xenograft Model ◽  
High Level

Abstract Background Hepatocellular carcinoma (HCC) has an extremely poor prognosis due to the development of chemoresistance, coupled with inherently increased stemness properties. Long non-coding RNAs (LncRNAs) are key regulators for tumor cell stemness and chemosensitivity. Currently the relevance between LINC00680 and tumor progression was still largely unknown, with only one study showing its significance in glioblastoma. The study herein was aimed at identifying the role of LINC00680 in the regulation HCC stemness and chemosensitivity. Methods QRT-PCR was used to detect the expression of LINC00680, miR-568 and AKT3 in tissue specimen and cell lines. Gain- or loss-of function assays were applied to access the function of LINC00680 in HCC cells, including cell proliferation and stemness properties. HCC stemness and chemosensitivity were determined by sphere formation, cell viability and colony formation. Luciferase reporter, RNA immunoprecipitation (RIP), and RNA pull-down assays were performed to examine the interaction between LINC00680 and miR-568 as well as that between miR-568 and AKT3. A nude mouse xenograft model was established for the in vivo study. Results We found that LINC00680 was remarkably upregulated in HCC tissues. Patients with high level of LINC00680 had poorer prognosis. LINC00680 overexpression significantly enhanced HCC cell stemness and decreased in vitro and in vivo chemosensitivity to 5-fluorouracil (5-Fu), whereas LINC00680 knockdown led to opposite results. Mechanism study revealed that LINC00680 regulated HCC stemness and chemosensitivity through sponging miR-568, thereby expediting the expression of AKT3, which further activated its downstream signaling molecules, including mTOR, elF4EBP1, and p70S6K. Conclusion LINC00680 promotes HCC stemness properties and decreases chemosensitivity through sponging miR-568 to activate AKT3, suggesting that LINC00680 might be a potentially important HCC diagnosis marker and therapeutic target.

scholarly journals p53 upregulated by HIF-1α promotes hypoxia-induced G2/M arrest and renal fibrosis in vitro and in vivo

2018 ◽  
Vol 11 (5) ◽  
pp. 371-382 ◽  
Author(s):  
Limin Liu ◽  
Peng Zhang ◽  
Ming Bai ◽  
Lijie He ◽  
Lei Zhang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Transcriptional Activation ◽  
Renal Fibrosis ◽  
Intraperitoneal Administration ◽  
Luciferase Reporter ◽  
Loss Of Function ◽  
P53 Expression ◽  
Tubular Cells

Abstract Hypoxia plays an important role in the genesis and progression of renal fibrosis. The underlying mechanisms, however, have not been sufficiently elucidated. We examined the role of p53 in hypoxia-induced renal fibrosis in cell culture (human and rat renal tubular epithelial cells) and a mouse unilateral ureteral obstruction (UUO) model. Cell cycle of tubular cells was determined by flow cytometry, and the expression of profibrogenic factors was determined by RT-PCR, immunohistochemistry, and western blotting. Chromatin immunoprecipitation and luciferase reporter experiments were performed to explore the effect of HIF-1α on p53 expression. We showed that, in hypoxic tubular cells, p53 upregulation suppressed the expression of CDK1 and cyclins B1 and D1, leading to cell cycle (G2/M) arrest (or delay) and higher expression of TGF-β, CTGF, collagens, and fibronectin. p53 suppression by siRNA or by a specific p53 inhibitor (PIF-α) triggered opposite effects preventing the G2/M arrest and profibrotic changes. In vivo experiments in the UUO model revealed similar antifibrotic results following intraperitoneal administration of PIF-α (2.2 mg/kg). Using gain-of-function, loss-of-function, and luciferase assays, we further identified an HRE3 region on the p53 promoter as the HIF-1α-binding site. The HIF-1α–HRE3 binding resulted in a sharp transcriptional activation of p53. Collectively, we show the presence of a hypoxia-activated, p53-responsive profibrogenic pathway in the kidney. During hypoxia, p53 upregulation induced by HIF-1α suppresses cell cycle progression, leading to the accumulation of G2/M cells, and activates profibrotic TGF-β and CTGF-mediated signaling pathways, causing extracellular matrix production and renal fibrosis.

scholarly journals LncRNA GAPLINC Promotes Renal Cell Cancer Tumorigenesis by Targeting the miR-135b-5p/CSF1 Axis

2021 ◽  
Vol 11 ◽  
Author(s):  
Siyuan Wang ◽  
Xiaorong Yang ◽  
Wenjie Xie ◽  
Shengqiang Fu ◽  
Qiang Chen ◽  
...  
Keyword(s):  
Renal Cell ◽  
Noncoding Rna ◽  
Cell Cancer ◽  
Luciferase Reporter ◽  
Loss Of Function ◽  
Long Intergenic Noncoding Rna ◽  
And Migration ◽  
Molecular Therapeutic

BackgroundLong noncoding RNAs (lncRNAs) are closely related to the occurrence and development of cancer. Gastric adenocarcinoma-associated, positive CD44 regulator, long intergenic noncoding RNA (GAPLINC) is a recently identified lncRNA that can actively participate in the tumorigenesis of various cancers. Here, we investigated the functional roles and mechanism of GAPLINC in renal cell carcinoma (RCC) development.MethodsDifferentially expressed lncRNAs between RCC tissues and normal kidney tissues were detected by using a microarray technique. RNA sequencing was applied to explore the mRNA expression profile changes after GAPLINC silencing. After gain- and loss-of-function approaches were implemented, the effect of GAPLINC on RCC in vitro and in vivo was assessed by cell proliferation and migration assays. Moreover, rescue experiments and luciferase reporter assays were used to study the interactions between GAPLINC, miR-135b-5p and CSF1.ResultsGAPLINC was significantly upregulated in RCC tissues and cell lines and was associated with a poor prognosis in RCC patients. Knockdown of GAPLINC repressed RCC growth in vitro and in vivo, while overexpression of GAPLINC exhibited the opposite effect. Mechanistically, we found that GAPLINC upregulates oncogene CSF1 expression by acting as a sponge of miR-135b-5p.ConclusionTaken together, our results suggest that GAPLINC is a novel prognostic marker and molecular therapeutic target for RCC.

scholarly journals LINC00839 Knockdown Restrains The Metastatic Behaviors of Nasopharyngeal Carcinoma By Sponging miR-454-3p

2021 ◽  
Author(s):  
Feng Ying Zhang ◽  
Xia Li ◽  
Ting Ting Huang ◽  
Mei Ling Xiang ◽  
Lin Lin Sun ◽  
...  
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Underlying Mechanism ◽  
Luciferase Reporter ◽  
Regulatory Function ◽  
Migration And Invasion ◽  
Luciferase Reporter Gene ◽  
Non Coding Rna ◽  
Invasive Capacity

Abstract Background Long intergenic non-coding RNA 00839 (LINC00839) has been verified as a cancer-promoting gene in malignancies. However, the significance of LINC00839 in nasopharyngeal carcinoma (NPC) has yet to be elaborated, as well as its underlying mechanism.Methods LINC00839 and miR-454-3p relative expression levels in NPC cells were examined by qRT-PCR. The growth of cells was examined by CCK-8 and colony formation assays. Cell migration and invasion were examined by wound healing and Transwell experiment, respectively. The binding sequence of LINC00839 and miR-454-3p was confirmed by the luciferase reporter gene experiment. The regulatory function of LINC00839 and miR-454-3p on c-Met was investigated by western blot.Results Here, we revealed that LINC00839 was elevated in NPC. Both LINC00839 knockdown and upregulation of miR-454-3p suppressed NPC cells proliferation, invasive capacity and EMT in vitro. Besides, LINC00839 was validated as a miR-454-3p “sponge”, and upregulation of LINC00839 could reverse miR-454-3p-mediated functions in NPC C666-1 and SUNE-1 cells. Furthermore, c-Met was determined to be targeted by miR-454-3p. Notably, c-Met was downregulated by LINC00839 knockdown through sponging miR-454-3p. In vivo, LINC00839 knockdown resulted in a slower tumor growth.Conclusions Altogether, knockdown of LINC00839 inhibits the aggressive properties of NPC cells via sponging miR-454-3p and regulating c-Met.

scholarly journals Circular RNA CDR1as Inhibits the Metastasis of Gastric Cancer through Targeting miR-876-5p/GNG7 Axis

2021 ◽  
Vol 2021 ◽  
pp. 1-13
Author(s):  
Jiajia Jiang ◽  
Rong Li ◽  
Junyi Wang ◽  
Jie Hou ◽  
Hui Qian ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Epithelial Mesenchymal Transition ◽  
Circular Rna ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Luciferase Reporter Gene ◽  
Mesenchymal Transition ◽  
Underlying Mechanisms

Circular RNA CDR1as has been demonstrated to participate in various cancer progressions as miRNA sponges. The exact underlying mechanisms of CDR1as on gastric cancer (GC) metastasis remain unknown. Here, we found that CDR1as knockdown facilitated GC cell migration and invasion while its overexpression inhibited the migration and invasion abilities of GC cells in vitro and in vivo. Moreover, epithelial-mesenchymal transition- (EMT-) associated proteins and MMP2 and MMP9 were downregulated by CDR1as. Bioinformatics analysis combined with dual-luciferase reporter gene assays, western blot, RT-qPCR analysis, and functional rescue experiments demonstrated that CDR1as served as a miR-876-5p sponge and upregulated the target gene GNG7 expression to suppress GC metastasis. In summary, our findings indicate that CDR1as suppresses GC metastasis through the CDR1as/miR-876-5p/GNG7 axis.

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