Dataset Information

MiR?494?3p regulates lipopolysaccharide?induced inflammatory responses in RAW264.7 cells by targeting PTEN.

ABSTRACT: MicroRNAs (miRNAs/miRs) serve important roles in regulating inflammatory responses at the post?transcriptional level. In the present study, the limma package was used to analyze the GSE43300 array dataset downloaded from the Gene Expression Omnibus database. It was identified that several miRNAs, including miR?494?3p, were upregulated in lipopolysaccharide (LPS)?treated RAW264.7 macrophages compared to control cells. Transfection experiments indicated that overexpressing miR?494?3p inhibited production of LPS?induced proinflammatory cytokines, including interleukin?1? and tumor necrosis factor??. Conversely, knockdown of miR?494?3p enhanced cytokine expression. Bioinformatics prediction and luciferase assay both revealed that miR?494?3p could directly target phosphatase and tensin homolog (PTEN) and upregulate protein kinase B activity. In addition, miR?494?3p mimics suppressed p65 translocation to the nucleus. Similar effects were observed following PTEN silencing. In conclusion, the results of the present study revealed that miR?494?3p may act as an important immune regulator in LPS?stimulated macrophages, and be an effective therapeutic target for treating infections in the future.

SUBMITTER: Zhang S

PROVIDER: S-EPMC6471187 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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miR‑494‑3p regulates lipopolysaccharide‑induced inflammatory responses in RAW264.7 cells by targeting PTEN.

Zhang Si S He Kang K Zhou Weiwei W Cao Jun J Jin Zuolin Z

Molecular medicine reports 20190326 5

MicroRNAs (miRNAs/miRs) serve important roles in regulating inflammatory responses at the post‑transcriptional level. In the present study, the limma package was used to analyze the GSE43300 array dataset downloaded from the Gene Expression Omnibus database. It was identified that several miRNAs, including miR‑494‑3p, were upregulated in lipopolysaccharide (LPS)‑treated RAW264.7 macrophages compared to control cells. Transfection experiments indicated that overexpressing miR‑494‑3p inhibited pro ...[more]

PMID: 30942409

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Project description:Gestational diabetes mellitus (GDM) is one of the most common pregnancy complications characterized by insulin resistance and pancreatic ?-cell dysfunction. Increasing evidence suggests that microRNAs (miRNAs) play key roles in the diverse types of diabetes, including GDM. However, the underlying mechanisms remain largely unknown. The purpose of this study is to investigate the role of microRNAs in GDM. The microarray data of dysregulated miRNAs in blood and placenta was retrieved in the GEO dataset under the accession number GSE19649. Quantitative reverse transcription PCR (qRT-PCR) was performed to analyze the expression levels of miR-494 in peripheral blood from twenty pairs of gestational diabetes (GDM) women and healthy women. Then, we investigated the effects of miR-494 on the insulin secretion of pancreatic ?-cells. Moreover, the role of this miR-494 in regulating the proliferation and apoptosis of pancreatic ?-cells were determined by MTT assay and flow cytometry, respectively in INS1 cells transfected with a miR-494 mimic or inhibitor. In addition, the direct target of miR-494 was confirmed using 3' untranslated region (UTR) luciferase reporter assay. Our data demonstrated that the miR-494 level was significantly decreased in the blood of GDM patients, and the low level was associated with a high concentration of blood glucose. Furthermore, overexpression of miR-494 improved pancreatic ?-cell dysfunction by enhancing insulin secretion and total insulin content, inducing cell proliferation, and inhibiting cell apoptosis, whereas miR-494 knockdown exhibited decreased insulin secretion and proliferation, as well as stimulated apoptosis. In addition, phosphatase and tensin homolog (PTEN) which has been shown to play a pivotal role in apoptosis, was proved to be a direct target of miR-494 in pancreatic ?-cells. More importantly, siRNA-induced downregulation of PTEN reversed the effects of miR-494 knockdown on insulin secretion, cell proliferation, and apoptosis of pancreatic ?-cells.

Project description:The present study aimed to investigate the effects of microRNA (miR)?21 on osteoclastogenesis and its underlying molecular mechanisms. The expression levels of tartrate?resistant acid phosphatase (TRAP) and miR?21 were detected during osteoclastogenesis in receptor activator of NF??B ligand (RANKL)?induced RAW264.7 cells via reverse transcription?quantitative PCR. Bioinformatics and dual luciferase reporter assays were performed to analyze the association between miR?21 and Pten. RANKL?induced RAW264.7 cells were divided into the following groups: MiR?negative control (NC), miR?21 mimic, miR?21 inhibitor and miR?21 mimic + LY294002. The effects of miR?21 on osteoclastogenesis and bone resorption were then detected using TRAP staining and a bone resorption assay. Pten, phosphorylated?Akt and nuclear factor of activated T cell (NFATc1) expression levels were measured by western blotting to analyze the effects of miR?21 on the PI3K/Akt signaling pathway. The present study revealed that miR?21 was upregulated during osteoclastogenesis in RANKL?induced RAW264.7 cells. Furthermore, miR?21 negatively regulated Pten. Compared with the miR?negative control (NC) group, the number of osteoclasts and the percentage of bone resorption were increased in the miR?21 mimic group, whereas they were decreased in the miR?21 inhibitor group. The number of osteoclasts and the percentage of bone resorption in the miR?21 mimic + LY294002 group were lower than in the miR?21 mimic group. Compared with the miR?NC group, the protein expression levels of Pten were decreased, whereas p?Akt and NFATc1 were increased in the miR?21 mimic group. Conversely, Pten protein expression was increased, whereas p?Akt and NFATc1 were decreased in the miR?21 inhibitor group. In the miR?21 mimic + LY294002 group, Pten protein expression was higher, and p?Akt and NFATc1 were lower than in the miR?21 mimic group. In conclusion, miR?21 is upregulated during osteoclastogenesis, and may promote osteoclastogenesis and bone resorption through activating the PI3K/Akt signaling pathway via targeting Pten.

Dataset Information

MiR?494?3p regulates lipopolysaccharide?induced inflammatory responses in RAW264.7 cells by targeting PTEN.

Publications

miR‑494‑3p regulates lipopolysaccharide‑induced inflammatory responses in RAW264.7 cells by targeting PTEN.

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