Dataset Information

MicroRNA in vivo precipitation identifies miR-151-3p as a computational unpredictable miRNA to target Stat3 and inhibits innate IL-6 production.

ABSTRACT: MicroRNAs (miRNAs) function as important regulators in the immune response and inflammation. Several approaches have been reported to computationally predict miRNAs and their potential targets. However, there are still many miRNA-target interactions that are unpredictable by using the current computational algorithms. We established a miRNA in vivo precipitation method (miRIP) to identify unpredictable miRNAs with definite targets in these cells. Because Stat3 is a well-known transcription factor involved in innate immunity and inflammation, we utilized the miRIP method to identify miRNAs that bind Stat3 mRNA in macrophages. Among the captured miRNAs, miR-151-3p was confirmed to interact with Stat3 mRNA 3'-UTR and downregulate the Stat3 protein levels. LPS stimulation decreased miR-151-3p expression, thereby increasing IL-6 production. Therefore, we found that miR-151-3p inhibited LPS-induced IL-6 production by targeting Stat3. These data further confirmed miRIP as an efficient method to identify unpredictable miRNAs and explore miRNAs-mediated regulation in innate immunity and inflammation.

SUBMITTER: Liu X

PROVIDER: S-EPMC5811685 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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MicroRNA in vivo precipitation identifies miR-151-3p as a computational unpredictable miRNA to target Stat3 and inhibits innate IL-6 production.

Liu Xiang X Su Xiaoping X Xu Sheng S Wang Huamin H Han Dan D Li Jiangxue J Huang Mingyan M Cao Xuetao X

Cellular & molecular immunology 20170911 2

MicroRNAs (miRNAs) function as important regulators in the immune response and inflammation. Several approaches have been reported to computationally predict miRNAs and their potential targets. However, there are still many miRNA-target interactions that are unpredictable by using the current computational algorithms. We established a miRNA in vivo precipitation method (miRIP) to identify unpredictable miRNAs with definite targets in these cells. Because Stat3 is a well-known transcription facto ...[more]

PMID: 28890541

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Project description:Gestational diabetes mellitus (GDM) is characterized by insulin resistance accompanied by low/absent beta-cell compensatory adaptation to the increased insulin demand. Although the molecular mechanisms and factors acting on beta-cell compensatory response during pregnancy have been partially elucidated and reported, those inducing an impaired beta-cell compensation and function, thus evolving in GDM, have yet to be fully addressed. MicroRNAs (miRNAs) are a class of small endogenous non-coding RNAs, which negatively modulate gene expression through their sequence-specific binding to 3'UTR of mRNA target. They have been described as potent modulators of cell survival and proliferation and, furthermore, as orchestrating molecules of beta-cell compensatory response and function in diabetes. Moreover, it has been reported that miRNAs can be actively secreted by cells and found in many biological fluids (e.g., serum/plasma), thus representing both optimal candidate disease biomarkers and mediators of tissues crosstalk(s). Here, we analyzed the expression profiles of circulating miRNAs in plasma samples obtained from n = 21 GDM patients and from n = 10 non-diabetic control pregnant women (24-33 weeks of gestation) using TaqMan array microfluidics cards followed by RT-real-time PCR single assay validation. The results highlighted the upregulation of miR-330-3p in plasma of GDM vs non-diabetics. Furthermore, the analysis of miR-330-3p expression levels revealed a bimodally distributed GDM patients group characterized by high or low circulating miR-330 expression and identified as GDM-miR-330high and GDM-miR-330low. Interestingly, GDM-miR-330high subgroup retained lower levels of insulinemia, inversely correlated to miR-330-3p expression levels, and a significant higher rate of primary cesarean sections. Finally, miR-330-3p target genes analysis revealed major modulators of beta-cell proliferation and of insulin secretion, such as the experimentally validated genes E2F1 and CDC42 as well as AGT2R2, a gene involved in the differentiation of mature beta-cells. In conclusion, we demonstrated that plasma miR-330-3p could be of help in identifying GDM patients with potential worse gestational diabetes outcome; in GDM, miR-330-3p may directly be transferred from plasma to beta-cells thus modulating key target genes involved in proliferation, differentiation, and insulin secretion.

Project description:BackgroundInflammatory bowel disease (IBD), including Crohn's disease and ulcerative colitis, is one of the most frequent gastrointestinal disorders worldwide. Although the actual etiology of IBD remains unclear, growing evidence suggests that CD4+ T cells-associated cytokines, including interferon (IFN)-γ, interleukin (IL)-10 and IL-17A, are crucial for the occurrence of IBD. It has been reported that there is a positive association between miRNAs and IBD development. In this study, we investigated the roles of hsa-miRNA-374b-5p(miRNA-374b-5p) and hsa-miRNA-106a-5p(miRNA-106a-5p) in regulating IBD development.MethodsSerum was obtained from vein blood of IBD patients and healthy controls, qRT-PCR was performed to study the expression of miRNA-374b-5p and miRNA-106a-5p. Furthermore, we investigate the effects of overexpression or inhibition of miRNA-374b-5p on naïve CD4 + T cell subsets differentiation from vein blood of healthy controls by RT-qPCR, flow cytometry and western blot. And more the prediction and confirmation of the targeting genes of miRNA-374b-5p and miRNA-106a-5p were performed by bioinformatics softwares and dual-luciferase reporter assay.ResultsThe results showed that miRNA-106a-5p and miRNA-374b-5p were significantly overexpressed in IBD patients. MiRNA-374b-5p could enhance Th1/Th17 cell differentiation and was related to IBD pathogenesis. MiRNA-374b-5p overexpression induced the mRNA expression of IL-17A and IFN-γ, and suppressed that of IL-10 in T cells. MiRNA-374b-5p inhibition decreased the mRNA expression of IL-17A and IFN-γ, while upregulated that of IL-10 in T cells. These qPCR data were further verified at protein level by western blotting and flow cytometry. In addition, dual-luciferase reporter (DLR) assay indicated that miRNA-374b-5p was directly targeted by IL-10, a key anti-inflammatory cytokine for preventing the occurrence of IBD. Meanwhile, STAT3 was identified as a target gene of miRNA-106a-5p by DLR assays. Further analysis revealed that miRNA-374b-5p regulated JAK1 and STAT3 pathways in CD4+ T cells via IL-10/STAT3 axis. MiRNA-374b-5p overexpression remarkably decreased the mRNA expression and phosphorylated (ser-727) protein levels of STAT3, while miRNA-374b-5p inhibition had the opposite effects.ConclusionMiRNA-374b-5p and miRNA-106a-5p may contribute to IBD development by regulating IL-10/STAT3 signal transduction.

Dataset Information

MicroRNA in vivo precipitation identifies miR-151-3p as a computational unpredictable miRNA to target Stat3 and inhibits innate IL-6 production.

Publications

MicroRNA in vivo precipitation identifies miR-151-3p as a computational unpredictable miRNA to target Stat3 and inhibits innate IL-6 production.

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