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miR-374b-5p is increased in deep vein thrombosis and negatively targets IL-10.
Journal of Molecular and Cellular Cardiology ( IF 5 ) Pub Date : 2020-05-21 , DOI: 10.1016/j.yjmcc.2020.05.011
Yunhong Zhang 1 , Xiuming Miao 2 , Zhen Zhang 3 , Ran Wei 3 , Shangwen Sun 4 , Gang Liang 2 , Huihan Li 2 , Chu Chu 1 , Lin Zhao 3 , Xiaoxiao Zhu 3 , Qiang Guo 3 , Bin Wang 2 , Xia Li 3
Affiliation  

BACKGROUND Deep venous thrombosis (DVT) is one of the most common venous thromboembolic (VTE) disorders and the third leading cardiovascular complication. Accumulating evidence has shown that decreased interleukin-10 (IL-10) was involved in DVT. However, the underlying molecular mechanisms are still largely unknown. Here, we proposed that the epigenetic modification of IL-10 at the post-transcriptional level may be a crucial trigger for IL-10 down-regulation in DVT. METHODS miRNA expression in DVT was profiled by miRNA microarray analysis. The upstream miRNA regulators of IL-10 were predicted by in silico target prediction tools. The expression of IL-10 mRNA and miR-374b-5p were examined by quantitative real-time PCR (qRT-PCR) and the protein expression of IL-10 was detected by enzyme-linked immunoassay. Dual luciferase reporter assay was used to identify the interaction between miR-374b-5p and IL10. A murine model of DVT was developed and the localization of miR-374b-5p was visualized in vitro by fluorescence in situ hybridization. The biological effects of miR-374b-5p on IL-10 was examined both in vitro and in vivo. RESULTS Microarray and qRT-PCR results showed that the IL-10 expression was decreased while miR-374b-5p level was increased substantially in peripheral blood mononuclear cells of DVT patients, and there was significant negative correlation between miR-374b-5p and IL-10. Experiments in vitro showed that overexpressed miR-374b-5p reduced IL-10 expression, while miR-374b-5p knockdown increased IL-10 expression. Moreover, in vivo studies revealed that DVT mice with anti-IL-10 antibody or agomiR-374b-5p delivery resulted in decreased IL-10 expression and aggravated DVT formation, whereas antagomiR-374b-5p acted inversely. Dual luciferase reporter assay identified direct binding between miR-374b-5p and IL10. CONCLUSIONS These findings suggest that increased miR-374b-5p promotes DVT formation by downregulating IL-10 expression. miR-374b-5p may be explored as a promising diagnostic marker and therapeutic target for DVT.

中文翻译:

miR-374b-5p在深静脉血栓形成中增加,并负向IL-10。

背景技术深静脉血栓形成(DVT)是最常见的静脉血栓栓塞(VTE)疾病之一,也是第三大心血管并发症。越来越多的证据表明,DVT与白细胞介素10(IL-10)降低有关。但是,基本的分子机制仍是未知的。在这里,我们提出在转录后水平上IL-10的表观遗传修饰可能是DVT中IL-10下调的关键触发因素。方法通过miRNA微阵列分析分析DVT中miRNA的表达。通过计算机靶标预测工具预测了IL-10的上游miRNA调节剂。通过定量实时PCR(qRT-PCR)检测IL-10 mRNA和miR-374b-5p的表达,并通过酶联免疫法检测IL-10的蛋白表达。使用双重萤光素酶报告基因分析鉴定miR-374b-5p与IL10之间的相互作用。建立了DVT的小鼠模型,并通过荧光原位杂交在体外可视化了miR-374b-5p的定位。在体外和体内均检测了miR-374b-5p对IL-10的生物学作用。结果微阵列和qRT-PCR结果显示DVT患者外周血单个核细胞中IL-10表达降低而miR-374b-5p水平显着升高,且miR-374b-5p与IL-T呈显着负相关。 10。体外实验表明,过表达的miR-374b-5p降低了IL-10的表达,而miR-374b-5p的降低则提高了IL-10的表达。此外,体内研究表明,具有抗IL-10抗体或agomiR-374b-5p递送的DVT小鼠导致IL-10表达降低并加重了DVT的形成,而antagomiR-374b-5p则起相反的作用。双重荧光素酶报告基因分析鉴定了miR-374b-5p和IL10之间的直接结合。结论这些发现表明,增加的miR-374b-5p通过下调IL-10表达来促进DVT的形成。miR-374b-5p可作为DVT的有前途的诊断标记和治疗靶点进行探索。
更新日期:2020-05-21
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