miR-374b-5p is increased in deep vein thrombosis and negatively targets IL-10
miR-374b-5p is increased in deep vein thrombosis and negatively targets IL-10
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miR-374b-5p 在深静脉血栓形成中增加并负向靶向 IL-10
DOI：
10.1016/j.yjmcc.2020.05.011
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发表时间：
2020-07-01
影响因子：
5
通讯作者：
Li, Xia
中科院分区：
文献类型：
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作者：
Zhang, Yunhong;Miao, Xiuming;Li, Xia
背景资料：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，白细胞介素-10（IL-10）的降低参与了DVT的发生。然而，其潜在的分子机制在很大程度上仍然未知。在此，我们提出IL-10在转录后水平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：采用miRNA芯片分析DVT中miRNA的表达。通过计算机靶标预测工具预测IL-10的上游miRNA调节剂。采用实时荧光定量PCR（qRT-PCR）检测IL-10 mRNA和miR-374 b-5 p的表达，采用酶联免疫法检测IL-10蛋白的表达。双荧光素酶报告基因测定用于鉴定miR-374 b-5 p与IL 10之间的相互作用。建立DVT小鼠模型，并通过荧光原位杂交在体外观察miR-374 b-5 p的定位。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-374 b-5 p水平显著升高，且miR-374 b-5 p与IL-10呈显著负相关。体外实验表明，过表达的miR-374 b-5 p降低IL-10的表达，而miR-374 b-5 p敲低则增加IL-10的表达。此外，体内研究显示，用抗IL-10抗体或agomiR-374 b-5 p递送的DVT小鼠导致IL-10表达降低并加重DVT形成，而agomiR-374 b-5 p起相反作用。结论：miR-374 b-5 p表达上调可能通过下调IL-10的表达促进DVT的形成。miR-374 b-5 p有望成为DVT的诊断标志物和治疗靶点。
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.