miR-203a is involved in HBx-induced inflammation by targeting Rap1a
miR-203a is involved in HBx-induced inflammation by targeting Rap1a
复制标题
miR-203a 通过靶向 Rap1a 参与 HBx 诱导的炎症。
DOI：
10.1016/j.yexcr.2016.10.016
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发表时间：
2016-11-15
影响因子：
3.7
通讯作者：
Zhang, JinPing
中科院分区：
文献类型：
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作者：
Wu, AiRong;Chen, Huo;Zhang, JinPing
B型肝炎病毒（HBV）引起急性和慢性肝炎，并且是肝硬化和肝细胞癌的主要原因之一。越来越多的证据表明，炎症是肝硬化和肝癌发生的关键因素。microRNA在许多生物过程中发挥着重要作用。本研究旨在探讨microRNA在HBX诱导的炎症反应中的作用。首先，微阵列实验显示，与HBV肝样品相比，HBV+肝样品表达更高水平的miR-203 a。为了验证这些改变，将HBx编码质粒转染到HepG 2细胞中以过表达HBx蛋白。Real-time PCR结果提示HBx的过表达可诱导miR-203 a的表达上调。为了确定上调miR-203 a如何诱导肝细胞炎症，我们在HepG 2细胞中过表达miR-203 a。Annexin V染色和BrdU染色结果显示，miR-203 a过表达可显著促进细胞凋亡和增殖，同时可导致G 0/G1期细胞减少，G2/M期细胞增加。在miR-203 a过表达的HepG 2细胞中，包括IL-6和11：8在内的一些细胞因子的产生显著增加，但TGF β和IFN γ降低。荧光素酶报告基因实验、蛋白质质谱实验和实时荧光定量PCR实验均证明Rap 1a是miR-203 a的靶基因。进一步的实验表明这些改变是通过PI 3 K/ERK/p38/NF κ B B途径调节的。提示HBV感染可上调miR-203 a的表达，进而下调Rap 1 a的表达，影响PI 3 K/ERK/p38/NF kappa B通路，最终导致肝炎炎症的发生。
Hepatitis B virus (HBV) causes acute and chronic hepatitis, and is one of the major causes of cirrhosis and hepatocellular carcinoma. Accumulating evidence suggests that inflammation is the key factor for liver cirrhosis and hepatocellular carcinoma. MicroRNAs play important roles in many biological processes. Here, we aim to explore the function of microRNAs in the HBX-induced inflammation. First, microarray experiment showed that HBV+ liver samples expressed higher level of miR-203a compared to HBV liver samples. To verify these alterations, HBx-coding plasmid was transfected into HepG2 cells to overexpress HBx protein. The real-time PCR results suggested that over-expression of HBx could induce up-regulation of miR-203a. To define how up regulation of miR-203a can induce liver cells inflammation, we over-expressed miR-203a in HepG2 cells. Annexin V staining and BrdU staining suggested that overexpression of miR-203a significantly increased the cell apoptosis and proliferation, meanwhile, over-expression of miR-203a could lead to a decrease in G0/G1 phase cells and an increase in G2/M phase cells. Some cytokines production including IL-6 and 11:8 were significantly increased, but TGF beta and IFN gamma were decreased in miR-203a over-expressed HepG2 cells. Luciferase reporter assay experiments, protein mass-spectrum assay and real-time PCR all together demonstrated that Rap1a was the target gene of miR-203a. Further experiments showed that these alterations were modulated through PI3K/ERK/p38/NF kappa B pathways. These data suggested that HBV-infection could up-regulate the expression of miR-203a, thus down regulated the expression of Rap1a and affected the PI3K/ERK/p38/NF kappa B pathways, finally induced the hepatitis inflammation.