肝癌进展中乙肝病毒x蛋白对miR-338-3p转录抑制调控的作用及其机制研究

Recurrence and metastasis is the leading cause of the poor prognosis in hepatocelleluar cancer(HCC). We've reported that the downregulation of the miR-338-3p in HCC, its high expression can inhibit or reverse the invasion of the liver cancer cell, but its downrgulation mechanism is unknown. We've found that: The HBV X protein (HBx) can Inhibition the expression of miR-338-3p, which may be related to the transcription factor ZNF263. We speculate that the HBx could inhibit the transcription of the miR-338-3p by the ZNF263 promoting the progression of liver cancer which has been validated in cells and preliminary clinical specimens. To draw on this, this project first intends to identify the HBx's in vivo and vitro effect on miR-338-3p and progression in liver cancer; then determine ZNF263's binding sites of the miR-338-3p promoter; illustrate the mechanism that the ZNF263 do mediated the regulate process of HBx to miR -338-3p; finally validate the HBx/ZNF263, miR-338-3p and their effect on progression and prognosis in liver cancer though the animals and the clinical specimen. This project aims to get the reliable evidence that HBx could downregulate miR-338-3p by ZNF263 promoting the development of liver cancer. Establish reliable experimental basis for the miR-338-3p as a new target in liver cancer prevention.

转移复发是肝癌预后差的主要原因。我们已报道miR-338-3p在肝癌中下调，其高表达可抑制或逆转肝癌细胞侵袭，但miR-338-3p下调机制不明。我们发现：乙肝X蛋白（HBx）可抑制miR-338-3p表达，其机制可能与转录因子ZNF263有关。我们推测，HBx通过ZNF263抑制miR-338-3p转录促进肝癌进展，并已在细胞和临床标本中获得初步验证。为此,本项目拟首先通过体内外实验明确HBx对miR-338-3p在肝癌进展中的作用；再确定ZNF263直接调控miR-338-3p启动子及结合位点；阐明ZNF263介导了HBx对miR-338-3p的调控；最后在动物和临床组织中验证HBx/ZNF263与miR-338-3p及肝癌进展、预后的关系。本项目旨在获得HBx通过ZNF263下调miR-338-3p促进肝癌进展的可靠证据，为确立miR-338-3p为肝癌防治新靶点提供可靠的实验依据。

研究表明乙肝病毒x蛋白（HBx）可调控miRNA表达，通过与miRNA的共同作用促进肝癌的发生发展。本课题组前期已报道miR-338-3p在肝癌中下调，其低表达可抑制或逆转肝癌细胞侵袭，但miR-338-3p下调机制不明。HBx 是否是调控miR-338-3p的表达促进肝癌的侵袭转移也不清楚。通过本项目的研究，我们发现通过对检测肝癌组织及肝癌细胞中HBx和miR-338-3p表达水平，并结合临床资料进行相关性分析。分别转染或共转染HBx、HBx siRNA及其相应阴性对照后，观察肝癌细胞中miR-338-3p表达量的变化；结合免疫沉淀等方法进一步明确HBx调控miR-338-3p具体机制。我们的研究结果发现HBx在肝癌组织和肝癌细胞中表达显著上调，其高表达与miR-338-3p低表达密切正相关。本研究证实肝癌细胞中高表达HBx可抑制miR -338-3p的表达。通过免疫沉淀等相关实验证实HBx可以靶向作用于ZNF263调控miR-338-3p的表达。本项目的结果不仅揭示HBx可通过ZNF263抑制miR-338-3p的表达，而且将为确定miR-338-3p作为抗乙肝-肝癌细胞的治疗新靶点提供科学依据和理论基础。